CAP Profiles

Bio

Clinical Focus

  • Cancer > Hematology
  • Hematology
  • Pathology
  • Pathology and Laboratory Medicine

Academic Appointments

Administrative Appointments

  • Director, Clinical Pathology, Department of Pathology, Stanford University School of Medicine (2016 - Present)
  • Director, Coagulation and Molecular Pathology Laboratories, Stanford Hospital (1995 - Present)
  • Attending Physician, Hematology Service, Stanford Hospital (1995 - Present)

Professional Education

  • Board Certification, American Board of Internal Medicine, Hematology (2016)
  • Board Certification: Hematology, American Board of Internal Medicine (1992)
  • Fellowship:Stanford University (1984) CA
  • Residency:Stanford University (1984) CA
  • Internship:Stanford University School of Medicine (1984) CA
  • Board Certification: Internal Medicine, American Board of Internal Medicine (1987)
  • Medical Education:Tufts University (1984) MA
  • MD, Tufts Univ. School of Medicine, Medicine (1984)
  • BA, UC Santa Cruz, Chemistry (1979)

Contact

    • Tel: (650) 723-9232
    Fax: (650) 736-1476
  • Mirela Puljic Administrative Associate
    • Tel: (650) 725-5604
  • Pathology Department 300 Pasteur Dr Rm L235 MC 5324 Stanford, CA 94305
    • Tel: (650) 723-9232
    Fax: (650) 736-1476

Research & Scholarship

Current Research and Scholarly Interests

Our laboratory focuses on translational research in 2 main areas - genomic approaches to diagnosis and minimal residual disease testing for patients with cancer, and molecular basis of disorders of thrombosis and hemostasis. My clinical focus is in molecular pathology, diagnosis and treatment of disorders of hemostasis and thrombosis and general hematology.

Clinical Trials

  • A Study to Test the Safety of the Investigational Drug Loxo-195 in Children and Adults That May Treat Cancer Recruiting

    This research study is done to test the safety of the new drug Loxo-195 in children and adults with cancer having a change in a particular gene (NTRK1, NTRK2 or NTRK3). The drug may treat cancer by interfering with the effect of the NTRK genes on cancer growth. The study also investigates how the drug is absorbed and processed in the human body, and how well and for how long the cancer responds to the drug. This is the first study to test Loxo-195 in humans with cancer, for whom no other effective therapy exists.

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  • Bone Marrow Grafting for Leukemia and Lymphoma Recruiting

    The purpose of this study is to obtain tissue samples for ongoing studies regarding transplant outcomes and complications.

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  • A Longitudinal Study of Plasma EBV DNA in Nasopharyngeal Carcinoma From Both Endemic and Non-Endemic Patient Populations Not Recruiting

    1. To determine the prognostic implication of plasma Epstein-Bar Virus (EBV) DNA concentrations, as measured by quantitative polymerase chain reaction (PCR) in patients with nasopharyngeal carcinoma (NPC). 2. To relate pretreatment plasma EBV DNA concentration to WHO classification of these tumors both in endemic and non-endemic areas. 3. To determine whether pretreatment plasma EBV DNA can serve as a prognostic factor for both endemic and non-endemic patient populations.

    Stanford is currently not accepting patients for this trial. For more information, please contact Quynh-Thu Le, 650-498-6184.

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  • Evaluation of Pathwork Tissue of Origin (TOO) Test for Human Malignancies Not Recruiting

    The pathworks tissue of origin test is a microarray-based test with the goal of identifying the tissue of origin in patients with metastatic tumors of unknown primary site.

    Stanford is currently not accepting patients for this trial. For more information, please contact James Zehnder, (650) 723 - 9232.

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  • Genome-Wide Gene Expression Profiling of Patients With ITP Receiving Thrombopoietin Mimetics Not Recruiting

    Introduction: Ineffective platelet production has been proven to play a role in the etiology of Immune Thrombocytopenia (ITP) in addition to increased platelet destruction. The second-generation thrombopoietin (TPO) mimetics have shown good efficacy in boosting platelet counts in the great majority of patients with chronic ITP in several clinical trials.1, 2 Nevertheless, about 20% of patients with ITP fail to respond to the TPO mimetic treatment. Those treatment-resistant patients are un-characterized and the reasons for the lack of response have not been studied. The identification of predictive blood biomarkers of patients' response to treatment will be useful in reducing both cost and potential side effects; and it will be of equal importance and interest to investigate the molecular mechanisms underlying the patients' heterogeneous responses to TPO mimetic treatment. Specific Aims: 1. To identify blood classifier genes which correlate with patients' response to TPO mimetic treatment. 2. To compare the blood gene expression changes in responders and non-responders after TPO mimetic treatment and explore the possible molecular mechanisms accounting for the non-responsiveness to the treatment.

    Stanford is currently not accepting patients for this trial. For more information, please contact James L Zehnder, MD, 650-723-9232.

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  • Integrated Whole-Genome Analysis of Hematologic Disorders Not Recruiting

    We will use new technologies to look at the DNA, RNA, proteins, and metabolites in the disease-containing blood, bone marrow, or tissue and normal cells from the skin. Our goal is to analyze all of the genes in the diseased and normal skin sample. By comparing the results of the diseased sample and normal skin cells and the results of the two types of genetic information (DNA and RNA), we should be able to identify genetic changes that are important for the initiation, progression, or treatment response of that particular disorder.

    Stanford is currently not accepting patients for this trial. For more information, please contact Jason D Merker, 650-922-1885.

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  • Phase 2 Fludarabine, Cytoxan and FCCAM <Alemtuzumab> in Untreated B-Cell Chronic Lymphocytic Leukemia Not Recruiting

    The primary objective of this study was to evaluate the safety and efficacy of the combination of fludarabine and cyclophosphamide in previously untreated CLL patients. Participants will receive fludarabine and cyclophosphamide on days 1, 2, and 3 of six 28-day cycles.

    Stanford is currently not accepting patients for this trial. For more information, please contact Nini Estevez, (650) 725 - 4041.

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  • Phase 2 Study of Temozolomide to Treat Poor Risk / Refractory Acute Myeloid Leukemia Not Recruiting

    Open-label, non-randomized, parallel assignment, phase 2 trial assessing the safety and efficacy of distinct temozolomide treatment regimens for patients with AML and poor prognosis

    Stanford is currently not accepting patients for this trial. For more information, please contact Richa Rajwanshi, (650) 736 - 4031.

