Robert West

All Publications

• MYB-activated models for testing therapeutic agents in adenoid cystic carcinoma. Oral oncology Jiang, Y., Gao, R., Cao, C., Forbes, L., Li, J., Freeberg, S., Fredenburg, K. M., Justice, J. M., Silver, N. L., Wu, L., Varma, S., West, R., Licht, J. D., Zajac-Kaye, M., Kentsis, A., Kaye, F. J. 2019; 98: 147?55

  Abstract

  OBJECTIVE: There are no effective systemic therapies for adenoid cystic cancer (ACC) and lack of tumor lines and mouse models have hindered drug development.We aim to develop MYB-activated models for testing new therapeutic agents.MATERIALS AND METHODS: We studied new ACC patient-derived xenograft (PDX) models and generated a matched cell line from one patient. In addition, we generated a genetically-engineered MYB-NFIB mouse model (GEMM) that was crossed with Ink4a+/-/Arf+/- mice to study tumor spectrum and obtain tumor lines. Using human and murine ACC-like tumor lines, we analyzed MYB expression by RNA-Seq and immunoblot and tested efficacy of new MYB inhibitors.RESULTS: We detected MYB-NFIB transcripts in both UFH1 and UFH2 PDX and observed tumor inhibition by MYB depletion using shRNA in vivo. We observed rapid loss of MYB expression when we cultured UFH1 in vitro, but were able to generate a UFH2 tumor cell line that retained MYB expression for 6?months. RNA-Seq expression detected an ACC-like mRNA signature in PDX samples and we confirmed an identical KMT2A/MLL variant in UFH2 PDX, matched cell line, and primary biopsy. Although the predominant phenotype of the MYB-NFIB GEMM was B-cell leukemia, we also generated a MYB-activated ACC-like mammary tumor cell line. We observed tumor inhibition using a novel MYB peptidomimetic in both human and murine tumor models.CONCLUSIONS: We generated and studied new murine and human MYB-activated tumor samples and detected growth inhibition with MYB peptidomimetics. These data provide tools to define treatment strategies for patients with advanced MYB-activated ACC.

  View details for DOI 10.1016/j.oraloncology.2019.09.005

  View details for PubMedID 31606723

• MIBI-TOF: A multiplexed imaging platform relates cellular phenotypes and tissue structure. Science advances Keren, L., Bosse, M., Thompson, S., Risom, T., Vijayaragavan, K., McCaffrey, E., Marquez, D., Angoshtari, R., Greenwald, N. F., Fienberg, H., Wang, J., Kambham, N., Kirkwood, D., Nolan, G., Montine, T. J., Galli, S. J., West, R., Bendall, S. C., Angelo, M. 2019; 5 (10): eaax5851

  Abstract

  Understanding tissue structure and function requires tools that quantify the expression of multiple proteins while preserving spatial information. Here, we describe MIBI-TOF (multiplexed ion beam imaging by time of flight), an instrument that uses bright ion sources and orthogonal time-of-flight mass spectrometry to image metal-tagged antibodies at subcellular resolution in clinical tissue sections. We demonstrate quantitative, full periodic table coverage across a five-log dynamic range, imaging 36 labeled antibodies simultaneously with histochemical stains and endogenous elements. We image fields of view up to 800 mum * 800 mum at resolutions down to 260 nm with sensitivities approaching single-molecule detection. We leverage these properties to interrogate intrapatient heterogeneity in tumor organization in triple-negative breast cancer, revealing regional variability in tumor cell phenotypes in contrast to a structured immune response. Given its versatility and sample back-compatibility, MIBI-TOF is positioned to leverage existing annotated, archival tissue cohorts to explore emerging questions in cancer, immunology, and neurobiology.

  View details for DOI 10.1126/sciadv.aax5851

  View details for PubMedID 31633026

• Gene-expression profiling of single cells from archival tissue with laser-capture microdissection and Smart-3SEQ. Genome research Foley, J. W., Zhu, C., Jolivet, P., Zhu, S. X., Lu, P., Meaney, M. J., West, R. B. 2019

  Abstract

  RNA sequencing (RNA-seq) is a sensitive and accurate method for quantifying gene expression. Small samples or those whose RNA is degraded, such as formalin-fixed, paraffin-embedded (FFPE) tissue, remain challenging to study with nonspecialized RNA-seq protocols. Here we present a new method, Smart-3SEQ, that accurately quantifies transcript abundance even with small amounts of total RNA and effectively characterizes small samples extracted by laser-capture microdissection (LCM) from FFPE tissue. We also obtain distinct biological profiles from FFPE single cells, which have been impossible to study with previous RNA-seq protocols, and we use these data to identify possible new macrophage phenotypes associated with the tumor microenvironment. We propose Smart-3SEQ as a highly cost-effective method to enable large gene-expression profiling experiments unconstrained by sample size and tissue availability. In particular, Smart-3SEQ's compatibility with FFPE tissue unlocks an enormous number of archived clinical samples, and combined with LCM it allows unprecedented studies of small cell populations and single cells isolated by their in situ context.

  View details for DOI 10.1101/gr.234807.118

  View details for PubMedID 31519740

• Race and risk of subsequent aggressive breast cancer following ductal carcinoma in situ. Cancer Liu, Y., West, R., Weber, J. D., Colditz, G. A. 2019

  Abstract

  BACKGROUND: General populations of black women have a higher risk of developing breast cancer negative for both estrogen receptor (ER) and progesterone receptor (PR) in comparison with white counterparts. Racial differences remain unknown in the risk of developing aggressive invasive breast cancer (IBC) that is characterized by negativity for both ER and PR (ER-PR-) or higher 21-gene recurrence scores after ductal carcinoma in situ (DCIS).METHODS: This study identified 163,892 women (10.5% black, 9.8% Asian, and 8.6% Hispanic) with incident DCIS between 1990 and 2015 from the Surveillance, Epidemiology, and End Results data sets. Cox proportional hazards regression was used to estimate hazards ratios (HRs) of subsequent IBC classified by the hormone receptor status and 21-gene recurrence scores.RESULTS: During a median follow-up of 90months, 8333 women developed IBC. In comparison with white women, the adjusted HR of subsequent ER-PR- breast cancer was 1.86 (95% confidence interval [CI], 1.57-2.20) for black women (absolute 10-year difference, 2.2%) and 1.40 (95% CI, 1.14-1.71) for Asian women (absolute 10-year difference, 0.4%); this was stronger than the associations for ER+ and/or PR+ subtypes (Pheterogeneity =.0004). The 21-gene recurrence scores of subsequent early-stage, ER+ IBCs varied by race/ethnicity (Pheterogeneity =.057); black women were more likely than white women to have a recurrence score of 26 or higher (HR, 1.38; 95% CI, 1.00-1.92). No significant difference was observed in the risks of subsequent IBC subtypes for Hispanic women.CONCLUSIONS: Black and Asian women with DCIS had higher risks of developing biologically aggressive IBC than white counterparts. This should be considered in treatment decisions for black and Asian patients with DCIS.

  View details for DOI 10.1002/cncr.32200

  View details for PubMedID 31120565

• Increased Galectin-1 Expression in Thymic Epithelial Tumors CLINICAL LUNG CANCER Riess, J. W., Kong, C. S., West, R. B., Padda, S. K., Neal, J. W., Wakelee, H. A., Le, Q. 2019; 20 (3): E356?E361

  View details for DOI 10.1016/j.cllc.2018.12.005

  View details for Web of Science ID 000467981200020

• YAP-independent mechanotransduction drives breast cancer progression NATURE COMMUNICATIONS Lee, J. Y., Chang, J. K., Dominguez, A. A., Lee, H., Nam, S., Chang, J., Varma, S., Qi, L. S., West, R. B., Chaudhuri, O. 2019; 10

  View details for DOI 10.1038/s41467-019-09755-0

  View details for Web of Science ID 000465200300008

• The HTN3-MSANTD3 Fusion Gene Defines a Subset of Acinic Cell Carcinoma of the Salivary Gland AMERICAN JOURNAL OF SURGICAL PATHOLOGY Andreasen, S., Varma, S., Barasch, N., Thompson, L. R., Miettinen, M., Rooper, L., Stelow, E. B., Agander, T. K., Seethala, R. R., Chiosea, S. I., Homoe, P., Wessel, I., Larsen, S. R., Erentaite, D., Bishop, J. A., Ulhoi, B. P., Kiss, K., Melchior, L. C., Pollack, J. R., West, R. B. 2019; 43 (4): 489?96

  View details for DOI 10.1097/PAS.0000000000001200

  View details for Web of Science ID 000462180400007

• Systematic Analysis of GATA3 Expression in 530 Primary Mesenchymal Neoplasms Wei, C., Charville, G., Bean, G., Troxell, M., Allison, K., Van de Rijn, M., West, R. NATURE PUBLISHING GROUP. 2019

  View details for Web of Science ID 000478915500094

• Systematic Analysis of GATA3 Expression in 530 Primary Mesenchymal Neoplasms Wei, C., Charville, G., Bean, G., Troxell, M., Allison, K., Van de Rijn, M., West, R. NATURE PUBLISHING GROUP. 2019

  View details for Web of Science ID 000478081100094

• Most canine ameloblastomas harbor HRAS mutations, providing a novel large-animal model of RAS-driven cancer. Oncogenesis Saffari, P. S., Vapniarsky, N., Pollack, A. S., Gong, X., Vennam, S., Pollack, A. J., Verstraete, F. J., West, R. B., Arzi, B., Pollack, J. R. 2019; 8 (2): 11

  Abstract

  Canine acanthomatous ameloblastomas (CAA), analogs of human ameloblastoma, are oral tumors of odontogenic origin for which the genetic drivers have remained undefined. By whole-exome sequencing, we have now discovered recurrent HRAS and BRAF activating mutations, respectively, in 63% and 8% of CAA. Notably, cell lines derived from CAA with HRAS mutation exhibit marked sensitivity to MAP kinase (MAPK) pathway inhibitors, which constrain cell proliferation and drive ameloblast differentiation. Our findings newly identify a large-animal spontaneous cancer model to study the progression and treatment of RAS-driven cancer. More broadly, our study highlights the translational potential of canine cancer genome sequencing to benefit both humans and their companion animals.

  View details for PubMedID 30741938

• ERBB2 copy number analysis of invasive breast carcinoma using digital droplet PCR and targeted next-generation sequencing: A focus on 'non-classical' HER2 FISH groups using the 2018 ASCO/CAP HER2 testing guideline Yang, S., Bouhlal, Y., De La Vega, F. M., Ballard, M., West, R. B., Sibley, R. K., Kuo, C. J., Vilborg, A., Allison, K. H. AMER ASSOC CANCER RESEARCH. 2019

  View details for DOI 10.1158/1538-7445.SABCS18-P3-10-12

  View details for Web of Science ID 000478677001200

• Development of a machine learning-based classifier for Oncotype DX (R) category prediction in a population of lymph node positive breast carcinoma patients Beca, F., Yang, S., Gruber, J. G., Barry-Holson, K., West, R., Wen, H. Y., Allison, K. H. AMER ASSOC CANCER RESEARCH. 2019

  View details for DOI 10.1158/1538-7445.SABCS18-P2-07-07

  View details for Web of Science ID 000478677000416

• Spatial integration of radiology and pathology images to characterize breast cancer aggressiveness on pre-surgical MRI Rusu, M., Daniel, B., West, R., Angelini, E. D., Landman, B. A. SPIE-INT SOC OPTICAL ENGINEERING. 2019

  View details for DOI 10.1117/12.2512670

  View details for Web of Science ID 000483012700032

• Framework for the co-registration of MRI and Histology Images in Prostate Cancer Patients with Radical Prostatectomy Rusu, M., Kunder, C., Fan, R., Ghanouni, P., West, R., Sonn, G., Brooks, J., Angelini, E. D., Landman, B. A. SPIE-INT SOC OPTICAL ENGINEERING. 2019

  View details for DOI 10.1117/12.2513099

  View details for Web of Science ID 000483012700057

• Cell cycle progression in confining microenvironments is regulated by a growth-responsive TRPV4-PI3K/Akt-p27Kip1 signaling axis. Science advances Nam, S., Gupta, V. K., Lee, H. P., Lee, J. Y., Wisdom, K. M., Varma, S., Flaum, E. M., Davis, C., West, R. B., Chaudhuri, O. 2019; 5 (8): eaaw6171

  Abstract

  In tissues, cells reside in confining microenvironments, which may mechanically restrict the ability of a cell to double in size as it prepares to divide. How confinement affects cell cycle progression remains unclear. We show that cells progressed through the cell cycle and proliferated when cultured in hydrogels exhibiting fast stress relaxation but were mostly arrested in the G0/G1 phase of the cell cycle when cultured in hydrogels that exhibit slow stress relaxation. In fast-relaxing gels, activity of stretch-activated channels (SACs), including TRPV4, promotes activation of the phosphatidylinositol 3-kinase (PI3K)/Akt pathway, which in turn drives cytoplasmic localization of the cell cycle inhibitor p27Kip1, thereby allowing S phase entry and proliferation. Cell growth during G1 activated the TRPV4-PI3K/Akt-p27Kip1 signaling axis, but growth is inhibited in the confining slow-relaxing hydrogels. Thus, in confining microenvironments, cells sense when growth is sufficient for division to proceed through a growth-responsive signaling axis mediated by SACs.

  View details for DOI 10.1126/sciadv.aaw6171

  View details for PubMedID 31457089

  View details for PubMedCentralID PMC6685709

• Genomic landscape of ductal carcinoma in situ and association with progression. Breast cancer research and treatment Lin, C. Y., Vennam, S., Purington, N., Lin, E., Varma, S., Han, S., Desa, M., Seto, T., Wang, N. J., Stehr, H., Troxell, M. L., Kurian, A. W., West, R. B. 2019

  Abstract

  The detection rate of breast ductal carcinoma in situ (DCIS) has increased significantly, raising the concern that DCIS is overdiagnosed and overtreated. Therefore, there is an unmet clinical need to better predict the risk of progression among DCIS patients. Our hypothesis is that by combining molecular signatures with clinicopathologic features, we can elucidate the biology of breast cancer progression, and risk-stratify patients with DCIS.Targeted exon sequencing with a custom panel of 223 genes/regions was performed for 125 DCIS cases. Among them, 60 were from cases having concurrent or subsequent invasive breast cancer (IBC) (DCIS?+?IBC group), and 65 from cases with no IBC development over a median follow-up of 13 years (DCIS-only group). Copy number alterations in chromosome 1q32, 8q24, and 11q13 were analyzed using fluorescence in situ hybridization (FISH). Multivariable logistic regression models were fit to the outcome of DCIS progression to IBC as functions of demographic and clinical features.We observed recurrent variants of known IBC-related mutations, and the most commonly mutated genes in DCIS were PIK3CA (34.4%) and TP53 (18.4%). There was an inverse association between PIK3CA kinase domain mutations and progression (Odds Ratio [OR] 10.2, p?

  View details for DOI 10.1007/s10549-019-05401-x

  View details for PubMedID 31420779

• Clonal replacement and heterogeneity in breast tumors treated with neoadjuvant HER2-targeted therapy. Nature communications Caswell-Jin, J. L., McNamara, K., Reiter, J. G., Sun, R., Hu, Z., Ma, Z., Ding, J., Suarez, C. J., Tilk, S., Raghavendra, A., Forte, V., Chin, S. F., Bardwell, H., Provenzano, E., Caldas, C., Lang, J., West, R., Tripathy, D., Press, M. F., Curtis, C. 2019; 10 (1): 657

  Abstract

  Genomic changes observed across treatment may result from either clonal evolution or geographically disparate sampling of heterogeneous tumors. Here we use computational modeling based on analysis of fifteen primary breast tumors and find that apparent clonal change between two tumor samples can frequently be explained by pre-treatment heterogeneity, such that at least two regions are necessary to detect treatment-induced clonal shifts. To assess for clonal replacement, we devise a summary statistic based on whole-exome sequencing of a pre-treatment biopsy and multi-region sampling of the post-treatment surgical specimen and apply this measure to five breast tumors treated with neoadjuvant HER2-targeted therapy. Two tumors underwent clonal replacement with treatment, and mathematical modeling indicates these two tumors had resistant subclones prior to treatment and rates of resistance-related genomic changes that were substantially larger than previous estimates. Our results provide a needed framework to incorporate primary tumor heterogeneity in investigating the evolution of resistance.

  View details for PubMedID 30737380

• YAP-independent mechanotransduction drives breast cancer progression. Nature communications Lee, J. Y., Chang, J. K., Dominguez, A. A., Lee, H. P., Nam, S., Chang, J., Varma, S., Qi, L. S., West, R. B., Chaudhuri, O. 2019; 10 (1): 1848

  Abstract

  Increased tissue stiffness is a driver of breast cancer progression. The transcriptional regulator YAP is considered a universal mechanotransducer, based largely on 2D culture studies. However, the role of YAP during in vivo breast cancer remains unclear. Here, we find that mechanotransduction occurs independently of YAP in breast cancer patient samples and mechanically tunable 3D cultures. Mechanistically, the lack of YAP activity in 3D culture and in vivo is associated with the absence of stress fibers and an order of magnitude decrease in nuclear cross-sectional area relative to 2D culture. This work highlights the context-dependent role of YAP in mechanotransduction, and establishes that YAP does not mediate mechanotransduction in breast cancer.

  View details for PubMedID 31015465

• Publisher Correction: Clonal replacement and heterogeneity in breast tumors treated with neoadjuvant HER2-targeted therapy. Nature communications Caswell-Jin, J. L., McNamara, K., Reiter, J. G., Sun, R., Hu, Z., Ma, Z., Ding, J., Suarez, C. J., Tilk, S., Raghavendra, A., Forte, V., Chin, S. F., Bardwell, H., Provenzano, E., Caldas, C., Lang, J., West, R., Tripathy, D., Press, M. F., Curtis, C. 2019; 10 (1): 2433

  Abstract

  The original version of this Article omitted from the Author Contributions statement that 'R.S. and J.G.R contributed equally to this work.' This has been corrected in both the PDF and HTML versions of the Article.

  View details for DOI 10.1038/s41467-019-10456-x

  View details for PubMedID 31147552

• Genomic analysis of benign prostatic hyperplasia implicates cellular re-landscaping in disease pathogenesis. JCI insight Middleton, L. W., Shen, Z., Varma, S., Pollack, A. S., Gong, X., Zhu, S., Zhu, C., Foley, J. W., Vennam, S., Sweeney, R. T., Tu, K., Biscocho, J., Eminaga, O., Nolley, R., Tibshirani, R., Brooks, J. D., West, R. B., Pollack, J. R. 2019; 5

  Abstract

  Benign prostatic hyperplasia (BPH) is the most common cause of lower urinary tract symptoms in men. Current treatments target prostate physiology rather than BPH pathophysiology and are only partially effective. Here, we applied next-generation sequencing to gain new insight into BPH. By RNAseq, we uncovered transcriptional heterogeneity among BPH cases, where a 65-gene BPH stromal signature correlated with symptom severity. Stromal signaling molecules BMP5 and CXCL13 were enriched in BPH while estrogen regulated pathways were depleted. Notably, BMP5 addition to cultured prostatic myofibroblasts altered their expression profile towards a BPH profile that included the BPH stromal signature. RNAseq also suggested an altered cellular milieu in BPH, which we verified by immunohistochemistry and single-cell RNAseq. In particular, BPH tissues exhibited enrichment of myofibroblast subsets, whilst depletion of neuroendocrine cells and an estrogen receptor (ESR1)-positive fibroblast cell type residing near epithelium. By whole-exome sequencing, we uncovered somatic single-nucleotide variants (SNVs) in BPH, of uncertain pathogenic significance but indicative of clonal cell expansions. Thus, genomic characterization of BPH has identified a clinically-relevant stromal signature and new candidate disease pathways (including a likely role for BMP5 signaling), and reveals BPH to be not merely a hyperplasia, but rather a fundamental re-landscaping of cell types.

  View details for DOI 10.1172/jci.insight.129749

  View details for PubMedID 31094703

• Blood transcriptome and clonal T cell correlates of response and nonresponse to eltrombopag therapy in a cohort of patients with chronic immune thrombocytopenia. Haematologica Zhang, H., Zhang, B. M., Guo, X., Xu, L., You, X., West, R. B., Bussel, J. B., Zehnder, J. L. 2019

  View details for DOI 10.3324/haematol.2019.226688

  View details for PubMedID 31296576

• Increased Galectin-1 Expression in Thymic Epithelial Tumors. Clinical lung cancer Riess, J. W., Kong, C. S., West, R. B., Padda, S. K., Neal, J. W., Wakelee, H. A., Le, Q. 2018

  Abstract

  INTRODUCTION: Thymic epithelial tumors (TET) are rare malignancies with a paucity of data on biology and therapeutics. Galectin-1 is a member of the beta-galactoside binding protein family and has been shown to mediate tumor growth via modulation of immune cell function. This study examined galectin-1 expression in TET.PATIENTS AND METHODS: A tissue microarray of 68 patients with TET and 8 benign thymus controls were stained for galectin-1 expression and scored by a pathologist blinded to patient clinical and pathologic data. Galectin-1 expression+1 or greater staining intensity was considered positive. Clinical and pathologic data were abstracted from institutional databases. Expression of galectin-1 in thymic tumor was compared to benign thymus controls and correlated with pertinent clinical and pathologic data.RESULTS: Galectin-1 expression was higher in TET compared to benign thymus controls (65% vs. 0%). No significant association between galectin-1 expression and the development of recurrent disease, paraneoplastic syndromes, or overall survival was noted.CONCLUSION: Galectin-1 is overexpressed in the majority of TET. Detection of galectin-1 may differentiate benign from neoplastic thymic processes. Additional studies are needed to assess the role of galectin-1 in the development of TET.

  View details for PubMedID 30773448

• The HTN3-MSANTD3 Fusion Gene Defines a Subset of Acinic Cell Carcinoma of the Salivary Gland. The American journal of surgical pathology Andreasen, S., Varma, S., Barasch, N., Thompson, L. D., Miettinen, M., Rooper, L., Stelow, E. B., Agander, T. K., Seethala, R. R., Chiosea, S. I., Homoe, P., Wessel, I., Larsen, S. R., Erentaite, D., Bishop, J. A., Ulhoi, B. P., Kiss, K., Melchior, L. C., Pollack, J. R., West, R. B. 2018

  Abstract

  The spectrum of tumors arising in the salivary glands is wide and has recently been shown to harbor a network of tumor-specific fusion genes. Acinic cell carcinoma (AciCC) is one of the more frequently encountered types of salivary gland carcinoma, but it has remained a genetic orphan until recently when a fusion between the HTN3 and MSANTD3 genes was described in one case. Neither of these 2 genes is known to be implicated in any other malignancy. This study was undertaken to investigate whether the HTN3-MSANTD3 fusion is a recurrent genetic event in AciCC and whether it is a characteristic of one of its histological variants. Of the 273 AciCCs screened, 9 cases showed rearrangement of MSANTD3 by break-apart fluorescence in situ hybridization, 2 had 1 to 2 extra signals, and 1 had gain, giving a total of 4.4% with MSANTD3 aberrations. In 6 of 7 available cases with MSANTD3 rearrangement, the HTN3-MSANTD3 fusion transcript was demonstrated with real-time polymerase chain reaction . Histologically, all fusion-positive cases were predominantly composed of serous tumor cells growing in solid sheets, with serous tumor cells expressing DOG-1 and the intercalated duct-like cell component being CK7 positive and S-100 positive in 6/9 cases. All but one case arose in the parotid gland, and none of the patients experienced a recurrence during follow-up. In contrast, the case with MSANTD3 gain metastasized to the cervical lymph nodes and lungs. In conclusion, we find the HTN3-MSANTD3 gene fusion to be a recurrent event in AciCC with prominent serous differentiation and an indolent clinical course.

  View details for PubMedID 30520817

• Matrix mechanical plasticity regulates cancer cell migration through confining microenvironments. Nature communications Wisdom, K. M., Adebowale, K., Chang, J., Lee, J. Y., Nam, S., Desai, R., Rossen, N. S., Rafat, M., West, R. B., Hodgson, L., Chaudhuri, O. 2018; 9 (1): 4144

  Abstract

  Studies of cancer cell migration have found two modes: one that is protease-independent, requiring micron-sized pores or channels for cells to squeeze through, and one that is protease-dependent, relevant for confining nanoporous matrices such as basement membranes (BMs). However, many extracellular matrices exhibit viscoelasticity and mechanical plasticity, irreversibly deforming in response to force, so that pore size may be malleable. Here we report the impact of matrix plasticity on migration. We develop nanoporous and BM ligand-presenting interpenetrating network (IPN) hydrogels in which plasticity could be modulated independent of stiffness. Strikingly, cells in high plasticity IPNs carry out protease-independent migration through the IPNs. Mechanistically, cells in high plasticity IPNs extend invadopodia protrusions to mechanically and plastically open up micron-sized channels and then migrate through them. These findings uncover a new mode of protease-independent migration, in which cells can migrate through confining matrix if it exhibits sufficient mechanical plasticity.

  View details for PubMedID 30297715

• A Structured Tumor-Immune Microenvironment in Triple Negative Breast Cancer Revealed by Multiplexed Ion Beam Imaging. Cell Keren, L., Bosse, M., Marquez, D., Angoshtari, R., Jain, S., Varma, S., Yang, S., Kurian, A., Van Valen, D., West, R., Bendall, S. C., Angelo, M. 2018; 174 (6): 1373

  Abstract

  The immune system is critical in modulating cancer progression, but knowledge of immune composition, phenotype, and interactions with tumor is limited. We used multiplexed ion beam imaging by time-of-flight (MIBI-TOF) to simultaneously quantify in situ expression of 36 proteins covering identity, function, and immune regulation at sub-cellular resolution in 41 triple-negative breast cancer patients. Multi-step processing, including deep-learning-based segmentation, revealed variability in the composition of tumor-immune populations across individuals, reconciled by overall immune infiltration and enriched co-occurrence of immune subpopulations and checkpoint expression. Spatial enrichment analysis showed immune mixed and compartmentalized tumors, coinciding with expression of PD1, PD-L1, and IDO in a cell-type- and location-specific manner. Ordered immune structures along the tumor-immune border were associated with compartmentalization and linked to survival. These data demonstrate organization in the tumor-immune microenvironment that is structured in cellular composition, spatial arrangement, and regulatory-protein expression and provide a framework to apply multiplexed imaging to immune oncology.

  View details for PubMedID 30193111

• A Structured Tumor-Immune Microenvironment in Triple Negative Breast Cancer Revealed by Multiplexed Ion Beam Imaging CELL Keren, L., Bosse, M., Marquez, D., Angoshtari, R., Jain, S., Varma, S., Yang, S., Kurian, A., Van Valen, D., West, R., Bendall, S. C., Angelo, M. 2018; 174 (6): 1373-+

  View details for DOI 10.1016/j.cell.2018.08.039

  View details for Web of Science ID 000443841000009

• Biological subtypes of nasopharyngeal carcinoma by genomic profiling Tay, K., Zhu, C., Vennam, S., Varma, S., Quynh-Thu Le, Sunwoo, J., West, R. AMER ASSOC CANCER RESEARCH. 2018

  View details for DOI 10.1158/1538-7445.AM2018-3411

  View details for Web of Science ID 000468819501213

• p300 and STAT3 drive YAP-independent mechanotransduction during breast cancer invasion Lee, J. Y., Chang, J., Nam, S., Lee, H., Dominguez, A. A., Varma, S., Qi, L. S., West, R. B., Chaudhuri, O. AMER ASSOC CANCER RESEARCH. 2018

  View details for DOI 10.1158/1538-7445.AM2018-185

  View details for Web of Science ID 000468818901255

• VISTA immune checkpoint deregulation in human triple-negative breast cancer Gruber, J. J., Juntilla, M. M., Yang, S., Geller, B., Jager, N., Lin, C., Lipchik, A. M., Chen, J., Ram, A., Vinayak, S., Telli, M. L., West, R. B., Ford, J. M., Snyder, M. P. AMER ASSOC CANCER RESEARCH. 2018

  View details for DOI 10.1158/1538-7445.AM2018-4749

  View details for Web of Science ID 000468819503369

• GFPT2-expressing cancer-associated fibroblasts mediate metabolic reprogramming in human lung adenocarcinoma. Cancer research Zhang, W., Bouchard, G., Yu, A., Shafiq, M., Jamali, M., Shrager, J. B., Ayers, K., Bakr, S., Gentles, A. J., Diehn, M., Quon, A., West, R. B., Nair, V., van de Rijn, M., Napel, S., Plevritis, S. K. 2018

  Abstract

  Metabolic reprogramming of the tumor microenvironment is recognized as a cancer hallmark. To identify new molecular processes associated with tumor metabolism, we analyzed the transcriptome of bulk and flow-sorted human primary non-small cell lung cancer (NSCLC) together with 18FDG-positron emission tomography scans, which provide a clinical measure of glucose uptake. Tumors with higher glucose uptake were functionally enriched for molecular processes associated with invasion in adenocarcinoma (AD) and cell growth in squamous cell carcinoma (SCC). Next, we identified genes correlated to glucose uptake that were predominately overexpressed in a single cell-type comprising the tumor microenvironment. For SCC, most of these genes were expressed by malignant cells, whereas in AD they were predominately expressed by stromal cells, particularly cancer-associated fibroblasts (CAFs). Among these AD genes correlated to glucose uptake, we focused on Glutamine-Fructose-6-Phosphate Transaminase 2 (GFPT2), which codes for the Glutamine-Fructose-6-Phosphate Aminotransferase 2 (GFAT2), a rate-limiting enzyme of the hexosamine biosynthesis pathway (HBP), which is responsible for glycosylation. GFPT2 was predictive of glucose uptake independent of GLUT1, the primary glucose transporter, and was prognostically significant at both gene and protein level. We confirmed that normal fibroblasts transformed to CAF-like cells, following TGF-beta treatment, upregulated HBP genes, including GFPT2, with less change in genes driving glycolysis, pentose phosphate pathway and TCA cycle. Our work provides new evidence of histology-specific tumor-stromal properties associated with glucose uptake in NSCLC and identifies GFPT2 as a critical regulator of tumor metabolic reprogramming in AD.

  View details for PubMedID 29760045

• Gene Expression in Nasopharyngeal Carcinoma by Laser Capture Microdissected Transcriptome Sequencing Tay, J. K., Zhu, C., Sunwoo, J., Le, Q. T., West, R. B. ELSEVIER SCIENCE INC. 2018: 1385

  View details for Web of Science ID 000428145600224

• Pathologic Features and Clinical Outcomes of Breast Cancers with HER2/CEP17 ratio < 2.0 and mean HER2 signals /cell > 6.0 by FISH; A Multi-Institutional Study Ballard, M., Toukatly, M., Bean, G., Jalikis, F., Krings, G., Schmidt, R., Chen, Y., Rendi, M., Dintzis, S., Troxell, M., West, R., Sibley, R., Allison, K. NATURE PUBLISHING GROUP. 2018: 48

  View details for Web of Science ID 000429308600140

• Recurrent MSANTD3 Aberrations Defines a Subset of Acinic Cell Carcinomas of the Salivary Gland Andreasen, S., Varma, S., Melchior, L. C., Agander, T. K., Kiss, K., Wessel, I., Homoe, P., Bishop, J., Pollack, J., West, R. NATURE PUBLISHING GROUP. 2018: 470?71

  View details for Web of Science ID 000429308603188

• Potential Impact of Proposed HER2 FISH Guideline Updates on FISH results; A Multi-Institutional Study Ballard, M., MacKerricher, W., Jalikis, F., Krings, G., Schmidt, R., Chen, Y., Rendi, M., Dintzis, S., Troxell, M., West, R., Sibley, R., Allison, K. NATURE PUBLISHING GROUP. 2018: 48

  View details for Web of Science ID 000429308600139

• Pathologic Features and Clinical Outcomes of HER2 FISH Cases with HER2: CEP17 ratio > 2.0 but < 4 HER2 signals/cell; A Multi-Institutional Study Ballard, M., Toukatly, M., Bean, G., Jalikis, F., Krings, G., Schmidt, R., Chen, Y., Dintzis, S., Troxell, M., West, R., Sibley, R., Allison, K. NATURE PUBLISHING GROUP. 2018: 47?48

  View details for Web of Science ID 000429308600138

• Higher Absolute Lymphocyte Counts Predict Lower Mortality from Early-Stage Triple-Negative Breast Cancer. Clinical cancer research : an official journal of the American Association for Cancer Research Afghahi, A., Purington, N., Han, S. S., Desai, M., Pierson, E., Mathur, M. B., Seto, T., Thompson, C. A., Rigdon, J., Telli, M. L., Badve, S. S., Curtis, C., West, R. B., Horst, K., Gomez, S. L., Ford, J. M., Sledge, G. W., Kurian, A. W. 2018

  Abstract

  Tumor-infiltrating lymphocytes (TILs) in pre-treatment biopsies are associated with improved survival in triple-negative breast cancer (TNBC). We investigated whether higher peripheral lymphocyte counts are associated with lower breast cancer-specific mortality (BCM) and overall mortality (OM) in TNBC.Data on treatments and diagnostic tests from electronic medical records of two healthcare systems were linked with demographic, clinical, pathologic, and mortality data from the California Cancer Registry. Multivariable regression models adjusted for age, race/ethnicity, socioeconomic status, cancer stage, grade, neoadjuvant/adjuvant chemotherapy use, radiotherapy use, and germline BRCA1/2 mutations were used to evaluate associations between absolute lymphocyte count (ALC), BCM and OM. For a subgroup with TILs data available, we explored the relationship between TILs and peripheral lymphocyte counts.1,463 Stage I-III TNBC patients were diagnosed from 2000-2014; 1113 (76%) received neoadjuvant/adjuvant chemotherapy within one year of diagnosis. Of 759 patients with available ALC data, 481 (63.4%) were ever lymphopenic (minimum ALC <1.0 K/?L). On multivariable analysis, higher minimum ALC, but not absolute neutrophil count, predicted lower OM (hazard ratio [HR]: 0.23, 95% confidence interval [CI]: 0.16-0.35) and BCM (HR: 0.19, CI: 0.11-0.34). Five-year probability of BCM was 15% for patients who were ever lymphopenic versus 4% for those who were not. An exploratory analysis (N=70) showed a significant association between TILs and higher peripheral lymphocyte counts during neoadjuvant chemotherapy.Higher peripheral lymphocyte counts predicted lower mortality from early-stage, potentially curable TNBC, suggesting that immune function may enhance the effectiveness of early TNBC treatment.

  View details for PubMedID 29581131

• QUANTIFYING CHROMOSOMAL COPY NUMBER ALTERATIONS IN BREAST DUCTAL CARCINOMA IN SITU: A DEEP LEARNING BASED APPROACH Sirinukunwattana, K., Lin, J., Lu, P., Beca, F., Peng, J., Tolwani, A., Stancu, A., Varma, S., West, R., IEEE IEEE. 2018: 186?90

  View details for Web of Science ID 000455045600040

• Genome-wide reconstruction of complex structural variants using read clouds NATURE METHODS Spies, N., Weng, Z., Bishara, A., McDaniel, J., Catoe, D., Zook, J. M., Salit, M., West, R. B., Batzoglou, S., Sidow, A. 2017; 14 (9): 915-+

  Abstract

  In read cloud approaches, microfluidic partitioning of long genomic DNA fragments and barcoding of shorter fragments derived from these fragments retains long-range information in short sequencing reads. This combination of short reads with long-range information represents a powerful alternative to single-molecule long-read sequencing. We develop Genome-wide Reconstruction of Complex Structural Variants (GROC-SVs) for SV detection and assembly from read cloud data and apply this method to Illumina-sequenced 10x Genomics sarcoma and breast cancer data sets. Compared with short-fragment sequencing, GROC-SVs substantially improves the specificity of breakpoint detection at comparable sensitivity. This approach also performs sequence assembly across multiple breakpoints simultaneously, enabling the reconstruction of events exhibiting remarkable complexity. We show that chromothriptic rearrangements occurred before copy number amplifications, and that rates of single-nucleotide variants and SVs are not correlated. Our results support the use of read cloud approaches to advance the characterization of large and complex structural variation.

  View details for PubMedID 28714986

• Circulating Tumor DNA Detects Residual Disease and Anticipates Tumor Progression Earlier Than CT Imaging Chaudhuri, A. A., Chabon, J. J., Lovejoy, A. F., Newman, A. M., Stehr, H., Azad, T. D., Zhou, L., Liu, C., Scherer, F., Kurtz, D. M., Esfahani, M. S., Say, C., Carter, J. N., Merriott, D., Dudley, J., Binkley, M. S., Modlin, L., Padda, S. K., Gensheimer, M., West, R. B., Shrager, J. B., Neal, J. W., Wakelee, H. A., Billy, W., Alizadeh, A. A., Diehn, M. ELSEVIER SCIENCE INC. 2017: E4

  View details for DOI 10.1016/j.ijrobp.2017.02.048

  View details for Web of Science ID 000403079100013

• 'Non-classical' HER2 FISH results in breast cancer: a multi-institutional study MODERN PATHOLOGY Ballard, M., Jalikis, F., Krings, G., Schmidt, R. A., Chen, Y., Rendi, M. H., Dintzis, S. M., Jensen, K. C., West, R. B., Sibley, R. K., Troxell, M. L., Allison, K. H. 2017; 30 (2): 227-235

  Abstract

  The 2013 CAP/ASCO HER2 Testing Guidelines Update modified HER2 FISH categories such that some cases with 'monosomy', 'co-amplification/polysomy', low-level increased HER2 signals or clustered heterogeneity now are considered amplified or equivocal. This study examines the frequency and clinico-pathologic characteristics of breast cancers with equivocal or 'non-classical' HER2 FISH results. Breast cancers (2001-2014) with HER2 FISH results, HER2 immunohistochemistry, ER, grade, and age from three institutions (Stanford, UCSF, UWMC) were collected. HER2 FISH was interpreted using the updated recommendations. Amplified cases with non-classical results were grouped into the following categories: (1) 'monosomy' (ratio ?2.0, mean HER2/cell<4.0); (2) 'co-amplified' (ratio<2.0, mean HER2/cell ?6.0); (3) 'low amplified' (ratio ?2.0, mean HER2/cell 4.0-5.9). Heterogeneous cases with clustered HER2-positive cells were also included. Of 8068 cases, 5.2% were equivocal and 4.6% had a 'non-classical' HER2 amplified result; 1.4% 'monosomy', 0.8% 'co-amplified', 2.1% 'low amplified', and 0.3% clustered heterogeneity. These cancers had a high frequency of ER positive (80.4%), Nottingham grade 3 (52.1%) results. The highest percentage of grade 3 cancers (66.7%) and positive HER2 immunohistochemistry (31.7%) was in the 'co-amplified' group. The 'monosomy' group had the highest percent grade 1 cancers (13.3%) and was most frequently HER2 immunohistochemistry negative (30.1%). Equivocal cases had very similar characteristics to the 'low-amplified' category. Cases with non-classical HER2 amplification or equivocal results are typically ER positive, higher grade cancers. 'Co-amplified' cases have the highest frequencies of aggressive characteristics and 'monosomy' cases the highest frequencies of lower risk features. With little clinical outcomes data currently available on these non-classical HER2 results, these results support the current classification scheme for HER2 FISH, with case-by-case correlation with additional clinical-pathologic factors when evaluating whether to offer HER2-targeted therapies in these non-classical cases.Modern Pathology advance online publication, 14 October 2016; doi:10.1038/modpathol.2016.175.