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Teaching

2018-19 Courses

Stanford Advisees

Publications

All Publications

  • Clonal Evolution and Changes in Two AML Patients Detected with A Novel Single-Cell DNA Sequencing Platform. Scientific reports Xu, L., Durruthy-Durruthy, R., Eastburn, D. J., Pellegrino, M., Shah, O., Meyer, E., Zehnder, J. 2019; 9 (1): 11119

    Abstract

    Next-generation sequencing (NGS) is used to detect gene variants in genetically complex cell populations of cancer patient samples. Traditional bulk analysis can only provide average variant allele frequencies of the targeted genes across all sampled cells. It fails to resolve mutational co-occurrences and may miss rare cancer cells. Genome analysis at the single cell level offers the opportunity to more fully resolve clonal architecture. Peripheral blood mononuclear cells were sampled from acute myeloid leukemia patients longitudinally and single-cell DNA sequencing libraries were generated with a novel droplet-based microfluidics approach. Molecular profiling of single nucleotide variants across thousands of cells revealed genetic chimerism in patients after bone marrow transplantation (BMT). Importantly, hierarchical clustering analysis of single nucleotide variants (SNVs) uncovered a distinct oncogenic clone of cells carrying mutated tumor-suppressor and/or oncogene(s). This novel single-cell DNA sequencing approach enabled precise monitoring of engraftment and revealed clonal evolution of oncogenic cells during the progression and treatment of the disease.

    View details for DOI 10.1038/s41598-019-47297-z

    View details for PubMedID 31366893

  • Indolent In Situ B-Cell Neoplasms With MYC Rearrangements Show Somatic Mutations in MYC and TNFRSF14 by Next-generation Sequencing. The American journal of surgical pathology Kumar, J., Butzmann, A., Wu, S., Easly, S., Zehnder, J. L., Warnke, R. A., Bangs, C. D., Jangam, D., Cherry, A., Lau, J., Nybakken, G., Ohgami, R. S. 2019

    Abstract

    Systemic high-grade B-cell lymphomas (HGBCLs) with MYC gene rearrangements are clinically aggressive. In situ lesions with indolent behavior have not been described to date. We have identified 2 cases of in situ B-cell neoplasms with MYC rearrangements (IS-BCN, MYC) occurring, and focally confined to ≤4 lymphoid follicles in otherwise healthy individuals and without clinical progression despite minimal intervention (surgical only). Morphologically similar to systemic HGBCLs, the low power view of these lesions showed a starry sky pattern with numerous mitotic figures. High power imaging demonstrated these cells to be medium-large in size with irregular nuclear contours, immature chromatin, and prominent nucleoli. Immunophenotypically these cells were light chain restricted, positive for CD20, CD10, c-Myc, and dim or negative for BCL2 with a Ki67 proliferative index of >95%. By fluorescence in situ hybridization studies, we detected MYC translocations in these cells but no rearrangements in BCL2 or BCL6. Microdissection of neoplastic cells in these patients followed by targeted next-generation sequencing identified a mutation in MYC, D2N, and an indel in TNFRSF14. Mutations in ID3 or TCF3 were not identified. Although rare, these lesions should be separated from HGBCLs involving follicles but with systemic spread which has been previously described. Unlike systemic lymphomas with MYC gene rearrangements, these in situ B-cell neoplasms with MYC rearrangements did not require systemic therapy and no progression has been seen in either patient beyond 1 year (29 and 16mo). Our work offers pathologic and biologic insight into the early process of B-cell neoplasia.

    View details for DOI 10.1097/PAS.0000000000001338

    View details for PubMedID 31368914

  • Coexistence of BRAF V600E and TERT Promoter Mutations in Low-grade Serous Carcinoma of Ovary Recurring as Carcinosarcoma in a Lymph Node: Report of a Case INTERNATIONAL JOURNAL OF GYNECOLOGICAL PATHOLOGY Tavallaee, M., Steiner, D. F., Zehnder, J. L., Folkins, A. K., Karam, A. K. 2019; 38 (4): 386–92

    View details for DOI 10.1097/PGP.0000000000000507

    View details for Web of Science ID 000471670300012

  • Complete and Prolonged Response to Immune Checkpoint Blockade in POLE-Mutated Colorectal Cancer JCO PRECISION ONCOLOGY Silberman, R., Steiner, D. F., Lo, A. A., Gomez, A., Zehnder, J. L., Chu, G., Suarez, C. J. 2019; 3: 1–5

    View details for DOI 10.1200/PO.18.00214

    View details for Web of Science ID 000473546800001

  • Revisiting diagnostic criteria for myelodysplastic/myeloproliferative neoplasms with ring sideroblasts and thrombocytosis: Borderline cases without anemia exist INTERNATIONAL JOURNAL OF LABORATORY HEMATOLOGY Li, P., Shahmarvand, N., Lynch, D., Gotlib, J. R., Merker, J. D., Zehnder, J. L., George, T., Ohgami, R. S. 2019; 41 (3): 345–52

    View details for DOI 10.1111/ijlh.12981

    View details for Web of Science ID 000468370800013

  • Update on diagnostic testing for platelet function disorders: What is practical and useful? International journal of laboratory hematology Hayward, C. P., Moffat, K. A., Brunet, J., Carlino, S. A., Plumhoff, E., Meijer, P., Zehnder, J. L. 2019; 41 Suppl 1: 26–32

    Abstract

    INTRODUCTION: Platelet function disorders (PFD) are an important group of bleeding disorders that require validated and practical laboratory strategies for diagnosis.METHODS: This review summarizes the authors' experiences, current literature, and an international survey to evaluate the practices of diagnostic laboratories that offer tests for PFD.RESULTS: Blood counts, blood film review, and aggregation tests are the most commonly performed investigations for PFD and help determine whether there is thrombocytopenia and/or defective platelet function due to a variety of causes. The performance characteristics of tests for PFD, and the level of evidence that these tests detect bleeding problems, are important issues to determine where tests are useful for diagnostic or correlative purposes, or research only uses. Platelet aggregation assays, and quantitative analysis of platelet dense granule numbers, are tests with good performance characteristics that detect abnormalities associated with increased bleeding in a significant proportion of individuals referred for PFD investigations. Lumiaggregometry estimates of platelet adenosine triphosphate release show greater variability which limits the diagnostic usefulness. Diagnostic laboratories report that fiscal and other constraints, including a lack of high-quality evidence, limit their ability to offer an expanded test menu for PFD.CONCLUSION: PFD are clinically important bleeding disorders that remain challenging for diagnostic laboratories to investigate. While some PFD tests are well validated for diagnostic purposes, gaps in scientific evidence and resource limitations influence diagnostic laboratory decisions on which PFD tests to offer.