  View details for DOI 10.1038/modpathol.2016.175

  View details for Web of Science ID 000393257400007

• Loss of MAdCAM-1 Expression in Colorectal Adenocarcinoma Devereaux, K., Gomez, A., Varma, S., Pai, R., Modiano, N., Longacre, T., West, R., Troxell, M. NATURE PUBLISHING GROUP. 2017: 170A

  View details for Web of Science ID 000394467300675

• Loss of MAdCAM-1 Expression in Colorectal Adenocarcinoma Devereaux, K., Gomez, A., Varma, S., Pai, R., Modiano, N., Longacre, T., West, R., Troxell, M. NATURE PUBLISHING GROUP. 2017: 170A

  View details for Web of Science ID 000393724400674

• Fibroepithelial Lesions of the Breast Involved by Atypical Epithelial Proliferations: A 12-Year Single Institution Study Singer, B., Lin, C., West, R. NATURE PUBLISHING GROUP. 2017: 71A

  View details for Web of Science ID 000393724400271

• Fibroepithelial Lesions of the Breast Involved by Atypical Epithelial Proliferations: A 12-Year Single Institution Study Singer, B., Lin, C., West, R. NATURE PUBLISHING GROUP. 2017: 71A

  View details for Web of Science ID 000394467300271

• Local estrogen axis in the human bone microenvironment regulates estrogen receptor-positive breast cancer cells. Breast cancer research : BCR Amanatullah, D. F., Tamaresis, J. S., Chu, P., Bachmann, M. H., Hoang, N. M., Collyar, D., Mayer, A. T., West, R. B., Maloney, W. J., Contag, C. H., King, B. L. 2017; 19 (1): 121

  Abstract

  Approximately 70% of all breast cancers express the estrogen receptor, and are regulated by estrogen. While the ovaries are the primary source of estrogen in premenopausal women, most breast cancer is diagnosed following menopause, when systemic levels of this hormone decline. Estrogen production from androgen precursors is catalyzed by the aromatase enzyme. Although aromatase expression and local estrogen production in breast adipose tissue have been implicated in the development of primary breast cancer, the source of estrogen involved in the regulation of estrogen receptor-positive (ER+) metastatic breast cancer progression is less clear.Bone is the most common distant site of breast cancer metastasis, particularly for ER+ breast cancers. We employed a co-culture model using trabecular  bone tissues obtained from total hip replacement (THR) surgery specimens to study ER+ and estrogen receptor-negative (ER-) breast cancer cells within the human bone microenvironment. Luciferase-expressing ER+ (MCF-7, T-47D, ZR-75) and ER- (SK-BR-3, MDA-MB-231, MCF-10A) breast cancer cells were cultured directly on bone tissue fragments or in bone tissue-conditioned media, and monitored over time with bioluminescence imaging (BLI). Bone tissue-conditioned media were generated in the presence vs. absence of aromatase inhibitors, and testosterone. Bone tissue fragments were analyzed for aromatase expression by immunohistochemistry.ER+ breast cancer cells were preferentially sustained in co-cultures with bone tissues and bone tissue-conditioned media relative to ER- cells. Bone fragments analyzed by immunohistochemistry revealed expression of the aromatase enzyme. Bone tissue-conditioned media generated in the presence of testosterone had increased estrogen levels and heightened capacity to stimulate ER+ breast cancer cell proliferation. Pretreatment of cultured bone tissues with aromatase inhibitors, which inhibited estrogen production, reduced the capacity of conditioned media to stimulate ER+ cell proliferation.These results suggest that a local estrogen signaling axis regulates ER+ breast cancer cell viability and proliferation within the bone metastatic niche, and that aromatase inhibitors modulate this axis. Although endocrine therapies are highly effective in the treatment of ER+ breast cancer, resistance to these treatments reduces their efficacy. Characterization of estrogen signaling networks within the bone microenvironment will identify new strategies for combating metastatic progression and endocrine resistance.

  View details for PubMedID 29141657

  View details for PubMedCentralID PMC5688761

• YAP-independent mechanotransduction drives breast cancer invasion. Lee, J. Y., Chang, J., Nam, S., Lee, H., Dominguez, A., Varma, S., Qi, L. S., West, R., Chaudhuri, O. AMER SOC CELL BIOLOGY. 2017

  View details for Web of Science ID 000426664300595

• YAP-independent mechanotransduction drives breast cancer invasion. Lee, J. Y., Chang, J., Nam, S., Lee, H., Dominguez, A., Varma, S., Qi, L. S., West, R., Chaudhuri, O. AMER SOC CELL BIOLOGY. 2017

  View details for Web of Science ID 000426660800265

• Circulating Tumor DNA Detects Minimal Residual Disease and Predicts Outcome in Localized Lung Cancer Chaudhuri, A., Lovejoy, A., Chabon, J., Newman, A., Stehr, H., Say, C., Carter, J., Zhou, L., West, R., Shrager, J., Neal, J., Wakelee, H., Loo, B., Alizadeh, A., Diehn, M. ELSEVIER SCIENCE INC. 2017: S445

  View details for Web of Science ID 000413055801024

• Early detection of molecular residual disease in localized lung cancer by circulating tumor DNA profiling. Cancer discovery Chaudhuri, A. A., Chabon, J. J., Lovejoy, A. F., Newman, A. M., Stehr, H., Azad, T. D., Khodadoust, M. S., Esfahani, M. S., Liu, C. L., Zhou, L., Scherer, F., Kurtz, D. M., Say, C., Carter, J. N., Merriott, D. J., Dudley, J. C., Binkley, M. S., Modlin, L., Padda, S. K., Gensheimer, M. F., West, R. B., Shrager, J. B., Neal, J. W., Wakelee, H. A., Loo, B. W., Alizadeh, A. A., Diehn, M. 2017

  Abstract

  Identifying molecular residual disease (MRD) after treatment of localized lung cancer could facilitate early intervention and personalization of adjuvant therapies. Here we apply Cancer Personalized Profiling by Deep Sequencing (CAPP-Seq) circulating tumor DNA (ctDNA) analysis to 255 samples from 40 patients treated with curative intent for stage I-III lung cancer and 54 healthy adults. In 94% of evaluable patients experiencing recurrence, ctDNA was detectable in the first post-treatment blood sample, indicating reliable identification of MRD. Post-treatment ctDNA detection preceded radiographic progression in 72% of patients by a median of 5.2 months and 53% of patients harbored ctDNA mutation profiles associated with favorable responses to tyrosine kinase inhibitors or immune checkpoint blockade. Collectively, these results indicate that ctDNA MRD in lung cancer patients can be accurately detected using CAPP-Seq and may allow personalized adjuvant treatment while disease burden is lowest.

  View details for PubMedID 28899864

• Role of KEAP1/NRF2 and TP53 Mutations in Lung Squamous Cell Carcinoma Development and Radiation Resistance CANCER DISCOVERY Jeong, Y., Hoang, N. T., Lovejoy, A., Stehr, H., Newman, A. M., Gentles, A. J., Kong, W., Diana Truong, D., Martin, S., Chaudhuri, A., Heiser, D., Zhou, L., Say, C., Carter, J. N., Hiniker, S. M., Loo, B. W., West, R. B., Beachy, P., Alizadeh, A. A., Diehn, M. 2017; 7 (1): 86-101

  Abstract

  Lung squamous cell carcinoma (LSCC) pathogenesis remains incompletely understood, and biomarkers predicting treatment response remain lacking. Here, we describe novel murine LSCC models driven by loss of Trp53 and Keap1, both of which are frequently mutated in human LSCCs. Homozygous inactivation of Keap1 or Trp53 promoted airway basal stem cell (ABSC) self-renewal, suggesting that mutations in these genes lead to expansion of mutant stem cell clones. Deletion of Trp53 and Keap1 in ABSCs, but not more differentiated tracheal cells, produced tumors recapitulating histologic and molecular features of human LSCCs, indicating that they represent the likely cell of origin in this model. Deletion of Keap1 promoted tumor aggressiveness, metastasis, and resistance to oxidative stress and radiotherapy (RT). KEAP1/NRF2 mutation status predicted risk of local recurrence after RT in patients with non-small lung cancer (NSCLC) and could be noninvasively identified in circulating tumor DNA. Thus, KEAP1/NRF2 mutations could serve as predictive biomarkers for personalization of therapeutic strategies for NSCLCs.We developed an LSCC mouse model involving Trp53 and Keap1, which are frequently mutated in human LSCCs. In this model, ABSCs are the cell of origin of these tumors. KEAP1/NRF2 mutations increase radioresistance and predict local tumor recurrence in radiotherapy patients. Our findings are of potential clinical relevance and could lead to personalized treatment strategies for tumors with KEAP1/NRF2 mutations. Cancer Discov; 7(1); 86-101. ©2016 AACR.This article is highlighted in the In This Issue feature, p. 1.

  View details for DOI 10.1158/2159-8290.CD-16-0127

  View details for Web of Science ID 000396017700024

  View details for PubMedCentralID PMC5222718

• Recurrent rearrangements of the Myb/SANT-like DNA-binding domain containing 3 gene (MSANTD3) in salivary gland acinic cell carcinoma. PloS one Barasch, N., Gong, X., Kwei, K. A., Varma, S., Biscocho, J., Qu, K., Xiao, N., Lipsick, J. S., Pelham, R. J., West, R. B., Pollack, J. R. 2017; 12 (2)

  Abstract

  Pathogenic gene fusions have been identified in several histologic types of salivary gland neoplasia, but not previously in acinic cell carcinoma (AcCC). To discover novel gene fusions, we performed whole-transcriptome sequencing surveys of three AcCC archival cases. In one specimen we identified a novel HTN3-MSANTD3 gene fusion, and in another a novel PRB3-ZNF217 gene fusion. The structure of both fusions was consistent with the promoter of the 5' partner (HTN3 or PRB3), both highly expressed salivary gland genes, driving overexpression of full-length MSANTD3 or ZNF217. By fluorescence in situ hybridization of an expanded AcCC case series, we observed MSANTD3 rearrangements altogether in 3 of 20 evaluable cases (15%), but found no additional ZNF217 rearrangements. MSANTD3 encodes a previously uncharacterized Myb/SANT domain-containing protein. Immunohistochemical staining demonstrated diffuse nuclear MSANTD3 expression in 8 of 27 AcCC cases (30%), including the three cases with MSANTD3 rearrangement. MSANTD3 displayed heterogeneous expression in normal salivary ductal epithelium, as well as among other histologic types of salivary gland cancer though without evidence of translocation. In a broader survey, MSANTD3 showed variable expression across a wide range of normal and neoplastic human tissue specimens. In preliminary functional studies, engineered MSANTD3 overexpression in rodent salivary gland epithelial cells did not enhance cell proliferation, but led to significant upregulation of gene sets involved in protein synthesis. Our findings newly identify MSANTD3 rearrangement as a recurrent event in salivary gland AcCC, providing new insight into disease pathogenesis, and identifying a putative novel human oncogene.

  View details for DOI 10.1371/journal.pone.0171265

  View details for PubMedID 28212443

• Loss of Expression of AZGP1 Is Associated With Worse Clinical Outcomes in a Multi-Institutional Radical Prostatectomy Cohort. Prostate Brooks, J. D., Wei, W., Pollack, J. R., West, R. B., Shin, J. H., Sunwoo, J. B., Hawley, S. J., Auman, H., Newcomb, L. F., Simko, J., Hurtado-Coll, A., Troyer, D. A., Carroll, P. R., Gleave, M. E., Lin, D. W., Nelson, P. S., Thompson, I. M., True, L. D., McKenney, J. K., Feng, Z., Fazli, L. 2016; 76 (15): 1409-1419

  Abstract

  Given the uncertainties inherent in clinical measures of prostate cancer aggressiveness, clinically validated tissue biomarkers are needed. We tested whether Alpha-2-Glycoprotein 1, Zinc-Binding (AZGP1) protein levels, measured by immunohistochemistry, and RNA expression, by RNA in situ hybridization (RISH), predict recurrence after radical prostatectomy independent of clinical and pathological parameters.AZGP1 IHC and RISH were performed on a large multi-institutional tissue microarray resource including 1,275 men with 5 year median follow-up. The relationship between IHC and RISH expression levels was assessed using the Kappa analysis. Associations with clinical and pathological parameters were tested by the Chi-square test and the Wilcoxon rank sum test. Relationships with outcome were assessed with univariable and multivariable Cox proportional hazards models and the Log-rank test.Absent or weak expression of AZGP1 protein was associated with worse recurrence free survival (RFS), disease specific survival, and overall survival after radical prostatectomy in univariable analysis. AZGP1 protein expression, along with pre-operative serum PSA levels, surgical margin status, seminal vesicle invasion, extracapsular extension, and Gleason score predicted RFS on multivariable analysis. Similarly, absent or low AZGP1 RNA expression by RISH predicted worse RFS after prostatectomy in univariable and multivariable analysis.In our large, rigorously designed validation cohort, loss of AZGP1 expression predicts RFS after radical prostatectomy independent of clinical and pathological variables. Prostate © 2016 Wiley Periodicals, Inc.

  View details for DOI 10.1002/pros.23225

  View details for PubMedID 27325561

• 'Non-classical' HER2 FISH results in breast cancer: a multi-institutional study. Modern pathology Ballard, M., Jalikis, F., Krings, G., Schmidt, R. A., Chen, Y., Rendi, M. H., Dintzis, S. M., Jensen, K. C., West, R. B., Sibley, R. K., Troxell, M. L., Allison, K. H. 2016

  Abstract

  The 2013 CAP/ASCO HER2 Testing Guidelines Update modified HER2 FISH categories such that some cases with 'monosomy', 'co-amplification/polysomy', low-level increased HER2 signals or clustered heterogeneity now are considered amplified or equivocal. This study examines the frequency and clinico-pathologic characteristics of breast cancers with equivocal or 'non-classical' HER2 FISH results. Breast cancers (2001-2014) with HER2 FISH results, HER2 immunohistochemistry, ER, grade, and age from three institutions (Stanford, UCSF, UWMC) were collected. HER2 FISH was interpreted using the updated recommendations. Amplified cases with non-classical results were grouped into the following categories: (1) 'monosomy' (ratio ?2.0, mean HER2/cell<4.0); (2) 'co-amplified' (ratio<2.0, mean HER2/cell ?6.0); (3) 'low amplified' (ratio ?2.0, mean HER2/cell 4.0-5.9). Heterogeneous cases with clustered HER2-positive cells were also included. Of 8068 cases, 5.2% were equivocal and 4.6% had a 'non-classical' HER2 amplified result; 1.4% 'monosomy', 0.8% 'co-amplified', 2.1% 'low amplified', and 0.3% clustered heterogeneity. These cancers had a high frequency of ER positive (80.4%), Nottingham grade 3 (52.1%) results. The highest percentage of grade 3 cancers (66.7%) and positive HER2 immunohistochemistry (31.7%) was in the 'co-amplified' group. The 'monosomy' group had the highest percent grade 1 cancers (13.3%) and was most frequently HER2 immunohistochemistry negative (30.1%). Equivocal cases had very similar characteristics to the 'low-amplified' category. Cases with non-classical HER2 amplification or equivocal results are typically ER positive, higher grade cancers. 'Co-amplified' cases have the highest frequencies of aggressive characteristics and 'monosomy' cases the highest frequencies of lower risk features. With little clinical outcomes data currently available on these non-classical HER2 results, these results support the current classification scheme for HER2 FISH, with case-by-case correlation with additional clinical-pathologic factors when evaluating whether to offer HER2-targeted therapies in these non-classical cases.Modern Pathology advance online publication, 14 October 2016; doi:10.1038/modpathol.2016.175.

  View details for DOI 10.1038/modpathol.2016.175

  View details for PubMedID 27739440

• BRAF inhibitor therapy of primary ameloblastoma ORAL SURGERY ORAL MEDICINE ORAL PATHOLOGY ORAL RADIOLOGY Tan, S., Pollack, J. R., Kaplan, M. J., Colevas, A., West, R. B. 2016; 122 (4): 518?19

  View details for PubMedID 27651290

• CAPP-Seq Circulating Tumor DNA Analysis for Early Detection of Tumor Progression After Definitive Radiation Therapy for Lung Cancer Chaudhuri, A. A., Lovejoy, A. F., Chabon, J. J., Newman, A., Stehr, H., Say, C., Aggarwal, S., Carter, J. N., West, R. B., Neal, J. W., Wakelee, H. A., Loo, B. W., Alizadeh, A., Diehn, M. ELSEVIER SCIENCE INC. 2016: S41?S42

  View details for DOI 10.1016/j.ijrobp.2016.06.112

  View details for Web of Science ID 000387655804429

• Role of KEAP1/NRF2 and TP53 Mutations in Lung Squamous Cell Carcinoma Development and Radiation Resistance. Cancer discovery Jeong, Y., Hoang, N. T., Lovejoy, A., Stehr, H., Newman, A. M., Gentles, A. J., Kong, W., Truong, D., Martin, S., Chaudhuri, A., Heiser, D., Zhou, L., Say, C., Carter, J. N., Hiniker, S. M., Loo, B. W., West, R. B., Beachy, P., Alizadeh, A. A., Diehn, M. 2016

  Abstract

  Lung squamous cell carcinoma (LSCC) pathogenesis remains incompletely understood, and biomarkers predicting treatment response remain lacking. Here, we describe novel murine LSCC models driven by loss of Trp53 and Keap1, both of which are frequently mutated in human LSCCs. Homozygous inactivation of Keap1 or Trp53 promoted airway basal stem cell (ABSC) self-renewal, suggesting that mutations in these genes lead to expansion of mutant stem cell clones. Deletion of Trp53 and Keap1 in ABSCs, but not more differentiated tracheal cells, produced tumors recapitulating histologic and molecular features of human LSCCs, indicating that they represent the likely cell of origin in this model. Deletion of Keap1 promoted tumor aggressiveness, metastasis, and resistance to oxidative stress and radiotherapy (RT). KEAP1/NRF2 mutation status predicted risk of local recurrence after RT in patients with non-small lung cancer (NSCLC) and could be noninvasively identified in circulating tumor DNA. Thus, KEAP1/NRF2 mutations could serve as predictive biomarkers for personalization of therapeutic strategies for NSCLCs.We developed an LSCC mouse model involving Trp53 and Keap1, which are frequently mutated in human LSCCs. In this model, ABSCs are the cell of origin of these tumors. KEAP1/NRF2 mutations increase radioresistance and predict local tumor recurrence in radiotherapy patients. Our findings are of potential clinical relevance and could lead to personalized treatment strategies for tumors with KEAP1/NRF2 mutations. Cancer Discov; 7(1); 86-101. ©2016 AACR.This article is highlighted in the In This Issue feature, p. 1.

  View details for PubMedID 27663899

• Ameloblastoma: a clinical review and trends in management EUROPEAN ARCHIVES OF OTO-RHINO-LARYNGOLOGY McClary, A. C., West, R. B., McClary, A. C., Pollack, J. R., Fischbein, N. J., Holsinger, C. F., Sunwoo, J., Colevas, A. D., Sirjani, D. 2016; 273 (7): 1649-1661

  Abstract

  Ameloblastoma is a rare odontogenic neoplasm of the mandible and maxilla, with multiple histologic variants, and high recurrence rates if improperly treated. The current mainstay of treatment is wide local excision with appropriate margins and immediate reconstruction. Here we review the ameloblastoma literature, using the available evidence to highlight the change in management over the past several decades. In addition, we explore the recent molecular characterization of these tumors which may point towards new potential avenues of personalized treatment.

  View details for DOI 10.1007/s00405-015-3631-8

  View details for PubMedID 25926124

• BRAF inhibitor treatment of primary BRAF-mutant ameloblastoma with pathologic assessment of response ORAL SURGERY ORAL MEDICINE ORAL PATHOLOGY ORAL RADIOLOGY Tan, S., Pollack, J. R., Kaplan, M. J., Colevas, A. D., West, R. B. 2016; 122 (1): E5-E7

  Abstract

  Molecular characterization of ameloblastoma has indicated a high frequency of driver mutations in BRAF and SMO. Preclinical data suggest that Food and Drug Administration-approved BRAF-targeted therapies may be immediately relevant for patients with ameloblastoma positive for the BRAF V600E mutation.A neoadjuvant treatment regime of dabrafenib was given to a patient with recurrent BRAF-mutant mandibular ameloblastoma. The patient subsequently underwent left mandible composite resection of the tumor and pathologic evaluation of treatment response.The ameloblastoma had a slow but dramatic response with >90% tumor volume reduction. The inner areas of the tumor underwent degeneration and squamous differentiation, and intact ameloblastoma was present in the outer areas associated with bone.Targeted neoadjuvant therapy for ameloblastoma may be useful in certain clinical settings of primary ameloblastoma. These might include tumors of advanced local stage when a neoadjuvant reduction could alter the extent of surgery and instances of local recurrence when surgical options are limited.

  View details for DOI 10.1016/j.oooo.2015.12.016

  View details for Web of Science ID 000377426600002

  View details for PubMedID 27209484

• Transcriptome sequencing analysis of four psammomatous meningiomas Samghabadi, P., Forgo, E., West, R., Vogel, H. LIPPINCOTT WILLIAMS & WILKINS. 2016: 575

  View details for Web of Science ID 000377665000038

• Integrated digital error suppression for improved detection of circulating tumor DNA NATURE BIOTECHNOLOGY Newman, A. M., Lovejoy, A. F., Klass, D. M., Kurtz, D. M., Chabon, J. J., Scherer, F., Stehr, H., Liu, C. L., Bratman, S. V., Say, C., Zhou, L., Carter, J. N., West, R. B., Sledge, G. W., Shrager, J. B., Loo, B. W., Neal, J. W., Wakelee, H. A., Diehn, M., Alizadeh, A. A. 2016; 34 (5): 547-555

  Abstract

  High-throughput sequencing of circulating tumor DNA (ctDNA) promises to facilitate personalized cancer therapy. However, low quantities of cell-free DNA (cfDNA) in the blood and sequencing artifacts currently limit analytical sensitivity. To overcome these limitations, we introduce an approach for integrated digital error suppression (iDES). Our method combines in silico elimination of highly stereotypical background artifacts with a molecular barcoding strategy for the efficient recovery of cfDNA molecules. Individually, these two methods each improve the sensitivity of cancer personalized profiling by deep sequencing (CAPP-Seq) by about threefold, and synergize when combined to yield ?15-fold improvements. As a result, iDES-enhanced CAPP-Seq facilitates noninvasive variant detection across hundreds of kilobases. Applied to non-small cell lung cancer (NSCLC) patients, our method enabled biopsy-free profiling of EGFR kinase domain mutations with 92% sensitivity and >99.99% specificity at the variant level, and with 90% sensitivity and 96% specificity at the patient level. In addition, our approach allowed monitoring of NSCLC ctDNA down to 4 in 10(5) cfDNA molecules. We anticipate that iDES will aid the noninvasive genotyping and detection of ctDNA in research and clinical settings.

  View details for DOI 10.1038/nbt.3520

  View details for PubMedID 27018799

• Validation of Digital Whole Slide Imaging System for Breast Sentinel Lymph Node Touch Prep Analysis: A Single Institution Experience Hoffmann, J., McGinnis, L., Mafnas, C. T., Ziskin, J., Folkins, A. K., Allison, K., West, R. B., Higgins, J. P., Kambham, N., Long, S. R., Tan, B. NATURE PUBLISHING GROUP. 2016: 496A

  View details for Web of Science ID 000369270703125

• HER2 FISH Equivocal Category: Does Retesting Resolve This Clinical Grey Zone? Ballard, M., Srivastava, S., West, R. B., Jensen, K. C., Sibley, R. K., Allison, K. NATURE PUBLISHING GROUP. 2016: 31A

  View details for Web of Science ID 000369270700115

• Genomic Profiling of Ductal Carcinoma In Situ: Can We Predict Outcome? Lin, C., Vennam, S., Sweeney, R. T., Zhu, S. X., Varma, S., West, R. B. NATURE PUBLISHING GROUP. 2016: 54A

  View details for Web of Science ID 000370302500212

• Correlation of Protein Expression with Chromosomal Copy Number Alterations in the Progression of Early Breast Neoplasia to DCIS Forgo, E., Varma, S., West, R. B. NATURE PUBLISHING GROUP. 2016: 41A

  View details for Web of Science ID 000369270700157

• Genomic Profiling of Ductal Carcinoma In Situ: Can We Predict Outcome? Lin, C., Vennam, S., Sweeney, R. T., Zhu, S. X., Varma, S., West, R. B. NATURE PUBLISHING GROUP. 2016: 54A

  View details for Web of Science ID 000369270700212

• Correlation of Protein Expression with Chromosomal Copy Number Alterations in the Progression of Early Breast Neoplasia to DCIS Forgo, E., Varma, S., West, R. B. NATURE PUBLISHING GROUP. 2016: 41A

  View details for Web of Science ID 000370302500157

• Validation of Digital Whole Slide Imaging System for Intraoperative Breast Sentinel Lymph Node Touch Prep Analysis: A Single Institution Experience Hoffmann, J., McGinnis, L., Mafnas, C. T., Ziskin, J., Folkins, A. K., Allison, K., West, R. B., Higgins, J. P., Kambham, N., Long, S. R., Tan, B. NATURE PUBLISHING GROUP. 2016: 496A

  View details for Web of Science ID 000370302503446

• The prognostic landscape of genes and infiltrating immune cells across human cancers Gentles, A. J., Newman, A. M., Liu, C., Bratman, S. V., Feng, W., Kim, D., Nair, V. S., Yue, X., Khuong, A., Hoang, C. D., Diehn, M., West, R. B., Plevritis, S. K., Alizadeh, A. A. AMER ASSOC CANCER RESEARCH. 2015

  View details for DOI 10.1158/1538-7445.TRANSCAGEN-PR09

  View details for Web of Science ID 000370972600122

• Integrating Tumor and Stromal Gene Expression Signatures With Clinical Indices for Survival Stratification of Early-Stage Non-Small Cell Lung Cancer. Journal of the National Cancer Institute Gentles, A. J., Bratman, S. V., Lee, L. J., Harris, J. P., Feng, W., Nair, R. V., Shultz, D. B., Nair, V. S., Hoang, C. D., West, R. B., Plevritis, S. K., Alizadeh, A. A., Diehn, M. 2015; 107 (10)

  Abstract

  Accurate survival stratification in early-stage non-small cell lung cancer (NSCLC) could inform the use of adjuvant therapy. We developed a clinically implementable mortality risk score incorporating distinct tumor microenvironmental gene expression signatures and clinical variables.Gene expression profiles from 1106 nonsquamous NSCLCs were used for generation and internal validation of a nine-gene molecular prognostic index (MPI). A quantitative polymerase chain reaction (qPCR) assay was developed and validated on an independent cohort of formalin-fixed paraffin-embedded (FFPE) tissues (n = 98). A prognostic score using clinical variables was generated using Surveillance, Epidemiology, and End Results data and combined with the MPI. All statistical tests for survival were two-sided.The MPI stratified stage I patients into prognostic categories in three microarray and one FFPE qPCR validation cohorts (HR = 2.99, 95% CI = 1.55 to 5.76, P < .001 in stage IA patients of the largest microarray validation cohort; HR = 3.95, 95% CI = 1.24 to 12.64, P = .01 in stage IA of the qPCR cohort). Prognostic genes were expressed in distinct tumor cell subpopulations, and genes implicated in proliferation and stem cells portended poor outcomes, while genes involved in normal lung differentiation and immune infiltration were associated with superior survival. Integrating the MPI with clinical variables conferred greatest prognostic power (HR = 3.43, 95% CI = 2.18 to 5.39, P < .001 in stage I patients of the largest microarray cohort; HR = 3.99, 95% CI = 1.67 to 9.56, P < .001 in stage I patients of the qPCR cohort). Finally, the MPI was prognostic irrespective of somatic alterations in EGFR, KRAS, TP53, and ALK.The MPI incorporates genes expressed in the tumor and its microenvironment and can be implemented clinically using qPCR assays on FFPE tissues. A composite model integrating the MPI with clinical variables provides the most accurate risk stratification.

  View details for DOI 10.1093/jnci/djv211

  View details for PubMedID 26286589

• Integrating Tumor and Stromal Gene Expression Signatures With Clinical Indices for Survival Stratification of Early-Stage Non-Small Cell Lung Cancer. Journal of the National Cancer Institute Gentles, A. J., Bratman, S. V., Lee, L. J., Harris, J. P., Feng, W., Nair, R. V., Shultz, D. B., Nair, V. S., Hoang, C. D., West, R. B., Plevritis, S. K., Alizadeh, A. A., Diehn, M. 2015; 107 (10)

  Abstract

  Accurate survival stratification in early-stage non-small cell lung cancer (NSCLC) could inform the use of adjuvant therapy. We developed a clinically implementable mortality risk score incorporating distinct tumor microenvironmental gene expression signatures and clinical variables.Gene expression profiles from 1106 nonsquamous NSCLCs were used for generation and internal validation of a nine-gene molecular prognostic index (MPI). A quantitative polymerase chain reaction (qPCR) assay was developed and validated on an independent cohort of formalin-fixed paraffin-embedded (FFPE) tissues (n = 98). A prognostic score using clinical variables was generated using Surveillance, Epidemiology, and End Results data and combined with the MPI. All statistical tests for survival were two-sided.The MPI stratified stage I patients into prognostic categories in three microarray and one FFPE qPCR validation cohorts (HR = 2.99, 95% CI = 1.55 to 5.76, P < .001 in stage IA patients of the largest microarray validation cohort; HR = 3.95, 95% CI = 1.24 to 12.64, P = .01 in stage IA of the qPCR cohort). Prognostic genes were expressed in distinct tumor cell subpopulations, and genes implicated in proliferation and stem cells portended poor outcomes, while genes involved in normal lung differentiation and immune infiltration were associated with superior survival. Integrating the MPI with clinical variables conferred greatest prognostic power (HR = 3.43, 95% CI = 2.18 to 5.39, P < .001 in stage I patients of the largest microarray cohort; HR = 3.99, 95% CI = 1.67 to 9.56, P < .001 in stage I patients of the qPCR cohort). Finally, the MPI was prognostic irrespective of somatic alterations in EGFR, KRAS, TP53, and ALK.The MPI incorporates genes expressed in the tumor and its microenvironment and can be implemented clinically using qPCR assays on FFPE tissues. A composite model integrating the MPI with clinical variables provides the most accurate risk stratification.

  View details for DOI 10.1093/jnci/djv211

  View details for PubMedID 26286589

• Read clouds uncover variation in complex regions of the human genome. Genome research Bishara, A., Liu, Y., Weng, Z., Kashef-Haghighi, D., Newburger, D. E., West, R., Sidow, A., Batzoglou, S. 2015; 25 (10): 1570-1580

  Abstract

  Although an increasing amount of human genetic variation is being identified and recorded, determining variants within repeated sequences of the human genome remains a challenge. Most population and genome-wide association studies have therefore been unable to consider variation in these regions. Core to the problem is the lack of a sequencing technology that produces reads with sufficient length and accuracy to enable unique mapping. Here, we present a novel methodology of using read clouds, obtained by accurate short-read sequencing of DNA derived from long fragment libraries, to confidently align short reads within repeat regions and enable accurate variant discovery. Our novel algorithm, Random Field Aligner (RFA), captures the relationships among the short reads governed by the long read process via a Markov Random Field. We utilized a modified version of the Illumina TruSeq synthetic long-read protocol, which yielded shallow-sequenced read clouds. We test RFA through extensive simulations and apply it to discover variants on the NA12878 human sample, for which shallow TruSeq read cloud sequencing data are available, and on an invasive breast carcinoma genome that we sequenced using the same method. We demonstrate that RFA facilitates accurate recovery of variation in 155 Mb of the human genome, including 94% of 67 Mb of segmental duplication sequence and 96% of 11 Mb of transcribed sequence, that are currently hidden from short-read technologies.

  View details for DOI 10.1101/gr.191189.115

  View details for PubMedID 26286554

  View details for PubMedCentralID PMC4579342

• Chromosomal copy number alterations for associations of ductal carcinoma in situ with invasive breast cancer BREAST CANCER RESEARCH Afghahi, A., Forgo, E., Mitani, A. A., Desai, M., Varma, S., Seto, T., Rigdon, J., Jensen, K. C., Troxell, M. L., Gomez, S. L., Das, A. K., Beck, A. H., Kurian, A. W., West, R. B. 2015; 17

  Abstract

  Screening mammography has contributed to a significant increase in the diagnosis of ductal carcinoma in situ (DCIS), raising concerns about overdiagnosis and overtreatment. Building on prior observations from lineage evolution analysis, we examined whether measuring genomic features of DCIS would predict association with invasive breast carcinoma (IBC). The long-term goal is to enhance standard clinicopathologic measures of low- versus high-risk DCIS and to enable risk-appropriate treatment.We studied three common chromosomal copy number alterations (CNA) in IBC and designed fluorescence in situ hybridization-based assay to measure copy number at these loci in DCIS samples. Clinicopathologic data were extracted from the electronic medical records of Stanford Cancer Institute and linked to demographic data from the population-based California Cancer Registry; results were integrated with data from tissue microarrays of specimens containing DCIS that did not develop IBC versus DCIS with concurrent IBC. Multivariable logistic regression analysis was performed to describe associations of CNAs with these two groups of DCIS.We examined 271 patients with DCIS (120 that did not develop IBC and 151 with concurrent IBC) for the presence of 1q, 8q24 and 11q13 copy number gains. Compared to DCIS-only patients, patients with concurrent IBC had higher frequencies of CNAs in their DCIS samples. On multivariable analysis with conventional clinicopathologic features, the copy number gains were significantly associated with concurrent IBC. The state of two of the three copy number gains in DCIS was associated with a risk of IBC that was 9.07 times that of no copy number gains, and the presence of gains at all three genomic loci in DCIS was associated with a more than 17-fold risk (P = 0.0013).CNAs have the potential to improve the identification of high-risk DCIS, defined by presence of concurrent IBC. Expanding and validating this approach in both additional cross-sectional and longitudinal cohorts may enable improved risk stratification and risk-appropriate treatment in DCIS.

  View details for DOI 10.1186/s13058-015-0623-y

  View details for Web of Science ID 000359348400001

  View details for PubMedID 26265211

  View details for PubMedCentralID PMC4534146

• Clinically Relevant Molecular Subtypes in Leiomyosarcoma. Clinical cancer research Guo, X., Jo, V. Y., Mills, A. M., Zhu, S. X., Lee, C., Espinosa, I., Nucci, M. R., Varma, S., Forgó, E., Hastie, T., Anderson, S., Ganjoo, K., Beck, A. H., West, R. B., Fletcher, C. D., van de Rijn, M. 2015; 21 (15): 3501-3511

  Abstract

  Leiomyosarcoma is a malignant neoplasm with smooth muscle differentiation. Little is known about its molecular heterogeneity and no targeted therapy currently exists for leiomyosarcoma. Recognition of different molecular subtypes is necessary to evaluate novel therapeutic options. In a previous study on 51 leiomyosarcomas, we identified three molecular subtypes in leiomyosarcoma. The current study was performed to determine whether the existence of these subtypes could be confirmed in independent cohorts.Ninety-nine cases of leiomyosarcoma were expression profiled with 3'end RNA-Sequencing (3SEQ). Consensus clustering was conducted to determine the optimal number of subtypes.We identified 3 leiomyosarcoma molecular subtypes and confirmed this finding by analyzing publically available data on 82 leiomyosarcoma from The Cancer Genome Atlas (TCGA). We identified two new formalin-fixed, paraffin-embedded tissue-compatible diagnostic immunohistochemical markers; LMOD1 for subtype I leiomyosarcoma and ARL4C for subtype II leiomyosarcoma. A leiomyosarcoma tissue microarray with known clinical outcome was used to show that subtype I leiomyosarcoma is associated with good outcome in extrauterine leiomyosarcoma while subtype II leiomyosarcoma is associated with poor prognosis in both uterine and extrauterine leiomyosarcoma. The leiomyosarcoma subtypes showed significant differences in expression levels for genes for which novel targeted therapies are being developed, suggesting that leiomyosarcoma subtypes may respond differentially to these targeted therapies.We confirm the existence of 3 molecular subtypes in leiomyosarcoma using two independent datasets and show that the different molecular subtypes are associated with distinct clinical outcomes. The findings offer an opportunity for treating leiomyosarcoma in a subtype-specific targeted approach. Clin Cancer Res; 21(15); 3501-11. ©2015 AACR.

  View details for DOI 10.1158/1078-0432.CCR-14-3141

  View details for PubMedID 25896974

• The prognostic landscape of genes and infiltrating immune cells across human cancers NATURE MEDICINE Gentles, A. J., Newman, A. M., Liu, C. L., Bratman, S. V., Feng, W., Kim, D., Nair, V. S., Xu, Y., Khuong, A., Hoang, C. D., Diehn, M., West, R. B., Plevritis, S. K., Alizadeh, A. A. 2015; 21 (8): 938-945

  Abstract

  Molecular profiles of tumors and tumor-associated cells hold great promise as biomarkers of clinical outcomes. However, existing data sets are fragmented and difficult to analyze systematically. Here we present a pan-cancer resource and meta-analysis of expression signatures from ?18,000 human tumors with overall survival outcomes across 39 malignancies. By using this resource, we identified a forkhead box MI (FOXM1) regulatory network as a major predictor of adverse outcomes, and we found that expression of favorably prognostic genes, including KLRB1 (encoding CD161), largely reflect tumor-associated leukocytes. By applying CIBERSORT, a computational approach for inferring leukocyte representation in bulk tumor transcriptomes, we identified complex associations between 22 distinct leukocyte subsets and cancer survival. For example, tumor-associated neutrophil and plasma cell signatures emerged as significant but opposite predictors of survival for diverse solid tumors, including breast and lung adenocarcinomas. This resource and associated analytical tools (http://precog.stanford.edu) may help delineate prognostic genes and leukocyte subsets within and across cancers, shed light on the impact of tumor heterogeneity on cancer outcomes, and facilitate the discovery of biomarkers and therapeutic targets.

  View details for DOI 10.1038/nm.3909

  View details for PubMedID 26193342

• Automated Analysis and Classification of Histological Tissue Features by Multi-Dimensional Microscopic Molecular Profiling PLOS ONE Riordan, D. P., Varma, S., West, R. B., Brown, P. O. 2015; 10 (7)

  Abstract

  Characterization of the molecular attributes and spatial arrangements of cells and features within complex human tissues provides a critical basis for understanding processes involved in development and disease. Moreover, the ability to automate steps in the analysis and interpretation of histological images that currently require manual inspection by pathologists could revolutionize medical diagnostics. Toward this end, we developed a new imaging approach called multidimensional microscopic molecular profiling (MMMP) that can measure several independent molecular properties in situ at subcellular resolution for the same tissue specimen. MMMP involves repeated cycles of antibody or histochemical staining, imaging, and signal removal, which ultimately can generate information analogous to a multidimensional flow cytometry analysis on intact tissue sections. We performed a MMMP analysis on a tissue microarray containing a diverse set of 102 human tissues using a panel of 15 informative antibody and 5 histochemical stains plus DAPI. Large-scale unsupervised analysis of MMMP data, and visualization of the resulting classifications, identified molecular profiles that were associated with functional tissue features. We then directly annotated H&E images from this MMMP series such that canonical histological features of interest (e.g. blood vessels, epithelium, red blood cells) were individually labeled. By integrating image annotation data, we identified molecular signatures that were associated with specific histological annotations and we developed statistical models for automatically classifying these features. The classification accuracy for automated histology labeling was objectively evaluated using a cross-validation strategy, and significant accuracy (with a median per-pixel rate of 77% per feature from 15 annotated samples) for de novo feature prediction was obtained. These results suggest that high-dimensional profiling may advance the development of computer-based systems for automatically parsing relevant histological and cellular features from molecular imaging data of arbitrary human tissue samples, and can provide a framework and resource to spur the optimization of these technologies.

  View details for DOI 10.1371/journal.pone.0128975

  View details for Web of Science ID 000358197600010

  View details for PubMedCentralID PMC4503351

• Cell-lineage heterogeneity and driver mutation recurrence in pre-invasive breast neoplasia GENOME MEDICINE Weng, Z., Spies, N., Zhu, S. X., Newburger, D. E., Kashef-Haghighi, D., Batzoglou, S., Sidow, A., West, R. B. 2015; 7

  Abstract

  All cells in an individual are related to one another by a bifurcating lineage tree, in which each node is an ancestral cell that divided into two, each branch connects two nodes, and the root is the zygote. When a somatic mutation occurs in an ancestral cell, all its descendants carry the mutation, which can then serve as a lineage marker for the phylogenetic reconstruction of tumor progression. Using this concept, we investigate cell lineage relationships and genetic heterogeneity of pre-invasive neoplasias compared to invasive carcinomas.We deeply sequenced over a thousand phylogenetically informative somatic variants in 66 morphologically independent samples from six patients that represent a spectrum of normal, early neoplasia, carcinoma in situ, and invasive carcinoma. For each patient, we obtained a highly resolved lineage tree that establishes the phylogenetic relationships among the pre-invasive lesions and with the invasive carcinoma.The trees reveal lineage heterogeneity of pre-invasive lesions, both within the same lesion, and between histologically similar ones. On the basis of the lineage trees, we identified a large number of independent recurrences of PIK3CA H1047 mutations in separate lesions in four of the six patients, often separate from the diagnostic carcinoma.Our analyses demonstrate that multi-sample phylogenetic inference provides insights on the origin of driver mutations, lineage heterogeneity of neoplastic proliferations, and the relationship of genomically aberrant neoplasias with the primary tumors. PIK3CA driver mutations may be comparatively benign inducers of cellular proliferation.

  View details for DOI 10.1186/s13073-015-0146-2

  View details for Web of Science ID 000353532600001

  View details for PubMedID 25918554

  View details for PubMedCentralID PMC4410742

• Diffuse High Intensity PD-L1 Staining in Thymic Epithelial Tumors. Journal of thoracic oncology Padda, S. K., Riess, J. W., Schwartz, E. J., Tian, L., Kohrt, H. E., Neal, J. W., West, R. B., Wakelee, H. A. 2015; 10 (3): 500-508

  Abstract

  Blockade of the immune checkpoint programmed death receptor ligand-1 (PD-L1)/PD-1 pathway has well-established clinical activity across many tumor types. PD-L1 protein expression by immunohistochemistry is emerging as a predictive biomarker of response to these therapies. Here, we examine PD-L1 expression in a thymic epithelial tumor (TET) tissue microarray (TMA).The TMA contained 69 TETs and 17 thymic controls, with each case represented by triplicate cores. The TMA was stained with rabbit monoclonal antibody (clone 15; Sino Biological, Beijing, China) to human PD-L1. PD-L1 staining was scored based on intensity as follows: 0 = none, 1 = equivocal/uninterpretable, 2 = weak, and 3 = intermediate-strong. Those cases with all cores scoring three in the epithelial component were categorized as PD-L1 and the remaining as PD-L1.PD-L1 scores were more frequent in TETs than in controls (68.1% versus 17.6%; p = 0.0036). PD-L1 scores and histology were significantly correlated, with higher intensity staining in World Health Organization (WHO). B2/B3/C TETs. Only 14.8% of TETs had PD-L1 staining of associated lymphocytes. In an adjusted analysis (age/sex), PD-L1 TETs had a significantly worse overall survival (hazard ratio: 5.40, 95% confidence interval: 1.13-25.89; p = 0.035) and a trend for worse event-free survival (hazard ratio: 2.94, 95% confidence interval: 0.94-9.24; p = 0.064).PD-L1 expression was present in all cases of TETs within the epithelial component but only in a minority in the lymphocytic component. TETs stained more intensely for PD-L1 than in controls, and PD-L1 TETs were associated with more aggressive histology and worse prognosis. This study lends rationale to a clinical trial with anti-PD-1/PD-L1 therapy in this rare tumor type.

  View details for DOI 10.1097/JTO.0000000000000429

  View details for PubMedID 25402569

• Novel mutations in neuroendocrine carcinoma of the breast: possible therapeutic targets. Applied immunohistochemistry & molecular morphology Ang, D., Ballard, M., Beadling, C., Warrick, A., Schilling, A., O'Gara, R., Pukay, M., Neff, T. L., West, R. B., Corless, C. L., Troxell, M. L. 2015; 23 (2): 97-103

  Abstract

  Primary neuroendocrine carcinoma of the breast is a rare variant, accounting for only 2% to 5% of diagnosed breast cancers, and may have relatively aggressive behavior. Mutational profiling of invasive ductal breast cancers has yielded potential targets for directed cancer therapy, yet most studies have not included neuroendocrine carcinomas. In a tissue microarray screen, we found a 2.4% prevalence (9/372) of neuroendocrine breast carcinoma, including several with lobular morphology. We then screened primary or metastatic neuroendocrine breast carcinomas (excluding papillary and mucinous) for mutations in common cancer genes using polymerase chain reaction-mass spectroscopy (643 hotspot mutations across 53 genes), or semiconductor-based next-generation sequencing analysis (37 genes). Mutations were identified in 5 of 15 tumors, including 3 with PIK3CA exon 9 E542K mutations, 2 of which also harbored point mutations in FGFR family members (FGFR1 P126S, FGFR4 V550M). Single mutations were found in each of KDR (A1065T) and HRAS (G12A). PIK3CA mutations are common in other types of breast carcinoma. However, FGFR and RAS family mutations are exceedingly rare in the breast cancer literature. Likewise, activating mutations in the receptor tyrosine kinase KDR (VEGFR2) have been reported in angiosarcomas and non-small cell lung cancers; the KDR A1065T mutation is reported to be sensitive to VEGFR kinase inhibitors, and fibroblast growth factor receptor inhibitors are in trials. Our findings demonstrate the utility of broad-based genotyping in the study of rare tumors such as neuroendocrine breast cancer.

  View details for DOI 10.1097/PDM.0b013e3182a40fd1

  View details for PubMedID 25679062

• Translocation and Expression of MSANTD3 in Neoplasia and Normal Tissue Barasch, N., Xue, G., Sweeney, R., Varma, S., Biscocho, J., Kwei, K., Pelham, R., Lipsick, J., West, R., Pollack, J. NATURE PUBLISHING GROUP. 2015: 320A

  View details for Web of Science ID 000348948002299

• Translocation and Expression of MSANTD3 in Neoplasia and Normal Tissue Barasch, N., Xue, G., Sweeney, R., Varma, S., Biscocho, J., Kwei, K., Pelham, R., Lipsick, J., West, R., Pollack, J. NATURE PUBLISHING GROUP. 2015: 320A

  View details for Web of Science ID 000349502201618

• Survey of SMO and BRAF Mutational Status in Head and Neck and Adnexal Neoplasms McClary, A., Biscocho, J., Varma, S., Pollack, J., West, R. NATURE PUBLISHING GROUP. 2015: 327A?328A

  View details for Web of Science ID 000348948002332

• Correlation of Protein Expression With Chromosomal Copy Number Alterations for Risk Classification of Ductal Carcinoma In Situ Forgo, E., Varma, S., West, R. NATURE PUBLISHING GROUP. 2015: 45A

  View details for Web of Science ID 000348948000169

• Characterization of Breast Cancers With Equivocal and Non-Classical HER2 FISH Results - A Multi-Institutional Study Ballard, M., Jalikis, F., Krings, G., Schmidt, R., Chen, Y., Rendi, M., Dintzis, S. M., Jensen, K., West, R., Sibley, R., Allison, K. NATURE PUBLISHING GROUP. 2015: 35A

  View details for Web of Science ID 000349502200129

• Genomic Profiling of Radial Scar Lin, C., Sweeney, R., Zhu, S., West, R. NATURE PUBLISHING GROUP. 2015: 54A

  View details for Web of Science ID 000348948000204

• Correlation of Protein Expression With Chromosomal Copy Number Alterations for Risk Classification of Ductal Carcinoma In Situ Forgo, E., Varma, S., West, R. NATURE PUBLISHING GROUP. 2015: 45A

  View details for Web of Science ID 000349502200169

• Genomic Profiling of Radial Scar Lin, C., Sweeney, R., Zhu, S., West, R. NATURE PUBLISHING GROUP. 2015: 54A

  View details for Web of Science ID 000349502200204

• Survey of SMO and BRAF Mutational Status in Head and Neck and Adnexal Neoplasms McClaty, A., Biscocho, J., Varma, S., Pollacic, J., West, R. NATURE PUBLISHING GROUP. 2015: 327A?328A

  View details for Web of Science ID 000349502202023

• GLI1, CTNNB1 and NOTCH1 Protein Expression in a Thymic Epithelial Malignancy Tissue Microarray. Anticancer research Riess, J. W., West, R., Dean, M., Klimowicz, A. C., Neal, J. W., Hoang, C., Wakelee, H. A. 2015; 35 (2): 669-676

  Abstract

  Thymic epithelial tumors (TET) are rare. Wingless and INT (WNT), NOTCH and sonic hedgehog pathway interactions between thymocytes and thymic stroma are important to thymus and T-cell development. We analyzed a thymoma tissue microarray (TMA) for glioma associated oncogene homolog 1 (GLI1), NOTCH1 and catenin (cadherin-associated protein, beta 1) (CTNNB1) expression as surrogate markers of sonic hedgehog, NOTCH and WNT pathway activity.GLI1, NOTCH1 and CTNNB1 expression were assayed in a tissue microarray of 68 TET and eight benign thymus by fluorescent immunohistochemistry (AQUA) as surrogates for activity of the sonic hedgehog, NOTCH and WNT pathways respectively.No difference in tumor GLI1 (mean 201 vs. 211, p=0.31), CTNNB1 (mean 222 vs. 306, p=0.66) or NOTCH1 expression (mean 317 vs. 325, p=0.82) was noted between thymic tumor and benign thymus.No evidence for preferential expression of GLI1, NOTCH1 or CTNNB1 was noted. High-throughput immunofluorescence using AQUA technology can help overcome limitations of small sample size and tissue heterogeneity when analyzing protein expression in thymic tumors.

  View details for PubMedID 25667444

• Novel mutations in neuroendocrine carcinoma of the breast: possible therapeutic targets. Applied immunohistochemistry & molecular morphology Ang, D., Ballard, M., Beadling, C., Warrick, A., Schilling, A., O'Gara, R., Pukay, M., Neff, T. L., West, R. B., Corless, C. L., Troxell, M. L. 2015; 23 (2): 97-103

  Abstract

  Primary neuroendocrine carcinoma of the breast is a rare variant, accounting for only 2% to 5% of diagnosed breast cancers, and may have relatively aggressive behavior. Mutational profiling of invasive ductal breast cancers has yielded potential targets for directed cancer therapy, yet most studies have not included neuroendocrine carcinomas. In a tissue microarray screen, we found a 2.4% prevalence (9/372) of neuroendocrine breast carcinoma, including several with lobular morphology. We then screened primary or metastatic neuroendocrine breast carcinomas (excluding papillary and mucinous) for mutations in common cancer genes using polymerase chain reaction-mass spectroscopy (643 hotspot mutations across 53 genes), or semiconductor-based next-generation sequencing analysis (37 genes). Mutations were identified in 5 of 15 tumors, including 3 with PIK3CA exon 9 E542K mutations, 2 of which also harbored point mutations in FGFR family members (FGFR1 P126S, FGFR4 V550M). Single mutations were found in each of KDR (A1065T) and HRAS (G12A). PIK3CA mutations are common in other types of breast carcinoma. However, FGFR and RAS family mutations are exceedingly rare in the breast cancer literature. Likewise, activating mutations in the receptor tyrosine kinase KDR (VEGFR2) have been reported in angiosarcomas and non-small cell lung cancers; the KDR A1065T mutation is reported to be sensitive to VEGFR kinase inhibitors, and fibroblast growth factor receptor inhibitors are in trials. Our findings demonstrate the utility of broad-based genotyping in the study of rare tumors such as neuroendocrine breast cancer.

  View details for DOI 10.1097/PDM.0b013e3182a40fd1

  View details for PubMedID 25679062

• GLI1, CTNNB1 and NOTCH1 protein expression in a thymic epithelial malignancy tissue microarray. Anticancer research Riess, J. W., West, R., Dean, M., Klimowicz, A. C., Neal, J. W., Hoang, C., Wakelee, H. A. 2015; 35 (2): 669-676

  Abstract

  Thymic epithelial tumors (TET) are rare. Wingless and INT (WNT), NOTCH and sonic hedgehog pathway interactions between thymocytes and thymic stroma are important to thymus and T-cell development. We analyzed a thymoma tissue microarray (TMA) for glioma associated oncogene homolog 1 (GLI1), NOTCH1 and catenin (cadherin-associated protein, beta 1) (CTNNB1) expression as surrogate markers of sonic hedgehog, NOTCH and WNT pathway activity.GLI1, NOTCH1 and CTNNB1 expression were assayed in a tissue microarray of 68 TET and eight benign thymus by fluorescent immunohistochemistry (AQUA) as surrogates for activity of the sonic hedgehog, NOTCH and WNT pathways respectively.No difference in tumor GLI1 (mean 201 vs. 211, p=0.31), CTNNB1 (mean 222 vs. 306, p=0.66) or NOTCH1 expression (mean 317 vs. 325, p=0.82) was noted between thymic tumor and benign thymus.No evidence for preferential expression of GLI1, NOTCH1 or CTNNB1 was noted. High-throughput immunofluorescence using AQUA technology can help overcome limitations of small sample size and tissue heterogeneity when analyzing protein expression in thymic tumors.

  View details for PubMedID 25667444

• Pemetrexed in patients with thymic malignancies previously treated with chemotherapy. Lung cancer Liang, Y., Padda, S. K., Riess, J. W., West, R. B., Neal, J. W., Wakelee, H. A. 2015; 87 (1): 34-38

  Abstract

  Thymic malignancies are rare, with limited published trials of chemotherapy activity. We performed a retrospective analysis of pemetrexed activity in patients with thymic malignancies.Patients with unresectable histologically confirmed invasive, recurrent, or metastatic thymoma or thymic carcinoma seen at the Stanford Cancer Center between January 2005 and November 2013 were identified, and those who were treated with pemetrexed in the second-line setting and beyond were included in this analysis.A total of 81 thymic malignancy patients were identified, of whom 16 received pemetrexed alone (N=14) or in combination (N=2). There were 10 patients (62.5%) with thymic carcinoma and 6 patients (37.5%) with thymoma. Among the 6 patients with thymoma, best response was 1 (17%) with a partial response (PR) and 5 (83%) with stable disease (SD). At a median follow-up of 21.2 months, the median PFS in the thymoma patients was 13.8 months (95% CI, 4.9-22.6 months) and the median OS was 20.1 months (95% CI, 16.4-23.9 months). Among the 10 patients with thymic carcinoma, best response to treatment was 1 (10%) PR, 5 (50%) SD, and 4 (40%) progressive disease (PD). At a median follow-up of 13.5 months, the median PFS in patients with thymic carcinoma was 6.5 months (95% CI, 0.2-12.8 months) and the median OS was 12.7 months (95% CI, 2.9-22.5 months).This small retrospective study demonstrates modest pemetrexed activity and disease stabilization in thymic malignancies with a clinically meaningful duration, and supports previous reports of pemetrexed efficacy in these rare diseases.

  View details for DOI 10.1016/j.lungcan.2014.11.006

  View details for PubMedID 25443273

• Chromosomal copy number alterations for associations of ductal carcinoma in situ with invasive breast cancer. Breast cancer research Afghahi, A., Forgó, E., Mitani, A. A., Desai, M., Varma, S., Seto, T., Rigdon, J., Jensen, K. C., Troxell, M. L., Gomez, S. L., Das, A. K., Beck, A. H., Kurian, A. W., West, R. B. 2015; 17: 108-?

  Abstract

  Screening mammography has contributed to a significant increase in the diagnosis of ductal carcinoma in situ (DCIS), raising concerns about overdiagnosis and overtreatment. Building on prior observations from lineage evolution analysis, we examined whether measuring genomic features of DCIS would predict association with invasive breast carcinoma (IBC). The long-term goal is to enhance standard clinicopathologic measures of low- versus high-risk DCIS and to enable risk-appropriate treatment.We studied three common chromosomal copy number alterations (CNA) in IBC and designed fluorescence in situ hybridization-based assay to measure copy number at these loci in DCIS samples. Clinicopathologic data were extracted from the electronic medical records of Stanford Cancer Institute and linked to demographic data from the population-based California Cancer Registry; results were integrated with data from tissue microarrays of specimens containing DCIS that did not develop IBC versus DCIS with concurrent IBC. Multivariable logistic regression analysis was performed to describe associations of CNAs with these two groups of DCIS.We examined 271 patients with DCIS (120 that did not develop IBC and 151 with concurrent IBC) for the presence of 1q, 8q24 and 11q13 copy number gains. Compared to DCIS-only patients, patients with concurrent IBC had higher frequencies of CNAs in their DCIS samples. On multivariable analysis with conventional clinicopathologic features, the copy number gains were significantly associated with concurrent IBC. The state of two of the three copy number gains in DCIS was associated with a risk of IBC that was 9.07 times that of no copy number gains, and the presence of gains at all three genomic loci in DCIS was associated with a more than 17-fold risk (P = 0.0013).CNAs have the potential to improve the identification of high-risk DCIS, defined by presence of concurrent IBC. Expanding and validating this approach in both additional cross-sectional and longitudinal cohorts may enable improved risk stratification and risk-appropriate treatment in DCIS.

  View details for DOI 10.1186/s13058-015-0623-y

  View details for PubMedID 26265211

• Read Clouds Uncover Variation in Complex Regions of the Human Genome Bishara, A., Liu, Y., Kashef-Haghighi, D., Weng, Z., Newburger, D. E., West, R., Sidow, A., Batzoglou, S., Przytycka, T. M. SPRINGER-VERLAG BERLIN. 2015: 30?31

  View details for DOI 10.1007/978-3-319-16706-0_5

  View details for Web of Science ID 000361983900005

• Cell-lineage heterogeneity and driver mutation recurrence in pre-invasive breast neoplasia. Genome medicine Weng, Z., Spies, N., Zhu, S. X., Newburger, D. E., Kashef-Haghighi, D., Batzoglou, S., Sidow, A., West, R. B. 2015; 7 (1): 28-?

  Abstract

  All cells in an individual are related to one another by a bifurcating lineage tree, in which each node is an ancestral cell that divided into two, each branch connects two nodes, and the root is the zygote. When a somatic mutation occurs in an ancestral cell, all its descendants carry the mutation, which can then serve as a lineage marker for the phylogenetic reconstruction of tumor progression. Using this concept, we investigate cell lineage relationships and genetic heterogeneity of pre-invasive neoplasias compared to invasive carcinomas.We deeply sequenced over a thousand phylogenetically informative somatic variants in 66 morphologically independent samples from six patients that represent a spectrum of normal, early neoplasia, carcinoma in situ, and invasive carcinoma. For each patient, we obtained a highly resolved lineage tree that establishes the phylogenetic relationships among the pre-invasive lesions and with the invasive carcinoma.The trees reveal lineage heterogeneity of pre-invasive lesions, both within the same lesion, and between histologically similar ones. On the basis of the lineage trees, we identified a large number of independent recurrences of PIK3CA H1047 mutations in separate lesions in four of the six patients, often separate from the diagnostic carcinoma.Our analyses demonstrate that multi-sample phylogenetic inference provides insights on the origin of driver mutations, lineage heterogeneity of neoplastic proliferations, and the relationship of genomically aberrant neoplasias with the primary tumors. PIK3CA driver mutations may be comparatively benign inducers of cellular proliferation.

  View details for DOI 10.1186/s13073-015-0146-2

  View details for PubMedID 25918554

  View details for PubMedCentralID PMC4410742

• Fast and scalable inference of multi-sample cancer lineages. Genome biology Popic, V., Salari, R., Hajirasouliha, I., Kashef-Haghighi, D., West, R. B., Batzoglou, S. 2015; 16 (1): 91-?

  Abstract

  Somatic variants can be used as lineage markers for the phylogenetic reconstruction of cancer evolution. Since somatic phylogenetics is complicated by sample heterogeneity, novel specialized tree-building methods are required for cancer phylogeny reconstruction. We present LICHeE (Lineage Inference for Cancer Heterogeneity and Evolution), a novel method that automates the phylogenetic inference of cancer progression from multiple somatic samples. LICHeE uses variant allele frequencies of somatic single nucleotide variants obtained by deep sequencing to reconstruct multi-sample cell lineage trees and infer the subclonal composition of the samples. LICHeE is open source and available at http://viq854.github.io/lichee .

  View details for DOI 10.1186/s13059-015-0647-8

  View details for PubMedID 25944252

  View details for PubMedCentralID PMC4501097

• Automated Analysis and Classification of Histological Tissue Features by Multi-Dimensional Microscopic Molecular Profiling. PloS one Riordan, D. P., Varma, S., West, R. B., Brown, P. O. 2015; 10 (7)

  Abstract

  Characterization of the molecular attributes and spatial arrangements of cells and features within complex human tissues provides a critical basis for understanding processes involved in development and disease. Moreover, the ability to automate steps in the analysis and interpretation of histological images that currently require manual inspection by pathologists could revolutionize medical diagnostics. Toward this end, we developed a new imaging approach called multidimensional microscopic molecular profiling (MMMP) that can measure several independent molecular properties in situ at subcellular resolution for the same tissue specimen. MMMP involves repeated cycles of antibody or histochemical staining, imaging, and signal removal, which ultimately can generate information analogous to a multidimensional flow cytometry analysis on intact tissue sections. We performed a MMMP analysis on a tissue microarray containing a diverse set of 102 human tissues using a panel of 15 informative antibody and 5 histochemical stains plus DAPI. Large-scale unsupervised analysis of MMMP data, and visualization of the resulting classifications, identified molecular profiles that were associated with functional tissue features. We then directly annotated H&E images from this MMMP series such that canonical histological features of interest (e.g. blood vessels, epithelium, red blood cells) were individually labeled. By integrating image annotation data, we identified molecular signatures that were associated with specific histological annotations and we developed statistical models for automatically classifying these features. The classification accuracy for automated histology labeling was objectively evaluated using a cross-validation strategy, and significant accuracy (with a median per-pixel rate of 77% per feature from 15 annotated samples) for de novo feature prediction was obtained. These results suggest that high-dimensional profiling may advance the development of computer-based systems for automatically parsing relevant histological and cellular features from molecular imaging data of arbitrary human tissue samples, and can provide a framework and resource to spur the optimization of these technologies.

  View details for DOI 10.1371/journal.pone.0128975

  View details for PubMedID 26176839

  View details for PubMedCentralID PMC4503351

• Effects of Thrombopoietin Mimetics on Patients with Chronic ITP: Perspectives from Blood Transcriptome Profiling Analysis Zhang, H., Zhang, B., Guo, X., Haq, N., West, R. B., Bussel, J. B., Zehnder, J. L. AMER SOC HEMATOLOGY. 2014

  View details for Web of Science ID 000349233800107

• Long noncoding RNA EWSAT1-mediated gene repression facilitates Ewing sarcoma oncogenesis JOURNAL OF CLINICAL INVESTIGATION Howarth, M. M., Simpson, D., Ngok, S. P., Nieves, B., Chen, R., Siprashvili, Z., Vaka, D., Breese, M. R., Crompton, B. D., Alexe, G., Hawkins, D. S., Jacobson, D., Brunner, A. L., West, R., Mora, J., Stegmaier, K., Khavari, P., Sweet-Cordero, E. A. 2014; 124 (12): 5275-5290

  Abstract

  Chromosomal translocation that results in fusion of the genes encoding RNA-binding protein EWS and transcription factor FLI1 (EWS-FLI1) is pathognomonic for Ewing sarcoma. EWS-FLI1 alters gene expression through mechanisms that are not completely understood. We performed RNA sequencing (RNAseq) analysis on primary pediatric human mesenchymal progenitor cells (pMPCs) expressing EWS-FLI1 in order to identify gene targets of this oncoprotein. We determined that long noncoding RNA-277 (Ewing sarcoma-associated transcript 1 [EWSAT1]) is upregulated by EWS-FLI1 in pMPCs. Inhibition of EWSAT1 expression diminished the ability of Ewing sarcoma cell lines to proliferate and form colonies in soft agar, whereas EWSAT1 inhibition had no effect on other cell types tested. Expression of EWS-FLI1 and EWSAT1 repressed gene expression, and a substantial fraction of targets that were repressed by EWS-FLI1 were also repressed by EWSAT1. Analysis of RNAseq data from primary human Ewing sarcoma further supported a role for EWSAT1 in mediating gene repression. We identified heterogeneous nuclear ribonucleoprotein (HNRNPK) as an RNA-binding protein that interacts with EWSAT1 and found a marked overlap in HNRNPK-repressed genes and those repressed by EWS-FLI1 and EWSAT1, suggesting that HNRNPK participates in EWSAT1-mediated gene repression. Together, our data reveal that EWSAT1 is a downstream target of EWS-FLI1 that facilitates the development of Ewing sarcoma via the repression of target genes.

  View details for DOI 10.1172/JC172124

  View details for Web of Science ID 000345677200020

  View details for PubMedID 25401475

• Ameloblastoma driver mutations revealed by next-generation sequencing of formalin-fixed paraffin-embedded specimens McClary, A. C., Sweeney, R. T., Biscocho, J., Myers, B. R., Neahring, L., Kwei, K. A., Qu, K., Gong, X., Ng, T., Jones, C. D., Varma, S., Odegaard, J. I., Rubin, B., Troxell, M. L., Pelham, R. J., Zehnder, J. L., Beachy, P. A., Pollack, J. R., West, R. B. AMER ASSOC CANCER RESEARCH. 2014

  View details for DOI 10.1158/1538-7445.AM2014-3436

  View details for Web of Science ID 000349910200435

• PEMETREXED IN PATIENTS WITH THYMIC MALIGNANCIES PREVIOUSLY TREATED WITH CHEMOTHERAPY Liang, Y., Padda, S., Riess, J. W., West, R., Neal, J. W., Wakelee, H. A. LIPPINCOTT WILLIAMS & WILKINS. 2014: S226

  View details for Web of Science ID 000344627500020

• PROGRAMMED DEATH RECEPTOR LIGAND-1 (PD-L1) EXPRESSION IN A THYMOMA TISSUE MICROARRAY (TMA) Padda, S., Riess, J. W., Schwartz, E., Tian, L., Kohrt, H., Neal, J. W., West, R., Wakelee, H. A. LIPPINCOTT WILLIAMS & WILKINS. 2014: S212

  View details for Web of Science ID 000344627500002

• Fingerprints of Epstein-Barr virus in nasopharyngeal carcinoma NATURE GENETICS West, R. B. 2014; 46 (8): 809-810

  Abstract

  The influence of Epstein-Barr virus (EBV) on cancer is not well understood. High-throughput sequencing of nasopharyngeal carcinoma (NPC) illustrates the influences of EBV on oncogenesis and identifies driver pathways that might be therapeutically useful for NPC treatment.

  View details for Web of Science ID 000339704400006

  View details for PubMedID 25070800

• Identification of recurrent SMO and BRAF mutations in ameloblastomas NATURE GENETICS Sweeney, R. T., McClary, A. C., Myers, B. R., Biscocho, J., Neahring, L., Kwei, K. A., Qu, K., Gong, X., Ng, T., Jones, C. D., Varma, S., Odegaard, J. I., Sugiyama, T., Koyota, S., Rubin, B. P., Troxell, M. L., Pelham, R. J., Zehnder, J. L., Beachy, P. A., Pollack, J. R., West, R. B. 2014; 46 (7): 722-725

  Abstract

  Here we report the discovery of oncogenic mutations in the Hedgehog and mitogen-activated protein kinase (MAPK) pathways in over 80% of ameloblastomas, locally destructive odontogenic tumors of the jaw, by genomic analysis of archival material. Mutations in SMO (encoding Smoothened, SMO) are common in ameloblastomas of the maxilla, whereas BRAF mutations are predominant in tumors of the mandible. We show that a frequently occurring SMO alteration encoding p.Leu412Phe is an activating mutation and that its effect on Hedgehog-pathway activity can be inhibited by arsenic trioxide (ATO), an anti-leukemia drug approved by the US Food and Drug Administration (FDA) that is currently in clinical trials for its Hedgehog-inhibitory activity. In a similar manner, ameloblastoma cells harboring an activating BRAF mutation encoding p.Val600Glu are sensitive to the BRAF inhibitor vemurafenib. Our findings establish a new paradigm for the diagnostic classification and treatment of ameloblastomas.

  View details for DOI 10.1038/ng.2986

  View details for Web of Science ID 000338093800013

• Identification of recurrent SMO and BRAF mutations in ameloblastomas. Nature genetics Sweeney, R. T., McClary, A. C., Myers, B. R., Biscocho, J., Neahring, L., Kwei, K. A., Qu, K., Gong, X., Ng, T., Jones, C. D., Varma, S., Odegaard, J. I., Sugiyama, T., Koyota, S., Rubin, B. P., Troxell, M. L., Pelham, R. J., Zehnder, J. L., Beachy, P. A., Pollack, J. R., West, R. B. 2014; 46 (7): 722-725

  Abstract

  Here we report the discovery of oncogenic mutations in the Hedgehog and mitogen-activated protein kinase (MAPK) pathways in over 80% of ameloblastomas, locally destructive odontogenic tumors of the jaw, by genomic analysis of archival material. Mutations in SMO (encoding Smoothened, SMO) are common in ameloblastomas of the maxilla, whereas BRAF mutations are predominant in tumors of the mandible. We show that a frequently occurring SMO alteration encoding p.Leu412Phe is an activating mutation and that its effect on Hedgehog-pathway activity can be inhibited by arsenic trioxide (ATO), an anti-leukemia drug approved by the US Food and Drug Administration (FDA) that is currently in clinical trials for its Hedgehog-inhibitory activity. In a similar manner, ameloblastoma cells harboring an activating BRAF mutation encoding p.Val600Glu are sensitive to the BRAF inhibitor vemurafenib. Our findings establish a new paradigm for the diagnostic classification and treatment of ameloblastomas.

  View details for DOI 10.1038/ng.2986

  View details for PubMedID 24859340

• Programmed death receptor ligand-1 (PD-L1) expression in a thymoma (T) tissue microarray (TMA) Padda, S., Riess, J., Schwartz, E., Tian, L., Kohrt, H., Neal, J. W., Natkunam, Y., West, R. B., Wakelee, H. A. AMER SOC CLINICAL ONCOLOGY. 2014

  View details for Web of Science ID 000358613204064

• Chromosomal copy number alterations (CNAs) for risk assessment of ductal carcinoma in situ (DCIS) Afghahi, A., Forgo, E., Mitani, A., Desai, M., Varma, S., Seto, T., Jensen, K. C., Gomez, S., Das, A. K., Beck, A. H., Kurian, A. W., West, R. B. AMER SOC CLINICAL ONCOLOGY. 2014

  View details for Web of Science ID 000358613202339

• Human papillomavirus 16 detected in nasopharyngeal carcinomas in white Americans but not in endemic Southern Chinese patients. Head & neck Lin, Z., Khong, B., Kwok, S., Cao, H., West, R. B., Le, Q., Kong, C. S. 2014; 36 (5): 709-714

  Abstract

  BACKGROUND: We evaluated the relationship of HPV and EBV with race in endemic and non-endemic cohorts of patients with nasopharyngeal carcinoma (NPC), and with smoking status in the non-endemic cohort. METHODS: Tissue microarrays (TMA) were constructed using samples from 86 patients treated in southern China and 108 patients from Stanford. TMAs were stained with p16, HPV ISH, and EBV ISH. PCR was used to confirm EBV(-) cases and HPV status in p16(+) cases. Survival data was available for the Stanford cohort only. RESULTS: No HPV(+) cases were detected in the Chinese cohort. In the Stanford cohort, 5/11 EBV(-) cases harbored HPV16, 10/10 occurred in Caucasians, and 8/11 were smokers. Patients with EBV(-) NPC also showed a trend towards worse survival. CONCLUSIONS: EBV(-) NPC shows an association with the presence of HPV, Caucasian race, and smoking. In contrast, EBV(-) NPC shows no association with HPV in the endemic cohort. Head Neck, 2013.

  View details for DOI 10.1002/hed.23362

  View details for PubMedID 23616441

• Stromal signatures in endometrioid endometrial carcinomas MODERN PATHOLOGY Espinosa, I., Catasus, L., D'angelo, E., Mozos, A., Pedrola, N., Bertolo, C., Ferrer, I., Zannoni, G. F., West, R. B., van de Rijn, M., Matias-Guiu, X., Prat, J. 2014; 27 (4): 631-639

  Abstract

  The pattern of myometrial invasion in endometrioid endometrial carcinomas varies considerably; ie, from widely scattered glands and cell nests, often associated with a fibromyxoid stromal reaction (desmoplasia) and/or a lymphocytic infiltrate, to invasive glands with little or no stromal response. Recently, two distinct stromal signatures derived from a macrophage response (colony-stimulating factor 1, CSF1) and a fibroblastic response (desmoid-type fibromatosis, DTF) were identified in breast carcinomas and correlated with clinicopathologic features including outcome. In this study, we explored whether these stromal signatures also apply to endometrioid carcinomas and how their expression patterns correlated with morphologic changes. We studied the stromal signatures both by immunohistochemistry and in situ hybridization in 98 primary endometrioid carcinomas with (87 cases) and without (11 cases) myometrial invasion as well as in the corresponding regional lymph nodes metatases of 9 myoinvasive tumors. Desmoplasia correlated positively with the DTF expression signature. Likewise, mononuclear infiltrates were found in the stroma of tumors expressing CSF1. Twenty-four out of eighty-seven (27%) myoinvasive endometrioid carcinomas were positive for the macrophage signature and thirteen out of eighty-seven (15%) expressed the fibroblast signature. Eleven additional cases were positive for both DTF and CSF1 signatures (11/87; 13%). However, over half of the cases (39/87; 45%) and the majority of the non-myoinvasive tumors (8/11; 73%) failed to express any of the two stromal signatures. The macrophage response (CSF1) was associated with higher tumor grade, lymphovascular invasion, and PIK3CA mutations (P<0.05). There was a concordance in the expression of the CSF1 signature in the primary tumors and their corresponding lymph node metastases. This study is the first characterization of stromal signatures in endometrioid carcinomas. Our findings shed new light on the relationship between genetically different endometrioid carcinomas and various stromal responses. Preservation of the CSF1 macrophage stromal response in the metastases leds support to targeting the CSF1 pathway in endometrioid endometrial carcinomas.

  View details for DOI 10.1038/modpathol.2013.131

  View details for PubMedID 24263966

• Stromal responses among carcinomas--response. Clinical cancer research West, R. B., van de Rijn, M., Chen, J. L. 2014; 20 (5): 1397-?

  View details for DOI 10.1158/1078-0432.CCR-13-3238

  View details for PubMedID 24590889

• Novel Mutations in Neuroendocrine Carcinoma of the Breast: Possible Therapeutic Targets. Diagnostic molecular pathology Ang, D., Ballard, M., Beadling, C., Warrick, A., Schilling, A., O'Gara, R., Pukay, M., Neff, T. L., West, R. B., Corless, C. L., Troxell, M. L. 2014: -?

  Abstract

  Primary neuroendocrine carcinoma of the breast is a rare variant, accounting for only 2% to 5% of diagnosed breast cancers, and may have relatively aggressive behavior. Mutational profiling of invasive ductal breast cancers has yielded potential targets for directed cancer therapy, yet most studies have not included neuroendocrine carcinomas. In a tissue microarray screen, we found a 2.4% prevalence (9/372) of neuroendocrine breast carcinoma, including several with lobular morphology. We then screened primary or metastatic neuroendocrine breast carcinomas (excluding papillary and mucinous) for mutations in common cancer genes using polymerase chain reaction-mass spectroscopy (643 hotspot mutations across 53 genes), or semiconductor-based next-generation sequencing analysis (37 genes). Mutations were identified in 5 of 15 tumors, including 3 with PIK3CA exon 9 E542K mutations, 2 of which also harbored point mutations in FGFR family members (FGFR1 P126S, FGFR4 V550M). Single mutations were found in each of KDR (A1065T) and HRAS (G12A). PIK3CA mutations are common in other types of breast carcinoma. However, FGFR and RAS family mutations are exceedingly rare in the breast cancer literature. Likewise, activating mutations in the receptor tyrosine kinase KDR (VEGFR2) have been reported in angiosarcomas and non-small cell lung cancers; the KDR A1065T mutation is reported to be sensitive to VEGFR kinase inhibitors, and fibroblast growth factor receptor inhibitors are in trials. Our findings demonstrate the utility of broad-based genotyping in the study of rare tumors such as neuroendocrine breast cancer.

  View details for PubMedID 24569783

• NF-?B protein expression associates with (18)F-FDG PET tumor uptake in non-small cell lung cancer: A radiogenomics validation study to understand tumor metabolism. Lung cancer Nair, V. S., Gevaert, O., Davidzon, G., Plevritis, S. K., West, R. 2014; 83 (2): 189-196

  Abstract

  We previously demonstrated that NF-?B may be associated with (18)F-FDG PET uptake and patient prognosis using radiogenomics in patients with non-small cell lung cancer (NSCLC). To validate these results, we assessed NF-?B protein expression in an extended cohort of NSCLC patients.We examined NF-?Bp65 by immunohistochemistry (IHC) using a Tissue Microarray. Staining intensity was assessed by qualitative ordinal scoring and compared to tumor FDG uptake (SUVmax and SUVmean), lactate dehydrogenase A (LDHA) expression (as a positive control) and outcome using ANOVA, Kaplan Meier (KM), and Cox-proportional hazards (CPH) analysis.365 tumors from 355 patients with long-term follow-up were analyzed. The average age for patients was 67±11 years, 46% were male and 67% were ever smokers. Stage I and II patients comprised 83% of the cohort and the majority had adenocarcinoma (73%). From 88 FDG PET scans available, average SUVmax and SUVmean were 8.3±6.6, and 3.7±2.4 respectively. Increasing NF-?Bp65 expression, but not LDHA expression, was associated with higher SUVmax and SUVmean (p=0.03 and 0.02 respectively). Both NF-?Bp65 and positive FDG uptake were significantly associated with more advanced stage, tumor histology and invasion. Higher NF-?Bp65 expression was associated with death by KM analysis (p=0.06) while LDHA was strongly associated with recurrence (p=0.04). Increased levels of combined NF-?Bp65 and LDHA expression were synergistic and associated with both recurrence (p=0.04) and death (p=0.03).NF-?B IHC was a modest biomarker of prognosis that associated with tumor glucose metabolism on FDG PET when compared to existing molecular correlates like LDHA, which was synergistic with NF-?B for outcome. These findings recapitulate radiogenomics profiles previously reported by our group and provide a methodology for studying tumor biology using computational approaches.

  View details for DOI 10.1016/j.lungcan.2013.11.001

  View details for PubMedID 24355259

• HER2 Positive Rates Vary by County and Geographic Region in California Independent of Stage and Age at Presentation Allison, K. H., Jensen, K. C., Dur, T. C., Gomez, S. L., West, R. B., Kurian, A. W. NATURE PUBLISHING GROUP. 2014: 34A?35A

  View details for Web of Science ID 000331502200130

• HER2 Heterogeneity by FISH in Breast Cancers and Matched Lymph Node Metastases: How Different Are They? Suarez, C. J., Schmidt, R. A., Dintzis, S. M., Rendi, M. H., Jensen, K. C., West, R. B., Allison, K. H. NATURE PUBLISHING GROUP. 2014: 83A

  View details for Web of Science ID 000331502200325

• Can We Predict Which Her2 Tests Change from Biopsy to Excision? Brown, R. A., Clay, M. R., Jensen, K. C., West, R. B., Allison, K. H. NATURE PUBLISHING GROUP. 2014: 39A

  View details for Web of Science ID 000331155800148

• Can We Predict Which Her2 Tests Change from Biopsy to Excision? Brown, R. A., Clay, M. R., Jensen, K. C., West, R. B., Allison, K. H. NATURE PUBLISHING GROUP. 2014: 39A

  View details for Web of Science ID 000331502200148

• CD5 Is Highly Sensitive and Specific for Pancreaticobiliary Adenocarcinoma Lewis, G. H., West, R. B., Pai, R. K., Longacre, T. A. NATURE PUBLISHING GROUP. 2014: 451A

  View details for Web of Science ID 000331155802410

• Mutational Profiling of Ameloblastoma Identifies Common Gain-of-Function Mutations McClary, A. C., Sweeney, R. T., Biscocho, J., Meyers, B. R., Neahring, L., Kwei, K. A., Qu, K., Gong, X., Jones, C. D., Varma, S., Odegaard, J. I., Rubin, B., Ng, T., Troxell, M. L., Pelham, R. J., Zehnder, J. L., Beachy, P. A., Pollack, J. R., West, R. B. NATURE PUBLISHING GROUP. 2014: 324A

  View details for Web of Science ID 000331155801553

• HER2 Positive Rates Vary by County and Geographic Region in California Independent of Stage and Age at Presentation Allison, K. H., Jensen, K. C., Dur, T. C., Gomez, S. L., West, R. B., Kurian, A. W. NATURE PUBLISHING GROUP. 2014: 34A?35A

  View details for Web of Science ID 000331155800130

• Mutational Profiling of Ameloblastoma Identifies Common Gain-of-Function Mutations McClary, A. C., Sweeney, R. T., Biscocho, J., Meyers, B. R., Neahring, L., Kwei, K. A., Qu, K., Gong, X., Jones, C. D., Varma, S., Odegaard, J. I., Rubin, B., Ng, T., Troxell, M. L., Pelham, R. J., Zehnder, J. L., Beachy, P. A., Pollack, J. R., West, R. B. NATURE PUBLISHING GROUP. 2014: 324A

  View details for Web of Science ID 000331502201650

• CD5 Is Highly Sensitive and Specific for Pancreaticobiliary Adenocarcinoma Lewis, G. H., West, R. B., Pai, R. K., Longacre, T. A. NATURE PUBLISHING GROUP. 2014: 451A

  View details for Web of Science ID 000331502202410

• HER2 Heterogeneity by FISH in Breast Cancers and Matched Lymph Node Metastases: How Different Are They? Suarez, C. J., Schmidt, R. A., Dintzis, S. M., Rendi, M. H., Jensen, K. C., West, R. B., Allison, K. H. NATURE PUBLISHING GROUP. 2014: 83A

  View details for Web of Science ID 000331155800325

• Discovery of recurrent structural variants in nasopharyngeal carcinoma GENOME RESEARCH Valouev, A., Weng, Z., Sweeney, R. T., Varma, S., Quynh-Thu Le, Q. T., Kong, C., Sidow, A., West, R. B. 2014; 24 (2): 300-309

  Abstract

  We present the discovery of genes recurrently involved in structural variation in nasopharyngeal carcinoma (NPC) and the identification of a novel type of somatic structural variant. We identified the variants with high complexity mate-pair libraries and a novel computational algorithm specifically designed for tumor-normal comparisons, SMASH. SMASH combines signals from split reads and mate-pair discordance to detect somatic structural variants. We demonstrate a >90% validation rate and a breakpoint reconstruction accuracy of 3 bp by Sanger sequencing. Our approach identified three in-frame gene fusions (YAP1-MAML2, PTPLB-RSRC1, and SP3-PTK2) that had strong levels of expression in corresponding NPC tissues. We found two cases of a novel type of structural variant, which we call "coupled inversion," one of which produced the YAP1-MAML2 fusion. To investigate whether the identified fusion genes are recurrent, we performed fluorescent in situ hybridization (FISH) to screen 196 independent NPC cases. We observed recurrent rearrangements of MAML2 (three cases), PTK2 (six cases), and SP3 (two cases), corresponding to a combined rate of structural variation recurrence of 6% among tested NPC tissues.