    View details for PubMedID 31069975

  • Structural. Variation Detection by Proximity Ligation from Formalin-Fixed, Paraffin-Embedded Tumor Tissue JOURNAL OF MOLECULAR DIAGNOSTICS Troll, C. J., Putnam, N. H., Hartley, P. D., Rice, B., Blanchette, M., Siddiqui, S., Ganbat, J., Powers, M. P., Ramakrishnan, R., Kunder, C. A., Bustamante, C. D., Zehnder, J. L., Green, R. E., Costa, H. A. 2019; 21 (3): 375–83

    View details for DOI 10.1016/j.jmoldx.2018.11.003

    View details for Web of Science ID 000467350300002

  • Update on diagnostic testing for platelet function disorders: What is practical and useful? INTERNATIONAL JOURNAL OF LABORATORY HEMATOLOGY Hayward, C. M., Moffat, K. A., Brunet, J., Carlino, S. A., Plumhoff, E., Meijer, P., Zehnder, J. L. 2019; 41: 26–32

    View details for DOI 10.1111/ijlh.12995

    View details for Web of Science ID 000468349600006

  • Indolent in situ High-Grade B-cell lymphoma with a MYC Translocation and Mutations Identified by Next Generation Sequencing Kumar, J., Wu, S., Butzmann, A., Lau, J., Zehnder, J., Warnke, R., Nybakken, G., Van de Rijn, M., Ohgami, R. NATURE PUBLISHING GROUP. 2019

    View details for Web of Science ID 000478081102331

  • Targeted deep sequencing of cell-free DNA from body cavity fluids with malignant, suspicious, and benign cytology Yang, S., Libiran, P., Jones, C., Joshi, R., Lau, H., Stehr, H., Longacre, T., Allison, K., Zehnder, J., Long, S., Berry, G., Kunder, C. NATURE PUBLISHING GROUP. 2019

    View details for Web of Science ID 000478081100461

  • Molecular Profiling of Intraductal Tubulopapillary Neoplasm Saleem, A., Stehr, H., Zehnder, J., Kunder, C., Lin, C. NATURE PUBLISHING GROUP. 2019

    View details for Web of Science ID 000478081103213

  • Next-Generation Sequencing for HER2 Status in Breast Cancer: Comparison with Immunohistochemistry and in situ Hybridization by 2018 Focused Update Mooney, K., Stehr, H., Kunder, C., Zehnder, J., Allison, K., Lin, C. NATURE PUBLISHING GROUP. 2019

    View details for Web of Science ID 000478915500217

  • Targeted deep sequencing of cell-free DNA from body cavity fluids with malignant, suspicious, and benign cytology Yang, S., Libiran, P., Jones, C., Joshi, R., Lau, H., Stehr, H., Longacre, T., Allison, K., Zehnder, J., Long, S., Berry, G., Kunder, C. NATURE PUBLISHING GROUP. 2019

    View details for Web of Science ID 000478915500443

  • Molecular Profiling of Intraductal Tubulopapillary Neoplasm Saleem, A., Stehr, H., Zehnder, J., Kunder, C., Lin, C. NATURE PUBLISHING GROUP. 2019

    View details for Web of Science ID 000478915501405

  • Next-Generation Sequencing for HER2 Status in Breast Cancer: Comparison with Immunohistochemistry and in situ Hybridization by 2018 Focused Update Mooney, K., Stehr, H., Kunder, C., Zehnder, J., Allison, K., Lin, C. NATURE PUBLISHING GROUP. 2019

    View details for Web of Science ID 000478081100217

  • Indolent in situ High-Grade B-cell lymphoma with a MYC Translocation and Mutations Identified by Next Generation Sequencing Kumar, J., Wu, S., Butzmann, A., Lau, J., Zehnder, J., Warnke, R., Nybakken, G., Van De Rijn, M., Ohgami, R. NATURE PUBLISHING GROUP. 2019

    View details for Web of Science ID 000478915501090

  • Revisiting diagnostic criteria for myelodysplastic/myeloproliferative neoplasms with ring sideroblasts and thrombocytosis: Borderline cases without anemia exist. International journal of laboratory hematology Li, P., Shahmarvand, N., Lynch, D., Gotlib, J. R., Merker, J. D., Zehnder, J. L., George, T. I., Ohgami, R. S. 2019

    Abstract

    INTRODUCTION: Myelodysplastic/myeloproliferative neoplasm with ring sideroblasts and thrombocytosis (MDS/MPN-RS-T) is a rare disease in the 2016 revised World Health Organization (WHO) classification. Diagnostic criteria include the following: persistent thrombocytosis (>450*109 /L) with clustering of atypical megakaryocytes, refractory anemia, dyserythropoiesis with ring sideroblasts, and the presence of the spliceosome factor 3b subunit (SF3B1) mutation. It is unclear if anemia should be a required criterion for this diagnosis as cases which show all other features of MDS/MPN-RS-T but without anemia exist.METHODS: We searched for borderline cases of MDS/MPN-RS-T in which refractory anemia was absent at diagnosis in two major academic institutes.RESULTS: Three cases without anemia were identified. These cases all showed other classic morphologic and clinical features of MDS/MPN-RS-T, including thrombocytosis, atypical megakaryocytes with clustering, and characteristic SF3B1 and JAK2 V617F mutations.CONCLUSION: Given these findings, the requirement of refractory anemia as a diagnostic criterion for MDS/MPN-RS-T should be re-evaluated. Removal of refractory anemia as a diagnostic criterion would incorporate current borderline cases and extend the spectrum of this disorder.