  View details for DOI 10.1101/gr.156224.113

  View details for Web of Science ID 000330696800012

  View details for PubMedID 24214394

  View details for PubMedCentralID PMC3912420

• Discovery of recurrent structural variants in nasopharyngeal carcinoma. Genome research Valouev, A., Weng, Z., Sweeney, R. T., Varma, S., Le, Q., Kong, C., Sidow, A., West, R. B. 2014; 24 (2): 300-309

  Abstract

  We present the discovery of genes recurrently involved in structural variation in nasopharyngeal carcinoma (NPC) and the identification of a novel type of somatic structural variant. We identified the variants with high complexity mate-pair libraries and a novel computational algorithm specifically designed for tumor-normal comparisons, SMASH. SMASH combines signals from split reads and mate-pair discordance to detect somatic structural variants. We demonstrate a >90% validation rate and a breakpoint reconstruction accuracy of 3 bp by Sanger sequencing. Our approach identified three in-frame gene fusions (YAP1-MAML2, PTPLB-RSRC1, and SP3-PTK2) that had strong levels of expression in corresponding NPC tissues. We found two cases of a novel type of structural variant, which we call "coupled inversion," one of which produced the YAP1-MAML2 fusion. To investigate whether the identified fusion genes are recurrent, we performed fluorescent in situ hybridization (FISH) to screen 196 independent NPC cases. We observed recurrent rearrangements of MAML2 (three cases), PTK2 (six cases), and SP3 (two cases), corresponding to a combined rate of structural variation recurrence of 6% among tested NPC tissues.

  View details for DOI 10.1101/gr.156224.113

  View details for PubMedID 24214394

• A shared transcriptional program in early breast neoplasias despite genetic and clinical distinctions GENOME BIOLOGY Brunner, A. L., Li, J., Guo, X., Sweeney, R. T., Varma, S., Zhu, S. X., Li, R., Tibshirani, R., West, R. B. 2014; 15 (5)

  Abstract

  The earliest recognizable stages of breast neoplasia are lesions that represent a heterogeneous collection of epithelial proliferations currently classified based on morphology. Their role in the development of breast cancer is not well understood but insight into the critical events at this early stage will improve efforts in breast cancer detection and prevention. These microscopic lesions are technically difficult to study so very little is known about their molecular alterations.To characterize the transcriptional changes of early breast neoplasia, we sequenced 3'- end enriched RNAseq libraries from formalin-fixed paraffin-embedded tissue of early neoplasia samples and matched normal breast and carcinoma samples from 25 patients. We find that gene expression patterns within early neoplasias are distinct from both normal and breast cancer patterns and identify a pattern of pro-oncogenic changes, including elevated transcription of ERBB2, FOXA1, and GATA3 at this early stage. We validate these findings on a second independent gene expression profile data set generated by whole transcriptome sequencing. Measurements of protein expression by immunohistochemistry on an independent set of early neoplasias confirms that ER pathway regulators FOXA1 and GATA3, as well as ER itself, are consistently upregulated at this early stage. The early neoplasia samples also demonstrate coordinated changes in long non-coding RNA expression and microenvironment stromal gene expression patterns.This study is the first examination of global gene expression in early breast neoplasia, and the genes identified here represent candidate participants in the earliest molecular events in the development of breast cancer.

  View details for DOI 10.1186/gb-2014-15-5-r71

  View details for Web of Science ID 000338981700005

  View details for PubMedCentralID PMC4072957

• A shared transcriptional program in early breast neoplasias despite genetic and clinical distinctions. Genome biology Brunner, A. L., Li, J., Guo, X., Sweeney, R. T., Varma, S., Zhu, S. X., Li, R., Tibshirani, R., West, R. B. 2014; 15 (5): R71-?

  Abstract

  The earliest recognizable stages of breast neoplasia are lesions that represent a heterogeneous collection of epithelial proliferations currently classified based on morphology. Their role in the development of breast cancer is not well understood but insight into the critical events at this early stage will improve efforts in breast cancer detection and prevention. These microscopic lesions are technically difficult to study so very little is known about their molecular alterations.To characterize the transcriptional changes of early breast neoplasia, we sequenced 3'- end enriched RNAseq libraries from formalin-fixed paraffin-embedded tissue of early neoplasia samples and matched normal breast and carcinoma samples from 25 patients. We find that gene expression patterns within early neoplasias are distinct from both normal and breast cancer patterns and identify a pattern of pro-oncogenic changes, including elevated transcription of ERBB2, FOXA1, and GATA3 at this early stage. We validate these findings on a second independent gene expression profile data set generated by whole transcriptome sequencing. Measurements of protein expression by immunohistochemistry on an independent set of early neoplasias confirms that ER pathway regulators FOXA1 and GATA3, as well as ER itself, are consistently upregulated at this early stage. The early neoplasia samples also demonstrate coordinated changes in long non-coding RNA expression and microenvironment stromal gene expression patterns.This study is the first examination of global gene expression in early breast neoplasia, and the genes identified here represent candidate participants in the earliest molecular events in the development of breast cancer.

  View details for DOI 10.1186/gb-2014-15-5-r71

  View details for PubMedID 24887547

• Neuregulin Autocrine Signaling Promotes Self-Renewal of Breast Tumor-Initiating Cells by Triggering HER2/HER3 Activation CANCER RESEARCH Lee, C. Y., Lin, Y., Bratman, S. V., Feng, W., Kuo, A. H., Scheeren, F. A., Engreitz, J. M., Varma, S., West, R. B., Diehn, M. 2014; 74 (1): 341-352

  Abstract

  Currently, only patients with HER2-positive tumors are candidates for HER2-targeted therapies. However, recent clinical observations suggest that the survival of patients with HER2-low breast cancers, who lack HER2 amplification, may benefit from adjuvant therapy that targets HER2. In this study, we explored a mechanism through which these benefits may be obtained. Prompted by the hypothesis that HER2/HER3 signaling in breast tumor-initiating cells (TIC) promotes self-renewal and survival, we obtained evidence that neuregulin 1 (NRG1) produced by TICs promotes their proliferation and self-renewal in HER2-low tumors, including in triple-negative breast tumors. Pharmacologic inhibition of EGFR, HER2, or both receptors reduced breast TIC survival and self-renewal in vitro and in vivo and increased TIC sensitivity to ionizing radiation. Through a tissue microarray analysis, we found that NRG1 expression and associated HER2 activation occurred in a subset of HER2-low breast cancers. Our results offer an explanation for why HER2 inhibition blocks the growth of HER2-low breast tumors. Moreover, they argue that dual inhibition of EGFR and HER2 may offer a useful therapeutic strategy to target TICs in these tumors. In generating a mechanistic rationale to apply HER2-targeting therapies in patients with HER2-low tumors, this work shows why these therapies could benefit a considerably larger number of patients with breast cancer than they currently reach.

  View details for DOI 10.1158/0008-5472.CAN-13-1055

  View details for Web of Science ID 000329297600033

  View details for PubMedID 24177178

  View details for PubMedCentralID PMC3917843

• Molecular pathological analysis of sarcomas using paraffin-embedded tissue: current limitations and future possibilities HISTOPATHOLOGY van de Rijn, M., Guo, X., Sweeney, R. T., Beck, A. H., West, R. B. 2014; 64 (1): 163-170

  Abstract

  Sarcomas of soft tissue and bone are rare neoplasms that can be separated into a large number of different diagnostic entities. Over the years, a number of diagnostic markers have been developed that aid pathologists in reaching the appropriate diagnoses. Many of these markers are sarcoma-specific proteins that can be detected by immunohistochemistry in formalin-fixed, paraffin-embedded (FFPE) sections. In addition, a wide range of molecular studies have been developed that can detect gene mutations, gene amplifications or chromosomal translocations in FFPE material. Until recently, most sequencing-based approaches relied on the availability of fresh frozen tissue. However, with the advent of next-generation sequencing technologies, FFPE material is increasingly being used as a tool to identify novel immunohistochemistry markers, gene mutations, and chromosomal translocations, and to develop diagnostic tests.

  View details for DOI 10.1111/his.12290

  View details for PubMedID 24107169

• Variation in HER2 positive rates in California by geographic region: Implications for setting pathology laboratory benchmarks Allison, K. H., Jensen, K. C., West, R. B., Clarke, C. A., Gomez, S. L., Kurian, A. W. AMER ASSOC CANCER RESEARCH. 2013

  View details for DOI 10.1158/0008-5472.SABCS13-P1-02-10

  View details for Web of Science ID 000209496900136

• MAST2 and NOTCH1 translocations in breast carcinoma and associated pre-invasive lesions. Human pathology Clay, M. R., Varma, S., West, R. B. 2013; 44 (12): 2837-2844

  Abstract

  There are several mutations and structural variations common to breast cancer. Many of these genomic changes are thought to represent driver mutations in oncogenesis. Less well understood is how and when these changes take place in breast cancer development. Previous studies have identified gene rearrangements in the microtubule-associated serine-threonine kinase (MAST) and NOTCH gene families in 5% to 7% of invasive breast cancers. Some of these translocations can be detected by fluorescence in situ hybridization (FISH) allowing for examination of the correlation between these genomic changes and concurrent morphologic changes in early breast neoplasia. NOTCH and MAST gene rearrangements were identified by FISH in a large series of breast cancer cases organized on tissue microarrays (TMA). When translocations were identified by TMA, we performed full cross-section FISH to evaluate concurrent pre-invasive lesions. FISH break-apart assays were designed for NOTCH1 and MAST2 gene rearrangements. Translocations were identified in 16 cases of invasive carcinoma; 10 with MAST2 translocations (2.0%) and 6 cases with NOTCH1 translocations (1.2%). Whole section FISH analysis of these cases demonstrated that the translocations are present in the majority of concurrent ductal carcinoma in situ (DCIS) (6/8). When DCIS wasn't associated with an invasive component, it was never translocated (0/170, P=.0048). We have confirmed the presence of MAST and NOTCH family gene rearrangements in invasive breast carcinoma, and show that FISH studies can effectively be used with TMAs to screen normal, pre-invasive, and coexisting invasive disease. Our findings suggest that these translocations occur during the transition to DCIS and/or invasive carcinoma.

  View details for DOI 10.1016/j.humpath.2013.08.001

  View details for PubMedID 24140425

• Inference of tumor phylogenies with improved somatic mutation discovery. Journal of computational biology Salari, R., Saleh, S. S., Kashef-Haghighi, D., Khavari, D., Newburger, D. E., West, R. B., Sidow, A., Batzoglou, S. 2013; 20 (11): 933-944

  Abstract

  Next-generation sequencing technologies provide a powerful tool for studying genome evolution during progression of advanced diseases such as cancer. Although many recent studies have employed new sequencing technologies to detect mutations across multiple, genetically related tumors, current methods do not exploit available phylogenetic information to improve the accuracy of their variant calls. Here, we present a novel algorithm that uses somatic single-nucleotide variations (SNVs) in multiple, related tissue samples as lineage markers for phylogenetic tree reconstruction. Our method then leverages the inferred phylogeny to improve the accuracy of SNV discovery. Experimental analyses demonstrate that our method achieves up to 32% improvement for somatic SNV calling of multiple, related samples over the accuracy of GATK's Unified Genotyper, the state-of-the-art multisample SNV caller.

  View details for DOI 10.1089/cmb.2013.0106

  View details for PubMedID 24195709

• INCREASED GALECTIN-1 EXPRESSION IN A THYMIC EPITHELIAL TUMOR TISSUE MICROARRAY (TMA) AND GALECTIN-1 KNOCKDOWN STUDIES IN A THYMOMA CELL LINE Riess, J. W., Kuo, P., Kong, C., Wakelee, H. A., West, R., Quynh Le LIPPINCOTT WILLIAMS & WILKINS. 2013: S269

  View details for Web of Science ID 000339624901100

• GLI1, NOTCH1 AND CTNNB1 EXPRESSION BY AUTOMATED QUANTITATIVE IMMUNOFLUORESCENCE (AQUA) IN A THYMIC MALIGNANCY TISSUE MICROARRAY (TMA) Riess, J. W., West, R., Padda, S. K., Dean, M., Klimowicz, A., Huong, C., Neal, J. W., Wakelee, H. A. LIPPINCOTT WILLIAMS & WILKINS. 2013: S944

  View details for Web of Science ID 000339624904251

• Inference of Tumor Phylogenies with Improved Somatic Mutation Discovery JOURNAL OF COMPUTATIONAL BIOLOGY Salari, R., Saleh, S. S., Kashef-Haghighi, D., Khavari, D., Newburger, D. E., West, R. B., Sidow, A., Batzoglou, S. 2013; 20 (11): 933-944

  Abstract

  Next-generation sequencing technologies provide a powerful tool for studying genome evolution during progression of advanced diseases such as cancer. Although many recent studies have employed new sequencing technologies to detect mutations across multiple, genetically related tumors, current methods do not exploit available phylogenetic information to improve the accuracy of their variant calls. Here, we present a novel algorithm that uses somatic single-nucleotide variations (SNVs) in multiple, related tissue samples as lineage markers for phylogenetic tree reconstruction. Our method then leverages the inferred phylogeny to improve the accuracy of SNV discovery. Experimental analyses demonstrate that our method achieves up to 32% improvement for somatic SNV calling of multiple, related samples over the accuracy of GATK's Unified Genotyper, the state-of-the-art multisample SNV caller.

  View details for DOI 10.1089/cmb.2013.0106

  View details for Web of Science ID 000326577600008

  View details for PubMedID 24195709

  View details for PubMedCentralID PMC3822366

• Stromal Responses among Common Carcinomas Correlated with Clinicopathologic Features. Clinical cancer research Chen, J. L., Espinosa, I., Lin, A. Y., Liao, O. Y., van de Rijn, M., West, R. B. 2013; 19 (18): 5127-5135

  Abstract

  We have previously characterized a tumor stroma expression signature in a subset of breast tumors that correlates with better clinical outcome. The purpose of this study is to determine whether this stromal signature, termed the DTF fibroblast signature, is specific to breast cancer or is a common stromal response found in different types of cancer. Experimental Designs: The DTF fibroblast signature was applied to gene expression profiles from five ovarian, five lung, two colon and three prostate cancer expression microarray datasets. Additionally, two different tissue microarrays of 204 ovarian tumors and 140 colon tumors were examined for the expression of previously characterized protein markers of DTF fibroblast signature. The DTF fibroblast stromal response was then correlated with clinicopathologic features.The DTF fibroblast signature is robustly present in ovarian, lung, and colon carcinomas. Both expression microarray data and immunohistochemistry show the subset of ovarian tumors with strong DTF fibroblast signature expression has statistically significant worse survival outcomes. No reproducible survival differences were found in either the lung or the colon cancers. The prostate cancers failed to demonstrate a DTF fibroblast signature. Multi-variant analysis showed that DTF fibroblast signature was significantly more prognostic than the proliferation status in ovarian carcinomas.Our results suggest that the DTF fibroblast signature is a common tumor stroma signature in different types of cancer including ovarian, lung and colon carcinomas. Our findings provide further insight into the DTF fibroblast stromal responses across different types of carcinomas and their potential as prognostic and therapeutic targets.

  View details for DOI 10.1158/1078-0432.CCR-12-3127

  View details for PubMedID 23804424

• GLI1, NOTCH1 AND CTNNB1 EXPRESSION BY AUTOMATED QUANTITATIVE IMMUNOFLUORESCENCE (AQUA) IN A THYMIC MALIGNANCY TISSUE MICROARRAY (TMA) Riess, J., West, R., Padda, S., Dean, M., Klimowicz, A., Huong, C., Neal, J., Wakelee, H. LIPPINCOTT WILLIAMS & WILKINS. 2013: 25

  View details for Web of Science ID 000339623500027

• INCREASED GALECTIN-1 EXPRESSION IN A THYMIC EPITHELIAL TUMOR TISSUE MICROARRAY (TMA) Riess, J., Kuo, P., Kong, C., West, R., Padda, S., Wakelee, H., Le, Q. LIPPINCOTT WILLIAMS & WILKINS. 2013: 25?26

  View details for Web of Science ID 000339623500028

• Genome evolution during progression to breast cancer GENOME RESEARCH Newburger, D. E., Kashef-Haghighi, D., Weng, Z., Salari, R., Sweeney, R. T., Brunner, A. L., Zhu, S. X., Guo, X., Varma, S., Troxell, M. L., West, R. B., Batzoglou, S., Sidow, A. 2013; 23 (7): 1097-1108

  Abstract

  Cancer evolution involves cycles of genomic damage, epigenetic deregulation, and increased cellular proliferation that eventually culminate in the carcinoma phenotype. Early neoplasias, which are often found concurrently with carcinomas and are histologically distinguishable from normal breast tissue, are less advanced in phenotype than carcinomas and are thought to represent precursor stages. To elucidate their role in cancer evolution we performed comparative whole-genome sequencing of early neoplasias, matched normal tissue, and carcinomas from six patients, for a total of 31 samples. By using somatic mutations as lineage markers we built trees that relate the tissue samples within each patient. On the basis of these lineage trees we inferred the order, timing, and rates of genomic events. In four out of six cases, an early neoplasia and the carcinoma share a mutated common ancestor with recurring aneuploidies, and in all six cases evolution accelerated in the carcinoma lineage. Transition spectra of somatic mutations are stable and consistent across cases, suggesting that accumulation of somatic mutations is a result of increased ancestral cell division rather than specific mutational mechanisms. In contrast to highly advanced tumors that are the focus of much of the current cancer genome sequencing, neither the early neoplasia genomes nor the carcinomas are enriched with potentially functional somatic point mutations. Aneuploidies that occur in common ancestors of neoplastic and tumor cells are the earliest events that affect a large number of genes and may predispose breast tissue to eventual development of invasive carcinoma.

  View details for DOI 10.1101/gr.151670.112

  View details for Web of Science ID 000321119900007

  View details for PubMedID 23568837

• Desktop transcriptome sequencing from archival tissue to identify clinically relevant translocations. American journal of surgical pathology Sweeney, R. T., Zhang, B., Zhu, S. X., Varma, S., Smith, K. S., Montgomery, S. B., van de Rijn, M., Zehnder, J., West, R. B. 2013; 37 (6): 796-803

  Abstract

  Somatic mutations, often translocations or single nucleotide variations, are pathognomonic for certain types of cancers and are increasingly of clinical importance for diagnosis and prediction of response to therapy. Conventional clinical assays only evaluate 1 mutation at a time, and targeted tests are often constrained to identify only the most common mutations. Genome-wide or transcriptome-wide high-throughput sequencing (HTS) of clinical samples offers an opportunity to evaluate for all clinically significant mutations with a single test. Recently a "desktop version" of HTS has become available, but most of the experience to date is based on data obtained from high-quality DNA from frozen specimens. In this study, we demonstrate, as a proof of principle, that translocations in sarcomas can be diagnosed from formalin-fixed paraffin-embedded (FFPE) tissue with desktop HTS. Using the first generation MiSeq platform, full transcriptome sequencing was performed on FFPE material from archival blocks of 3 synovial sarcomas, 3 myxoid liposarcomas, 2 Ewing sarcomas, and 1 clear cell sarcoma. Mapping the reads to the "sarcomatome" (all known 83 genes involved in translocations and mutations in sarcoma) and using a novel algorithm for ranking fusion candidates, the pathognomonic fusions and the exact breakpoints were identified in all cases of synovial sarcoma, myxoid liposarcoma, and clear cell sarcoma. The Ewing sarcoma fusion gene was detectable in FFPE material only with a sequencing platform that generates greater sequencing depth. The results show that a single transcriptome HTS assay, from FFPE, has the potential to replace conventional molecular diagnostic techniques for the evaluation of clinically relevant mutations in cancer.

  View details for DOI 10.1097/PAS.0b013e31827ad9b2

  View details for PubMedID 23598961

• Reconsidering the Diagnostic and Prognostic Utility of LN-2 for Undifferentiated Pleomorphic Sarcoma and Atypical Fibroxanthoma AMERICAN JOURNAL OF DERMATOPATHOLOGY Hollmig, S. T., Rieger, K. E., Henderson, M. T., West, R. B., Sundram, U. N. 2013; 35 (2): 176-179

  Abstract

  The topic of distinguishing atypical fibroxanthoma (AFX) from undifferentiated pleomorphic sarcoma (UPS), formerly malignant fibrous histiocytoma, is highly controversial. Although their clinical behavior is disparate, AFX and UPS commonly appear nearly identical on routine histopathologic examination. Although conceptually useful, subcategorization of UPS into superficial (confined to the dermis and subcutaneous tissue) and deep (involvement of fascia and deeper structures) types has not improved our ability to differentiate UPS from AFX. Numerous authors have purported LN-2 (CD74) immunopositivity as able to distinguish UPS from AFX and to predict those rare AFX likely to behave aggressively, although only a single prior study has been dedicated to evaluating this marker. We performed LN-2 staining of 14 AFX, 8 superficial UPS, and 65 deep UPS specimens using an identical protocol as described by prior authors. Of the 73 total UPS specimens, only 1 (1.4%) stained strongly with LN-2, as compared with 3 of 14 (21%) AFX (P = 0.012). One of 2 (50%) clinically aggressive AFX tumors that later exhibited both local recurrence and metastasis stained strongly for LN-2, whereas 2 of 12 (17%) of the more indolent tumors stained strongly with this marker (P = 0.40). Our data do not replicate prior reports of LN-2 as a sensitive and specific marker for UPS, or as indicative of prognosis for AFX, and therefore does not support the use of LN-2 as either a diagnostic or prognostic marker.

  View details for DOI 10.1097/DAD.0b013e318265fb9e

  View details for Web of Science ID 000316941200009

  View details for PubMedID 23000905

• Breakpoint analysis of transcriptional and genomic profiles uncovers novel gene fusions spanning multiple human cancer types. PLoS genetics Giacomini, C. P., Sun, S., Varma, S., Shain, A. H., Giacomini, M. M., Balagtas, J., Sweeney, R. T., Lai, E., Del Vecchio, C. A., Forster, A. D., Clarke, N., Montgomery, K. D., Zhu, S., Wong, A. J., van de Rijn, M., West, R. B., Pollack, J. R. 2013; 9 (4)

  View details for DOI 10.1371/journal.pgen.1003464

  View details for PubMedID 23637631

• Breakpoint analysis of transcriptional and genomic profiles uncovers novel gene fusions spanning multiple human cancer types. PLoS genetics Giacomini, C. P., Sun, S., Varma, S., Shain, A. H., Giacomini, M. M., Balagtas, J., Sweeney, R. T., Lai, E., Del Vecchio, C. A., Forster, A. D., Clarke, N., Montgomery, K. D., Zhu, S., Wong, A. J., van de Rijn, M., West, R. B., Pollack, J. R. 2013; 9 (4)

  Abstract

  Gene fusions, like BCR/ABL1 in chronic myelogenous leukemia, have long been recognized in hematologic and mesenchymal malignancies. The recent finding of gene fusions in prostate and lung cancers has motivated the search for pathogenic gene fusions in other malignancies. Here, we developed a "breakpoint analysis" pipeline to discover candidate gene fusions by tell-tale transcript level or genomic DNA copy number transitions occurring within genes. Mining data from 974 diverse cancer samples, we identified 198 candidate fusions involving annotated cancer genes. From these, we validated and further characterized novel gene fusions involving ROS1 tyrosine kinase in angiosarcoma (CEP85L/ROS1), SLC1A2 glutamate transporter in colon cancer (APIP/SLC1A2), RAF1 kinase in pancreatic cancer (ATG7/RAF1) and anaplastic astrocytoma (BCL6/RAF1), EWSR1 in melanoma (EWSR1/CREM), CDK6 kinase in T-cell acute lymphoblastic leukemia (FAM133B/CDK6), and CLTC in breast cancer (CLTC/VMP1). Notably, while these fusions involved known cancer genes, all occurred with novel fusion partners and in previously unreported cancer types. Moreover, several constituted druggable targets (including kinases), with therapeutic implications for their respective malignancies. Lastly, breakpoint analysis identified new cell line models for known rearrangements, including EGFRvIII and FIP1L1/PDGFRA. Taken together, we provide a robust approach for gene fusion discovery, and our results highlight a more widespread role of fusion genes in cancer pathogenesis.

  View details for DOI 10.1371/journal.pgen.1003464

  View details for PubMedID 23637631

• Anti-KIT monoclonal antibody inhibits imatinib-resistant gastrointestinal stromal tumor growth PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Edris, B., Willingham, S. B., Weiskopf, K., Volkmer, A. K., Volkmer, J., Muehlenberg, T., Montgomery, K. D., Contreras-Trujillo, H., Czechowicz, A., Fletcher, J. A., West, R. B., Weissman, I. L., van de Rijn, M. 2013; 110 (9): 3501-3506

  Abstract

  Gastrointestinal stromal tumor (GIST) is the most common sarcoma of the gastrointestinal tract and arises from the interstitial cells of Cajal. It is characterized by expression of the receptor tyrosine kinase CD117 (KIT). In 70-80% of GIST cases, oncogenic mutations in KIT are present, leading to constitutive activation of the receptor, which drives the proliferation of these tumors. Treatment of GIST with imatinib, a small-molecule tyrosine kinase inhibitor, inhibits KIT-mediated signaling and initially results in disease control in 70-85% of patients with KIT-positive GIST. However, the vast majority of patients eventually develop resistance to imatinib treatment, leading to disease progression and posing a significant challenge in the clinical management of these tumors. Here, we show that an anti-KIT monoclonal antibody (mAb), SR1, is able to slow the growth of three human GIST cell lines in vitro. Importantly, these reductions in cell growth were equivalent between imatinib-resistant and imatinib-sensitive GIST cell lines. Treatment of GIST cell lines with SR1 reduces cell-surface KIT expression, suggesting that mAb-induced KIT down-regulation may be a mechanism by which SR1 inhibits GIST growth. Furthermore, we also show that SR1 treatment enhances phagocytosis of GIST cells by macrophages, indicating that treatment with SR1 may enhance immune cell-mediated tumor clearance. Finally, using two xenotransplantation models of imatinib-sensitive and imatinib-resistant GIST, we demonstrate that SR1 is able to strongly inhibit tumor growth in vivo. These results suggest that treatment with mAbs targeting KIT may represent an alternative, or complementary, approach for treating GIST.

  View details for DOI 10.1073/pnas.1222893110

  View details for PubMedID 23382202

• Desktop Transcriptome Sequencing from Archival Tissue To Identify Clinically Relevant Translocations 102nd Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology (USCAP) SWEENEY, R. T., Zhang, B., Zhu, S. X., Varma, S., Smith, K., Montgomery, S. B., van de Rijn, M., Zehnder, J., West, R. B. NATURE PUBLISHING GROUP. 2013: 438A?438A

  View details for Web of Science ID 000314789302442

• Two New Stromal Signatures Stratify Breast Cancers with Different Prognosis Guo, X., Zhu, S. X., Montgomery, K., van de Rijn, M., West, R. B. NATURE PUBLISHING GROUP. 2013: 434A?435A

  View details for Web of Science ID 000314789302425

• Desktop Transcriptome Sequencing from Archival Tissue To Identify Clinically Relevant Translocations Sweeney, R. T., Zhang, B., Zhu, S. X., Varma, S., Smith, K., Montgomery, S. B., de Rijn, A., Zehnder, J., West, R. B. NATURE PUBLISHING GROUP. 2013: 438A

  View details for Web of Science ID 000314444402452

• Positive SOX10 Expression in a Broad Range of Salivary Gland Tumors Ng, T. L., West, R., Kwok, S., Berry, G., Kong, C. NATURE PUBLISHING GROUP. 2013: 310A

  View details for Web of Science ID 000314444401700

• BRST2, ER, e-Cadherin, CDX2, and NKX3.1 Utility in Distinguishing Diffuse Gastric from Lobular Carcinoma Lewis, G. H., West, R., Pai, R. K., Longacre, T. A., Jensen, K. C. NATURE PUBLISHING GROUP. 2013: 52A?53A

  View details for Web of Science ID 000314789300212

• Positive SOX10 Expression in a Broad Range of Salivary Gland Tumors Ng, T. L., West, R., Kwok, S., Berry, G., Kong, C. NATURE PUBLISHING GROUP. 2013: 310A

  View details for Web of Science ID 000314789301620

• Novel Mutations in Neuroendocrine Carcinoma of the Breast: Possible Therapeutic Targets Ballard, M., Ang, D., Beadling, C., Warrick, A., Schilling, A., O'Gara, R., Pukay, M., West, R., Corless, C., Troxell, M. NATURE PUBLISHING GROUP. 2013: 29A

  View details for Web of Science ID 000314444400114

• BRST2, ER, e-Cadherin, CDX2, and NKX3.1 Utility in Distinguishing Diffuse Gastric from Lobular Carcinoma Lewis, G. H., West, R., Pai, R. K., Longacre, T. A., Jensen, K. C. NATURE PUBLISHING GROUP. 2013: 52A?53A

  View details for Web of Science ID 000314444400212

• ROR2 Expression in Breast Cancer Guo, X., Sweeny, P., Varma, S., Montgomery, K., West, R. B., van de Rijn, M. NATURE PUBLISHING GROUP. 2013: 45A

  View details for Web of Science ID 000314444400179

• Two New Stromal Signatures Stratify Breast Cancers with Different Prognosis 102nd Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology (USCAP) Guo, X., Zhu, S. X., Montgomery, K., van de Rijn, M., West, R. B. NATURE PUBLISHING GROUP. 2013: 434A?435A

  View details for Web of Science ID 000314444402435

• ROR2 Expression in Breast Cancer 102nd Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology (USCAP) Guo, X., Sweeny, P., Varma, S., Montgomery, K., West, R. B., van de Rijn, M. NATURE PUBLISHING GROUP. 2013: 45A?45A

  View details for Web of Science ID 000314789300179

• Next generation sequencing-based expression profiling identifies signatures from benign stromal proliferations that define stromal components of breast cancer BREAST CANCER RESEARCH Guo, X., Zhu, S. X., Brunner, A. L., van de Rijn, M., West, R. B. 2013; 15 (6)

  Abstract

  Multiple studies have shown that the tumor microenvironment (TME) of carcinomas can play an important role in the initiation, progression, and metastasis of cancer. Here we test the hypothesis that specific benign fibrous soft tissue tumor gene expression profiles may represent distinct stromal fibroblastic reaction types that occur in different breast cancers. The discovered stromal profiles could classify breast cancer based on the type of stromal reaction patterns in the TME.Next generation sequencing-based gene expression profiling (3SEQ) was performed on formalin fixed, paraffin embedded (FFPE) samples of 10 types of fibrous soft tissue tumors. We determined the extent to which these signatures could identify distinct subsets of breast cancers in four publicly available breast cancer datasets.A total of 53 fibrous tumors were sequenced by 3SEQ with an average of 29 million reads per sample. Both the gene signatures derived from elastofibroma (EF) and fibroma of tendon sheath (FOTS) demonstrated robust outcome results for survival in the four breast cancer datasets. The breast cancers positive for the EF signature (20-33% of the cohort) demonstrated significantly better outcome for survival. In contrast, the FOTS signature-positive breast cancers (11-35% of the cohort) had a worse outcome.We defined and validated two new stromal signatures in breast cancer (EF and FOTS), which are significantly associated with prognosis. Our group has previously identified novel cancer stromal gene expression signatures associated with outcome differences in breast cancer by gene expression profiling of three soft tissue tumors, desmoid-type fibromatosis (DTF), solitary fibrous tumor (SFT), and tenosynovial giant cell tumor (TGCT/CSF1), as surrogates for stromal expression patterns. By combining the stromal signatures of EF and FOTS, with our previously identified DTF and TGCT/CSF1 signatures we can now characterize clinically relevant stromal expression profiles in the TME for between 74% to 90% of all breast cancers.

  View details for DOI 10.1186/bcr3586

  View details for Web of Science ID 000331544200013

  View details for PubMedCentralID PMC3978842

• Integrative Bioinformatics Links HNF1B with Clear Cell Carcinoma and Tumor-Associated Thrombosis. PloS one Cuff, J., Salari, K., Clarke, N., Esheba, G. E., Forster, A. D., Huang, S., West, R. B., Higgins, J. P., Longacre, T. A., Pollack, J. R. 2013; 8 (9)

  View details for DOI 10.1371/journal.pone.0074562

  View details for PubMedID 24040285

• Integrative bioinformatics links HNF1B with clear cell carcinoma and tumor-associated thrombosis. PloS one Cuff, J., Salari, K., Clarke, N., Esheba, G. E., Forster, A. D., Huang, S., West, R. B., Higgins, J. P., Longacre, T. A., Pollack, J. R. 2013; 8 (9)

  Abstract

  Clear cell carcinoma (CCC) is a histologically distinct carcinoma subtype that arises in several organ systems and is marked by cytoplasmic clearing, attributed to abundant intracellular glycogen. Previously, transcription factor hepatocyte nuclear factor 1-beta (HNF1B) was identified as a biomarker of ovarian CCC. Here, we set out to explore more broadly the relation between HNF1B and carcinomas with clear cell histology. HNF1B expression, evaluated by immunohistochemistry, was significantly associated with clear cell histology across diverse gynecologic and renal carcinomas (P<0.001), as was hypomethylation of the HNF1B promoter (P<0.001). From microarray analysis, an empirically-derived HNF1B signature was significantly enriched for computationally-predicted targets (with HNF1 binding sites) (P<0.03), as well as genes associated with glycogen metabolism, including glucose-6-phophatase, and strikingly the blood clotting cascade, including fibrinogen, prothrombin and factor XIII. Enrichment of the clotting cascade was also evident in microarray data from ovarian CCC versus other histotypes (P<0.01), and HNF1B-associated prothrombin expression was verified by immunohistochemistry (P?=?0.015). Finally, among gynecologic carcinomas with cytoplasmic clearing, HNF1B immunostaining was linked to a 3.0-fold increased risk of clinically-significant venous thrombosis (P?=?0.043), and with a 2.3-fold increased risk (P?=?0.011) in a combined gynecologic and renal carcinoma cohort. Our results define HNF1B as a broad marker of clear cell phenotype, and support a mechanistic link to glycogen accumulation and thrombosis, possibly reflecting (for gynecologic CCC) derivation from secretory endometrium. Our findings also implicate a novel mechanism of tumor-associated thrombosis (a major cause of cancer mortality), based on the direct production of clotting factors by cancer cells.

  View details for DOI 10.1371/journal.pone.0074562

  View details for PubMedID 24040285

  View details for PubMedCentralID PMC3767734

• Whole-genome progression of breast cancer from early neoplasia to invasive carcinoma West, R. B., Kashef-Haghighi, D., Newburger, D., Weng, Z., Brunner, A., Salari, R., Guo, X., Troxell, M., Zhu, S., Varma, S., Sidow, A., Batzoglou, S. AMER ASSOC CANCER RESEARCH. 2012

  View details for DOI 10.1158/0008-5472.SABCS12-PD05-09

  View details for Web of Science ID 000209704901392

• Biphasic Papillary and Lobular Breast Carcinoma With PIK3CA and IDH1 Mutations DIAGNOSTIC MOLECULAR PATHOLOGY Ang, D., VanSandt, A. M., Beadling, C., Warrick, A., West, R. B., Corless, C. L., Troxell, M. L. 2012; 21 (4): 221-224

  Abstract

  Morphologic "special types" of breast carcinomas have been recognized for many years, and their molecular and genetic properties have not been specifically studied until recently. Lobular carcinoma lacks functional E-cadherin expression but shares molecular similarities with low-grade invasive ductal carcinomas. Papillary carcinoma is relatively rare, and molecular features are just being elucidated. We report a case of concurrent invasive lobular and papillary carcinoma, the latter with extensive nodal involvement. Multiplex screening for activating point mutations identified different point mutations in the distinct morphologic components: lobular PIK3CA H1047R, papillary; PIK3CA Q546P, and IDH1 R132H. These molecular data favor coincidental "collision tumors" over clonal evolution. The IDH1 R132H point mutation is common in gliomas and acute myelogenous leukemia, but this has not been previously reported in breast carcinoma. The characterization of activating point mutations in morphologic special types of breast carcinoma may suggest avenues amenable to targeted therapy.

  View details for DOI 10.1097/PDM.0b013e31826ddbd1

  View details for PubMedID 23111200

• TdT(+) T-lymphoblastic Populations Are Increased in Castleman Disease, in Castleman Disease in Association With Follicular Dendritic Cell Tumors, and in Angioimmunoblastic T-cell Lymphoma AMERICAN JOURNAL OF SURGICAL PATHOLOGY Ohgami, R. S., Zhao, S., Ohgami, J. K., Leavitt, M. O., Zehnder, J. L., West, R. B., Arber, D. A., Natkunam, Y., Warnke, R. A. 2012; 36 (11): 1619-1628

  Abstract

  T-lymphoblastic lymphoma is an aggressive neoplasm requiring prompt clinical treatment. Conversely, indolent T-lymphoblastic proliferation mimics T-lymphoblastic lymphoma but consists of a proliferation of non-neoplastic TdT+ T cells, requiring no treatment. Recently, we identified several cases of indolent T-lymphoblastic proliferations in extrathymic lymphoid tissues: 1 in a patient suffering from Castleman disease (CD) associated with a follicular dendritic cell sarcoma/tumor, 1 in a patient with a history of angioimmunoblastic T-cell lymphoma (AITL), and 1 in association with acinic cell carcinoma. Interestingly, in the case of the patient with a history of AITL, these TdT+ T cells were seen in multiple anatomic sites over the span of 5 years. Here we review these 3 cases and extend our findings by demonstrating that TdT+ T-lymphoblastic populations are increased in lymph nodes of patients with CD (P=0.011), CD in association with follicular dendritic cell tumors, and AITL (P<0.01) compared with other T-cell or B-cell lymphomas or reactive lymph nodes. Finally, analysis of 352 nonhematolymphoid tumors including carcinomas, melanomas, and sarcomas demonstrates that TdT+ T cells are not increased in these tumors. Our studies not only present several detailed cases of indolent T-lymphoblastic proliferations, but also correlate these populations with specific hematologic diseases.

  View details for DOI 10.1097/PAS.0b013e318264e223

  View details for Web of Science ID 000310059600004

  View details for PubMedID 23060347

• beta-Adrenergic receptor expression in vascular tumors MODERN PATHOLOGY Chisholm, K. M., Chang, K. W., Truong, M. T., Kwok, S., West, R. B., Heerema-McKenney, A. E. 2012; 25 (11): 1446-1451

  Abstract

  Propranolol has recently emerged as an effective therapy for infantile hemangiomas causing regression. The ?-adrenergic receptor (AR) antagonist is thought to cause vasoconstriction by its effect on nitric oxide, block angiogenesis by its effect on vascular endothelial growth factor (VEGF), and induce apoptosis. In a prior report, we identified expression of ?2-AR (B2-AR) and its phosphorylated form (B2-ARP) in a case of infantile hemangioma that responded to propranolol treatment. We now explore the expression of ?ARs on a variety of vascular lesions utilizing a tissue microarray containing 141 lesions, including infantile hemangiomas, angiosarcomas, hemangiomas, hemangioendotheliomas, and various vascular malformations. The array was immunostained for B2-AR, B2-ARP, and ?3-AR (B3-AR), and the results scored for the intensity of endothelial cell expression as negative, weak positive, or strong positive. All phases of infantile hemangiomas had strong expression of all three receptors, with the exception of only weak expression of B2-ARP in the proliferative phase infantile hemangioma. Strong expression of all three receptors was present in many hemangiomas, hemangioendotheliomas, and vascular malformations. Absent to weak expression of all three receptors was seen in glomus tumor, hobnail hemangioendothelioma, pyogenic granuloma, and reactive vascular proliferations. This is the first study to report ?-AR expression in a variety of vascular lesions. Although immunohistochemical expression of the receptors does not necessarily indicate that similar pathways of responsiveness to ?-blockade are present, it does raises the possibility that ?-blockade could potentially affect apoptosis and decrease responsiveness to VEGF. Additional study is warranted, as therapeutic options are limited for some patients with these lesions.

  View details for DOI 10.1038/modpathol.2012.108

  View details for PubMedID 22743651

• Detection of Long Non-Coding RNA in Archival Tissue: Correlation with Polycomb Protein Expression in Primary and Metastatic Breast Carcinoma PLOS ONE Chisholm, K. M., Wan, Y., Li, R., Montgomery, K. D., Chang, H. Y., West, R. B. 2012; 7 (10)

  Abstract

  A major function of long non-coding RNAs (lncRNAs) is regulating gene expression through changes in chromatin state. Experimental evidence suggests that in cancer, they can influence Polycomb Repressive Complexes (PRC) to retarget to an occupancy pattern resembling that of the embryonic state. We have previously demonstrated that the expression level of lncRNA in the HOX locus, including HOTAIR, is a predictor of breast cancer metastasis. In this current project, RNA in situ hybridization of probes to three different lncRNAs (HOTAIR, ncHoxA1, and ncHoxD4), as well a immunohistochemical staining of EZH2, is undertaken in formalin-fixed paraffin-embedded breast cancer tissues in a high throughput tissue microarray format to correlate expression with clinicopathologic features. Though overall EZH2 and HOTAIR expression levels were highly correlated, the subset of cases with strong HOTAIR expression correlated with ER and PR positivity, while the subset of cases with strong EZH2 expression correlated with an increased proliferation rate, ER and PR negativity, HER2 underexpression, and triple negativity. Co-expression of HOTAIR and EZH2 trended with a worse outcome. In matched primary and metastatic cancers, both HOTAIR and EZH2 had increased expression in the metastatic carcinomas. This is the first study to show that RNA in situ hybridization of formalin fixed paraffin-embedded clinical material can be used to measure levels of long non-coding RNAs. This approach offers a method to make observations on lncRNAs that may influence the cancer epigenome in a tissue-based technique.

  View details for DOI 10.1371/journal.pone.0047998

  View details for PubMedID 23133536

• Sox10 and S100 in the Diagnosis of Soft-tissue Neoplasms APPLIED IMMUNOHISTOCHEMISTRY & MOLECULAR MORPHOLOGY Karamchandani, J. R., Nielsen, T. O., van de Rijn, M., West, R. B. 2012; 20 (5): 445-450

  Abstract

  Despite a well-characterized lack of specificity, pathologists routinely employ S100 in the diagnosis of neural crest-derived tumors. Recent studies have shown that Sox10 is a reliable marker of neural crest differentiation that is consistently expressed in schwannian and melanocytic tumors. We sought to validate these results in a larger series of soft tissue neoplasms of both neural crest and non-neural crest origin, and to further characterize the sensitivity and specificity of Sox10 for use in clinical diagnosis. We evaluated Sox10 and S100 mRNA levels in 122 cases of peripheral nerve sheath tumors and synovial sarcoma and used immunohistochemistry for Sox10 and S100 protein expression in 1012 tissue specimens. This study includes 174 tissue microarray cases previously reported by Nonaka and colleagues, which include cases of melanoma, dermatofibrosarcoma protuberans, neurofibroma, synovial sarcoma, clear-cell sarcoma, malignant peripheral nerve sheath tumor (MPNST), perineurioma, and schwannoma. Synovial sarcomas expressed significantly higher levels of S100B than Sox10 (P=7.9×10), and no significant Sox10 mRNA expression was identified in synovial sarcoma (n=40), whereas 18/40 cases showed comparatively increased levels of S100 mRNA. The majority of schwannomas (n=26) and neurofibromas (n=28) showed relatively an increased expression of both Sox10 and S100 mRNA. MPNSTs (n=28) showed variable levels of Sox10 and S100 mRNA expression, and these expression levels were highly correlated (Pearson correlation coefficient r=0.79). In contrast, immunohistochemistry performed on a larger and more varied number of cases highlighted significant differences between the 2 proteins. We identified 5 non-neural, nonmelanocytic sarcoma types in which a subset of cases showed S100 protein expression: synovial sarcoma (12/79, 15%), Ewing sarcoma (3/14, 21%), rhabdomyosarcoma (4/17, 24%), chondrosarcoma (3/4, 75%), and extraskeletal myxoid chondrosarcoma (5/11, 45%). For each of these entities, we identified cases with strong and diffuse S100 staining. Of these cases, only 1 case of rhabdomyosarcoma showed focal Sox10 positivity. In 78 cases of MPNST, S100 increased the sensitivity (31/78, 40%) as compared with Sox10 (21/78, 27%), but the majority of these cases were negative for both Sox10 and S100 (44/78, 56%). Sox10 proved superior to S100 in the detection of desmoplastic melanoma (7/9, 78%) and clear-cell sarcoma (4/7, 57%). We also report for the first time Sox10 expression in 26 cases of granular cell tumor, further supporting the neural crest derivation of this tumor. Excluding MPNST, S100 and Sox10 showed similar sensitivity in tumors of neural crest origin (140/148, 95% and 137/148, 93%, respectively). In summary, Sox10 shows an increased specificity for tumors of neural crest origin compared with S100: Sox10 was positive in only 5 of 668 cases (99% specificity) in nonschwannian, nonmelanocytic tumors, whereas S100 was positive in 53 of 668 cases (91% specificity). Sox10 should be used in the place of or along with S100 in soft tissue tumor diagnosis.

  View details for DOI 10.1097/PAI.0b013e318244ff4b

  View details for PubMedID 22495377

• Pathologic Features and Immunophenotype of Estrogen Receptor-positive Breast Cancers in BRCA1 Mutation Carriers AMERICAN JOURNAL OF SURGICAL PATHOLOGY Kaplan, J. S., Schnitt, S. J., Collins, L. C., Wang, Y., Garber, J. E., Montgomery, K., West, R. B., Krag, K., Fetten, K., Lincoln, A., Tung, N. M. 2012; 36 (10): 1483-1488

  Abstract

  Although most breast cancers in BRCA1 mutation carriers are estrogen receptor negative (ER-) with a basal-like phenotype, up to one third are ER positive (ER+). Little is known about the characteristics of this subgroup. To address this, we compared histologic and immunophenotypic features of 60 BRCA1-related ER+ breast cancers with those of 85 BRCA1-related ER- cancers and 174 matched ER+ sporadic cancers. ER+ BRCA1-related cancers were significantly less likely than ER- BRCA1-related cancers to be of pure invasive ductal type (P<0.001) and to be of histologic grade 3 (P<0.001), and less frequently to have a high mitotic rate (P<0.001), pushing (or unknown) margins (P<0.001), a moderate/marked lymphocytic infiltrate (P=0.003), or geographic necrosis/fibrotic focus (P<0.001). In addition, ER+ BRCA1-related cancers less often expressed CK5/6 (P<0.0001), CK14 (P<0.0001), and epidermal growth factor receptor (P<0.0001) and more often expressed progesterone receptor (P<0.0001). In contrast, when compared with ER+ sporadic cancers, ER+ BRCA1-related cancers were significantly more often of invasive ductal type (P=0.005) and of histologic grade 3 (P=0.006), more frequently had a high mitotic rate (P=0.0003), and were more often CK14+ (P=0.03). On unsupervised cluster analysis, some ER+ BRCA1 cancers clustered more closely with sporadic ER+ cancers, whereas others clustered more closely with ER- BRCA1-related cancers. Nuclear expression levels of poly(ADP) ribose polymerase 1 in ER+ BRCA1-related cancers were similar to those in ER- BRCA1-related cancers but significantly higher than in ER+ sporadic cancers. We conclude that ER+ BRCA1-related breast cancers show several morphologic and immunophenotypic differences from ER+ sporadic breast cancers as well as some similarities to ER- BRCA1-related cancers.

  View details for DOI 10.1097/PAS.0b013e31825789ed

  View details for Web of Science ID 000309115100008

  View details for PubMedID 22613997

• Chest Wall Leiomyosarcoma After Breast-Conservative Therapy for Early-Stage Breast Cancer in a Young Woman With Li-Fraumeni Syndrome JOURNAL OF THE NATIONAL COMPREHENSIVE CANCER NETWORK Henry, E., Villalobos, V., Million, L., Jensen, K. C., West, R., Ganjoo, K., Lebensohn, A., Ford, J. M., Telli, M. L. 2012; 10 (8): 939-942

  Abstract

  Li-Fraumeni syndrome (LFS) is one of the most penetrant forms of familial cancer susceptibility syndromes, characterized by early age at tumor onset and a wide spectrum of malignant tumors. Identifying LFS in patients with cancer is clinically imperative because they have an increased sensitivity to ionizing radiation and are more likely to develop radiation-induced secondary malignancies. This case report describes a young woman whose initial presentation of LFS was early-onset breast cancer and whose treatment of this primary malignancy with breast conservation likely resulted in a secondary malignancy arising in her radiation field. As seen in this case, most breast cancers in patients with LFS exhibit a triple-positive phenotype (estrogen receptor-positive/progesterone receptor-positive/HER2-positive). Although this patient met classic LFS criteria based on age and personal and family history of cancer, the NCCN Clinical Practice Guidelines in Oncology for Genetic/Familial High-Risk Assessment: Breast and Ovarian Cancer endorse genetic screening for TP53 mutations in a subset of patients with early-onset breast cancer, even in the absence of a suggestive family history, because of the potential for de novo TP53 mutations.

  View details for PubMedID 22878818

• Phosphatidylinositol-3-kinase pathway mutations are common in breast columnar cell lesions MODERN PATHOLOGY Troxell, M. L., Brunner, A. L., Neff, T., Warrick, A., Beadling, C., Montgomery, K., Zhu, S., Corless, C. L., West, R. B. 2012; 25 (7): 930-937

  Abstract

  The phosphatidylinositol-3-kinase pathway is one of the most commonly mutated pathways in invasive breast carcinoma, with PIK3CA mutations in ?25% of invasive carcinomas, and AKT1 mutations in up to 5%. Ductal carcinoma in situ, and benign papillomas harbor similar mutations. However, activating point mutations in breast columnar cell lesions have been infrequently studied. Twenty-three breast resection specimens containing columnar cell lesions were identified; 14 with associated invasive carcinoma or carcinoma in situ. DNA extracts were prepared from formalin-fixed paraffin-embedded tissue and screened for a panel of point mutations (321 mutations in 30 genes) using a multiplex PCR panel with mass-spectroscopy readout. PIK3CA mutations were identified in 13/24 columnar cell lesions (54%) and 3/8 invasive carcinomas (37%). The mutation status of columnar cell lesions and associated carcinoma was frequently discordant. Of the 14 cases, only 5 demonstrated the same genotype in matched samples of columnar cell lesions and carcinoma (4 wild type, 1 PIK3CA H1047R). Interestingly, five patients had mutations in columnar cell lesions with wild-type carcinoma; two patients had different point mutations in columnar cell lesions and carcinoma. Only three cases had wild-type columnar cell lesion and mutated carcinoma. The 50% PIK3CA mutation prevalence in columnar cell lesions is greater than reported in most studies of invasive breast cancer. Further, columnar cell lesions and carcinoma were frequently discordant for PIK3CA/AKT1 mutation status. These findings raise interesting questions about the role of PIK3CA/AKT pathway in breast carcinogenesis, and the biologic/precursor potential of columnar cell lesions.

  View details for DOI 10.1038/modpathol.2012.55

  View details for PubMedID 22460814

• ROR2 is a novel prognostic biomarker and a potential therapeutic target in leiomyosarcoma and gastrointestinal stromal tumour JOURNAL OF PATHOLOGY Edris, B., Espinosa, I., Muehlenberg, T., Mikels, A., Lee, C., Steigen, S. E., Zhu, S., Montgomery, K. D., Lazar, A. J., Lev, D., Fletcher, J. A., Beck, A. H., West, R. B., Nusse, R., van de Rijn, M. 2012; 227 (2): 223-233

  Abstract

  Soft-tissue sarcomas are a group of malignant tumours whose clinical management is complicated by morphological heterogeneity, inadequate molecular markers and limited therapeutic options. Receptor tyrosine kinases (RTKs) have been shown to play important roles in cancer, both as therapeutic targets and as prognostic biomarkers. An initial screen of gene expression data for 48 RTKs in 148 sarcomas showed that ROR2 was expressed in a subset of leiomyosarcoma (LMS), gastrointestinal stromal tumour (GIST) and desmoid-type fibromatosis (DTF). This was further confirmed by immunohistochemistry (IHC) on 573 tissue samples from 59 sarcoma tumour types. Here we provide evidence that ROR2 expression plays a role in the invasive abilities of LMS and GIST cells in vitro. We also show that knockdown of ROR2 significantly reduces tumour mass in vivo using a xenotransplantation model of LMS. Lastly, we show that ROR2 expression, as measured by IHC, predicts poor clinical outcome in patients with LMS and GIST, although it was not independent of other clinico-pathological features in a multivariate analysis, and that ROR2 expression is maintained between primary tumours and their metastases. Together, these results show that ROR2 is a useful prognostic indicator in the clinical management of these soft-tissue sarcomas and may represent a novel therapeutic target.

  View details for DOI 10.1002/path.3986

  View details for PubMedID 22294416

• Antibody therapy targeting the CD47 protein is effective in a model of aggressive metastatic leiomyosarcoma PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Edris, B., Weiskopf, K., Volkmer, A. K., Volkmer, J., Willingham, S. B., Contreras-Trujillo, H., Liu, J., Majeti, R., West, R. B., Fletcher, J. A., Beck, A. H., Weissman, I. L., van de Rijn, M. 2012; 109 (17): 6656-6661

  Abstract

  Antibodies against CD47, which block tumor cell CD47 interactions with macrophage signal regulatory protein-?, have been shown to decrease tumor size in hematological and epithelial tumor models by interfering with the protection from phagocytosis by macrophages that intact CD47 bestows upon tumor cells. Leiomyosarcoma (LMS) is a tumor of smooth muscle that can express varying levels of colony-stimulating factor-1 (CSF1), the expression of which correlates with the numbers of tumor-associated macrophages (TAMs) that are found in these tumors. We have previously shown that the presence of TAMs in LMS is associated with poor clinical outcome and the overall effect of TAMs in LMS therefore appears to be protumorigenic. However, the use of inhibitory antibodies against CD47 offers an opportunity to turn TAMs against LMS cells by allowing the phagocytic behavior of resident macrophages to predominate. Here we show that interference with CD47 increases phagocytosis of two human LMS cell lines, LMS04 and LMS05, in vitro. In addition, treatment of mice bearing subcutaneous LMS04 and LMS05 tumors with a novel, humanized anti-CD47 antibody resulted in significant reductions in tumor size. Mice bearing LMS04 tumors develop large numbers of lymph node and lung metastases. In a unique model for neoadjuvant treatment, mice were treated with anti-CD47 antibody starting 1 wk before resection of established primary tumors and subsequently showed a striking decrease in the size and number of metastases. These data suggest that treatment with anti-CD47 antibodies not only reduces primary tumor size but can also be used to inhibit the development of, or to eliminate, metastatic disease.

  View details for DOI 10.1073/pnas.1121629109

  View details for PubMedID 22451919

• Identification of Pathogens in Archival Tissues Using a High-Throughput Sequencing Approach, 35EQ 101st Annual Meeting of United-States-and-Canadian-Academy-of-Pathology (USCAP) SWEENEY, R. T., Brunner, A. L., Montgomery, K. D., Zhu, S. X., Kong, C., Le, Q., West, R. B. NATURE PUBLISHING GROUP. 2012: 467A?467A

  View details for Web of Science ID 000299799902459

• Long Non-Coding RNA and Polycomb Protein Expression Levels Are Increased in Metastatic Versus Primary Breast Carcinoma Chisholm, K. M., Li, R., Wan, Y., Chang, H., West, R. B. NATURE PUBLISHING GROUP. 2012: 29A

  View details for Web of Science ID 000299799900111

• Long Non-Coding RNA and Polycomb Protein Expression Levels Are Increased in Metastatic Versus Primary Breast Carcinoma Chisholm, K. M., Li, R., Wan, Y., Chang, H., West, R. B. NATURE PUBLISHING GROUP. 2012: 29A

  View details for Web of Science ID 000299986900111

• Comparison of Immunohistochemical Stains for Myoepithelial Cells Versus Collagen Type IV in Invasive Ductal Carcinomas and Ductal Carcinoma In Situ of the Breast Kitayama, J. M., West, R. B., Jensen, K. C. NATURE PUBLISHING GROUP. 2012: 46A

  View details for Web of Science ID 000299986900182

• Comparison of Immunohistochemical Stains for Myoepithelial Cells Versus Collagen Type IV in Invasive Ductal Carcinomas and Ductal Carcinoma In Situ of the Breast Kitayama, J. M., West, R. B., Jensen, K. C. NATURE PUBLISHING GROUP. 2012: 46A

  View details for Web of Science ID 000299799900182

• Identification of Pathogens in Archival Tissues Using a High-Throughput Sequencing Approach, 3SEQ 101st Annual Meeting of United-States-and-Canadian-Academy-of-Pathology (USCAP) SWEENEY, R. T., Brunner, A. L., Montgomery, K. D., Zhu, S. X., Kong, C., Le, Q., West, R. B. NATURE PUBLISHING GROUP. 2012: 467A?467A

  View details for Web of Science ID 000299986902459

• 14-3-3 fusion oncogenes in high-grade endometrial stromal sarcoma PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Lee, C., Ou, W., Marino-Enriquez, A., Zhu, M., Mayeda, M., Wang, Y., Guo, X., Brunner, A. L., Amant, F., French, C. A., West, R. B., McAlpine, J. N., Gilks, C. B., Yaffe, M. B., Prentice, L. M., McPherson, A., Jones, S. J., Marra, M. A., Shah, S. P., van de Rijn, M., Huntsman, D. G., Dal Cin, P., Debiec-Rychter, M., Nucci, M. R., Fletcher, J. A. 2012; 109 (3): 929-934

  Abstract

  14-3-3 proteins are ubiquitously expressed regulators of various cellular functions, including proliferation, metabolism, and differentiation, and altered 14-3-3 expression is associated with development and progression of cancer. We report a transforming 14-3-3 oncoprotein, which we identified through conventional cytogenetics and whole-transcriptome sequencing analysis as a highly recurrent genetic mechanism in a clinically aggressive form of uterine sarcoma: high-grade endometrial stromal sarcoma (ESS). The 14-3-3 oncoprotein results from a t(10;17) genomic rearrangement, leading to fusion between 14-3-3? (YWHAE) and either of two nearly identical FAM22 family members (FAM22A or FAM22B). Expression of YWHAE-FAM22 fusion oncoproteins was demonstrated by immunoblot in t(10;17)-bearing frozen tumor and cell line samples. YWHAE-FAM22 fusion gene knockdowns were performed with shRNAs and siRNAs targeting various FAM22A exons in an t(10;17)-bearing ESS cell line (ESS1): Fusion protein expression was inhibited, with corresponding reduction in cell growth and migration. YWHAE-FAM22 maintains a structurally and functionally intact 14-3-3? (YWHAE) protein-binding domain, which is directed to the nucleus by a FAM22 nuclear localization sequence. In contrast to classic ESS, harboring JAZF1 genetic fusions, YWHAE-FAM22 ESS display high-grade histologic features, a distinct gene-expression profile, and a more aggressive clinical course. Fluorescence in situ hybridization analysis demonstrated absolute specificity of YWHAE-FAM22A/B genetic rearrangement for high-grade ESS, with no fusions detected in other uterine and nonuterine mesenchymal tumors (55 tumor types, n = 827). These discoveries reveal diagnostically and therapeutically relevant models for characterizing aberrant 14-3-3 oncogenic functions.

  View details for DOI 10.1073/pnas.1115528109

  View details for Web of Science ID 000299154000058

  View details for PubMedID 22223660

  View details for PubMedCentralID PMC3271913

• Transcriptional profiling of long non-coding RNAs and novel transcribed regions across a diverse panel of archived human cancers. Genome biology Brunner, A. L., Beck, A. H., Edris, B., Sweeney, R. T., Zhu, S. X., Li, R., Montgomery, K., Varma, S., Gilks, T., Guo, X., Foley, J. W., Witten, D. M., Giacomini, C. P., Flynn, R. A., Pollack, J. R., Tibshirani, R., Chang, H. Y., van de Rijn, M., West, R. B. 2012; 13 (8): R75-?

  Abstract

  BACKGROUND: Molecular characterization of tumors has been critical for identifying important genes in cancer biology and for improving tumor classification and diagnosis. Long non-coding RNAs, as a new, relatively unstudied class of transcripts, provide a rich opportunity to identify both functional drivers and cancer-type-specific biomarkers. However, despite the potential importance of long non-coding RNAs to the cancer field, no comprehensive survey of long non-coding RNA expression across various cancers has been reported. RESULTS: We performed a sequencing-based transcriptional survey of both known long non-coding RNAs and novel intergenic transcripts across a panel of 64 archival tumor samples comprising 17 diagnostic subtypes of adenocarcinomas, squamous cell carcinomas and sarcomas. We identified hundreds of transcripts from among the known 1,065 long non-coding RNAs surveyed that showed variability in transcript levels between the tumor types and are therefore potential biomarker candidates. We discovered 1,071 novel intergenic transcribed regions and demonstrate that these show similar patterns of variability between tumor types. We found that many of these differentially expressed cancer transcripts are also expressed in normal tissues. One such novel transcript specifically expressed in breast tissue was further evaluated using RNA in situ hybridization on a panel of breast tumors. It was shown to correlate with low tumor grade and estrogen receptor expression, thereby representing a potentially important new breast cancer biomarker. CONCLUSIONS: This study provides the first large survey of long non-coding RNA expression within a panel of solid cancers and also identifies a number of novel transcribed regions differentially expressed across distinct cancer types that represent candidate biomarkers for future research.

  View details for DOI 10.1186/gb-2012-13-8-r75

  View details for PubMedID 22929540

• Comparative gene expression profiling of benign and malignant lesions reveals candidate therapeutic compounds for leiomyosarcoma. Sarcoma Edris, B., Fletcher, J. A., West, R. B., van de Rijn, M., Beck, A. H. 2012; 2012: 805614-?

  Abstract

  Leiomyosarcoma (LMS) is a malignant, soft-tissue tumor for which few effective therapies exist. Previously, we showed that there are three molecular subtypes of LMS. Here, we analyzed genes differentially expressed in each of the three LMS subtypes as compared to benign leiomyomas and then used the Connectivity Map (cmap) to calculate enrichment scores for the 1309 cmap drugs in order to identify candidate molecules with the potential to induce a benign, leiomyoma-like phenotype in LMS cells. 11 drugs were selected and tested for their ability to inhibit the growth of three human LMS cell lines. We identified two drugs with in vitro efficacy against LMS, one of which had a strongly negative enrichment score (Cantharidin) and the other of which had a strongly positive enrichment score (MG-132). Given MG-132's strong inhibitory effect on LMS cell viability, we hypothesized that LMS cells may be sensitive to treatment with other proteasome inhibitors and demonstrated that bortezomib, a clinically-approved proteasome inhibitor not included in the original cmap screen, potently inhibited the viability of the LMS cell lines. These findings suggest that systematically linking LMS subtype-specific expression signatures with drug-associated expression profiles represents a promising approach for the identification of new drugs for LMS.

  View details for DOI 10.1155/2012/805614

  View details for PubMedID 22919280

• Transcriptional profiling of long non-coding RNAs and novel transcribed regions across a diverse panel of archived human cancers GENOME BIOLOGY Brunner, A. L., Beck, A. H., Edris, B., Sweeney, R. T., Zhu, S. X., Li, R., Montgomery, K., Varma, S., Gilks, T., Guo, X., Foley, J. W., Witten, D. M., Giacomini, C. P., Flynn, R. A., Pollack, J. R., Tibshirani, R., Chang, H. Y., van de Rijn, M., West, R. B. 2012; 13 (8)

  Abstract

  BACKGROUND: Molecular characterization of tumors has been critical for identifying important genes in cancer biology and for improving tumor classification and diagnosis. Long non-coding RNAs, as a new, relatively unstudied class of transcripts, provide a rich opportunity to identify both functional drivers and cancer-type-specific biomarkers. However, despite the potential importance of long non-coding RNAs to the cancer field, no comprehensive survey of long non-coding RNA expression across various cancers has been reported. RESULTS: We performed a sequencing-based transcriptional survey of both known long non-coding RNAs and novel intergenic transcripts across a panel of 64 archival tumor samples comprising 17 diagnostic subtypes of adenocarcinomas, squamous cell carcinomas and sarcomas. We identified hundreds of transcripts from among the known 1,065 long non-coding RNAs surveyed that showed variability in transcript levels between the tumor types and are therefore potential biomarker candidates. We discovered 1,071 novel intergenic transcribed regions and demonstrate that these show similar patterns of variability between tumor types. We found that many of these differentially expressed cancer transcripts are also expressed in normal tissues. One such novel transcript specifically expressed in breast tissue was further evaluated using RNA in situ hybridization on a panel of breast tumors. It was shown to correlate with low tumor grade and estrogen receptor expression, thereby representing a potentially important new breast cancer biomarker. CONCLUSIONS: This study provides the first large survey of long non-coding RNA expression within a panel of solid cancers and also identifies a number of novel transcribed regions differentially expressed across distinct cancer types that represent candidate biomarkers for future research.

  View details for Web of Science ID 000315867500009

• Phosphatidylinositol-3-Kinase Pathway Mutations Are Common in Breast Columnar Cell Lesions. Troxell, M. L., Brunner, A. L., Montgomery, K., Zhu, S. X., Neff, T., Warrick, A., Beadling, C., Corless, C. L., West, R. B. AMER ASSOC CANCER RESEARCH. 2011

  View details for DOI 10.1158/0008-5472.SABCS11-P2-06-04

  View details for Web of Science ID 000209699802230

• Increased midkine expression correlates with desmoid tumour recurrence: a potential biomarker and therapeutic target JOURNAL OF PATHOLOGY Colombo, C., Creighton, C. J., Ghadimi, M. P., Bolshakov, S., Warneke, C. L., Zhang, Y., Lusby, K., Zhu, S., Lazar, A. J., West, R. B., van de Rijn, M., Lev, D. 2011; 225 (4): 574-582

  Abstract

  Desmoid tumours (DTs) are soft tissue monoclonal neoplasms exhibiting a unique phenotype, consisting of aggressive local invasiveness without metastatic capacity. While DTs can infrequently occur as part of familial adenomatosis polyposis, most cases arise sporadically. Sporadic DTs harbour a high prevalence of CTNNB1 mutations and hence increased ?-catenin signalling. However, ?-catenin downstream transcriptional targets and other molecular deregulations operative in DT inception and progression are currently not well defined, contributing to the lack of sensitive molecular prognosticators and efficacious targeted therapeutic strategies. We compared the gene expression profiles of 14 sporadic DTs to those of five corresponding normal tissues and six solitary fibrous tumour specimens. A DT expression signature consisting of 636 up- and 119 down-regulated genes highly enriched for extracellular matrix, cell adhesion and wound healing-related proteins was generated. Furthermore, 98 (15%) of the over-expressed genes were demonstrated to contain a TCF/LEF consensus binding site in their promoters, possibly heralding direct ?-catenin downstream targets relevant to DT. The protein products of three of the up-regulated DT genes: ADAM12, MMP2 and midkine, were found to be commonly expressed in a large cohort of human DT samples assembled on a tissue microarray. Interestingly, enhanced midkine expression significantly correlated with a higher propensity and decreased time for primary DT recurrence (log-rank p = 0.0025). Finally, midkine was found to enhance the migration and invasion of primary DT cell cultures. Taken together, these studies provide insights into potential DT molecular aberrations and novel ?-catenin transcriptional targets. Further studies to confirm the utility of midkine as a clinical DT molecular prognosticator and a potential therapeutic target are therefore warranted. Raw gene array data can be found at: http://smd.stanford.edu/

  View details for DOI 10.1002/path.2951

  View details for Web of Science ID 000297299500011

  View details for PubMedID 21826666

• Systematic Analysis of Breast Cancer Morphology Uncovers Stromal Features Associated with Survival SCIENCE TRANSLATIONAL MEDICINE Beck, A. H., Sangoi, A. R., Leung, S., Marinelli, R. J., Nielsen, T. O., van de Vijver, M. J., West, R. B., van de Rijn, M., Koller, D. 2011; 3 (108)

  Abstract

  The morphological interpretation of histologic sections forms the basis of diagnosis and prognostication for cancer. In the diagnosis of carcinomas, pathologists perform a semiquantitative analysis of a small set of morphological features to determine the cancer's histologic grade. Physicians use histologic grade to inform their assessment of a carcinoma's aggressiveness and a patient's prognosis. Nevertheless, the determination of grade in breast cancer examines only a small set of morphological features of breast cancer epithelial cells, which has been largely unchanged since the 1920s. A comprehensive analysis of automatically quantitated morphological features could identify characteristics of prognostic relevance and provide an accurate and reproducible means for assessing prognosis from microscopic image data. We developed the C-Path (Computational Pathologist) system to measure a rich quantitative feature set from the breast cancer epithelium and stroma (6642 features), including both standard morphometric descriptors of image objects and higher-level contextual, relational, and global image features. These measurements were used to construct a prognostic model. We applied the C-Path system to microscopic images from two independent cohorts of breast cancer patients [from the Netherlands Cancer Institute (NKI) cohort, n = 248, and the Vancouver General Hospital (VGH) cohort, n = 328]. The prognostic model score generated by our system was strongly associated with overall survival in both the NKI and the VGH cohorts (both log-rank P ? 0.001). This association was independent of clinical, pathological, and molecular factors. Three stromal features were significantly associated with survival, and this association was stronger than the association of survival with epithelial characteristics in the model. These findings implicate stromal morphologic structure as a previously unrecognized prognostic determinant for breast cancer.

  View details for DOI 10.1126/scitranslmed.3002564

  View details for PubMedID 22072638

• CSF1 Expression in Nongynecological Leiomyosarcoma Is Associated with Increased Tumor Angiogenesis AMERICAN JOURNAL OF PATHOLOGY Espinosa, I., Edris, B., Lee, C., Cheng, H. W., Gilks, C. B., Wang, Y., Montgomery, K. D., Varma, S., Li, R., Marinelli, R. J., West, R. B., Nielsen, T., Beck, A. H., van de Rijn, M. 2011; 179 (4): 2100-2107

  Abstract

  Leiomyosarcoma (LMS) is a malignant tumor of smooth muscle cells for which few effective therapies exist. A subset of LMS cases express macrophage colony-stimulating factor (CSF1) and the resultant tumor-associated macrophage (TAM) infiltration predicts poor clinical outcome. Further, TAMs have been shown to increase tumor angiogenesis. Here, we analyzed 149 LMS cases by immunohistochemistry for vascular marker CD34 and show that high microvessel density (MVD) in nongynecological LMS cases significantly predicts poor patient outcome. The majority of high MVD cases were also CSF1-positive, and when combining high MVD with CSF1 expression, an even stronger prognostic correlation with patient outcome was obtained. Gene expression profiling revealed that MVD has a stronger correlation with CSF1 expression than with expression of vascular endothelial growth factor isoforms, which have traditionally been used as markers of angiogenesis and as anti-angiogenic therapeutic targets. Finally, patterns of CSF1 expression and TAM recruitment remained consistent between primary tumors and their metastases, and between primary tumors and those grown as xenografts in mice, highlighting the stability of these features to the biology of LMS tumors. Together, these findings suggest an important role for CSF1 and the resulting TAM infiltration in the pathological neovascularization of LMS tumors and provide a rationale for CSF1-targeted therapies in LMS.

  View details for DOI 10.1016/j.ajpath.2011.06.021

  View details for PubMedID 21854753

• Identification of a Disease-Defining Gene Fusion in Epithelioid Hemangioendothelioma SCIENCE TRANSLATIONAL MEDICINE Tanas, M. R., Sboner, A., Oliveira, A. M., Erickson-Johnson, M. R., Hespelt, J., Hanwright, P. J., Flanagan, J., Luo, Y., Fenwick, K., Natrajan, R., Mitsopoulos, C., Zvelebil, M., Hoch, B. L., Weiss, S. W., Debiec-Rychter, M., Sciot, R., West, R. B., Lazar, A. J., Ashworth, A., Reis-Filho, J. S., Lord, C. J., Gerstein, M. B., Rubin, M. A., Rubin, B. P. 2011; 3 (98)

  Abstract

  Integrating transcriptomic sequencing with conventional cytogenetics, we identified WWTR1 (WW domain-containing transcription regulator 1) (3q25) and CAMTA1 (calmodulin-binding transcription activator 1) (1p36) as the two genes involved in the t(1;3)(p36;q25) chromosomal translocation that is characteristic of epithelioid hemangioendothelioma (EHE), a vascular sarcoma. This WWTR1/CAMTA1 gene fusion is under the transcriptional control of the WWTR1 promoter and encodes a putative chimeric transcription factor that joins the amino terminus of WWTR1, a protein that is highly expressed in endothelial cells, in-frame to the carboxyl terminus of CAMTA1, a protein that is normally expressed only in brain. Thus, CAMTA1 expression is activated inappropriately through a promoter-switch mechanism. The gene fusion is present in virtually all EHEs tested but is absent from all other vascular neoplasms, demonstrating it to be a disease-defining genetic alteration. A sensitive and specific break-apart fluorescence in situ hybridization assay was also developed to detect the translocation and will assist in the evaluation of this diagnostically challenging neoplasm. The chimeric WWTR1/CAMTA1 transcription factor may represent a therapeutic target for EHE and offers the opportunity to shed light on the functions of two poorly characterized proteins.

  View details for DOI 10.1126/scitranslmed.3002409

  View details for Web of Science ID 000294462100004

  View details for PubMedID 21885404

• Immunohistochemical Distinction of Primary Adrenal Cortical Lesions From Metastatic Clear Cell Renal Cell Carcinoma: A Study of 248 Cases AMERICAN JOURNAL OF SURGICAL PATHOLOGY Sangoi, A. R., Fujiwara, M., West, R. B., Montgomery, K. D., Bonventre, J. V., Higgins, J. P., Rouse, R. V., Gokden, N., McKenney, J. K. 2011; 35 (5): 678-686

  Abstract

  The diagnosis of metastatic clear cell renal cell carcinoma (CC-RCC) can be difficult because of its morphologic heterogeneity and the increasing use of small image-guided biopsies that yield scant diagnostic material. This is further complicated by the degree of morphologic and immunophenotypic overlap with nonrenal neoplasms and tissues, such as adrenal cortex. In this study, a detailed immunoprofile of 63 adrenal cortical lesions, which included 54 cortical neoplasms, was compared with 185 metastatic CC-RCCs using traditional [anticalretinin, CD10, antichromogranin, antiepithelial membrane antigen, anti-inhibin, antimelanA, anticytokeratins (AE1/AE3 and AE1/CAM5.2), antirenal cell carcinoma marker, and antisynaptophysin)] and novel [anticarbonic anhydrase-IX, antihepatocyte nuclear factor-1b, antihuman kidney injury molecule-1 (hKIM-1), anti-PAX-2, anti-PAX-8, antisteroidogenic factor-1 (SF-1), and anti-T-cell immunoglobulin mucin-1] antibodies. Tissue microarray methodology was used to simulate small image-guided biopsies. Staining extent and intensity were scored semiquantitatively for each antibody. In comparing different intensity thresholds required for a "positive" result, a value of ?2+ was identified as optimal for diagnostic sensitivity/specificity. For the distinction of adrenal cortical lesions from metastatic CC-RCCs, immunoreactivity for the adrenal cortical antigens SF-1 (86% adrenal; 0% CC-RCC), calretinin (89% adrenal; 10% CC-RCC), inhibin (86% adrenal; 9% CC-RCC), and melanA (86% adrenal; 10% CC-RCC) and for the renal epithelial antigens hKIM-1 (0% adrenal; 83% CC-RCC), PAX-8 (0% adrenal; 83% CC-RCC), hepatocyte nuclear factor-1b (0% adrenal; 76% CC-RCC), epithelial membrane antigen (0% adrenal; 78% CC-RCC), and carbonic anhydrase-IX (3% adrenal; 87% CC-RCC) had the most potential use. Use of novel renal epithelial markers hKIM-1 (clone AKG7) and/or PAX-8 and the adrenocortical marker SF-1 in an immunohistochemical panel for distinguishing adrenal cortical lesions from metastatic CC-RCC offers improved diagnostic sensitivity and specificity.

  View details for DOI 10.1097/PAS.0b013e3182152629

  View details for Web of Science ID 000289506600007

  View details for PubMedID 21490444

• Expression of Subtype-Specific Group 1 Leiomyosarcoma Markers in a Wide Variety of Sarcomas by Gene Expression Analysis and Immunohistochemistry AMERICAN JOURNAL OF SURGICAL PATHOLOGY Mills, A. M., Beck, A. H., Montgomery, K. D., Zhu, S. X., Espinosa, I., Lee, C., Subramanian, S., Fletcher, C. D., van de Rijn, M., West, R. B. 2011; 35 (4): 583-589

  Abstract

  Leiomyosarcomas (LMSs) constitute approximately one quarter of all sarcomas and are usually defined by morphologic criteria and/or immunoreactivity for actin or desmin. Among high-grade lesions, the distinction from undifferentiated pleomorphic sarcoma (UPS) can be problematic, and previous studies have shown that a significant number of LMS cases may be hiding under the diagnosis of UPS. We recently described 3 novel molecular LMS subtypes that are distributed similarly over LMSs of gyneocologic and non-gyneocologic origins. The group 1 subtype shows an improved disease-specific survival compared with the other 2 groups that is independent of histologic grade. Group 1 comprises approximately 25% of all LMSs, and is defined by a shared pattern of gene expression, a distinct pattern of genomic changes, and reactivity for at least 3 of 5 immunohistochemistry (IHC) markers (smooth muscle gamma actin, calsequestrin 2, human muscle cofilin2, myosin light chain kinase, and sarcolemmal membrane associated protein), as tested on 271 cases of LMS in tissue microarrays. These IHC markers have not been well characterized in non-LMS sarcomas. Here we provide a characterization of these 5 markers across normal tissues, an additional 59 cases of LMS, and a wide range of 565 non-LMS soft tissue tumors from 44 diagnostic categories, with a focus on UPS. When analyzed individually, the 5 markers were found to be expressed in many sarcomas other than LMSs. However, when analyzed by the same criteria used for the recognition of group 1 LMSs, in which a case is scored positive when at least 3 of 5 markers reacted, coordinate expression was seen in significant numbers of cases from only 3 diagnostic groups that included 22% of leiomyomas (n=22), 16% of gastrointestinal stromal tumors (n=43), and 18% of endometrial stromal sarcomas (n=11). In addition, 5% (n=57) of UPSs showed a staining pattern similar to that seen in group 1 LMSs. To further examine the possibility that group 1 LMS constitutes a small part of cases diagnosed as UPS, we examined the expression of the top 500 genes from the group 1 LMS expression signature in 29 UPSs by complementary DNA microarray. Unsupervised hierarchical clustering of 29 UPS expression showed that 2 (7%) had coordinated high levels of expression of genes from the group 1 LMS signature, a rate similar to that seen by IHC analysis. These findings show that group 1 LMS IHC markers smooth muscle gamma actin, calsequestrin 2, human muscle cofilin2, myosin light chain kinase, and sarcolemmal membrane associated protein when coordinately expressed have specificity for a subset of LMS when compared with other sarcomas, and may be useful for the recognition of group 1 LMS cases within cases diagnosed as UPS.

  View details for DOI 10.1097/PAS.0b013e318211abd6

  View details for PubMedID 21412072

• The Prognostic Value of Tumor-Associated Macrophages in Leiomyosarcoma A Single Institution Study AMERICAN JOURNAL OF CLINICAL ONCOLOGY-CANCER CLINICAL TRIALS Ganjoo, K. N., Witten, D., Patel, M., Espinosa, I., La, T., Tibshirani, R., van de Rijn, M., Jacobs, C., West, R. B. 2011; 34 (1): 82-86

  Abstract

  High numbers of tumor-associated macrophages (TAMs) have been associated with poor outcome in several solid tumors. In 2 previous studies, we showed that colony stimulating factor-1 (CSF1) is secreted by leiomyosarcoma (LMS) and that the increase in macrophages and CSF1 associated proteins are markers for poor prognosis in both gynecologic and nongynecologic LMS in a multicentered study. The purpose of this study is to evaluate the outcome of patients with LMS from a single institution according to the number of TAMs evaluated through 3 CSF1 associated proteins.Patients with LMS treated at Stanford University with adequate archived tissue and clinical data were eligible for this retrospective study. Data from chart reviews included tumor site, size, grade, stage, treatment, and disease status at the time of last follow-up. The 3 CSF1 associated proteins (CD163, CD16, and cathepsin L) were evaluated by immunohistochemistry on tissue microarrays. Kaplan-Meier survival curves and univariate Cox proportional hazards models were fit to assess the association of clinical predictors as well as CSF1 associated proteins with overall survival.A total of 52 patients diagnosed from 1983 to 2007 were evaluated. Univariate Cox proportional hazards models were fit to assess the significance of grade, size, stage, and the 3 CSF1 associated proteins in predicting OS. Grade, size, and stage were not significantly associated with survival in the full patient cohort, but grade and stage were significant predictors of survival in the gynecologic (GYN) LMS samples (P = 0.038 and P = 0.0164, respectively). Increased cathepsin L was associated with a worse outcome in GYN LMS (P = 0.049). Similar findings were seen with CD16 (P < 0.0001). In addition, CSF1 response enriched (all 3 stains positive) GYN LMS had a poor overall survival when compared with CSF1 response poor tumors (P = 0.001). These results were not seen in non-GYN LMS.Our data form an independent confirmation of the prognostic significance of TAMs and the CSF1 associated proteins in LMS. More aggressive or targeted therapies could be considered in the subset of LMS patients that highly express these markers.