    View details for PubMedID 30811101

  • Molecular profiling of clear cell adenocarcinoma of the urinary tract. Virchows Archiv : an international journal of pathology Lin, C. Y., Saleem, A., Stehr, H., Zehnder, J. L., Pinsky, B. A., Kunder, C. A. 2019

    Abstract

    Clear cell adenocarcinoma (CCA) of the urinary tract is a rare type of malignancy whose molecular profiles remain undefined. Here we reported an integrated clinicopathologic and molecular profiling analysis of four cases of clear cell adenocarcinoma arising in the urethra or the bladder. Utilizing a clinically validated 130-gene exon-sequencing assay, we identified recurrent pathogenic PIK3CA (p. E545K) and KRAS (p.G12D) variants in three of four (75%) of the cases. In addition, an APC variant (P.S2310X), a TP53 variant (p.R273C), and a MYC amplification event were identified. The only CCA case without either PIK3CA or KRAS variants has a distinct pathogenesis through BK virus, demonstrated by positive BK virus PCR and SV40 immunohistochemistry. The novel finding of recurrent variants in the PI3K/AKT/mTOR pathway provides not only insights into oncogenesis but also potential clinical therapeutic targets for patients with clear cell adenocarcinoma of the urinary tract.

    View details for DOI 10.1007/s00428-019-02634-5

    View details for PubMedID 31372739

  • Implementation of Whole-Blood Impedance Aggregometry for Heparin-Induced Thrombocytopenia Functional Assay and Case Discussion. American journal of clinical pathology Jin, J., Baker, S. A., Hall, E. T., Gombar, S., Bao, A., Zehnder, J. L. 2019; 152 (1): 50–58

    Abstract

    The diagnosis of heparin-induced thrombocytopenia (HIT) ideally requires a functional assay to confirm. 14C-serotonin release assay (SRA) as "gold standard" is technically challenging and unsuitable for routine use. We conducted a study to assess the performance of whole-blood impedance aggregometry (WBIA) as a simple and rapid HIT functional assay.Platelet factor 4 (PF4)/immunoglobulin G (IgG) antibody, WBIA, and SRA were tested on 70 patients suspected of having HIT. Patients with a 4Ts score of 4 or more, positive PF4/IgG, and positive SRA were considered HIT positive; others were designated HIT negative.WBIA had 85.7% (6/7) sensitivity and 98.4% (61/62) specificity, which were not statistically different compared with SRA. Sixty-two of 70 patients had concordant results (five positive and 57 negative) by both WBIA and SRA. Eight discordant cases revealed the importance of recognizing donor effect, interferences, and the presence of heparin-independent or non-heparin-dependent antibodies in functional assays.Implementation of WBIA could facilitate timely diagnosis and management of HIT.

    View details for DOI 10.1093/ajcp/aqz013

    View details for PubMedID 31165165

  • Morphologic, Immunophenotypic and Molecular Features of Hypermutation in Colorectal Carcinomas with Mutations in DNA Polymerase Ɛ (POLE). Histopathology Forgó, E., Gomez, A. J., Steiner, D., Zehnder, J., Longacre, T. A. 2019

    Abstract

    Colorectal carcinomas (CRC) with mismatch repair (MMR) deficiency have increased tumor mutation burden and respond to immune checkpoint inhibitor therapy. The Cancer Genome Atlas identified hypermutated CRCs with somatic mutations in DNA polymerase ε (POLE) with mutation burdens exceeding that of MMR-deficient CRCs.To identify the morphologic, immunophenotypic and molecular features of POLE mutated CRCs, 63 consecutive MMR-intact CRCs were evaluated by Sanger sequencing for POLE exonuclease domain mutations in exons 9, 11, 13 and 14 and confirmed by next generation sequencing. Tumor immune microenvironment and immunoscores were assessed in POLE-mutated CRCs using immunohistochemistry to detect CD3+/CD8+ tumor-infiltrating lymphocytes and compared to 59 non-POLE mutated MMR-intact CRC, 10 non-POLE mutated MMR-deficient CRCs, and 223 normal colonic mucosa.4.8% CRC (4 MMR-intact primary and 1 MMR-intact metastasis) harbored POLE mutations in amino acid 286 in exon 9 (p.P286R) or exon 13 (p.V411L). POLE mutated CRCs arose in the transverse colon and rectum, were male-predominant, younger, and showed increased tumor-infiltrating lymphocytes and immune cells at the tumor-stromal interface. The patient with metastatic POLE mutated CRC was placed on PD-1 inhibitor treatment with marked and sustained response. These data indicate that POLE mutated CRCs have hypermutated phenotypes despite MMR-intact status, with mutation burdens higher than that in microsatellite unstable CRCs. Given the recent approval for treatment of microsatellite unstable cancer with immune checkpoint inhibitors, assessment of POLE status may help guide therapeutic decisions for hypermutated tumors with intact MMR that would otherwise be missed by routine testing.

    View details for DOI 10.1111/his.13984

    View details for PubMedID 31479159

  • 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

  • Structural Variation Detection by Proximity Ligation from Formalin-Fixed, Paraffin-Embedded Tumor Tissue. The Journal of molecular diagnostics : JMD Troll, C. J., Putnam, N. H., Hartley, P. D., Rice, B., Blanchette, M., Siddiqui, S., Ganbat, J., Powers, M. P., Ramakrishnan, R., Kunder, C. A., Bustamante, C. D., Zehnder, J. L., Green, R. E., Costa, H. A. 2018

    Abstract

    The clinical management and therapy of many solid tumor malignancies is dependent on detection of medically actionable or diagnostically relevant genetic variation. However, a principal challenge for genetic assays from tumors is the fragmented and chemically damaged state of DNA in formalin-fixed, paraffin-embedded (FFPE) samples. From highly fragmented DNA and RNA there is no current technology for generating long-range DNA sequence data as is required to detect genomic structural variation or long-range genotype phasing. We have developed a high-throughput chromosome conformation capture approach for FFPE samples that we call "Fix-C", which is similar in concept to Hi-C. Fix-C enables structural variation detection from archival FFPE samples. This method was applied to 15 clinical adenocarcinoma and sarcoma positive control specimens spanning a broad range of tumor purities. In this panel, Fix-C analysis achieves a 90% concordance rate with FISH assays - the current clinical gold standard. Additionally, novel structural variation undetected by other methods could be identified and long-range chromatin configuration information recovered from these FFPE samples harboring highly degraded DNA. This powerful approach will enable detailed resolution of global genome rearrangement events during cancer progression from FFPE material and inform the development of targeted molecular diagnostic assays for patient care.

    View details for PubMedID 30605765

  • Clinical Implementation of Mutational Signature Analysis Lawrence, L., Kunder, C., Stehr, H., Zehnder, J. ELSEVIER SCIENCE INC. 2018: 975

    View details for Web of Science ID 000448637200267

  • Tumor Molecular Profiling Aids in Determining Tissue of Origin and Therapy for Metastatic Adenocarcinoma in a Patient With Multiple Primary Malignancies JCO PRECISION ONCOLOGY Costa, H. A., Reyes, R., Mills, M., Zehnder, J. L., Sledge, G., Curtis, C., Ford, J. M., Suarez, C. J. 2018; 2

    View details for DOI 10.1200/PO.18.00177

    View details for Web of Science ID 000462187500001

  • GENOMIC LANDSCAPE OF NON-DRIVER MUTATIONS IN MYELOPROLIFERATIVE NEOPLASMS: THE STANFORD EXPERIENCE Robert, H., Steiner, D., Zehnder, J. WILEY. 2018: 122

    View details for Web of Science ID 000444677800226

  • PLATELET TRANSCRIPTOMIC SIGNATURES IN MYELOPROLIFERATIVE NEOPLASMS Krishnan, A., Perkins, C., Gotlib, J., Zehnder, J. WILEY. 2018: E34

    View details for Web of Science ID 000446055700086

  • CASE REPORT: MONOCLONAL IGM & LAMBDA; COAGULATION INHIBITOR WITH PHOSPHATIDYLSERINE SPECIFICITY INTERFERING WITH PLASMA, BUT NOT WHOLE-BLOOD BASED COAGULATION TESTING Jin, J., Baker, S. A., Shokry, M., Zehnder, J. L. WILEY. 2018: E30

    View details for Web of Science ID 000446055700078

  • Single-cell analysis of mutational heterogeneity in acute myeloid leukemia tumors with high-throughput droplet microfluidics Eastburn, D. J., Pellegrino, M., Sciambi, A., Treusch, S., Xu, L., Durruthy-Durruthy, R., Gokhale, K., Jacob, J., Chen, T. X., Oldham, W., Matthews, J., Kantarjian, H., Futreal, A., Patel, K., Jones, K. W., Zehnder, J. L., Takahashi, K. AMER ASSOC CANCER RESEARCH. 2018

    View details for DOI 10.1158/1538-7445.AM2018-5348

    View details for Web of Science ID 000468819504334

  • MLL leukemia induction by t(9;11) chromosomal translocation in human hematopoietic stem cells using genome editing BLOOD ADVANCES Schneidawind, C., Jeong, J., Schneidawind, D., Kim, I., Duque-Afonso, J., Wong, S., Iwasaki, M., Breese, E. H., Zehnder, J. L., Porteus, M., Cleary, M. L. 2018; 2 (8): 832–45

    Abstract

    Genome editing provides a potential approach to model de novo leukemogenesis in primary human hematopoietic stem and progenitor cells (HSPCs) through induction of chromosomal translocations by targeted DNA double-strand breaks. However, very low efficiency of translocations and lack of markers for translocated cells serve as barriers to their characterization and model development. Here, we used transcription activator-like effector nucleases to generate t(9;11) chromosomal translocations encoding MLL-AF9 and reciprocal AF9-MLL fusion products in CD34+ human cord blood cells. Selected cytokine combinations enabled monoclonal outgrowth and immortalization of initially rare translocated cells, which were distinguished by elevated MLL target gene expression, high surface CD9 expression, and increased colony-forming ability. Subsequent transplantation into immune-compromised mice induced myeloid leukemias within 48 weeks, whose pathologic and molecular features extensively overlap with de novo patient MLL-rearranged leukemias. No secondary pathogenic mutations were revealed by targeted exome sequencing and whole genome RNA-sequencing analyses, suggesting the genetic sufficiency of t(9;11) translocation for leukemia development from human HSPCs. Thus, genome editing enables modeling of human acute MLL-rearranged leukemia in vivo, reflecting the genetic simplicity of this disease, and provides an experimental platform for biological and disease-modeling applications.

    View details for PubMedID 29650777

    View details for PubMedCentralID PMC5916000

  • Coexistence of BRAF V600E and TERT Promoter Mutations in Low-grade Serous Carcinoma of Ovary Recurring as Carcinosarcoma in a Lymph Node: Report of a Case. International journal of gynecological pathology : official journal of the International Society of Gynecological Pathologists Tavallaee, M., Steiner, D. F., Zehnder, J. L., Folkins, A. K., Karam, A. K. 2018

    Abstract

    Low-grade serous carcinomas only rarely coexist with or progress to high-grade tumors. We present a case of low-grade serous carcinoma with transformation to carcinosarcoma on recurrence in the lymph node. Identical BRAF V600E and telomerase reverse transcriptase promoter mutations were identified in both the original and recurrent tumor. Given that telomerase reverse transcriptase promotor mutations are thought to play a role in progression of other tumor types, the function of telomerase reverse transcriptase mutations in BRAF mutated low-grade serous carcinoma deserves investigation.

    View details for PubMedID 29620581

  • Next-generation sequencing of idiopathic multicentric and unicentric Castleman disease and follicular dendritic cell sarcomas BLOOD ADVANCES Nagy, A., Bhaduri, A., Shahmarvand, N., Shahryari, J., Zehnder, J. L., Warnke, R. A., Mughal, T., Ali, S., Ohgami, R. S. 2018; 2 (5): 481–91