  View details for DOI 10.1097/COC.0b013e3181d26d5e

  View details for PubMedID 23781555

• Long Non-Coding RNA Expression Levels Correlate with Proliferation Index in Breast Carcinoma Chisholm, K. M., Li, R., Wan, Y., Chang, H. Y., West, R. B. NATURE PUBLISHING GROUP. 2011: 31A?32A

  View details for Web of Science ID 000287011400122

• Beta-Adrenergic Receptor Expression in Adult Vascular Tumors Chisholm, K. M., Chang, K. W., Truong, M. T., Kwok, S., West, R., Heerema-McKenney, A. E. NATURE PUBLISHING GROUP. 2011: 10A?11A

  View details for Web of Science ID 000287282300028

• Beta-Adrenergic Receptor Expression in Adult Vascular Tumors Chisholm, K. M., Chang, K. W., Truong, M. T., Kwok, S., West, R., Heerema-McKenney, A. E. NATURE PUBLISHING GROUP. 2011: 10A?11A

  View details for Web of Science ID 000287011400028

• ROR2: A Potential Therapeutic Target in Soft Tissue Tumors Edris, B., Espinosa, I., Beck, A. H., Lee, C., Lazar, A. J., Lev, D., Fletcher, J. A., West, R. B., Nusse, R., van de Rijn, M. NATURE PUBLISHING GROUP. 2011: 13A

  View details for Web of Science ID 000287282300038

• SOX10 and S100 in the Diagnosis of Soft Tissue Neoplasms. Karamchandani, J. R., Nielsen, T., Van de Rijn, M., West, R. B. NATURE PUBLISHING GROUP. 2011: 16A

  View details for Web of Science ID 000287282300054

• SOX10 and S100 in the Diagnosis of Soft Tissue Neoplasms Karamchandani, J. R., Nielsen, T., de Rijn, M., West, R. B. NATURE PUBLISHING GROUP. 2011: 16A

  View details for Web of Science ID 000287011400054

• Long Non-Coding RNA Expression Levels Correlate with Proliferation Index in Breast Carcinoma. Chisholm, K. M., Li, R., Wan, Y., Chang, H. Y., West, R. B. NATURE PUBLISHING GROUP. 2011: 31A?32A

  View details for Web of Science ID 000287282300122

• PAX8 and WT1 Are Superior to PAX2 and BRST2 in Distinguishing Mullerian Tract Tumors from Breast Carcinomas 100th Annual Meeting United States-and-Canadian-Academy-of-Pathology DiMaio, M. A., Beck, A. H., Montgomery, K. D., West, R. B., Longacre, T. A., Kong, C. S. NATURE PUBLISHING GROUP. 2011: 243A?243A

  View details for Web of Science ID 000287282301183

• PAX8 and WT1 Are Superior to PAX2 and BRST2 in Distinguishing Mullerian Tract Tumors from Breast Carcinomas. 100th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology DiMaio, M. A., Beck, A. H., Montgomery, K. D., West, R. B., Longacre, T. A., Kong, C. S. NATURE PUBLISHING GROUP. 2011: 243A?243A

  View details for Web of Science ID 000291285000468

• Computational Image Analysis Identifies New Morphologic Features That Predict Breast Cancer Outcome. 100th Annual Meeting United States-and-Canadian-Academy-of-Pathology Beck, A. H., West, R. B., van De Vijver, M., Koller, D. NATURE PUBLISHING GROUP. 2011: 28A?28A

  View details for Web of Science ID 000287282300106

• CSF1-Response Signature Is Associated with Tumor Angiogenesis in Non-Gynecological Leiomyosarcoma 100th Annual Meeting United States-and-Canadian-Academy-of-Pathology Espinosa, I., Beck, A., Edris, B., Lee, C., West, R., van de Rijn, M. NATURE PUBLISHING GROUP. 2011: 13A?13A

  View details for Web of Science ID 000287282300041

• CSF1-Response Signature Is Associated with Tumor Angiogenesis in Non-Gynecological Leiomyosarcoma 100th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Espinosa, I., Beck, A., Edris, B., Lee, C., West, R., van de Rijn, M. NATURE PUBLISHING GROUP. 2011: 13A?13A

  View details for Web of Science ID 000287011400041

• Computational Image Analysis Identifies New Morphologic Features That Predict Breast Cancer Outcome 100th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Beck, A. H., West, R. B., van De Vijver, M., Koller, D. NATURE PUBLISHING GROUP. 2011: 28A?28A

  View details for Web of Science ID 000287011400106

• Endogenous Versus Tumor-Specific Host Response to Breast Carcinoma: A Study of Stromal Response in Synchronous Breast Primaries and Biopsy Site Changes CLINICAL CANCER RESEARCH Wu, J. M., Beck, A. H., Pate, L. L., Witten, D., Zhu, S. X., Montgomery, K. D., Allison, K. H., van de Rijn, M., West, R. B. 2011; 17 (3): 437-446

  Abstract

  We recently described two types of stromal response in breast cancer derived from gene expression studies of tenosynovial giant cell tumors and fibromatosis. The purpose of this study is to elucidate the basis of this stromal response--whether they are elicited by individual tumors or whether they represent an endogenous host reaction produced by the patient.Stromal signatures from patients with synchronous dual primaries were analyzed by immunohistochemistry on a tissue microarray (n = 26 pairs) to evaluate the similarity of stromal responses in different tumors within the same patient. We also characterized the extent to which the stromal signatures were conserved between stromal response to injury compared to the stromal response to carcinoma using gene expression profiling and tissue microarray immunohistochemistry.The two stromal response signatures showed divergent associations in synchronous primaries: the DTF fibroblast response is more likely to be similar in a patient with multiple breast primaries (permutation analysis P = 0.0027), whereas CSF1 macrophage response shows no significant concordance in separate tumors within a given patient. The DTF fibroblast signature showed more concordance across normal, cancer, and biopsy site samples from within a patient, than across normal, cancer, and biopsy site samples from a random group of patients, whereas the CSF1 macrophage response did not.The results suggest that the DTF fibroblast response is host-specific, whereas the CSF1 response may be tumor-elicited. Our findings provide further insight into stromal response and may facilitate the development of therapeutic strategies to target particular stromal subtypes.

  View details for DOI 10.1158/1078-0432.CCR-10-1709

  View details for PubMedID 21098336

• MYB Expression and Translocation in Adenoid Cystic Carcinomas and Other Salivary Gland Tumors With Clinicopathologic Correlation AMERICAN JOURNAL OF SURGICAL PATHOLOGY West, R. B., Kong, C., Clarke, N., Gilks, T., Lipsick, J. S., Cao, H., Kwok, S., Montgomery, K. D., Varma, S., Le, Q. 2011; 35 (1): 92-99

  Abstract

  Adenoid cystic carcinoma is a locally aggressive salivary gland neoplasm, which has a poor long-term prognosis. A chromosomal translocation involving the genes encoding the transcription factors, MYB and NFIB, has been recently discovered in these tumors.MYB translocation and protein expression were studied in 37 adenoid cystic carcinomas, 112 other salivary gland neoplasms, and 409 nonsalivary gland neoplasms by fluorescence in situ hybridization and immunohistochemistry. MYB translocation and expression status in adenoid cystic carcinoma was correlated with clinicopathologic features including outcome, with a median follow-up of 77.1 months (range, 23.2 to 217.5 mo) for living patients.A balanced translocation between MYB and NFIB is present in 49% of adenoid cystic carcinomas but is not identified in other salivary gland tumors or nonsalivary gland neoplasms. There is no apparent translocation of MYB in 35% of the cases. Strong Myb immunostaining is very specific for adenoid cystic carcinomas but is only present in 65% of all cases. It is interesting to note that Myb immunostaining is confined to the basal cell component although the translocation is present in all the cells. Neoplasms with MYB translocation show a trend toward higher local relapse rates, but the results are not statistically significant with the current number of cases.MYB translocation and expression are useful diagnostic markers for a subset of adenoid cystic carcinomas. The presence of the translocation may be indicative of local aggressive behavior, but a larger cohort may be required to show statistical significance.

  View details for DOI 10.1097/PAS.0b013e3182002777

  View details for PubMedID 21164292

• The Relationship between Histologic Grade, Ethnicity, EBV, and HPV Status in Nasopharyngeal Carcinoma Khong, B., Cao, H., Kwok, S., West, R., Kong, C. S., Le, Q. ELSEVIER SCIENCE INC. 2011: S177

  View details for DOI 10.1016/j.ijrobp.2011.06.1692

  View details for Web of Science ID 000296411700356

• Variations in stromal signatures in breast and colorectal cancer metastases JOURNAL OF PATHOLOGY Webster, J. A., Beck, A. H., Sharma, M., Espinosa, I., Weigelt, B., Schreuder, M., Montgomery, K. D., Jensen, K. C., van de Rijn, M., West, R. 2010; 222 (2): 158-165

  Abstract

  The tumour microenvironment (TME) plays an important role in tumour survival and growth, but little is known about the degree of preservation between different stromal response patterns found in primary tumours and their metastases. We have previously identified gene expression profiles for two distinct stromal signatures in breast carcinoma of fibroblast (aka DTF) and macrophage (aka CSF1) response and found them to be correlated with clinicopathological features, including outcome. In this study, we compare the DTF fibroblast and CSF1 macrophage stromal response patterns in primary breast and colorectal cancers to their matched lymph node metastases. In both breast and colorectal cancer, there was a significant positive correlation between the CSF1 macrophage signature in the primary tumours and the matched lymph node metastases, as assessed by immunohistochemical markers. No such correlation was observed for the DTF fibroblast signature. A similar result was seen in independent analysis of two published gene expression microarray datasets. The variations of these stromal reaction patterns from the primary to the metastasis shed light on the relationship between the neoplastic cells and the non-neoplastic cells in the TME. The preservation of the CSF1 macrophage response pattern in metastases lends support to targeting the CSF1 pathway in cancer.

  View details for DOI 10.1002/path.2738

  View details for PubMedID 20593409

• Analysis of stromal signatures in the tumor microenvironment of ductal carcinoma in situ BREAST CANCER RESEARCH AND TREATMENT Sharma, M., Beck, A. H., Webster, J. A., Espinosa, I., Montgomery, K., Varma, S., van de Rijn, M., Jensen, K. C., West, R. B. 2010; 123 (2): 397-404

  Abstract

  Recent advances in the study of the tumor microenvironment have revealed significant interaction between tumor cells and their surrounding stroma in model systems. We have previously shown that two distinct stromal signatures derived from a macrophage (CSF1) response and a fibroblastic (DTF-like) response are present in subsets of invasive breast cancers and show a correlation with clinical outcome. In the present study we explore whether these signatures also exist in the stroma of ductal carcinoma in situ (DCIS). We studied the signatures by both gene expression profile analysis of a publically available data set of DCIS and by immunohistochemistry (IHC) on a tissue microarray of DCIS and invasive breast cancer cases. Both the gene expression and immunohistochemical data show that the macrophage response and fibroblast expression signatures are present in the stroma of subsets of DCIS cases. The incidence of the stromal signatures in DCIS is similar to the incidence in invasive breast cancer that we have previously reported. We also find that the macrophage response signature is associated with higher grade DCIS and cases which are ER and PR negative, whereas the fibroblast signature was not associated with any clinicopathologic features in DCIS. A comparison of 115 matched cases of DCIS and invasive breast cancer found a correlation between the type of stromal response in DCIS and invasive ductal carcinoma (IDC) within the same patient for both the macrophage response and the fibroblast stromal signatures (P = 0.03 and 0.08, respectively). This study is a first characterization of these signatures in DCIS. These signatures have significant clinicopathologic associations and tend to be conserved as the tumor progresses from DCIS to invasive breast cancer.

  View details for DOI 10.1007/s10549-009-0654-0

  View details for Web of Science ID 000280807900008

  View details for PubMedID 19949854

  View details for PubMedCentralID PMC2976659

• Expression Profiling in Soft Tissue Sarcomas With Emphasis on Synovial Sarcoma, Gastrointestinal Stromal Tumor, and Leiomyosarcoma ADVANCES IN ANATOMIC PATHOLOGY West, R. B. 2010; 17 (5): 366-373

  Abstract

  Sarcomas are defined as malignant neoplasms derived from mesenchymal tissues. A variety of different molecular approaches, including gene expression profiling, have identified candidate biomarkers and insights into sarcoma biology that will aid in the diagnosis and treatment of these tumors. Many gene expression profiling findings have been translated into immunohistochemical tests for diagnostic, prognostic, or predictive purposes. This review details gene expression studies done in 3 sarcomas, synovial sarcoma, gastrointestinal stromal tumor, and leiomyosarcoma.

  View details for DOI 10.1097/PAP.0b013e3181ec7428

  View details for Web of Science ID 000281182300005

  View details for PubMedID 20733355

• DOG1 for the Diagnosis of Gastrointestinal Stromal Tumor (GIST): Comparison Between 2 Different Antibodies APPLIED IMMUNOHISTOCHEMISTRY & MOLECULAR MORPHOLOGY Lopes, L. F., West, R. B., Bacchi, L. M., van de Rijn, M., Bacchi, C. E. 2010; 18 (4): 333-337

  Abstract

  Gastrointestinal stromal tumor (GIST) is the most common mesenchymal neoplasm of the gastrointestinal tract. Discovered on GIST-1 (DOG1) is a recently described protein expressed in GISTs irrespective of mutation status. The aim of this study was to investigate the immunohistochemical expression of DOG1 using 2 different monoclonal antibodies (DOG1.1 and the commercially available K9 antibody) in 668 GIST cases and to compare the results with the expression of KIT. DOG1 and KIT expression also were studied in most human normal tissues and several nonmesenchymal and mesenchymal tumors other than GIST. KIT was expressed in 643 (96.3%) GISTs. DOG1.1 and K9 were positive in 538 (80.5%) and 642 (96.1%) GIST cases, respectively. In 25 (3.7%) KIT-negative GIST cases, DOG1 was expressed in 5 (20.0%) and 19 (76.0%) using DOG1.1 and K9 antibodies, respectively. Only 0.9% of GISTs were negative for KIT, DOG1.1, and K9. Most normal human tissues did not reveal KIT and DOG1 expression. DOG1.1 was positive in only 2 of 57 synovial sarcomas and 1 of 61 soft tissue leiomyosarcomas. K9 was positive in 5 of 57 synovial sarcomas, 1 of 14 angiosarcomas, 1 of 61 soft tissue leiomyosarcomas, 3 of 4 adenoid cystic carcinomas of the head and neck, and in myoepithelial cells of 9 of 11 fibroadenomas of the breast. In conclusion, the commercially available K9 is of great utility for the diagnosis of most KIT-negative GISTs, and the combination of both KIT and K9 antibody in a panel of immunohistochemistry can define the diagnosis of GIST in more than 99% of cases.

  View details for DOI 10.1097/PAI.0b013e3181d27ec8

  View details for Web of Science ID 000279059900004

  View details for PubMedID 20571340

• Long non-coding RNA HOTAIR reprograms chromatin state to promote cancer metastasis NATURE Gupta, R. A., Shah, N., Wang, K. C., Kim, J., Horlings, H. M., Wong, D. J., Tsai, M., Hung, T., Argani, P., Rinn, J. L., Wang, Y., Brzoska, P., Kong, B., Li, R., West, R. B., van de Vijver, M. J., Sukumar, S., Chang, H. Y. 2010; 464 (7291): 1071-U148

  Abstract

  Large intervening non-coding RNAs (lincRNAs) are pervasively transcribed in the genome yet their potential involvement in human disease is not well understood. Recent studies of dosage compensation, imprinting, and homeotic gene expression suggest that individual lincRNAs can function as the interface between DNA and specific chromatin remodelling activities. Here we show that lincRNAs in the HOX loci become systematically dysregulated during breast cancer progression. The lincRNA termed HOTAIR is increased in expression in primary breast tumours and metastases, and HOTAIR expression level in primary tumours is a powerful predictor of eventual metastasis and death. Enforced expression of HOTAIR in epithelial cancer cells induced genome-wide re-targeting of Polycomb repressive complex 2 (PRC2) to an occupancy pattern more resembling embryonic fibroblasts, leading to altered histone H3 lysine 27 methylation, gene expression, and increased cancer invasiveness and metastasis in a manner dependent on PRC2. Conversely, loss of HOTAIR can inhibit cancer invasiveness, particularly in cells that possess excessive PRC2 activity. These findings indicate that lincRNAs have active roles in modulating the cancer epigenome and may be important targets for cancer diagnosis and therapy.

  View details for DOI 10.1038/nature08975

  View details for Web of Science ID 000276635000045

  View details for PubMedID 20393566

  View details for PubMedCentralID PMC3049919

• Translating Gene Expression Into Clinical Care: Sarcomas As a Paradigm JOURNAL OF CLINICAL ONCOLOGY Nielsen, T. O., West, R. B. 2010; 28 (10): 1796-1805

  Abstract

  Whereas most solid tumors are characterized by considerable genetic instability and molecular heterogeneity, sarcomas include many subtypes with very specific underlying molecular events driving oncogenesis. Gene expression profiling and other modern techniques have consequently had particular success in identifying the critical biologic pathways active in specific sarcomas, yielding insights which can be translated into useful diagnostic biomarkers. Public availability of data sets and new sequencing-based technologies will accelerate this process. Molecular studies have also identified oncogenic pathways of particular importance in sarcomas which can be targeted by investigational drugs. Examples include histone deacetylases in translocation-associated sarcomas of young adults, Akt/mammalian target of rapamycin in pleomorphic sarcomas, and macrophage colony-stimulating factor in tenosynovial giant cell tumor. Despite challenges in organization and accrual, future clinical trials of sarcomas need to be designed that take into account specific molecular subtypes as distinct diseases.

  View details for DOI 10.1200/JCO.2009.26.1917

  View details for Web of Science ID 000276152200028

  View details for PubMedID 20194847

• Discovery of molecular subtypes in leiomyosarcoma through integrative molecular profiling ONCOGENE Beck, A. H., Lee, C., WITTEN, D. M., Gleason, B. C., Edris, B., Espinosa, I., Zhu, S., Li, R., Montgomery, K. D., Marinelli, R. J., Tibshirani, R., Hastie, T., Jablons, D. M., Rubin, B. P., Fletcher, C. D., West, R. B., van de Rijn, M. 2010; 29 (6): 845-854

  Abstract

  Leiomyosarcoma (LMS) is a soft tissue tumor with a significant degree of morphologic and molecular heterogeneity. We used integrative molecular profiling to discover and characterize molecular subtypes of LMS. Gene expression profiling was performed on 51 LMS samples. Unsupervised clustering showed three reproducible LMS clusters. Array comparative genomic hybridization (aCGH) was performed on 20 LMS samples and showed that the molecular subtypes defined by gene expression showed distinct genomic changes. Tumors from the 'muscle-enriched' cluster showed significantly increased copy number changes (P=0.04). A majority of the muscle-enriched cases showed loss at 16q24, which contains Fanconi anemia, complementation group A, known to have an important role in DNA repair, and loss at 1p36, which contains PRDM16, of which loss promotes muscle differentiation. Immunohistochemistry (IHC) was performed on LMS tissue microarrays (n=377) for five markers with high levels of messenger RNA in the muscle-enriched cluster (ACTG2, CASQ2, SLMAP, CFL2 and MYLK) and showed significantly correlated expression of the five proteins (all pairwise P<0.005). Expression of the five markers was associated with improved disease-specific survival in a multivariate Cox regression analysis (P<0.04). In this analysis that combined gene expression profiling, aCGH and IHC, we characterized distinct molecular LMS subtypes, provided insight into their pathogenesis, and identified prognostic biomarkers.

  View details for DOI 10.1038/onc.2009.381

  View details for Web of Science ID 000274397800007

  View details for PubMedID 19901961

  View details for PubMedCentralID PMC2820592

• Gene expression profiling for the investigation of soft tissue sarcoma pathogenesis and the identification of diagnostic, prognostic, and predictive biomarkers VIRCHOWS ARCHIV Beck, A. H., West, R. B., van de Rijn, M. 2010; 456 (2): 141-151

  Abstract

  Soft tissue sarcomas are malignant neoplasms derived from mesenchymal tissues. Their pathogenesis is poorly understood and there are few effective treatment options for advanced disease. In the past decade, gene expression profiling has been applied to sarcomas to facilitate understanding of sarcoma pathogenesis and to identify diagnostic, prognostic, and predictive markers. In this paper, we review this body of work and discuss how gene expression profiling has led to advancements in the understanding of sarcoma pathobiology, the identification of clinically useful biomarkers, and the refinement of sarcoma classification schemes. Lastly, we conclude with a discussion of strategies to further optimize the translation of gene expression data into a greater understanding of sarcoma pathogenesis and improved clinical outcomes for sarcoma patients.

  View details for DOI 10.1007/s00428-009-0774-2

  View details for PubMedID 19412622

• 3 '-End Sequencing for Expression Quantification (3SEQ) from Archival Tumor Samples Beck, A. H., Weng, Z., Witten, D. M., Zhu, S., Foley, J. W., Lacroute, P., Smith, C. L., Tibshirani, R., van de Rijn, M., Sidow, A., West, R. B. NATURE PUBLISHING GROUP. 2010: 384A

  View details for Web of Science ID 000274337301728

• 3 '-End Sequencing for Expression Quantification (3SEQ) from Archival Tumor Samples Beck, A. H., Weng, Z., Witten, D. M., Zhu, S., Foley, J. W., Lacroute, P., Smith, C. L., Tibshirani, R., van de Rijn, M., Sidow, A., West, R. B. NATURE PUBLISHING GROUP. 2010: 384A

  View details for Web of Science ID 000274582502394

• Meta-Mining of Gene Expression Profiles Identifies MG132 and Cantharidin as Therapeutic Agents with In Vitro Activity Against Leiomyosarcoma Edris, B., West, R. B., Lee, C. H., Zhu, M., Fletcher, J., Van de Rijn, M., Beck, A. H. NATURE PUBLISHING GROUP. 2010: 19A

  View details for Web of Science ID 000274582500073

• Expression of Group I Leiomyosarcoma Markers by Gene Expression Analysis and Immunohistochemistry in a Subset of Undifferentiated Pleomorphic Sarcomas Mills, A. M., Beck, A. H., van de Rijn, M., West, R. B. NATURE PUBLISHING GROUP. 2010: 24A?25A

  View details for Web of Science ID 000274582500098

• Expression of Group I Leiomyosarcoma Markers by Gene Expression Analysis and Immunohistochemistry in a Subset of Undifferentiated Pleomorphic Sarcomas Mills, A. M., Beck, A. H., van de Rijn, M., West, R. B. NATURE PUBLISHING GROUP. 2010: 24A?25A

  View details for Web of Science ID 000274337300098

• Loss of Stromal Caveolin-1 Independently Predicts Poor Disease-Free Survival and Time to Recurrence in Patients with Prostate Cancer Wagner, M., Dasgupta, A., Sotgia, F., West, R. B., Lisanti, M. P., Witkiewicz, A. K. NATURE PUBLISHING GROUP. 2010: 226A

  View details for Web of Science ID 000274582501346

• Histologic and Immunophenotypic Comparison of Estrogen Receptor (ER)-Positive Breast Cancers in BRCA1 Mutation Carriers and Sporadic ER-Positive Breast Cancers: A Case-Control Study Kaplan, J., Schnitt, S., Collins, L., Wang, Y., Garber, J., Montgomery, K., West, R., Tung, N. NATURE PUBLISHING GROUP. 2010: 55A

  View details for Web of Science ID 000274582500235

• Variations in Stromal Signatures in Breast Cancer Metastases Webster, J. A., Beck, A. H., Sharma, M., Espinosa, I., Schreuder, M., Montgomery, K. D., Jensen, K. C., van de Rijn, M., West, R. B. NATURE PUBLISHING GROUP. 2010: 77A

  View details for Web of Science ID 000274337300335

• Loss of Stromal Caveolin-1 Independently Predicts Poor Disease-Free Survival and Time to Recurrence in Patients with Prostate Cancer 99th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Wagner, M., Dasgupta, A., Sotgia, F., West, R. B., Lisanti, M. P., Witkiewicz, A. K. NATURE PUBLISHING GROUP. 2010: 226A?226A

  View details for Web of Science ID 000274337301027

• Meta-Mining of Gene Expression Profiles Identifies MG132 and Cantharidin as Therapeutic Agents with In Vitro Activity Against Leiomyosarcoma 99th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Edris, B., West, R. B., Lee, C. H., Zhu, M., Fletcher, J., van de Rijn, M., Beck, A. H. NATURE PUBLISHING GROUP. 2010: 19A?19A

  View details for Web of Science ID 000274337300073

• Fibroblast-Like Stromal Response Is Host-Dependent While Macrophage-Associated Stromal Response Is Tumor-Dependent: A Study of Stromal Response in Paired Breast Carcinomas from Patients with Dual Primaries 99th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Wu, J. M., Beck, A. H., WITTEN, D., Allison, K., van de Rijn, M., West, R. B. NATURE PUBLISHING GROUP. 2010: 79A?79A

  View details for Web of Science ID 000274337300346

• Histologic and Immunophenotypic Comparison of Estrogen Receptor (ER)-Positive Breast Cancers in BRCA1 Mutation Carriers and Sporadic ER-Positive Breast Cancers: A Case-Control Study 99th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Kaplan, J., Schnitt, S., Collins, L., Wang, Y., Garber, J., Montgomery, K., West, R., Tung, N. NATURE PUBLISHING GROUP. 2010: 55A?55A

  View details for Web of Science ID 000274337300235

• Variations in Stromal Signatures in Breast Cancer Metastases 99th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Webster, J. A., Beck, A. H., Sharma, M., Espinosa, I., Schreuder, M., Montgomery, K. D., Jensen, K. C., van de Rijn, M., West, R. B. NATURE PUBLISHING GROUP. 2010: 77A?77A

  View details for Web of Science ID 000274582500335

• 3 '-End Sequencing for Expression Quantification (3SEQ) from Archival Tumor Samples PLOS ONE Beck, A. H., Weng, Z., Witten, D. M., Zhu, S., Foley, J. W., Lacroute, P., Smith, C. L., Tibshirani, R., van de Rijn, M., Sidow, A., West, R. B. 2010; 5 (1)

  Abstract

  Gene expression microarrays are the most widely used technique for genome-wide expression profiling. However, microarrays do not perform well on formalin fixed paraffin embedded tissue (FFPET). Consequently, microarrays cannot be effectively utilized to perform gene expression profiling on the vast majority of archival tumor samples. To address this limitation of gene expression microarrays, we designed a novel procedure (3'-end sequencing for expression quantification (3SEQ)) for gene expression profiling from FFPET using next-generation sequencing. We performed gene expression profiling by 3SEQ and microarray on both frozen tissue and FFPET from two soft tissue tumors (desmoid type fibromatosis (DTF) and solitary fibrous tumor (SFT)) (total n = 23 samples, which were each profiled by at least one of the four platform-tissue preparation combinations). Analysis of 3SEQ data revealed many genes differentially expressed between the tumor types (FDR<0.01) on both the frozen tissue (approximately 9.6K genes) and FFPET (approximately 8.1K genes). Analysis of microarray data from frozen tissue revealed fewer differentially expressed genes (approximately 4.64K), and analysis of microarray data on FFPET revealed very few (69) differentially expressed genes. Functional gene set analysis of 3SEQ data from both frozen tissue and FFPET identified biological pathways known to be important in DTF and SFT pathogenesis and suggested several additional candidate oncogenic pathways in these tumors. These findings demonstrate that 3SEQ is an effective technique for gene expression profiling from archival tumor samples and may facilitate significant advances in translational cancer research.

  View details for DOI 10.1371/journal.pone.0008768

  View details for PubMedID 20098735

• Genome-wide transcriptome analyses reveal p53 inactivation mediated loss of miR-34a expression in malignant peripheral nerve sheath tumours JOURNAL OF PATHOLOGY Subramanian, S., Thayanithy, V., West, R. B., Lee, C., Beck, A. H., Zhu, S., Downs-Kelly, E., Montgomery, K., Goldblum, J. R., Hogendoom, P. C., Corless, C. L., Oliveira, A. M., Dry, S. M., Nielsen, T. O., Rubin, B. P., Fletcher, J. A., Fletcher, C. D., van de Rijn, M. 2010; 220 (1): 58-70

  Abstract

  Malignant peripheral nerve sheath tumours (MPNSTs) are aggressive soft tissue tumours that occur either sporadically or in patients with neurofibromatosis type 1. The malignant transformation of the benign neurofibroma to MPNST is incompletely understood at the molecular level. We have determined the gene expression signature for benign and malignant PNSTs and found that the major trend in malignant transformation from neurofibroma to MPNST consists of the loss of expression of a large number of genes, rather than widespread increase in gene expression. Relatively few genes are expressed at higher levels in MPNSTs and these include genes involved in cell proliferation and genes implicated in tumour metastasis. In addition, a gene expression signature indicating p53 inactivation is seen in the majority of MPNSTs. Subsequent microRNA profiling of benign and malignant PNSTs indicated a relative down-regulation of miR-34a in most MPNSTs compared to neurofibromas. In vitro studies using the cell lines MPNST-14 (NF1 mutant) and MPNST-724 (from a non-NF1 individual) show that exogenous expression of p53 or miR-34a promotes apoptotic cell death. In addition, exogenous expression of p53 in MPNST cells induces miR-34a and other miRNAs. Our data show that p53 inactivation and subsequent loss of expression of miR-34a may significantly contribute to the MPNST development. Collectively, our findings suggest that deregulation of miRNAs has a potential role in the malignant transformation process in peripheral nerve sheath tumours.

  View details for DOI 10.1002/path.2633

  View details for Web of Science ID 000273186100008

  View details for PubMedID 19890883

• Breast Angiosarcoma: Case Series and Expression of Vascular Endothelial Growth Factor. Case reports in oncology Brar, R., West, R., Witten, D., Raman, B., Jacobs, C., Ganjoo, K. 2009; 2 (3): 242-250

  Abstract

  PURPOSE: Angiosarcoma of the breast is a rare, malignant tumor for which little is known regarding prognostic indicators and optimal therapeutic regimens. To address this issue, we performed a retrospective analysis of breast angiosarcoma cases seen at Stanford University along with immunohistochemical analysis for markers of angiogenesis. METHODS: Breast angiosarcoma cases seen between 1980 and 2008 were examined. Viable tissue blocks were analyzed for expression of vascular endothelial growth factor and its receptors. RESULTS: A total of 16 cases were identified. Data was collected regarding epidemiology, treatment, response rates, disease-free survival, and the use of various imaging modalities. Five tissue blocks remained viable for immunohistochemical analysis. Vascular endothelial growth factor-A was positively expressed in 3 of these samples. CONCLUSION: Angiosarcoma of the breast is an aggressive malignancy with a propensity for both local recurrence and distant metastases. Angiogenesis inhibition may represent a novel therapeutic modality in this rare, vascular malignancy.

  View details for PubMedID 20737044

• HER2 Intermediate Breast Cancers AMERICAN JOURNAL OF SURGICAL PATHOLOGY Jensen, K. C., Nielsen, T. O., Gilks, C. B., West, R. B. 2009; 33 (11): 1739-1739

  View details for Web of Science ID 000271795800022

  View details for PubMedID 19745698

• Microtubule-associated Protein-2 is a Sensitive Marker of Primary and Metastatic Neuroblastoma AMERICAN JOURNAL OF SURGICAL PATHOLOGY Krishnan, C., Higgins, J. P., West, R. B., Natkunam, Y., Heerema-McKenney, A., Arber, D. A. 2009; 33 (11): 1695-1704

  Abstract

  Microtubule-associated protein-2 (MAP-2) is a protein expressed in high levels in cells derived from the neural crest. To the best of our knowledge, MAP-2 expression has not been thoroughly evaluated in tissues outside of the central nervous tissue. We examined the diagnostic utility of MAP-2 as a marker of neuroblastoma and attempted to characterize the expression of this protein in other tumors in the morphologic differential diagnosis of neuroblastoma.MAP-2 showed significant cytoplasmic reactivity in 95% of primary and 100% of metastatic neuroblastomas. Included within this set of tumors were 3 undifferentiated neuroblastomas, all of which showed strong staining. MAP-2 did not show significant staining in the majority of other small round blue cell tumors within the morphologic differential. Additionally, MAP-2 showed comparable sensitivity in staining primary neuroblastomas as compared with synaptophysin, chromogranin, CD56, and beta-catenin. In contrast to other markers of neuroblastoma, MAP-2 did not show significant cross reactivity to native bone marrow precursors, thus eliminating a potential source of confusion. In normal tissues, MAP-2 staining was essentially restricted to organs derived from the neural crest (adrenal medulla, endocrine organs). Variant patterns of staining were seen in exocrine organs, monocyte/macrophages and solitary fibrous tumor/hemangiopericytoma family of tumors. Rarely, high-grade adult sarcomas exhibiting strong cytoplasmic MAP-2 staining were seen.MAP-2 is a sensitive and specific marker of neuroblastoma, both in the primary tumor and bone marrow biopsy settings. We think that MAP-2, in conjunction with synaptophysin, is a very powerful immunohistochemical marker in differentiating neuroblastoma from its morphologic mimics.

  View details for Web of Science ID 000271795800016

  View details for PubMedID 19701075

• EXTERNAL BEAM RADIATION THERAPY ENHANCES LOCAL CONTROL IN PIGMENTED VILLONODULAR SYNOVITIS INTERNATIONAL JOURNAL OF RADIATION ONCOLOGY BIOLOGY PHYSICS Horoschak, M., Tran, P. T., Bachireddy, P., West, R. B., Mohler, D., Beaulieu, C. F., Kapp, D. S., Donaldson, S. S. 2009; 75 (1): 183-187

  Abstract

  Pigmented villonodular synovitis (PVNS) is a rare proliferative disorder of the synovium with locally aggressive behavior. We reviewed our experience using radiation therapy in the treatment of PVNS.Seventeen patients with 18 sites of PVNS were treated with radiation between 1993 and 2007. Cases were retrospectively reviewed for patient information, treatment parameters, complications, and outcome. Seven sites were primary presentations and 11 were recurrent with an average of 2.5 prior surgical interventions. The most common location was the knee joint (67%). Cytoreductive surgery was performed before radiation therapy in 16/18 sites with all having proven or suspected residual disease. Radiation was delivered using 4-15 MV photons with an average total dose 34 Gy (range, 20-36 Gy). Seventeen of 18 sites (94%) had postradiotherapy imaging.With average follow-up of 46 months (range, 8-181 months), initial local control was achieved in 75% (12/16) of the sites with prior cytoreductive surgery (mean time to recurrence, 38 months). Ultimate local control was 100% after repeat resection (mean follow-up, 61 months). Two additional sites without prior cytoreductive surgery showed growth after radiotherapy (mean time to documented growth, 10.5 months). Seventeen of the 18 involved joints (94%) were scored as excellent or good PVNS-related function, one site (5%) as fair function, and no site with poor function. No patient required amputation; and there were no Grade 3/4 treatment-related complications.Postoperative external beam radiation is effective in preventing disease recurrence and should be offered following maximal cytoreduction to enhance local control in PVNS.

  View details for DOI 10.1016/j.ijrobp.2008.10.058

  View details for Web of Science ID 000269328700031

  View details for PubMedID 19211195

• Expression of insulin-like growth factor 2 in mesenchymal neoplasms MODERN PATHOLOGY Steigen, S. E., Schaeffer, D. F., West, R. B., Nielsen, T. O. 2009; 22 (7): 914-921

  Abstract

  The insulin-like growth factor (IGF) system plays an important role in the growth and development of cells and has been implicated in oncogenesis and tumor progression. Gene expression profiling studies on limited numbers of specimens have shown high expression of IGF2, encoding the activating ligand for this system, in gastrointestinal stromal tumors (GISTs) and in synovial sarcomas. This data may have concrete clinical implications, as several reports exist of patients with GISTs suffering from severe hypoglycemia, a predicted effect of IGF2. Furthermore, new drugs targeting IGF signaling are entering clinical trials. The purpose of this study is to survey IGF2 expression at the protein level on a broad number of mesenchymal tumors representing all major diagnostic classes. By immunostaining tissue microarrays, results were obtained for 51 diagnostic categories of bone and soft-tissue tumors representing 1288 cases. Distinct membranous and/or cytoplasmic IGF2 immunoreactivity was assessed according to published criteria. Solitary fibrous tumors had the highest expression. Of 20 tumor types represented by more than 10 cases, synovial sarcomas, myxoid liposarcomas, GISTs, malignant peripheral nerve sheath tumors, chondrosarcomas, undifferentiated pleomorphic sarcomas (MFH), Ewing's sarcomas and tenosynovial giant cell tumors showed high levels of expression in more than 20% of cases. Of the 445 GIST cases with clinical information, those with high expression of IGF2 had a significantly worse outcome than those with low or no expression. IGF2 protein expression among mesenchymal tumors is largely consistent with gene expression studies and suggests a potential for molecular therapy targeting the IGF signaling pathway system in these neoplasms.

  View details for DOI 10.1038/modpathol.2009.48

  View details for Web of Science ID 000267575400008

  View details for PubMedID 19407853

• Coordinate Expression of Colony-Stimulating Factor-1 and Colony-Stimulating Factor-1-Related Proteins Is Associated with Poor Prognosis in Gynecological and Nongynecological Leiomyosarcoma AMERICAN JOURNAL OF PATHOLOGY Espinosa, I., Beck, A. H., Lee, C., Zhu, S., Montgomery, K. D., Marinelli, R. J., Ganjoo, K. N., Nielsen, T. O., Gilks, C. B., West, R. B., van de Rijn, M. 2009; 174 (6): 2347-2356

  Abstract

  Previously, we showed that the presence of high numbers of macrophages correlates with poor prognosis in nongynecological leiomyosarcoma (LMS). In gynecological LMS, a similar trend was noted but did not reach statistical significance. Colony-stimulating factor-1 (CSF1) is a major chemoattractant for macrophages. Here we show that in a subset of LMS cases, CSF1 is expressed by the malignant cells. Previously, we found that CSF1 is translocated and highly expressed in tenosynovial giant cell tumors (TGCTs), and this observation allowed us to identify genes that showed a coordinate expression with CSF1. Here, we evaluated the expression of CSF1 and TGCT-associated proteins in 149 cases of LMS. The coordinate expression of CSF1 and three TGCT-associated proteins (CD163, FCGR3a, and CTSL1) identified cases with poor prognosis in both gynecological LMS (P = 0.00006) and nongynecological LMS (P = 0.03). In gynecological LMS, the coordinate expression of these four markers was the only independent prognosticator in multivariate analysis (hazard ratio, 4.2; 95% CI, 1.12 to 16; P = 0.03). Our findings indicate that CSF1 may play an important role in the clinical behavior of LMS that may open a window for new therapeutic reagents.

  View details for DOI 10.2353/ajpath.2009.081037

  View details for PubMedID 19443701

• Ano1 is a selective marker of interstitial cells of Cajal in the human and mouse gastrointestinal tract AMERICAN JOURNAL OF PHYSIOLOGY-GASTROINTESTINAL AND LIVER PHYSIOLOGY Gomez-Pinilla, P. J., Gibbons, S. J., Bardsley, M. R., Lorincz, A., Pozo, M. J., Pasricha, P. J., van de Rijn, M., West, R. B., Sarr, M. G., Kendrick, M. L., Cima, R. R., Dozois, E. J., Larson, D. W., Ordog, T., Farrugia, G. 2009; 296 (6): G1370-G1381

  Abstract

  Populations of interstitial cells of Cajal (ICC) are altered in several gastrointestinal neuromuscular disorders. ICC are identified typically by ultrastructure and expression of Kit (CD117), a protein that is also expressed on mast cells. No other molecular marker currently exists to independently identify ICC. The expression of ANO1 (DOG1, TMEM16A), a Ca(2+)-activated Cl(-) channel, in gastrointestinal stromal tumors suggests it may be useful as an ICC marker. The aims of this study were therefore to determine the distribution of Ano1 immunoreactivity compared with Kit and to establish whether Ano1 is a reliable marker for human and mouse ICC. Expression of Ano1 in human and mouse stomach, small intestine, and colon was investigated by immunofluorescence labeling using antibodies to Ano1 alone and in combination with antibodies to Kit. Colocalization of immunoreactivity was demonstrated by epifluorescence and confocal microscopy. In the muscularis propria, Ano1 immunoreactivity was restricted to cells with the morphology and distribution of ICC. All Ano1-positive cells in the muscularis propria were also Kit positive. Kit-expressing mast cells were not Ano1 positive. Some non-ICC in the mucosa and submucosa of human tissues were Ano1 positive but Kit negative. A few (3.2%) Ano1-positive cells in the human gastric muscularis propria were labeled weakly for Kit. Ano1 labels all classes of ICC and represents a highly specific marker for studying the distribution of ICC in mouse and human tissues with an advantage over Kit since it does not label mast cells.

  View details for DOI 10.1152/ajpgi.00074.2009

  View details for Web of Science ID 000266453300023

  View details for PubMedID 19372102

  View details for PubMedCentralID PMC2697941

• Inter-observer reproducibility of HER2 immunohistochemical assessment and concordance with fluorescent in situ hybridization (FISH): pathologist assessment compared to quantitative image analysis BMC CANCER Turashvili, G., Leung, S., Turbin, D., Montgomery, K., Gilks, B., West, R., Carrier, M., Huntsman, D., Aparicio, S. 2009; 9

  Abstract

  In breast cancer patients, HER2 overexpression is routinely assessed by immunohistochemistry (IHC) and equivocal cases are subject to fluorescent in situ hybridization (FISH). Our study compares HER2 scoring by histopathologists with automated quantitation of staining, and determines the concordance of IHC scores with FISH results.A tissue microarray was constructed from 1,212 invasive breast carcinoma cases with linked treatment and outcome information. IHC slides were semi-quantitatively scored by two independent pathologists on a range of 0 to 3+, and also analyzed with an Ariol automated system by two operators. 616 cases were scorable by both IHC and FISH.Using data from unequivocal positive (3+) or negative (0, 1+) results, both visual and automated scores were highly consistent: there was excellent concordance between two pathologists (kappa = 1.000, 95% CI: 1-1), between two machines (kappa = 1.000, 95% CI: 1-1), and between both visual and both machine scores (kappa = 0.898, 95% CI: 0.775-0.979). Two pathologists successfully distinguished negative, positive and equivocal cases (kappa = 0.929, 95% CI: 0.909-0.946), with excellent agreement with machine 1 scores (kappa = 0.835, 95% CI: 0.806-0.862; kappa = 0.837, 95% CI: 0.81-0.862), and good agreement with machine 2 scores (kappa = 0.698, 95% CI: 0.6723-0.723; kappa = 0.709, 95% CI: 0.684-0.732), whereas the two machines showed good agreement (kappa = 0.806, 95% CI: 0.785-0.826). When comparing categorized IHC scores and FISH results, the agreement was excellent for visual 1 (kappa = 0.814, 95% CI: 0.768-0.856), good for visual 2 (kappa = 0.763, 95% CI: 0.712-0.81) and machine 1 (kappa = 0.665, 95% CI: 0.609-0.718), and moderate for machine 2 (kappa = 0.535, 95% CI: 0.485-0.584).A fully automated image analysis system run by an experienced operator can provide results consistent with visual HER2 scoring. Further development of such systems will likely improve the accuracy of detection and categorization of membranous staining, making this technique suitable for use in quality assurance programs and eventually in clinical practice.

  View details for DOI 10.1186/1471-2407-9-165

  View details for Web of Science ID 000267639900001

  View details for PubMedID 19476653

  View details for PubMedCentralID PMC2698924

• MOC-31 Exhibits Superior Reactivity Compared With Ber-EP4 in Invasive Lobular and Ductal Carcinoma of the Breast A Tissue Microarray Study APPLIED IMMUNOHISTOCHEMISTRY & MOLECULAR MORPHOLOGY Pai, R. K., West, R. B. 2009; 17 (3): 202-206

  Abstract

  Distinguishing between reactive mesothelial proliferations and adenocarcinoma is often very difficult. Ancillary studies, in particular immunohistochemistry, are often critical in detecting malignant epithelial cells, especially in serous effusion specimens. MOC-31 and Ber-EP4 are antibodies which target the epithelial cell adhesion molecule (Ep-CAM, TACSTD1) expressed in epithelial cells, and both are useful in distinguishing metastatic adenocarcinoma from reactive mesothelial cells. However, the reactivity of MOC-31 and Ber-EP4 with breast carcinoma, one of the more common carcinomas involving serous effusions, has not been extensively studied. We analyzed the immunohistochemical expression of MOC-31 and Ber-EP4 using tissue microarrays containing invasive ductal carcinoma (191 cases), invasive lobular carcinoma (44 cases), and 102 other carcinoma types comprising primary carcinomas of lung, gynecologic tract, pancreas, colon, gastric, esophageal, prostate, head and neck, hepatic, and renal origin. For MOC-31, 184 of 191 (96%) invasive ductal carcinomas and 39 of 44 (89%) invasive lobular carcinomas exhibited diffuse positive staining. In contrast, for Ber-EP4, 121 of 183 (66%) invasive ductal carcinomas and 11 of 40 (27.5%) invasive lobular carcinomas exhibited diffuse positive staining. With the exception of 1 case of esophageal adenocarcinoma, all other adenocarcinomas (86 of 87 cases) exhibited diffuse staining with both Ber-EP4 and MOC-31. MOC-31 and Ber-EP4 exhibited identical staining with all other carcinoma types. Our findings indicate that MOC-31 is superior to Ber-EP4 in detecting both invasive lobular and ductal carcinoma of the breast.

  View details for Web of Science ID 000265459300004

  View details for PubMedID 19391212

• LC3-mediated fibronectin mRNA translation induces fibrosarcoma growth by increasing connective tissue growth factor JOURNAL OF CELL SCIENCE Ying, L., Lau, A., Alvira, C. M., West, R., Cann, G. M., Zhou, B., Kinnear, C., Jan, E., Sarnow, P., van de Rijn, M., Rabinovitch, M. 2009; 122 (9): 1441-1451

  Abstract

  Previously, we related fibronectin (Fn1) mRNA translation to an interaction between an AU-rich element in the Fn1 3' UTR and light chain 3 (LC3) of microtubule-associated proteins 1A and 1B. Since human fibrosarcoma (HT1080) cells produce little fibronectin and LC3, we used these cells to investigate how LC3-mediated Fn1 mRNA translation might alter tumor growth. Transfection of HT1080 cells with LC3 enhanced fibronectin mRNA translation. Using polysome analysis and RNA-binding assays, we show that elevated levels of translation depend on an interaction between a triple arginine motif in LC3 and the AU-rich element in Fn1 mRNA. Wild-type but not mutant LC3 accelerated HT1080 cell growth in culture and when implanted in SCID mice. Comparison of WT LC3 with vector-transfected HT1080 cells revealed increased fibronectin-dependent proliferation, adhesion and invasion. Microarray analysis of genes differentially expressed in WT and vector-transfected control cells indicated enhanced expression of connective tissue growth factor (CTGF). Using siRNA, we show that enhanced expression of CTGF is fibronectin dependent and that LC3-mediated adhesion, invasion and proliferation are CTGF dependent. Expression profiling of soft tissue tumors revealed increased expression of both LC3 and CTGF in some locally invasive tumor types.

  View details for DOI 10.1242/jcs.025957

  View details for PubMedID 19366727

• A compact VEGF signature associated with distant metastases and poor outcomes BMC MEDICINE Hu, Z., Fan, C., Livasy, C., He, X., Oh, D. S., Ewend, M. G., Carey, L. A., Subramanian, S., West, R., Ikpatt, F., Olopade, O. I., van de Rijn, M., Perou, C. M. 2009; 7

  Abstract

  Tumor metastases pose the greatest threat to a patient's survival, and thus, understanding the biology of disseminated cancer cells is critical for developing effective therapies.Microarrays and immunohistochemistry were used to analyze primary breast tumors, regional (lymph node) metastases, and distant metastases in order to identify biological features associated with distant metastases.When compared with each other, primary tumors and regional metastases showed statistically indistinguishable gene expression patterns. Supervised analyses comparing patients with distant metastases versus primary tumors or regional metastases showed that the distant metastases were distinct and distinguished by the lack of expression of fibroblast/mesenchymal genes, and by the high expression of a 13-gene profile (that is, the 'vascular endothelial growth factor (VEGF) profile') that included VEGF, ANGPTL4, ADM and the monocarboxylic acid transporter SLC16A3. At least 8 out of 13 of these genes contained HIF1alpha binding sites, many are known to be HIF1alpha-regulated, and expression of the VEGF profile correlated with HIF1alpha IHC positivity. The VEGF profile also showed prognostic significance on tests of sets of patients with breast and lung cancer and glioblastomas, and was an independent predictor of outcomes in primary breast cancers when tested in models that contained other prognostic gene expression profiles and clinical variables.These data identify a compact in vivo hypoxia signature that tends to be present in distant metastasis samples, and which portends a poor outcome in multiple tumor types.This signature suggests that the response to hypoxia includes the ability to promote new blood and lymphatic vessel formation, and that the dual targeting of multiple cell types and pathways will be needed to prevent metastatic spread.

  View details for DOI 10.1186/1741-7015-7-9

  View details for Web of Science ID 000265841300001

  View details for PubMedID 19291283

  View details for PubMedCentralID PMC2671523

• The Macrophage Colony-Stimulating Factor 1 Response Signature in Breast Carcinoma CLINICAL CANCER RESEARCH Beck, A. H., Espinosa, I., Edris, B., Li, R., Montgomery, K., Zhu, S., Varma, S., Marinelli, R. J., van de Rijn, M., West, R. B. 2009; 15 (3): 778-787

  Abstract

  Macrophages play an important role in breast carcinogenesis. The pathways that mediate the macrophage contribution to breast cancer and the heterogeneity that exists within macrophages are incompletely understood. Macrophage colony-stimulating factor 1 (CSF1) is the primary regulator of tissue macrophages. The purpose of this study was to define a novel CSF1 response signature and to evaluate its clinical and biological significance in breast cancer.We defined the CSF1 response signature by identifying genes overexpressed in tenosynovial giant cell tumor and pigmented villonodular synovitis (tumors composed predominantly of macrophages recruited in response to the overexpression of CSF1) compared with desmoid-type fibromatosis and solitary fibrous tumor. To characterize the CSF1 response signature in breast cancer, we analyzed the expression of CSF1 response signature genes in eight published breast cancer gene expression data sets (n = 982) and did immunohistochemistry and in situ hybridization for CSF1 response genes on a breast cancer tissue microarray (n = 283).In both the gene microarray and tissue microarray analyses, a consistent subset (17-25%) of breast cancers shows the CSF1 response signature. The signature is associated with higher tumor grade, decreased expression of estrogen receptor, decreased expression of progesterone receptor, and increased TP53 mutations (P < 0.001).Our data show that the CSF1 response signature is consistently seen in a subset of breast carcinomas and correlates with biological features of the tumor. Our findings provide insight into macrophage biology and may facilitate the development of personalized therapy for patients most likely to benefit from CSF1-targeted treatments.

  View details for DOI 10.1158/1078-0432.CCR-08-1283

  View details for PubMedID 19188147

• The Transcription Factor LMO2 Is a Robust Marker of Vascular Endothelium and Vascular Neoplasms and Selected Other Entities AMERICAN JOURNAL OF CLINICAL PATHOLOGY Gratzinger, D., Zhao, S., West, R., Rouse, R. V., Vogel, H., Gil, E. C., Levy, R., Lossos, I. S., Natkunam, Y. 2009; 131 (2): 264-278

  Abstract

  The transcription factor LMO2 is involved in vascular and hematopoietic development and hematolymphoid neoplasia. We have demonstrated that LMO2 is expressed nearly ubiquitously in native and neoplastic vasculature, including lymphatics. LMO2 reactivity is otherwise virtually absent in nonhematolymphoid tissues except in breast myoepithelium, prostatic basal cells, and secretory phase endometrial glands. Vasculature is LMO2- in adult and fetal heart, brain of older adults, hepatic sinusoids, and hepatocellular carcinoma. LMO2 is uniformly expressed in benign vascular and lymphatic neoplasms and in most malignant vascular neoplasms with the exception of epithelioid vascular neoplasms of pleura and bone. Among nonvascular neoplasms, LMO2 reactivity is present in giant cell tumor of tendon sheath, juvenile xanthogranuloma, a subset of gastrointestinal stromal tumors, small round blue cell tumors, and myoepithelial-derived neoplasms. The restricted expression pattern, nuclear localization, and crisp staining of LMO2 in paraffin blocks make it an attractive candidate for the diagnostic immunohistochemistry laboratory.

  View details for DOI 10.1309/AJCP5FP3NAXAXRJE

  View details for PubMedID 19141387

• Discovery of Molecular Subtypes in Leiomyosarcoma through Integrative Molecular Profiling 98th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Beck, A. H., Lee, C. H., WITTEN, D. M., Zhou, S., Montgomery, K., Tibshirani, R., Hastie, T., West, R. B., van de Rijn, M. NATURE PUBLISHING GROUP. 2009: 368A?368A

  View details for Web of Science ID 000262486301668

• Breast Scar Gene Expression Profiles Overlap with Tumor Stroma Response Signatures Pate, L. L., West, R. B., van de Rijn, M. NATURE PUBLISHING GROUP. 2009: 62A

  View details for Web of Science ID 000262371500269

• CSF-1 and Fibromatosis Expression in Stroma of Ductal Carcinoma In Situ Sharma, M., Espinosa, I., Beck, A. H., Webster, J. A., Montgomery, K. D., van de Rijn, M., Jensen, K. C., West, R. B. NATURE PUBLISHING GROUP. 2009: 67A

  View details for Web of Science ID 000262371500290

• Breast Scar Gene Expression Profiles Overlap with Tumor Stroma Response Signatures Pate, L. L., West, R. B., van de Rijn, M. NATURE PUBLISHING GROUP. 2009: 62A

  View details for Web of Science ID 000262486300269

• Breast Angiosarcoma: Case Series and Expression of Vascular Endothelial Growth Factor CASE REPORTS IN ONCOLOGY Brar, R., West, R., Witten, D., Raman, B., Jacobs, C., Ganjoo, K. 2009; 2 (3): 242?50

  View details for DOI 10.1159/000264637

  View details for Web of Science ID 000216899300014

• Discovery of Molecular Subtypes in Leiomyosarcoma through Integrative Molecular Profiling 98th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Beck, A. H., Lee, C. H., WITTEN, D. M., Zhou, S., Montgomery, K., Tibshirani, R., Hastie, T., West, R. B., van de Rijn, M. NATURE PUBLISHING GROUP. 2009: 368A?368A

  View details for Web of Science ID 000262371501667

• CSF-1 and Fibromatosis Expression in Stroma of Ductal Carcinoma In Situ 98th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Sharma, M., Espinosa, I., Beck, A. H., Webster, J. A., Montgomery, K. D., van de Rijn, M., Jensen, K. C., West, R. B. NATURE PUBLISHING GROUP. 2009: 67A?67A

  View details for Web of Science ID 000262486300290

• Characterization of a novel anti-fatty acid synthase (FASN) antiserum in breast tissue 97th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Jensen, K. C., Schaeffer, D. F., Cheang, M., Montgomery, K., West, R. B., Gilks, C. B., Ross, D., Turashvili, G., Schnitt, S., van de Rijn, M. NATURE PUBLISHING GROUP. 2008: 1413?20

  Abstract

  Fatty acid synthase (FASN) expression has been reported in many different tumors, including breast cancer. In gene microarray studies, the fatty acid synthase gene co-clustered with cytokeratins 5 and 17 and other genes that defined the basal-like subset of breast cancers. To define the use of this marker in breast pathology, a rabbit polyclonal antiserum (S143) to a peptide fragment of this gene was produced and compared with a commercially available monoclonal antibody by immunohistochemistry on various tissue microarrays and whole tissue sections. The tissue microarrays included 1090 breast cancers and 244 normal breast tissues. Whole tissue sections consisted of benign and malignant tissues from breast resection specimens. In contrast to other 'basal' markers identified by gene expression profiling data, the fatty acid synthase (FASN) expression pattern in normal breast was notable for its expression in only a small subset of basal and suprabasal cells. Dual staining experiments revealed that the subpopulation of cells labeling with FASN did not coexpress myoepithelial markers CK5/6 or p63, but did coexpress e-cadherin. In addition to staining a subset of basal and suprabasal cells, the antiserum highlighted apocrine differentiation, and stained 106/144 (74%) cases of columnar cell lesions and five of five cases of flat epithelial atypia. Despite its association with basal keratins in gene array studies, FASN expression did not correlate significantly with the outcome in breast cancer. We describe an expression pattern that highlights only a subset of basal and suprabasal cells in normal breast ducts and we show by dual expression studies that this subset of cells is different from myoepithelial and basal cytokeratin-positive cells. In addition, FASN expression is described in apocrine metaplasia, columnar cell lesions, and flat epithelial atypia.

  View details for DOI 10.1038/modpathol.2008.163

  View details for PubMedID 18820672

• New cutpoints to identify increased HER2 copy number: analysis of a large, population-based cohort with long-term follow-up BREAST CANCER RESEARCH AND TREATMENT Jensen, K. C., Turbin, D. A., Leung, S., Miller, M. A., Johnson, K., Norris, B., Hastie, T., McKinney, S., Nielsen, T. O., Huntsman, D. G., Gilks, C. B., West, R. B. 2008; 112 (3): 453-459

  Abstract

  HER2 gene amplification and/or protein overexpression in breast cancer is associated with a poor prognosis and predicts response to anti-HER2 therapy. We examine the natural history of breast cancers in relationship to increased HER2 copy numbers in a large population-based study.HER2 status was measured by fluorescence in situ hybridization (FISH) and immunohistochemistry (IHC) in approximately 1,400 breast cancer cases with greater than 15 years of follow-up. Protein expression was evaluated with two different commercially-available antibodies.We looked for subgroups of breast cancer with different clinical outcomes, based on HER2 FISH amplification ratio. The current HER2 ratio cut point for classifying HER2 positive and negative cases is 2.2. However, we found an increased risk of disease-specific death associated with FISH ratios of >1.5. An 'intermediate' group of cases with HER2 ratios between 1.5 and 2.2 was found to have a significantly better outcome than the conventional 'amplified' group (HER2 ratio >2.2) but a significantly worse outcome than groups with FISH ratios less than 1.5.Breast cancers with increased HER2 copy numbers (low level HER2 amplification), below the currently accepted positive threshold ratio of 2.2, showed a distinct, intermediate outcome when compared to HER2 unamplified tumors and tumors with HER2 ratios greater than 2.2. These findings suggest that a new cut point to determine HER2 positivity, at a ratio of 1.5 (well below the current recommended cut point of 2.2), should be evaluated.

  View details for DOI 10.1007/s10549-007-9887-y

  View details for Web of Science ID 000261951000007

  View details for PubMedID 18193353

• KIT Gene Mutations and Copy Number in Melanoma Subtypes CLINICAL CANCER RESEARCH Beadling, C., Jacobson-Dunlop, E., Hodi, F. S., Le, C., Warrick, A., Patterson, J., Town, A., Harlow, A., Cruz, F., Azar, S., Rubin, B. P., Muller, S., West, R., Heinrich, M. C., Corless, C. L. 2008; 14 (21): 6821-6828

  Abstract

  We recently identified a KIT exon 11 mutation in an anorectal melanoma of a patient who had an excellent response to treatment with imatinib. To determine the frequency of KIT mutations across melanoma subtypes, we surveyed a large series of tumors.One hundred eighty-nine melanomas were screened for mutations in KIT exons 11, 13, and 17. KIT copy number was assessed by quantitative PCR. A subset of cases was evaluated for BRAF and NRAS mutations. Immunohistochemistry was done to assess KIT (CD117) expression.KIT mutations were detected in 23% (3 of 13) of acral melanomas, 15.6% (7 of 45) of mucosal melanomas, 7.7% (1 of 13) of conjunctival melanomas, 1.7% (1 of 58) of cutaneous melanomas, and 0% (0 of 60) of choroidal melanomas. Almost all the KIT mutations were of the type predicted to be imatinib sensitive. There was no overlap with NRAS mutations (11.1% of acral and 24.3% of mucosal tumors) or with BRAF mutations (absent in mucosal tumors). Increased KIT copy number was detected in 27.3% (3 of 11) of acral and 26.3% (10 of 38) of mucosal melanomas, but was less common among cutaneous (6.7%; 3 of 45), conjunctival (7.1%; 1 of 14), and choroidal melanomas (0 of 28). CD117 expression, present in 39% of 105 tumors representing all melanoma types, did not correlate with either KIT mutation status or KIT copy number.Our findings confirm that KIT mutations are most common in acral and mucosal melanomas but do not necessarily correlate with KIT copy number or CD117 expression. Screening for KIT mutations may open up new treatment options for melanoma patients.

  View details for DOI 10.1158/1078-0432.CCR-08-0575

  View details for PubMedID 18980976

• Immunohistochemical and Biogenetic Features of Diffuse-Type Tenosynovial Giant Cell Tumors: The Potential Roles of Cyclin A, P53, and Deletion of 15q in Sarcomatous Transformation CLINICAL CANCER RESEARCH Huang, H., West, R. B., Tzeng, C., van de Rijn, M., Wang, J., Chou, S., Huang, W., Eng, H., Lin, C., Yu, S., Wu, J., Lu, C., Li, C. 2008; 14 (19): 6023-6032

  Abstract

  Diffuse-type tenosynovial giant cell tumor (D-TSGCT) is an aggressive proliferation of synovial-like mononuclear cells with inflammatory infiltrates. Despite the COL6A3-CSF1 gene fusion discovered in benign lesions, molecular aberrations of malignant D-TSGCTs remain unidentified.We used fluorescent in situ hybridization and in situ hybridization to evaluate CSF1 translocation and mRNA expression in six malignant D-TSGCTs, which were further immunohistochemically compared with 24 benign cases for cell cycle regulators involving G(1) phase and G(1)-S transition. Comparative genomic hybridization, real-time reverse transcription-PCR, and a combination of laser microdissection and sequencing were adopted to assess chromosomal imbalances, cyclin A expression, and TP53 gene, respectively.Five of six malignant D-TSGCTs displayed CSF1 mRNA expression by in situ hybridization, despite only one having CSF1 translocation. Cyclin A (P = 0.008) and P53 (P < 0.001) could distinguish malignant from benign lesions without overlaps in labeling indices. Cyclin A transcripts were more abundant in malignant D-TSGCTs (P < 0.001). All malignant cases revealed a wild-type TP53 gene, which was validated by an antibody specifically against wild-type P53 protein. Chromosomal imbalances were only detected in malignant D-TSGCTs, with DNA losses predominating over gains. Notably, -15q was recurrently identified in five malignant D-TSGCTs, four of which showed a minimal overlapping deletion at 15q22-24.Deregulated CFS1 overexpression is frequent in malignant D-TSGCTs. The sarcomatous transformation involves aberrations of cyclin A, P53, and chromosome arm 15q. Cyclin A mRNA is up-regulated in malignant D-TSGCTs. Non-random losses at 15q22-24 suggest candidate tumor suppressor gene(s) in this region. However, P53 overexpression is likely caused by alternative mechanisms rather than mutations in hotspot exons.

  View details for DOI 10.1158/1078-0432.CCR-08-0252

  View details for Web of Science ID 000260142500013

  View details for PubMedID 18829481

• Diagnostic implications of podoplanin expression in peripheral nerve sheath neoplasms AMERICAN JOURNAL OF CLINICAL PATHOLOGY Jokinen, C. H., Dadras, S. S., Goldblum, J. R., van de Rijn, M., West, R. B., Rubin, B. P. 2008; 129 (6): 886-893

  Abstract

  By using the D2-40 antibody, we have observed podoplanin expression in Schwann cells and perineurial cells. Podoplanin expression has not been well characterized in peripheral nerve sheath tumors. Because neoplasms of neural crest lineage, including peripheral nerve sheath and melanocytic neoplasms, may share histologic and immunohistochemical characteristics, we evaluated podoplanin and S-100 expression in these lesions to determine the usefulness of podoplanin as a diagnostic marker. Diffuse podoplanin and S-100 expression was observed in 16 (76%) of 21 classical schwannomas, 6 (100%) of 6 cellular schwannomas, and 3 (75%) of 4 epithelioid malignant peripheral nerve sheath tumors (EMPNSTs). Podoplanin was expressed in 3 (7%) of 43 neurofibromas, 16 (21%) of 75 spindle cell MPNSTs (SMPNSTs), and 1 (10%) of 10 spindle cell melanomas but was absent in conventional melanoma. Only rare neurofibromas and SMPNSTs showed strong coexpression of podoplanin and S-100. These results suggest diffuse podoplanin expression or coexpression of podoplanin and S-100 is limited to schwannoma and EMPNST and may be useful in the evaluation of these neoplasms.

  View details for DOI 10.1309/M7D5KTVYYE51XYQA

  View details for Web of Science ID 000255959700006

  View details for PubMedID 18480004

• The fibromatosis signature defines a robust stromal response in breast carcinoma LABORATORY INVESTIGATION Beck, A. H., Espinosa, I., Gilks, C. B., van de Rijn, M., West, R. B. 2008; 88 (6): 591-601

  Abstract

  Breast cancer is a heterogeneous disease, and the influence of stromal gene and protein expression patterns on the biological and clinical heterogeneity of the disease is poorly understood. We previously demonstrated that evaluation of the gene expression patterns of two soft-tissue tumors (desmoid-type fibromatosis (DTF) and solitary fibrous tumor) could be used to identify distinct stromal reaction patterns in breast carcinoma. In the current study, we examined four additional data sets obtained from four different institutions and containing gene expression data from a total of 561 breast cancer patients. We identified a core set of 66 DTF-associated genes that were consistently coordinately expressed in a subset of 25-35% of breast cancers. Breast carcinomas defined by high levels of coordinated expression of DTF core genes tend to be lower grade, express estrogen receptor, and show significantly longer survival across the four data sets. Using multiple tissue microarrays of archival breast cancer specimens obtained from a total of 745 patients, we demonstrated that a subset of breast cancers show coordinate expression of DTF core proteins by stromal cells in the tumor microenvironment. We evaluated the protein expression of a single DTF core protein (SPARC) on a tissue microarray with clinical outcome data and demonstrated that breast cancers with strong stromal protein expression of SPARC show a trend for increased survival. Our data demonstrate that the DTF core gene set is a robust descriptor of a distinct stromal response that is associated with improved clinical outcome in breast cancer patients.

  View details for DOI 10.1038/labinvest.2008.31

  View details for PubMedID 18414401

• Predictive value of tumor associated histiocytes in patients with leiomyosarcoma Ganjoo, K. N., WITTEN, D., Patel, M., Espinosa, I., La, T., West, R., Jacobs, C., van de Rijn, M. AMER SOC CLINICAL ONCOLOGY. 2008

  View details for Web of Science ID 000208457403816

• Gene expression profiling identifies p63 as a diagnostic marker for giant cell tumor of the bone MODERN PATHOLOGY Lee, C., Espinosa, I., Jensen, K. C., Subramanian, S., Zhu, S. X., Varma, S., Montgomery, K. D., Nielsen, T. O., van de Rijn, M., West, R. B. 2008; 21 (5): 531-539

  Abstract

  Giant cell tumor of the bone (GCTOB) is a primary bone tumor that occurs mainly in young adults and is capable of locally aggressive growth. Its histologic appearance can resemble a number of benign and malignant tumors but no useful diagnostic marker is known currently. To identify diagnostic markers for this tumor, global gene expression profiling using cDNA microarray was performed on 6 fresh-frozen GCTOB, 3 aneurysmal bone cysts, 4 fibrous dysplasias and 12 giant cell tumors of tendon sheath/diffuse-type giant cell tumors. Unsupervised hierarchical clustering separated the tumors based on their histopathologic types, and significance analysis of microarray identified several genes including TP73L (encoding the p63 protein) that are significantly highly expressed in GCTOB relative to these other tumors. The diagnostic utility of p63 was subsequently confirmed using anti-p63 antibody on a series of 26 GCTOB, 25 aneurysmal bone cysts, 15 chondroblastomas, 13 giant cell reparative granulomas, 13 chondromyxoid fibromas, 4 brown tumors, 4 fibrous dysplasias, 53 giant cell tumors of tendon sheath/diffuse-type giant cell tumors and 385 additional mesenchymal tumors in tissue microarrays. Strong p63 nuclear staining was present in 18 of 26 (69%) GCTOB, 3 of 15 (20%) chondroblastomas and in 1 of 25 (4%) aneurysmal bone cysts while none of the other tumors commonly considered in the differential diagnosis of GCTOB showed any detectable p63 staining. Strong p63 staining is rare in bone and soft-tissue tumors in general. In contrast to the pattern of p63 staining, the majority of the chondroblastomas (70%) demonstrated S-100 immunoreactivity while only a minority of the GCTOB (8%) was immunoreactive for S-100. These findings altogether show that p63 can be used as a diagnostic marker to aid the clinical diagnosis of GCTOB.

  View details for DOI 10.1038/modpathol.3801023

  View details for PubMedID 18192965

• Prognostic significance of macrophage infiltration in leiomyosarcomas CLINICAL CANCER RESEARCH Lee, C., Espinosa, I., Vrijaldenhoven, S., Subramanian, S., Montgomery, K. D., Zhu, S., Marinelli, R. J., Peterse, J. L., Poulin, N., Nielsen, T. O., West, R. B., Gilks, C. B., van de Rijn, M. 2008; 14 (5): 1423-1430

  Abstract

  Macrophages are migratory cells that are frequently recruited to the site of tumors. Their presence is associated with poor clinical outcome in a variety of epithelial malignancies. The aim of this study is to examine the prognostic significance of tumor-associated macrophages in sarcomas.Global gene expression profiling data of a series of soft tissue tumors were analyzed for macrophage-associated gene expression. Immunohistochemistry on tissue microarrays containing leiomyosarcoma cases with known clinical outcome was used to verify the presence of macrophages and to examine the relationship between tumor-associated macrophages and clinical outcome.Gene expression profiling revealed high-level expression of several macrophage-associated genes such as CD163 and CD68 in a subset of leiomyosarcomas, indicating the presence of variable numbers of tumor-infiltrating macrophages. This was confirmed by CD68 and CD163 immunostaining of a tissue microarray containing 149 primary leiomyosarcomas. Kaplan-Meier survival analysis showed that high density of tumor-infiltrating macrophages as identified by CD163 or CD68 staining is associated with a significantly worse disease-specific survival in nongynecologic leiomyosarcomas, whereas leiomyosarcomas arising from the gynecologic tract showed no significant association between macrophage infiltration and survival. The presence of tumor necrosis did not correlate significantly with outcome.An increased density of CD163- or CD68-positive tumor-infiltrating macrophages is associated with poor outcome in nongynecologic leiomyosarcomas. This may help the clinical management of patients with leiomyosarcomas.

  View details for DOI 10.1158/1078-0432.CCR-07-1712

  View details for PubMedID 18316565

• A novel monoclonal antibody against DOG1 is a sensitive and specific marker for gastrointestinal stromal tumors AMERICAN JOURNAL OF SURGICAL PATHOLOGY Espinosa, I., Lee, C., Kim, M. K., Rouse, B., Subramanian, S., Montgomery, K., Varma, S., Corless, C. L., Heinrich, M. C., Smith, K. S., Wang, Z., Rubin, B., Nielsen, T. O., Seitz, R. S., Ross, D. T., West, R. B., Cleary, M. L., van de Rijn, M. 2008; 32 (2): 210-218

  Abstract

  Gastrointestinal stromal tumors (GIST) occur primarily in the wall of the intestine and are characterized by activating mutations in the receptor tyrosine kinases genes KIT or PDGFRA. The diagnosis of GIST relies heavily on the demonstration of KIT/CD117 protein expression by immunohistochemistry. However, KIT expression is absent in approximately 4% to 15% of GIST and this can complicate the diagnosis of GIST in patients who may benefit from treatment with receptor tyrosine kinase inhibitors. We previously identified DOG1/TMEM16A as a novel marker for GIST using a conventional rabbit antipeptide antiserum and an in situ hybridization probe. Here, we describe 2 new monoclonal antibodies against DOG1 (DOG1.1 and DOG1.3) and compare their staining profiles with KIT and CD34 antibodies on 447 cases of GIST. These included 306 cases with known mutational status for KIT and PDGFRA from a molecular consultation service. In addition, 935 other mesenchymal tumors and 432 nonsarcomatous tumors were studied. Both DOG1 antibodies showed high sensitivity and specificity for GIST, with DOG1.1 showing some advantages. This antibody yielded positive staining in 370 of 425 (87%) scorable GIST, whereas CD117 was positive in 317 of 428 (74%) GIST and CD34 in 254 of 430 (59%) GIST. In GIST with mutations in PDGFRA, 79% (23/29) showed DOG1.1 immunoreactivity while only 9% (3/32) and 27% (9/33) stained for CD117 and CD34, respectively. Only 1 of 326 (0.3%) leiomyosarcomas and 1 of 39 (2.5%) synovial sarcomas among the 935 soft tissue tumors examined showed positive immunostaining for DOG1.1. In addition, DOG1.1 immunoreactivity was seen in fewer cases of carcinoma, melanoma, and seminoma as compared with KIT.

  View details for PubMedID 18223323

• The Stanford Tissue Microarray Database NUCLEIC ACIDS RESEARCH Marinelli, R. J., Montgomery, K., Liu, C. L., Shah, N. H., Prapong, W., Nitzberg, M., Zachariah, Z. K., Sherlock, G. J., Natkunam, Y., West, R. B., van de Rijn, M., Brown, P. O., Ball, C. A. 2008; 36: D871-D877

  Abstract

  The Stanford Tissue Microarray Database (TMAD; http://tma.stanford.edu) is a public resource for disseminating annotated tissue images and associated expression data. Stanford University pathologists, researchers and their collaborators worldwide use TMAD for designing, viewing, scoring and analyzing their tissue microarrays. The use of tissue microarrays allows hundreds of human tissue cores to be simultaneously probed by antibodies to detect protein abundance (Immunohistochemistry; IHC), or by labeled nucleic acids (in situ hybridization; ISH) to detect transcript abundance. TMAD archives multi-wavelength fluorescence and bright-field images of tissue microarrays for scoring and analysis. As of July 2007, TMAD contained 205 161 images archiving 349 distinct probes on 1488 tissue microarray slides. Of these, 31 306 images for 68 probes on 125 slides have been released to the public. To date, 12 publications have been based on these raw public data. TMAD incorporates the NCI Thesaurus ontology for searching tissues in the cancer domain. Image processing researchers can extract images and scores for training and testing classification algorithms. The production server uses the Apache HTTP Server, Oracle Database and Perl application code. Source code is available to interested researchers under a no-cost license.

  View details for DOI 10.1093/nar/gkm861

  View details for PubMedID 17989087

• The fibromatosis stromal signature in breast carcinoma Beck, A. H., Espinosa, I., Cheang, M. C., Nielsen, T. O., De Rijn, J., West, R. B. NATURE PUBLISHING GROUP. 2008: 23A

  View details for Web of Science ID 000252181100091

• The CSF-1 response signature in breast carcinoma Beck, A. H., Espinosa, I., De Rijn, M., West, R. B. NATURE PUBLISHING GROUP. 2008: 22A

  View details for Web of Science ID 000252181100090

• The fibromatosis stromal signature in breast carcinoma Beck, A. H., Espinosa, L., Cheang, M. C., Nielsen, T. O., van de Rijn, M., West, R. B. NATURE PUBLISHING GROUP. 2008: 23A

  View details for Web of Science ID 000252180200091

• Coordinate expression of colony stimulating factor-1 (CSF1) and CSF1 related proteins is associated with poor prognosis in leiomyosarcoma Espinosa, I., Beck, A., Lee, C., Vielsen, T. O., Blake, G., West, R. B., van de Rijn, M. NATURE PUBLISHING GROUP. 2008: 12A

  View details for Web of Science ID 000252180200040

• HER2 status in a large, population-based cohort: Analysis of distinct HER2 subgroups Jensen, K. C., Turbin, D. A., Leung, S., Miller, M. A., Johnson, K., Norris, B., Hastie, T., McKinney, S., Nielsen, T. O., Huntsman, D. G., Gilks, C. B., West, R. B. NATURE PUBLISHING GROUP. 2008: 39A

  View details for Web of Science ID 000252180200167

• Periostin (POSTN, OSF2) expression in stroma of normal breast tissue and pre-invasive breast lesions Espinosa, I., Collins, L. C., West, R. B., Schnitt, S. J., van de Ryn, M. NATURE PUBLISHING GROUP. 2008: 30A

  View details for Web of Science ID 000252181100124

• Periostin (POSTN, OSF2) expression in stroma of normal breast tissue and pre-invasive breast lesions Espinosa, I., Collins, L. C., West, R. B., Schnitt, S. J., van de Rijn, M. NATURE PUBLISHING GROUP. 2008: 30A

  View details for Web of Science ID 000252180200124

• The CSF-1 response signature in breast carcinoma 97th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Beck, A. B., Espinosa, I., van de Rijn, M., West, R. B. NATURE PUBLISHING GROUP. 2008: 22A?22A

  View details for Web of Science ID 000252180200090

• A novel breast carcinoma stromal response defined by the Nodular Fasciitis gene signature 50th Annual Meeting of the American-Society-for-Therapeutic-Radiology-and-Oncology (ASTRO) Cho, N. K., Beck, A. H., Espinosa, I., Montgomery, K., Zhu, S., van de Rijn, M., West, R. B. ELSEVIER SCIENCE INC. 2008: S686?S686

  View details for Web of Science ID 000258805302436

• HER2 status in a large, population-based cohort: Analysis of distinct HER2 subgroups 97th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Jensen, K. C., Turbin, D. A., Leung, S., Miller, M. A., Johnson, K., Norris, B., Hastie, T., McKinney, S., Nielsen, T. O., Huntsman, D. G., Gilks, C. B., West, R. B. NATURE PUBLISHING GROUP. 2008: 39A?39A

  View details for Web of Science ID 000252181100167

• Experimental approaches to the study of cancer-stroma interactions: recent findings suggest a pivotal role for stroma in carcinogenesis LABORATORY INVESTIGATION West, R. B., De Rijn, M. V. 2007; 87 (10): 967-970

  Abstract

  An increasing body of research indicates that stroma surrounding cancer cells plays an important role in the development and subsequent behavior of the tumor. Studies using a wide range of techniques, including stromal cell isolation, modification of stromal-specific gene expression, and recreation of specific microenvironment conditions in culture, have demonstrated that stroma can promote cancer and that the expression patterns within the stroma can influence clinical outcome. Major hurdles in the study of the cancer stroma revolve around the cellular complexity of the tumor microenvironment, both in modeling the microenvironment and discovering/isolating pure populations of stromal cell types.

  View details for DOI 10.1038/labinvest.3700666

  View details for PubMedID 17700561

• Translocation and expression of CSF1 in pigmented villonodular synovitis, tenosynovial giant cell tumor, rheumatoid arthritis and other reactive synovitides AMERICAN JOURNAL OF SURGICAL PATHOLOGY Cupp, J. S., Miller, M. A., Montgomery, K. D., Nielsen, T. O., O'Connell, J. X., Huntsman, D., van de Rijn, M., Gilks, C. B., West, R. B. 2007; 31 (6): 970-976

  Abstract

  We recently demonstrated that CSF1, the ligand of the tyrosine kinase receptor, CSF1R, can be translocated in pigmented villonodular synovitis (PVNS) and tenosynovial giant cell tumor (TGCT). In this study, we evaluated the staining characteristics of PVNS/TGCT and reactive synovitides for CSF1 and CSF1R by in situ hybridization and immunohistochemistry on tissue microarrays and correlated these findings with the recently described translocation. We collected specimens of TGCT/PVNS from 60 patients and of rheumatoid arthritis and other reactive synovitides from 74 patients. We identify 2 groups of PVNS and TGCT cases by the presence of CSF1 translocation and CSF1 expression. The first group (35 of 57 cases; 61%) had both the CSF1 translocation and high expression of CSF1 RNA, confirming our previous findings. Interestingly, a second group (22 of 57 cases; 39%) was identified that showed high expression of CSF1 RNA or CSF1 protein but did not have the translocation. The rheumatoid arthritis and reactive synovitis specimens showed localization of CSF1 RNA and protein to the synovial lining cells, implying a possible role for CSF1 in the pathogenesis of these lesions. As the CSF1 translocation is postulated to play an important role in the biology of PVNS/TGCT, the consistent presence of CSF1 expression in translocation-negative cases implies that other mechanisms can lead to CSF1 up-regulation. The consistent presence of CSF1 overexpression in all cases of PVNS/TGCT and reactive synovitides suggests both an important role for CSF1 in the spectrum of synovial pathologies and the possibility of targeting the CSF1/CSF1R interaction therapeutically.

  View details for PubMedID 17527089

• Oncogenic regulators and substrates of the anaphase promoting complex/cyclosome are frequently overexpressed in malignant tumors AMERICAN JOURNAL OF PATHOLOGY Lehman, N. L., Tibshirani, R., Hsu, J. Y., Natkunam, Y., Harris, B. T., West, R. B., Masek, M. A., Montgomery, K., van de Rijn, M., Jackson, P. K. 2007; 170 (5): 1793-1805

  Abstract

  The fidelity of cell division is dependent on the accumulation and ordered destruction of critical protein regulators. By triggering the appropriately timed, ubiquitin-dependent proteolysis of the mitotic regulatory proteins securin, cyclin B, aurora A kinase, and polo-like kinase 1, the anaphase promoting complex/cyclosome (APC/C) ubiquitin ligase plays an essential role in maintaining genomic stability. Misexpression of these APC/C substrates, individually, has been implicated in genomic instability and cancer. However, no comprehensive survey of the extent of their misregulation in tumors has been performed. Here, we analyzed more than 1600 benign and malignant tumors by immunohistochemical staining of tissue microarrays and found frequent overexpression of securin, polo-like kinase 1, aurora A, and Skp2 in malignant tumors. Positive and negative APC/C regulators, Cdh1 and Emi1, respectively, were also more strongly expressed in malignant versus benign tumors. Clustering and statistical analysis supports the finding that malignant tumors generally show broad misregulation of mitotic APC/C substrates not seen in benign tumors, suggesting that a "mitotic profile" in tumors may result from misregulation of the APC/C destruction pathway. This profile of misregulated mitotic APC/C substrates and regulators in malignant tumors suggests that analysis of this pathway may be diagnostically useful and represent a potentially important therapeutic target.

  View details for DOI 10.2353/ajpath.2007.060767

  View details for PubMedID 17456782

• Comparison of a new mouse monoclonal Anti-HER2/neu antibody to a rabbit polyclonal antibody 96th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Turbin, D. A., Jensen, K., Leung, S., McKinney, S. E., Huntsman, D. G., Gilks, C. B., West, R. B. NATURE PUBLISHING GROUP. 2007: 52A?52A

  View details for Web of Science ID 000244935300217

• Evaluation of Her-2/neu status in carcinomas with amplified chromosome 17 centromere locus AMERICAN JOURNAL OF CLINICAL PATHOLOGY Troxell, M. L., Bangs, C. D., Lawce, H. J., Galperin, I. B., Baiyee, D., West, R. B., Olson, S. B., Cherry, A. M. 2006; 126 (5): 709-716

  Abstract

  Accurate assessment of Her-2/neu (erb-b2) status in breast carcinoma is essential for therapy planning. Clinical assays are targeted at protein overexpression (immunohistochemical analysis) or gene amplification (fluorescence in situ hybridization [FISH]). Cases with aberrant FISH signal patterns are problematic and may lead to underreporting of Her-2/neu amplification. We performed FISH with additional chromosome 17 probes, SMS (Smith-Magenis syndrome critical region) and RARA (retinoic acid receptor), on 7 cases with unusual Her-2/CEP17 (chromosome 17 centromere control probe) results to assess whether different measurements of chromosome 17 copy number might clarify the Her-2/neu amplicon status. Although the Her-2/CEP17 ratio scores were within normal range (<2.0), the Her-2/SMS or Her-2/RARA ratio revealed amplification of Her-2/neu in 5 of 7 cases. Immunohistochemical analysis demonstrated Her-2/neu protein overexpression in the same 5 cases only. We describe novel application of SMS/RARA FISH probes for assessing cases with complex Her-2/CEP17 FISH patterns. Such additional data, correlated with immunohistochemical analysis, may help guide therapy in patients with breast carcinoma.