    Abstract

    Castleman disease (CD) is a rare lymphoproliferative disorder subclassified as unicentric CD (UCD) or multicentric CD (MCD) based on clinical features and the distribution of enlarged lymph nodes with characteristic histopathology. MCD can be further subtyped based on human herpes virus 8 (HHV8) infection into HHV8-associated MCD, HHV8-/idiopathic MCD (iMCD), and polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy, and skin change (POEMS)-associated MCD. In a subset of cases of UCD, an associated follicular dendritic cell sarcoma (FDCS) may be seen. Although numerous reports of the clinical and histologic features of UCD, MCD, and FDCS exist, an understanding of the genetic and epigenetic landscape of these rare diseases is lacking. Given this paucity of knowledge, we analyzed 15 cases of UCD and 3 cases of iMCD by targeted next-generation sequencing (NGS; 405 genes) and 3 cases of FDCS associated with UCD hyaline vascular variant (UCD-HVV) by whole-exome sequencing. Common amplifications of ETS1, PTPN6, and TGFBR2 were seen in 1 iMCD and 1 UCD case; the iMCD case also had a somatic DNMT3A L295Q mutation. This iMCD patient also showed clinicopathologic features consistent with a specific subtype known as Castleman-Kojima disease (thrombocytopenia, anasarca, fever, reticulin fibrosis, and organomegaly [TAFRO] clinical subtype). Additionally, 1 case of UCD-HVV showed amplification of the cluster of histone genes on chromosome 6p. FDCS associated with UCD-HVV showed mutations and copy number changes in known oncogenes, tumor suppressors, and chromatin structural-remodeling proteins.

    View details for PubMedID 29496669

  • Detection of Large Chromosomal Abnormalities in Tumors through Analysis of Off-Target Next-Generation Sequencing (NGS) Read Data Chen, S. B., Stehr, H., Zehnder, J. L., Dudley, J. C. NATURE PUBLISHING GROUP. 2018: 804

    View details for Web of Science ID 000429308605111

  • A whole exome sequencing analysis of diffuse large B-cell lymphoma arising from topologically adjacent follicular lymphoma reveals an intimate and distant relationship Lin, C., Nagy, A., Zehnder, J. L., O'Malley, D. P., Ohgami, R. NATURE PUBLISHING GROUP. 2018: 533

    View details for Web of Science ID 000459341002362

  • The Significance of Dim Cytoplasmic CD3 Expression in Acute Myeloid Leukemia: A Long-Term Retrospective Study Identifies an Association with Acute Promyelocytic Leukemia with FLT3-ITD Mutations Kumar, J., Nagy, A., Lacayo, N., Gotlib, J., Zehnder, J. L., Ohgami, R. NATURE PUBLISHING GROUP. 2018: 529

    View details for Web of Science ID 000459341002354

  • Molecular Characterization of Aggressive Estrogen Receptor Positive Breast Cancer Resistant to Palbociclib Therapy Lei, L., Lin, C., Steiner, D., Ford, J. M., Zehnder, J. L., Suarez, C. J. NATURE PUBLISHING GROUP. 2018: 703

    View details for Web of Science ID 000459341003278

  • Detection of Large Chromosomal Abnormalities in Tumors through Analysis of Off-Target Next-Generation Sequencing (NGS) Read Data Chen, S. B., Stehr, H., Zehnder, J. L., Dudley, J. C. NATURE PUBLISHING GROUP. 2018: 804

    View details for Web of Science ID 000459341004013

  • Validation and Application of Predicted Tumor Mutational Burden (pTMB) Using a 130-Gene Sequencing Panel in Lung Adenocarcinoma Yang, S., Steiner, D., Stehr, H., Berry, G., Zehnder, J. L., Kunder, C. NATURE PUBLISHING GROUP. 2018: 758–59

    View details for Web of Science ID 000459341003440

  • The Significance of Dim Cytoplasmic CD3 Expression in Acute Myeloid Leukemia: A Long-Term Retrospective Study Identifies an Association with Acute Promyelocytic Leukemia with FLT3-ITD Mutations Kumar, J., Nagy, A., Lacayo, N., Gotlib, J., Zehnder, J. L., Ohgami, R. NATURE PUBLISHING GROUP. 2018: 529

    View details for Web of Science ID 000429308603354

  • Molecular Characterization of Aggressive Estrogen Receptor Positive Breast Cancer Resistant to Palbociclib Therapy Lei, L., Lin, C., Steiner, D., Ford, J. M., Zehnder, J. L., Suarez, C. J. NATURE PUBLISHING GROUP. 2018: 703

    View details for Web of Science ID 000429308604325

  • Validation and Application of Predicted Tumor Mutational Burden (pTMB) Using a 130-Gene Sequencing Panel in Lung Adenocarcinoma Yang, S., Steiner, D., Stehr, H., Berry, G., Zehnder, J. L., Kunder, C. NATURE PUBLISHING GROUP. 2018: 758–59

    View details for Web of Science ID 000429308604486

  • A whole exome sequencing analysis of diffuse large B-cell lymphoma arising from topologically adjacent follicular lymphoma reveals an intimate and distant relationship Lin, C., Nagy, A., Zehnder, J. L., O'Malley, D. P., Ohgami, R. NATURE PUBLISHING GROUP. 2018: 533

    View details for Web of Science ID 000429308603362

  • Molecular and Clinicopathologic Features Associated with PD-L1 Expression in Lung Adenocarcinoma Yang, S., Steiner, D., Berry, G. J., Neal, J. W., Zehnder, J. L., Kunder, C. A. ELSEVIER SCIENCE INC. 2017: 1009–10

    View details for Web of Science ID 000414275900290

  • Clinical Validation and Implementation of a Targeted Sequencing Panel for Myeloid Neoplasms Steiner, D., Devereaux, K., Dudley, J., Jones, C., Gojenola, L., Zehnder, J., Kunder, C. ELSEVIER SCIENCE INC. 2017: 958

    View details for Web of Science ID 000414275900079

  • Detection of Clonal TRG and TRB Gene Rearrangements Using Next Generation Sequencing Shah, P., Huang, Y., Huang, Y., Hutt, K., Panganiban, J., Bob, R., Ewalt, M. D., Zehnder, J. L., Stenzel, T., Miller, J. E. ELSEVIER SCIENCE INC. 2017: 1055

    View details for Web of Science ID 000414275900475

  • in silico Long-Read Sequencing from FFPE Solid Tumor Tissue for Structural Variation Detection and Phasing in Archival Specimens Costa, H. A., Blanchette, M., Bustamante, C. D., Green, R. E., Hadley, P. D., Kunder, C., Putnam, N., Rice, B., Trolf, C., Zehnder, J. L. ELSEVIER SCIENCE INC. 2017: 1034–35

    View details for Web of Science ID 000414275900390

  • A novel TRIP11-FLT3 fusion in a patient with a myeloid/lymphoid neoplasm with eosinophilia CANCER GENETICS Chung, A., Hou, Y., Ohgami, R. S., Von Gehr, A., Fisk, D. G., Roskin, K. M., Li, X., Gojenola, L., Bangs, C. D., Arber, D. A., Fire, A. Z., Cherry, A. M., Zehnder, J. L., Gotlib, J., Merker, J. D. 2017; 216: 10–15

    Abstract

    FLT3 fusions are associated with myeloid and lymphoid neoplasms with eosinophilia. We describe a patient presenting with clinicopathologic features of both chronic eosinophilic leukemia, not otherwise specified (CEL, NOS) and systemic mastocytosis (SM). The bone marrow demonstrated a myeloproliferative neoplasm with eosinophilia and aggregates of atypical mast cells. Cytogenetic analysis revealed a t(13;14)(q12;q32), which was subsequently molecularly characterized as a novel TRIP11-FLT3 rearrangement. A KIT D816V mutation was also identified. The patient rapidly transformed to T-lymphoblastic leukemia/lymphoma and expired shortly after diagnosis. This is the fifth FLT3 fusion gene described in the literature; the presence of both myeloid and lymphoid neoplasms implicates involvement of an early hematopoietic progenitor by rearranged FLT3. We suggest that leukemias and lymphomas with FLT3 fusion genes exhibit similar clinicopathologic features to, and should be included in, the WHO category of "Myeloid and lymphoid neoplasms with eosinophilia and abnormalities of PDGFRA, PDGFRB, or FGFR1, or with PCM1-JAK2."

    View details for PubMedID 29025582

  • IDH2 Mutation in a Patient with Metastatic Colon Cancer NEW ENGLAND JOURNAL OF MEDICINE Zhang, B. M., Zehnder, J. L., Suarez, C. J. 2017; 376 (20): 1991-1992

    View details for DOI 10.1056/NEJMc1701072

    View details for PubMedID 28514606

  • IMPLEMENTATION OF WHOLEBLOOD IMPEDANCE AGGREGOMETRYFOR HEPARIN-INDUCED THROMBOCYTOPENIAFUNCTIONAL ASSAY Jin, J., Hartinger, S., Wei, S., Zehnder, J. WILEY. 2017: 50

    View details for Web of Science ID 000402838000083

  • IMPROVING CLINICAL DECISION MAKING FOR HEPARINIZED PATIENTS WITH DISCORDANT ANTI-FACTOR XA ACTIVITY AND ACTIVATED PARTIAL THROMBOPLASTIN TIME. Gombar, S., Krishnan, A., Zehnder, J. WILEY. 2017: 40

    View details for Web of Science ID 000402838000064

  • NOVEL MARKERS FOR THE DIAGNOSIS OF ANTIPHOSPHOLIPID SYNDROME: ANTIBODIES AGAINST PHOSPHATIDYLSERINE/PROTHROMBIN COMPLEX Jin, J., Hartinger, S., Wei, S., Zehnder, J. WILEY. 2017: 50

    View details for Web of Science ID 000402838000082

  • Identification of a Novel Somatic Mutation Leading to Allele Dropout for EGFR L858R Genotyping in Non-Small Cell Lung Cancer. Molecular diagnosis & therapy Costa, H. A., Neal, J. W., Bustamante, C. D., Zehnder, J. L. 2017

    Abstract

    While PCR-based genotyping methods abound in molecular testing for lung cancer therapy, these approaches may not provide the robust sensitivity to detect accurate genotypes in a variable cancer genomic background.Here, we describe a study of a clinical tumor specimen containing a novel somatic single nucleotide variant that caused allele drop-out in EGFR L858R genotyping, resulting in a false-negative interpretation and impacting patient clinical management.We demonstrate that a subsequent unbiased next-generation sequencing approach correctly identified the driver mutation, and therefore may be more reliable for somatic variant detection.These findings magnify the potential pitfalls of PCR amplification-based approaches and stress the importance of unbiased and sensitive molecular testing strategies for therapeutic marker detection as molecular testing becomes the standard for determining clinical management of cancer patients.

    View details for DOI 10.1007/s40291-017-0275-y

    View details for PubMedID 28357677

  • A Retrospective Study of 305 Cases of Angioimmunoblastic T-Cell Lymphoma with Emphasis on Rare Lymphoplasmacytic and Plasma Cell Proliferations Oak, J., Hoffmann, J. C., Chisholm, K. M., Chen, J., Zehnder, J., Arber, D. A., Natkunam, Y., Warnke, R., Ohgami, R. NATURE PUBLISHING GROUP. 2017: 367A–368A

    View details for Web of Science ID 000393724401748

  • Detection of Mutations in DNA Polymerase epsilon (POLE) in Colorectal Carcinoma with Intact Mismatch Repair Proteins: Immunotherapeutic Implications of Ultramutation Gomez, A. J., Burton, A., Steiner, D., Zehnder, J., Longacre, T. NATURE PUBLISHING GROUP. 2017: 173A

    View details for Web of Science ID 000394467300687

  • Detection of Mutations in DNA Polymerase epsilon (POLE) in Colorectal Carcinoma with Intact Mismatch Repair Proteins: Immunotherapeutic Implications of Ultramutation Gomez, A. J., Burton, A., Steiner, D., Zehnder, J., Longacre, T. NATURE PUBLISHING GROUP. 2017: 173A

    View details for Web of Science ID 000393724400686

  • A Study of Disseminated Intravascular Coagulation in Acute Leukemia Reveals Markedly Elevated D-Dimer Levels Are a Sensitive Indicator of Acute Promyelocytic Leukemia Oak, J., Shahmarvand, N., Alcasid, M., Cascio, M., Goodman, E., Medeiros, B., Arber, D. A., Zehnder, J., Ohgami, R. NATURE PUBLISHING GROUP. 2017: 367A

    View details for Web of Science ID 000393724401747

  • A Study of Disseminated Intravascular Coagulation in Acute Leukemia Reveals Markedly Elevated D-Dimer Levels Are a Sensitive Indicator of Acute Promyelocytic Leukemia Oak, J., Shahmarvand, N., Alcasid, M., Cascio, M., Goodman, E., Medeiros, B., Arben, D. A., Zehnder, J., Ohgami, R. NATURE PUBLISHING GROUP. 2017: 367A