  View details for DOI 10.1309/9EYM6VE58F2YCD9F

  View details for PubMedID 17050068

• Bone morphogenetic protein antagonist gremlin 1 is widely expressed by cancer-associated stromal cells and can promote tumor cell proliferation PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Sneddon, J. B., Zhen, H. H., Montgomery, K., van de Rijn, M., Tward, A. D., West, R., Gladstone, H., Chang, H. Y., Morganroth, G. S., Oro, A. E., Brown, P. O. 2006; 103 (40): 14842-14847

  Abstract

  Although tissue microenvironments play critical roles in epithelial development and tumorigenesis, the factors mediating these effects are poorly understood. In this work, we used a genomic approach to identify factors produced by cells in the microenvironment of basal cell carcinoma (BCC) of the skin, one of the most common human cancers. The global gene expression programs of stromal cell cultures derived from human BCCs showed consistent, systematic differences from those derived from nontumor skin. The gene most consistently expressed at a higher level in BCC tumor stromal cells compared with those from nontumor skin was GREMLIN 1, which encodes a secreted antagonist of the bone morphogenetic protein (BMP) pathway. BMPs and their antagonists are known to play a crucial role in stem and progenitor cell biology as regulators of the balance between expansion and differentiation. Consistent with the hypothesis that BMP antagonists might have a similar role in cancer, we found GREMLIN 1 expression in the stroma of human BCC tumors but not in normal skin in vivo. Furthermore, BMP 2 and 4 are expressed by BCC cells. Ex vivo, BMP inhibits, and Gremlin 1 promotes, proliferation of cultured BCC cells. We further found that GREMLIN 1 is expressed by stromal cells in many carcinomas but not in the corresponding normal tissue counterparts that we examined. Our data suggest that BMP antagonists may be important constituents of tumor stroma, providing a favorable microenvironment for cancer cell survival and expansion in many cancers.

  View details for DOI 10.1073/pnas.0606857103

  View details for PubMedID 17003113

• Upregulation of histidine decarboxylase expression in superficial cortical nephrons during pregnancy in mice and women KIDNEY INTERNATIONAL Morgan, T. K., Montgomery, K., Mason, V., West, R. B., Wang, L., van de Rijn, M., Higgins, J. P. 2006; 70 (2): 306-314

  Abstract

  Mechanisms regulating pregnancy-induced changes in renal function are incompletely understood. Few candidate genes have been identified and data suggest that alternate mechanisms remain to be elucidated. Our objective was to screen thousands of genes expressed in kidneys from mice throughout gestation to identify possible key regulators of renal function during pregnancy. Mouse complementary DNA microarrays were used to screen for differences in expression during pregnancy in C57BL/6 mice. Interesting candidate genes whose expression varied with pregnancy were further analyzed by reverse transcription-PCR and Northern blot. Expression was localized by in situ hybridization and immunohistochemistry. Follow-up immunohistochemical analyses in archival human kidney sections from the fetus, non-pregnant, and pregnant women were also performed. Histidine decarboxylase (HDC), the enzyme that synthesizes histamine, was markedly upregulated in the mouse kidney during pregnancy. HDC expression localized to proximal tubule cells of fetal and adult mice. Females showed strong expression in the juxtamedullary zone before pregnancy and upregulation in the superficial cortical zone (SCZ) by mid-gestation. Histamine colocalized with HDC. Male mice showed only low HDC expression. Similar expression patterns were observed in human kidneys. Our results show that HDC expression and histamine production are increased in the SCZ during pregnancy. If histamine acts as a vasodilator, we speculate that increasing production in the SCZ may increase renal blood flow to this zone and recruit superficial cortical nephrons during pregnancy.

  View details for DOI 10.1038/sj.ki.5001553

  View details for Web of Science ID 000239118900011

  View details for PubMedID 16760908

• A landscape effect in tenosynovial giant-cell tumor from activation of CSF1 expression by a translocation in a minority of tumor cells PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA West, R. B., Rubin, B. P., Miller, M. A., Subramanian, S., Kaygusuz, G., Montgomery, K., Zhu, S., Marinelli, R. J., De Luca, A., Downs-Kelly, E., Goldblum, J. R., Corless, C. L., Brown, P. O., Gilks, C. B., Nielsen, T. O., Huntsman, D., van de Rijn, M. 2006; 103 (3): 690-695

  Abstract

  Tenosynovial giant-cell tumor (TGCT) and pigmented villonodular synovitis (PVNS) are related conditions with features of both reactive inflammatory disorders and clonal neoplastic proliferations. Chromosomal translocations involving chromosome 1p13 have been reported in both TGCT and PVNS. We confirm that translocations involving 1p13 are present in a majority of cases of TGCT and PVNS and show that CSF1 is the gene at the chromosome 1p13 breakpoint. In some cases of both TGCT and PVNS, CSF1 is fused to COL6A3 (2q35). The CSF1 translocations result in overexpression of CSF1. In cases of TGCT and PVNS carrying this translocation, it is present in a minority of the intratumoral cells, leading to CSF1 expression only in these cells, whereas the majority of cells express CSF1R but not CSF1, suggesting a tumor-landscaping effect with aberrant CSF1 expression in the neoplastic cells, leading to the abnormal accumulation of nonneoplastic cells that form a tumorous mass.

  View details for DOI 10.1073/pnas.0507321103

  View details for PubMedID 16407111

• The role of microarray technologies in the study of soft tissue tumours HISTOPATHOLOGY West, R. B., van de Rijn, M. 2006; 48 (1): 22-31

  Abstract

  Array technologies (gene array, tissue microarray and others) are being used in a growing number of research projects involving soft tissue tumours. Gene array techniques allow for measurements of RNA expression levels or gene copy number changes for a large number of genes in a single specimen. A complementary technique, tissue microarrays, allows for the measurement of expression of a single gene in a large number of specimens. These techniques and similar ones have created a fundamentally new approach to the investigation of soft tissue tumours. This review addresses some of the advantages, problems, and solutions to those problems that come with these technologies.

  View details for DOI 10.1111/j.1365-2559.2005.02286.x

  View details for Web of Science ID 000234030700004

  View details for PubMedID 16359534

• Epidermal growth factor receptor (EGFR) expression and gene amplification in a spectrum of spindle cell soft tissue neoplasms: A fluorescence in situ hybridization (FISH) and immunohistochemical (IHC) study 95th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Larson, A. J., Downs-Kelley, E., Skacel, M., Tubbs, R. R., Rubin, B. P., van de Rijn, M., West, R. B., Corless, C., Chiesa, A., Goldblum, J. R. NATURE PUBLISHING GROUP. 2006: 14A?14A

  View details for Web of Science ID 000234094500049

• Translocation and expression of CSF1 in pigmented villonodular synovitis, tenosynovial giant cell tumors, and reactive synovial lesions 95th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Cupp, J. S., Rubin, B. P., Miller, M. A., Subramanian, S., Montgomery, K., Marinelli, R. J., De Luca, A., Nielsen, T. O., O'Connell, J. X., Huntsman, D., van de Rijn, M., Gilks, C. B., West, R. B. NATURE PUBLISHING GROUP. 2006: 10A?10A

  View details for Web of Science ID 000234094500031

• Heat-shock protein-90 (HSP-90) expression in a spectrum of benign and malignant spindle cell neoplasms: An immunohistochemical study 95th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Larson, A. J., Downs-Kelley, E., Skacel, M., Nielsen, T. O., Ruttan, C., van de Rijn, M., West, R. B., Rubin, B. P., Corless, C., Goldblum, J. R. NATURE PUBLISHING GROUP. 2006: 13A?14A

  View details for Web of Science ID 000234207600048

• TMA-combiner, a simple software tool to permit analysis of replicate cores on tissue microarrays MODERN PATHOLOGY Liu, C. L., Montgomery, K. D., Natkunam, Y., West, R. B., Nielsen, T. O., Cheang, M. C., Turbin, D. A., Marinelli, R. J., van de Rijn, M., Higgins, J. P. 2005; 18 (12): 1641-1648

  Abstract

  We have previously published a suite of software tools that facilitates the reformulation of tissue microarray (TMA) data so that it may be analyzed using techniques originally devised for analysis of cDNA microarray data. However, current microarray data often feature multiple scores for a given tissue sample and antibody combination. Furthermore, an efficient and systematic method for combining scores that takes into account the differing staining properties of tissue epitopes has not been described. We thus present the TMA-Combiner, a new Microsoft Excel-based macro that permits analysis of data for which tissues may have two or more scores per antibody, and permits combination of data from multiple different tissue microarrays. It accomplishes this by rendering one score per tissue per antibody from two or more scores, using one of multiple user-selectable combination rules developed to account for the differing staining properties of tissue epitopes. This greatly facilitates analysis of tissue microarrays, particularly for users with large repositories of data, and may facilitate discovery of biological trends and help refine diagnostic accuracy of tissue markers in clinical samples.

  View details for DOI 10.1038/modpathol.3800491

  View details for Web of Science ID 000233372100016

  View details for PubMedID 16258508

• The gene expression profile of extraskeletal myxoid chondrosarcoma JOURNAL OF PATHOLOGY Subramanian, S., West, R. B., Marinelli, R. J., Nielsen, T. O., Rubin, B. P., Goldblum, J. R., Patel, R. M., Zhu, S., Montgomery, K., Ng, T. L., Corless, C. L., Heinrich, M. C., van de Rijn, M. 2005; 206 (4): 433-444

  Abstract

  Extraskeletal myxoid chondrosarcoma (EMC) is a soft tissue tumour that occurs primarily in the extremities and is characterized by a balanced translocation most commonly involving t(9;22) (q22;q12). The morphological spectrum of EMC is broad and thus a diagnosis based on histology alone can be difficult. Currently, no systemic therapy exists that improves survival in patients with EMC. In the present study, gene expression profiling has been performed to discover new diagnostic markers and potential therapeutic targets for this tumour type. Global gene expression profiling of ten EMCs and 26 other sarcomas using 42,000 spot cDNA microarrays revealed that the cases of EMC were closely related to each other and distinct from the other tumours profiled. Significance analysis of microarrays (SAM) identified 86 genes that distinguished EMC from the other sarcomas with 0.25% likelihood of false significance. NMB, DKK1, DNER, CLCN3, and DEF6 were the top five genes in this analysis. In situ hybridization for NMB gene expression on tissue microarrays (TMAs) containing a total of 1164 specimens representing 62 different sarcoma types and 15 different carcinoma types showed that NMB was highly expressed in 17 of 22 EMC cases and very rarely expressed in other tumours and thus could function as a novel diagnostic marker. High levels of expression of PPARG and the gene encoding its interacting protein, PPARGC1A, in most EMCs suggest activation of lipid metabolism pathways in this tumour. Small molecule inhibitors for PPARG exist and PPARG could be a potential therapeutic target for EMC.

  View details for DOI 10.1002/path.1792

  View details for PubMedID 15920699

• Determination of stromal signatures in breast carcinoma PLOS BIOLOGY West, R. B., Nuyten, D. S., Subramanian, S., Nielsen, T. O., Corless, C. L., Rubin, B. P., Montgomery, K., Zhu, S., Patel, R., Hernandez-Boussard, T., Goldblum, J. R., Brown, P. O., van De Vijver, M., van de Rijn, M. 2005; 3 (6): 1101-1110

  Abstract

  Many soft tissue tumors recapitulate features of normal connective tissue. We hypothesize that different types of fibroblastic tumors are representative of different populations of fibroblastic cells or different activation states of these cells. We examined two tumors with fibroblastic features, solitary fibrous tumor (SFT) and desmoid-type fibromatosis (DTF), by DNA microarray analysis and found that they have very different expression profiles, including significant differences in their patterns of expression of extracellular matrix genes and growth factors. Using immunohistochemistry and in situ hybridization on a tissue microarray, we found that genes specific for these two tumors have mutually specific expression in the stroma of nonneoplastic tissues. We defined a set of 786 gene spots whose pattern of expression distinguishes SFT from DTF. In an analysis of DNA microarray gene expression data from 295 previously published breast carcinomas, we found that expression of this gene set defined two groups of breast carcinomas with significant differences in overall survival. One of the groups had a favorable outcome and was defined by the expression of DTF genes. The other group of tumors had a poor prognosis and showed variable expression of genes enriched for SFT type. Our findings suggest that the host stromal response varies significantly among carcinomas and that gene expression patterns characteristic of soft tissue tumors can be used to discover new markers for normal connective tissue cells.

  View details for DOI 10.1371/journal.pbio.0030187

  View details for PubMedID 15869330

• Expression of CD163 (hemoglobin scavenger receptor) in normal tissues, lymphomas, carcinomas, and sarcomas is largely restricted to the monocyte/macrophage lineage AMERICAN JOURNAL OF SURGICAL PATHOLOGY Nguyen, T. D., Schwartz, E. J., West, R. B., Warnke, R. A., Arber, D. A., Natkunam, Y. 2005; 29 (5): 617-624

  Abstract

  CD163, a hemoglobin scavenger receptor, is expressed in monocytes and macrophages. We tested the expression of the CD163 protein in 1,105 human malignancies and normal tissues using tissue microarrays and conventional paraffin-embedded tissue sections. Besides staining nonneoplastic monocytes and histiocytes (tissue macrophages), membranous/cytoplasmic staining for CD163 was primarily limited to neoplasms with monocytic/histiocytic differentiation. CD163 reactivity was not observed in normal tissues, lymphomas, carcinomas, and in a majority of mesenchymal neoplasms, including follicular dendritic cell tumors (0 of 4), although it stained admixed histiocytes. Staining for CD163 was seen in Rosai-Dorfman disease (5 of 6), histiocytic sarcoma (3 of 4), littoral cell angioma (6 of 6), and Langerhans cell histiocytosis (3 of 5). A subset of atypical fibrous histiocytomas (9 of 16), benign fibrous histiocytomas (6 of 9), and atypical fibroxanthomas (1 of 3) also showed CD163 staining. Our studies also confirm earlier work showing that CD163 is expressed in acute myeloid leukemia with monocytic differentiation (AML, FAB subtype M5) (2 of 6), as well as a majority of giant cell tenosynovial tumors (7 of 8). Its limited range of expression and tissue specificity indicate that CD163 may have significant diagnostic utility in separating specific tumors with monocytic and histiocytic derivation from other entities in their differential diagnosis.

  View details for Web of Science ID 000228707200007

  View details for PubMedID 15832085

• Nuclear beta-catenin in mesenchymal tumors MODERN PATHOLOGY Ng, T. L., GOWN, A. M., Barry, T. S., Cheang, M. C., Chan, A. K., Turbin, D. A., Hsu, F. D., West, R. B., Nielsen, T. O. 2005; 18 (1): 68-74

  Abstract

  Beta-catenin is a crucial part of the Wnt and E-cadherin signalling pathways, which are involved in tumorigenesis. Dysregulation of these pathways allow beta-catenin to accumulate and translocate to the nucleus, where it may activate oncogenes. Such nuclear accumulation can be detected by immunohistochemistry, which may be useful in diagnosis. Although the role of beta-catenin has been established in various types of carcinomas, relatively little is known about its status in mesenchymal tumors. A number of studies suggest that beta-catenin dysregulation is important in desmoid-type fibromatosis, as well as in synovial sarcoma. We wished to determine whether nuclear beta-catenin expression is specific to and sensitive for particular bone and soft-tissue tumors, including sporadic desmoid-type fibromatosis. We studied the nuclear expression of beta-catenin using tissue microarrays in a comprehensive range of bone and soft-tissue tumor types. A total of 549 cases were included in our panel. Nuclear immunohistochemical staining was determined to be either high level (>25% of cells), low level (0-25%) or none. High-level nuclear beta-catenin staining was seen in a very limited subset of tumor types, including desmoid-type fibromatosis (71% of cases), solitary fibrous tumor (40%), endometrial stromal sarcoma (40%) and synovial sarcoma (28%). Although occasional cases of fibrosarcoma, clear cell sarcoma and carcinosarcoma had high-level staining, no high-level nuclear beta-catenin expression was seen in any of 381 fibrohistocytic, muscular, adipocytic, chondroid or osseous tumor cases representing 42 diagnostic categories. All primary immunostain tissue microarray images are made publicly accessible in a searchable database. High-level nuclear beta-catenin staining serves as a useful diagnostic tool, as it is specific to a small subset of mesenchymal tumors.

  View details for DOI 10.1038/modpathol.3800272

  View details for Web of Science ID 000226109900010

  View details for PubMedID 15375433

• Characterization of anti-fatty acid synthase expression in normal breast and breast cancer. 28th Annual San Antonio Breast Cancer Symposium Jensen, K. C., Montgomery, K., West, R. B., van de Rijn, J. M. SPRINGER. 2005: S282?S282

  View details for Web of Science ID 000233407100765

• CSF1R expression in soft tissue tumors 94th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology West, R. B., Kaygusuz, G., Subramanian, S., Corless, C. L., Rubin, B. P., Montgometry, K., Zhu, S. X., Nielsen, T. I., Patel, R., Goldblum, J. R., Brown, P. O. NATURE PUBLISHING GROUP. 2005: 23A?23A

  View details for Web of Science ID 000226117900090

• CSF1 expression signature identifies a subset of breast carcinomas and influences outcome. 28th Annual San Antonio Breast Cancer Symposium West, R. B., Horlings, H., Nuyten, D. S., Subramanian, S., Zhu, S. X., Miller, M., Rubin, B. P., Nielsen, T. O., Gilks, C. B., Huntsman, D. G., Tibshirani, R., van De Vijver, M., van de Rijn, M. SPRINGER. 2005: S135?S135

  View details for Web of Science ID 000233407100364

• Stromal expression signatures predict outcome in breast carcinoma 94th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology West, R. B., Nuyten, D. S., Subramanian, S., Corless, C., Rubin, B. P., Montgomergy, K., Zhu, S. X., Nielsen, T. O., Patel, R., Goldblum, J. R., Brown, P. O., van De Vijver, M., van de Rijn, M. NATURE PUBLISHING GROUP. 2005: 55A?55A

  View details for Web of Science ID 000226238600241

• Novel gene ANKS1 expression in gastrointestinal stromal tumors 94th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Kalof, A. N., West, R. B., Subramanian, S., Zhu, S., Montgomery, K., Nielsen, T. O., Goldblum, J. R., Patel, R., Rubin, B. P., Van vde Rijn, M. NATURE PUBLISHING GROUP. 2005: 16A?16A

  View details for Web of Science ID 000226117900060

• Gastrointestinal stromal tumors (GISTs) with KIT and PDGFRA mutations have distinct gene expression profiles ONCOGENE Subramanian, S., West, R. B., Corless, C. L., Ou, W. B., Rubin, B. P., Chu, K. M., Leung, S. Y., Yuen, S. T., Zhu, S., Hernandez-Boussard, T., Montgomery, K., Nielsen, T. O., Patel, R. M., Goldblum, J. R., Heinrich, M. C., Fletcher, J. A., van de Rijn, M. 2004; 23 (47): 7780-7790

  Abstract

  Most GISTs require oncogenic activation of the KIT or PDGFRA receptor tyrosine kinase proteins, and the genomic mechanisms of oncogene activation are heterogeneous. Notably, the kinase mutation type correlates with both tumor biology and imatinib response. For example, GISTs with KIT exon 11 mutations are typically gastric and have excellent imatinib response, whereas those with KIT exon 9 mutations generally arise in the small bowel and are less responsive to imatinib. To identify genes that might contribute to these biological differences, we carried out gene expression profiling of 26 GISTs with known KIT and PDGFRA mutational status. Expression differences were then evaluated further by RNA in situ hybridization, immunohistochemistry, and immunoblotting. Unsupervised hierarchical clustering grouped tumors with similar mutations together, but the distinction between the different groups was not absolute. Differentially expressed genes included ezrin, p70S6K, and PKCs, which are known to have key roles in KIT or PDGFRA signaling, and which might therefore contribute to the distinctive clinicopathological features in GISTs with different mutation types. These gene products could serve as highly selective therapeutic targets in GISTs containing the KIT or PDGFRA mutational types with which they are associated.

  View details for DOI 10.1038/sj.onc.1208056

  View details for PubMedID 15326474

• CD117 expression in mesothelioma MODERN PATHOLOGY Arber, D. A., Weiss, L. M., West, R. B. 2004; 17 (8): 1021-1021

  View details for DOI 10.1038/modpathol.3800159

  View details for Web of Science ID 000223118300016

  View details for PubMedID 15263910

• Apo D in soft tissue tumors - A novel marker for dermatofibrosarcoma protuberans AMERICAN JOURNAL OF SURGICAL PATHOLOGY West, R. B., Harvell, J., Linn, S. C., Lui, C. L., Prapong, W., Hernandez-Boussard, T., Montgomery, K., Nielsen, T. O., Rubin, B. P., Patel, R., Goldblum, J. R., Brown, P. O., van de Rijn, M. 2004; 28 (8): 1063-1069

  Abstract

  Using gene microarray expression profiling, we previously found that apolipoprotein D (Apo D) was highly expressed in dermatofibrosarcoma protuberans (DFSP). In this study, we confirm that Apo D is highly and relatively specifically expressed in DFSP using immunohistochemistry. A tissue microarray containing 421 soft tissue tumors was constructed and stained with antibodies against Apo D and CD34. Cytoplasmic immunostaining for Apo D was found in 9 of 10 typical DFSPs. In addition, 3 of 3 Bednar tumors and 2 of 3 giant cell fibroblastomas stained in conventional sections. In contrast, Apo D was immunoreactive in only a very small subset of a diverse collection of other soft tissue tumors, including Malignant Fibrous Histiocytoma (MFH), glomus tumor, neurofibroma, and malignant peripheral nerve sheath tumors. Immunostains for Apo D were negative in conventional sections of 16 fibrous histiocytomas, and an additional 12 variants of fibrous histiocytoma. Digital images of all immunohistochemical and hematoxylin and eosin tissue microarray stains are available at the accompanying website (http://microarray-pubs.stanford.edu/tma_portal/apod/). We conclude that Apo D is strongly expressed in DFSPs and neural lesions and may be useful in differentiating DFSP from fibrous histiocytoma.

  View details for PubMedID 15252314

• The novel marker, DOG1, is expressed ubiquitously in gastrointestinal stromal tumors irrespective of KIT or PDGFRA mutation status AMERICAN JOURNAL OF PATHOLOGY West, R. B., Corless, C. L., Chen, X., Rubin, B. P., Subramanian, S., Montgomery, K., Zhu, S., Ball, C. A., Nielsen, T. O., Patel, R., Goldblum, J. R., Brown, P. O., Heinrich, M. C., van de Rijn, M. 2004; 165 (1): 107-113

  Abstract

  We recently characterized gene expression patterns in gastrointestinal stromal tumors (GISTs) using cDNA microarrays, and found that the gene FLJ10261 (DOG1, discovered on GIST-1), encoding a hypothetical protein, was specifically expressed in GISTs. The immunoreactivity of a rabbit antiserum to synthetic DOG1 peptides was assessed on two soft tissue tumor microarrays. The tissue microarrays included 587 soft tissue tumors, with 149 GISTs, including 127 GIST cases for which the KIT and PDGFRA mutation status was known. Immunoreactivity for DOG1 was found in 136 of 139 (97.8%) of scorable GISTs. All seven GIST cases with a PDGFRA mutation were DOG1-positive, while most of these failed to react for KIT. The immunohistochemical findings were confirmed with in situ hybridization probes for DOG1, KIT, and PDGFRA. Other neoplasms in the differential diagnosis of GIST, including desmoid fibromatosis (0 of 17) and Schwannoma (0 of 3), were immunonegative for DOG1. Only 4 of 438 non-GIST cases were immunoreactive for DOG1. DOG1, a protein of unknown function, is expressed strongly on the cell surface of GISTs and is rarely expressed in other soft tissue tumors. Reactivity for DOG1 may aid in the diagnosis of GISTs, including PDGFRA mutants that fail to express KIT antigen, and lead to appropriate treatment with imatinib mesylate, an inhibitor of the KIT tyrosine kinase.

  View details for PubMedID 15215166

• Gene expression signature of fibroblast serum response predicts human cancer progression: similarities between tumors and wounds. PLoS biology Chang, H. Y., Sneddon, J. B., Alizadeh, A. A., Sood, R., West, R. B., Montgomery, K., Chi, J., van de Rijn, M., Botstein, D., Brown, P. O. 2004; 2 (2): E7-?

  Abstract

  Cancer invasion and metastasis have been likened to wound healing gone awry. Despite parallels in cellular behavior between cancer progression and wound healing, the molecular relationships between these two processes and their prognostic implications are unclear. In this study, based on gene expression profiles of fibroblasts from ten anatomic sites, we identify a stereotyped gene expression program in response to serum exposure that appears to reflect the multifaceted role of fibroblasts in wound healing. The genes comprising this fibroblast common serum response are coordinately regulated in many human tumors, allowing us to identify tumors with gene expression signatures suggestive of active wounds. Genes induced in the fibroblast serum-response program are expressed in tumors by the tumor cells themselves, by tumor-associated fibroblasts, or both. The molecular features that define this wound-like phenotype are evident at an early clinical stage, persist during treatment, and predict increased risk of metastasis and death in breast, lung, and gastric carcinomas. Thus, the transcriptional signature of the response of fibroblasts to serum provides a possible link between cancer progression and wound healing, as well as a powerful predictor of the clinical course in several common carcinomas.

  View details for PubMedID 14737219

• Gene expression signature of fibroblast serum response predicts human cancer progression: Similarities between tumors and wounds PLOS BIOLOGY Chang, H. Y., Sneddon, J. B., Alizadeh, A. A., Sood, R., West, R. B., Montgomery, K., Chi, J. T., van de Rijn, M., Botstein, D., Brown, P. O. 2004; 2 (2): 206-214

  Abstract

  Cancer invasion and metastasis have been likened to wound healing gone awry. Despite parallels in cellular behavior between cancer progression and wound healing, the molecular relationships between these two processes and their prognostic implications are unclear. In this study, based on gene expression profiles of fibroblasts from ten anatomic sites, we identify a stereotyped gene expression program in response to serum exposure that appears to reflect the multifaceted role of fibroblasts in wound healing. The genes comprising this fibroblast common serum response are coordinately regulated in many human tumors, allowing us to identify tumors with gene expression signatures suggestive of active wounds. Genes induced in the fibroblast serum-response program are expressed in tumors by the tumor cells themselves, by tumor-associated fibroblasts, or both. The molecular features that define this wound-like phenotype are evident at an early clinical stage, persist during treatment, and predict increased risk of metastasis and death in breast, lung, and gastric carcinomas. Thus, the transcriptional signature of the response of fibroblasts to serum provides a possible link between cancer progression and wound healing, as well as a powerful predictor of the clinical course in several common carcinomas.

  View details for DOI 10.1371/journal.pbio.0020007

  View details for Web of Science ID 000189314400013

  View details for PubMedCentralID PMC314300

• Gene expression patterns and gene copy number changes in dermatofibrosarcoma protuberans AMERICAN JOURNAL OF PATHOLOGY Linn, S. C., West, R. B., Pollack, J. R., Zhu, S., Hernandez-Boussard, T., Nielsen, T. O., Rubin, B. P., Patel, R., Goldblum, J. R., Siegmund, D., Botstein, D., Brown, P. O., Gilks, C. B., van de Rijn, M. 2003; 163 (6): 2383-2395

  Abstract

  Dermatofibrosarcoma protuberans (DFSP) is an aggressive spindle cell neoplasm. It is associated with the chromosomal translocation, t(17:22), which fuses the COL1A1 and PDGFbeta genes. We determined the characteristic gene expression profile of DFSP and characterized DNA copy number changes in DFSP by array-based comparative genomic hybridization (array CGH). Fresh frozen and formalin-fixed, paraffin-embedded samples of DFSP were analyzed by array CGH (four cases) and DNA microarray analysis of global gene expression (nine cases). The nine DFSPs were readily distinguished from 27 other diverse soft tissue tumors based on their gene expression patterns. Genes characteristically expressed in the DFSPs included PDGF beta and its receptor, PDGFRB, APOD, MEOX1, PLA2R, and PRKCA. Array CGH of DNA extracted either from frozen tumor samples or from paraffin blocks yielded equivalent results. Large areas of chromosomes 17q and 22q, bounded by COL1A1 and PDGF beta, respectively, were amplified in DFSP. Expression of genes in the amplified regions was significantly elevated. Our data shows that: 1) DFSP has a distinctive gene expression profile; 2) array CGH can be applied successfully to frozen or formalin-fixed, paraffin-embedded tumor samples; 3) a characteristic amplification of sequences from chromosomes 17q and 22q, demarcated by the COL1A1 and PDGF beta genes, respectively, was associated with elevated expression of the amplified genes.

  View details for PubMedID 14633610

• Tissue microarray validation of epidermal growth factor receptor and SALL2 in synovial sarcoma with comparison to tumors of similar histology AMERICAN JOURNAL OF PATHOLOGY Nielsen, T. O., Hsu, F. D., O'Connell, J. X., Gilks, C. B., Sorensen, P. H., Linn, S., West, R. B., Liu, C. L., Botstein, D., Brown, P. O., van de Rijn, M. 2003; 163 (4): 1449-1456

  Abstract

  Histological diagnosis of synovial sarcoma can be difficult. Genome-wide expression profiling has identified a number of genes expressed at higher levels in synovial sarcoma than in other soft tissue tumors, representing excellent candidates for diagnostic immunohistochemical markers. A tissue microarray comprising 77 sarcomas, including 46 synovial sarcomas, was constructed to validate identified markers and investigate their expression in tumors in the differential diagnosis of synovial sarcoma. Immunostaining was performed for two such markers, epidermal growth factor receptor and SAL (drosophila)-like 2 (SALL2), and for fifteen established markers used in the differential diagnosis of sarcomas. As predicted by expression profiling, epidermal growth factor receptor (a potential therapeutic target) and SALL2 stained most cases of synovial sarcoma; staining was significantly less common among other tested sarcomas. Hierarchical clustering analysis applied to immunostaining results for all 18 antibodies showed that synovial sarcomas, leiomyosarcomas, hemangiopericytomas, and solitary fibrous tumors cluster distinctly, and assigned one case with indeterminate histology as a Ewing sarcoma. Digital images from over 2500 immunostained cores analyzed in this study were captured and are made accessible through the accompanying website: http://microarray-pubs.stanford.edu/tma_portal/synsarc.

  View details for Web of Science ID 000185517500022

  View details for PubMedID 14507652

  View details for PubMedCentralID PMC1868308

• Array-based comparative genomic hybridization (ACGH) of dermatofibrosarcoma protuberans (DFSP) on cDNA micorarrays using DNA isolated from fresh frozen and paraffin embedded tissue 92nd Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Jensen, K., West, R. B., Zhu, S. X., Linn, S. C., Nielsen, T. O., Goldblum, J. R., Patel, R., Rubin, B. P., Botstein, D., BROWN, P., Pollack, J., Gilks, B., van de Rijn, M. NATURE PUBLISHING GROUP. 2003: 15A?15A

  View details for Web of Science ID 000180732500067

• Array-based comparative genomic hybridization (ACGH) of dermatofibrosarconta protuberans (DFSP) on cDNA micorarrays using DNA isolated from fresh frozen and paraffin embedded tissue 92nd Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Jensen, K., West, R. B., Zhu, S. X., Linn, S. C., Nielsen, T. O., Goldblum, J. R., Patel, R., Rubin, B. P., Botstein, D., BROWN, P., Pollack, J., Gilks, B., van de Rijn, M. NATURE PUBLISHING GROUP. 2003: 15A?15A

  View details for Web of Science ID 000180720100066

• Expression profiling of fibromatosis by cDNA gene array analysis 92nd Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Cheng, L., West, R. B., Zhu, S., Linn, S. C., Nielsen, T. O., Goldblum, J. R., Patel, R., Rubin, B. P., BROWN, P., Botstein, D., van de Rijn, M. NATURE PUBLISHING GROUP. 2003: 10A?10A

  View details for Web of Science ID 000180720100042

• Novel gene FLJ10261 in gastrointestinal stromal tumors 92nd Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology West, R. B., Linn, S. C., Nielsen, T. O., Montgomery, K., Goldblum, J. R., Patel, R., Rubin, B. P., BROWN, P., Botstein, D., van de Rijn, M. NATURE PUBLISHING GROUP. 2003: 20A?20A

  View details for Web of Science ID 000180720100089

• Molecular characterisation of soft tissue tumours: a gene expression study LANCET Nielsen, T. O., West, R. B., Linn, S. C., Alter, O., Knowling, M. A., O'Connell, J. X., Zhu, S., Fero, M., Sherlock, G., Pollack, J. R., Brown, P. O., Botstein, D., van de Rijn, M. 2002; 359 (9314): 1301-1307

  Abstract

  Soft-tissue tumours are derived from mesenchymal cells such as fibroblasts, muscle cells, or adipocytes, but for many such tumours the histogenesis is controversial. We aimed to start molecular characterisation of these rare neoplasms and to do a genome-wide search for new diagnostic markers.We analysed gene-expression patterns of 41 soft-tissue tumours with spotted cDNA microarrays. After removal of errors introduced by use of different microarray batches, the expression patterns of 5520 genes that were well defined were used to separate tumours into discrete groups by hierarchical clustering and singular value decomposition.Synovial sarcomas, gastrointestinal stromal tumours, neural tumours, and a subset of the leiomyosarcomas, showed strikingly distinct gene-expression patterns. Other tumour categories--malignant fibrous histiocytoma, liposarcoma, and the remaining leiomyosarcomas--shared molecular profiles that were not predicted by histological features or immunohistochemistry. Strong expression of known genes, such as KIT in gastrointestinal stromal tumours, was noted within gene sets that distinguished the different sarcomas. However, many uncharacterised genes also contributed to the distinction between tumour types.These results suggest a new method for classification of soft-tissue tumours, which could improve on the method based on histological findings. Large numbers of uncharacterised genes contributed to distinctions between the tumours, and some of these could be useful markers for diagnosis, have prognostic significance, or prove possible targets for treatment.

  View details for Web of Science ID 000174989700013

  View details for PubMedID 11965276

• The usefulness of immunohistochemistry in the diagnosis of follicular lymphoma in bone marrow biopsy specimens AMERICAN JOURNAL OF CLINICAL PATHOLOGY West, R. B., Warnke, R. A., Natkunam, Y. 2002; 117 (4): 636-643

  Abstract

  We used a panel of paraffin antibodies to determine whether neoplastic and nonneoplastic lymphoid aggregates in the bone marrow can be distinguished reliably. Formalin-fixed, paraffin-embedded bone marrow core biopsy specimens with lymphoid aggregates were stained using primary antibodies directed against bcl-2, bcl-6, CD5, CD10, CD20, and CD23. We studied 61 cases (26 follicular lymphoma and 35 benign or atypical aggregates). We found that no single stain is sufficient for identification of neoplastic lymphoid aggregates. However, this distinction was made possible by using a panel of antibodies. Under the conditions we tested, the most useful antibodies were CD10, bcl-2, CD5, and CD20. Most benign or atypical aggregates do not express CD10 and CD23. In addition, nonneoplastic aggregates had a large population of T cells. bcl-2 was useful in an architectural context for distinguishing neoplastic aggregates. bcl-6 often was expressed in both neoplastic and nonneoplastic aggregates and, thus, poorly discriminated between these processes. We studied the expression of CD10 and bcl-6 in selected lymph nodes in some cases.

  View details for Web of Science ID 000174715000018

  View details for PubMedID 11939740

• The mRNA expression signature of solitary fibrous tumors West, R. B., Linn, S. C., Foxman, E., Neilson, T., Zhu, S., Alter, O., Goldblum, J. R., Patel, R., Rubin, B. P., Brown, P. O., Botstein, D., van de Rijn, M. NATURE PUBLISHING GROUP. 2002: 24A?24A

  View details for Web of Science ID 000173379700100

• Tissue Microarray analysis of known and novel synovial sarcoma markers Nielsen, T. O., O'Connell, J. X., Hsu, F. D., West, R. B., Linn, S. C., van de Rijn, M. NATURE PUBLISHING GROUP. 2002: 20A?20A

  View details for Web of Science ID 000173379700084

• Genome-wide mRNA expression profiling of dermatofibrosarcoma protuberans using cDNA microarrays Linn, S. C., West, R. B., Zhu, S., Nielsen, T., Goldblum, J. R., Patel, R., Rubin, B. P., Alter, O., Brown, P. O., Botstein, D., van de Rijn, M. NATURE PUBLISHING GROUP. 2002: 18A?18A

  View details for Web of Science ID 000173388900077

• Tissue microarray analysis of CD44, p53, Ki-67 and bcl-2 as prognostic markers in rhabdomyosarcoma Linn, S. C., West, R. B., Wijnaendts, L. C., Montgomery, K., Mitchell, J., van de Rijn, M. NATURE PUBLISHING GROUP. 2002: 312A?313A

  View details for Web of Science ID 000173388901322

• Distinction between low grade endometrial stromal sarcoma and smooth muscle tumors by cDNA gene array analysis West, R. B., Linn, S. C., Nielsen, T., Zhu, S., Longacre, T., Husain, A., Alter, O., Patel, R., Brown, P. O., Botstein, D., Rubin, B. P., Goldblum, J. R., van de Rijn, M. NATURE PUBLISHING GROUP. 2002: 213A?214A

  View details for Web of Science ID 000173379700909

• The RAG-HMG1 complex enforces the 12/23 rule of V(D)J recombination specifically at the double-hairpin formation step MOLECULAR AND CELLULAR BIOLOGY West, R. B., Lieber, M. R. 1998; 18 (11): 6408-6415

  Abstract

  A central unanswered question concerning the initial phases of V(D)J recombination has been at which step the 12/23 rule applies. This rule, which governs which variable (V), diversity (D), and joining (J) segments are able to pair during recombination, could operate at the level of signal sequence synapsis after RAG-HMG1 complex binding, signal nicking, or signal hairpin formation. It has also been unclear whether additional proteins are required to achieve adherence to the 12/23 rule. We developed a novel system for the detailed biochemical analysis of the 12/23 rule by using an oligonucleotide-based substrate that can include two signals. Under physiologic conditions, we found that the complex of RAG1, RAG2, and HMG1 can successfully recapitulate the 12/23 rule with the same specificity as that seen intracellularly and in crude extracts. The cleavage complex can bind and nick 12x12 and 23x23 substrates as well as 12x23 substrates. However, hairpin formation occurs at both of the signals only on 12x23 substrates. Moreover, under physiologic conditions, the presence of a partner 23-bp spacer suppresses single-site hairpin formation at a 12-bp spacer and vice versa. Hence, this study illustrates that synapsis suppresses single-site reactions, thereby explaining the high physiologic ratio of paired versus unpaired V(D)J recombination events in lymphoid cells.

  View details for Web of Science ID 000076512900023

  View details for PubMedID 9774656

• Productive and nonproductive complexes of Ku and DNA-dependent protein kinase at DNA termini MOLECULAR AND CELLULAR BIOLOGY West, R. B., Yaneva, M., Lieber, M. R. 1998; 18 (10): 5908-5920

  Abstract

  DNA-dependent protein kinase (DNA-PK) is the only eukaryotic protein kinase known to be specifically activated by double-stranded DNA (dsDNA) termini, accounting for its importance in repair of dsDNA breaks and its role in physiologic processes involving dsDNA breaks, such as V(D)J recombination. In this study we conducted kinase and binding analyses using DNA-PK on DNA termini of various lengths in the presence and absence of Ku. We confirmed our previous observations that DNA-PK can bind DNA termini in the absence of Ku, and we determined rate constants for binding. However, in the presence of Ku, DNA-PK can assume either a productive or a nonproductive configuration, depending on the length of the DNA terminus. For dsDNA greater than 26 bp, the productive mode is achieved and Ku increases the affinity of the DNA-PK for the Ku:DNA complex. The change in affinity is achieved by increases in both the kinetic association rate and reduction in the kinetic dissociation rate. For dsDNA smaller than 26 bp, the nonproductive mode, in which DNA-PK is bound to Ku:DNA but is inactive as a kinase, is assumed. Both the productive and nonproductive configurations are likely to be of physiologic importance, depending on the distance of the dsDNA break site to other protein complexes, such as nucleosomes.

  View details for Web of Science ID 000075980900030

  View details for PubMedID 9742108

• Antigen receptor gene rearrangement CURRENT OPINION IN IMMUNOLOGY Grawunder, U., West, R. B., Lieber, M. R. 1998; 10 (2): 172-180

  Abstract

  Two specialized forms of site-directed double-strand (ds) DNA breakage and rejoining are part of the physiologic program of lymphocytes. One is recombination of the V, D and J gene sequences, termed V(D)J recombination, occurring during early B- and T-cell development, and the other is class-switch recombination occurring exclusively in mature B cells. For V(D)J recombination significant progress has been made recently elucidating the biochemistry of the reaction. In particular our understanding of how DNA ds breaks are both generated and rejoined has increased. For class-switch recombination no definitive information is known about the nucleases required for making the ds breaks, but recent evidence suggests that the joining phase shares activities also required for V(D)J recombination and general DNA ds break repair.

  View details for Web of Science ID 000073402300009

  View details for PubMedID 9602306