    View details for Web of Science ID 000394467302050

  • A Retrospective Study of 305 Cases of Angioimmunoblastic T-Cell Lymphoma with Emphasis on Rare Lymphoplasmacytic and Plasma Cell Proliferations Oak, J., Hoffinann, J. C., Chisholm, K. M., Chen, J., Zehnder, J., Arber, D. A., Natkunam, Y., Warnke, R., Ohgami, R. NATURE PUBLISHING GROUP. 2017: 367A–368A

    View details for Web of Science ID 000394467302051

  • The Significance of CD56 Expression and the RAM Immunophenotype, a Recurrent Immunophenotype Seen in Children, in Adult Acute Myeloid Leukemia Lazzareschi, D., Cherry, A., Zehnder, J., Gotlib, J., Arber, D. A., Lacayo, N., Ohgami, R. NATURE PUBLISHING GROUP. 2017: 358A–359A

    View details for Web of Science ID 000394467302016

  • The Significance of CD56 Expression and the RAM Immunophenotype, a Recurrent Immunophenotype Seen in Children, in Adult Acute Myeloid Leukemia Lazzareschi, D., Cherry, A., Zehnder, J., Gotlib, J., Arber, D. A., Lacayo, N., Ohgami, R. NATURE PUBLISHING GROUP. 2017: 358A–359A

    View details for Web of Science ID 000393724401713

  • Methodologic Considerations in the Application of Next-Generation Sequencing of Human TRB Repertoires for Clinical Use JOURNAL OF MOLECULAR DIAGNOSTICS Xu, L., You, X., Zheng, P., Zhang, B. M., Gupta, P. K., Lavori, P., Meyer, E., Zehnder, J. L. 2017; 19 (1): 72-83

    Abstract

    Next-generation sequencing (NGS) of immune receptors has become a standard tool to assess minimal residual disease (MRD) in patients treated for lymphoid malignancy, and it is being used to study the T-cell repertoire in many clinical settings. To better understanding the potential clinical utility and limitations of this application outside of MRD, we developed a BIOMED-2 primer-based NGS method and characterized its performance in controls and patients with graft-versus-host disease (GVHD) after allogeneic hematopoietic transplant. For controls and patients with GVHD, replicate sequencing of the same T-cell receptor β (TRB) libraries was highly reproducible. Higher variability was observed in sequencing of different TRB libraries made from the same DNA stock. Variability was increased in patients with GVHD compared with controls; patients with GVHD also had lower diversity than controls. In the T-cell repertoire of a healthy person, approximately 99.6% of the CDR3 clones were in low abundance, with frequency <10(-3). A single library could identify >93% of the clones with frequency ≥10(-3) in the repertoire. Sequencing in duplicate increased the average detection rate to >97%. This work demonstrates that NGS reliably and robustly characterizes TRB populations in healthy individuals and patients with GVHD with frequency ≥10(-3) and provides a methodologic framework for applying NGS immune repertoire methods to clinical testing applications beyond MRD.

    View details for DOI 10.1016/j.jmoldx.2016.07.009

    View details for Web of Science ID 000390983100008

  • A study of disseminated intravascular coagulation in acute leukemia reveals markedly elevated D-dimer levels are a sensitive indicator of acute promyelocytic leukemia. International journal of laboratory hematology Shahmarvand, N., Oak, J. S., Cascio, M. J., Alcasid, M., Goodman, E., Medeiros, B. C., Arber, D. A., Zehnder, J. L., Ohgami, R. S. 2017

    Abstract

    While the presence of disseminated intravascular coagulation (DIC) has been implicated in worse clinical outcome in acute leukemia, the relationship between different subtypes of acute leukemia and the clinicopathologic features of DIC has not been systematically well studied.In this study, we retrospectively reviewed 149 cases of newly diagnosed acute leukemia and assessed the utility of evaluating red blood cell morphologic features, and coagulation parameters in determining the presence of DIC as well as differentiating subtypes of acute leukemia.Review of our cohort demonstrates a novel finding, that elevated D-dimer concentrations ≥19 000 ng/mL fibrinogen equivalent units (FEU) are a sensitive diagnostic indicator of acute promyelocytic leukemia (APL) with moderate specificity, sensitivity 96%, specificity 92% in acute leukemia subtyping. Similar to other studies, APL showed an increased incidence of DIC (P < 0.01) compared to other subtypes of acute leukemia. Surprisingly, the presence of schistocytes on the peripheral blood smear was not a statistically significant indicator of DIC, sensitivity of 36% and specificity of 89%. Finally, the presence of DIC was not a significant indicator of poorer prognosis amongst all patients with AML.Overall we identify elevated D-dimer concentrations ≥19 000 ng/mL FEU are a sensitive indicator of acute promyelocytic leukemia (APL), with a sensitivity of 96% and specificity of 92% in the subtyping of acute leukemias, and that the presence of schistocytes in peripheral blood smears is not a diagnostically sensitive screening test for DIC with a sensitivity of 36%.

    View details for PubMedID 28422420

  • Comprehensive Genomic Profiling of Malignant Effusions in Patients with Metastatic Lung Adenocarcinoma. The Journal of molecular diagnostics : JMD Yang, S. R., Lin, C. Y., Stehr, H., Long, S. R., Kong, C. S., Berry, G. J., Zehnder, J. L., Kunder, C. A. 2017

    Abstract

    Cytology samples are being increasingly utilized for comprehensive molecular testing. Although fine-needle aspirates are adequate substrates for high-throughput sequencing, the suitability of malignant body fluids remains largely unexplored. Herein, we investigated the adequacy and utility of performing targeted next-generation sequencing (NGS) on malignant effusions from patients with metastatic lung adenocarcinoma. Thirty-two effusion samples that were submitted for hybrid capture-based NGS using a clinically validated solid tumor genotyping panel were examined. All cases showed ≥5% tumor cellularity; however, 28 (88%) provided sufficient DNA for NGS (≥1 ng/μL). The sequencing reads showed satisfactory quality control statistics, and the variant allele frequencies were correlated with tumor cellularity. Furthermore, pathogenic or likely pathogenic genomic alterations were identified in 26/28 samples (93%), whereas clinically actionable alterations were present in 18 (64%). Notably, nine patients had additional molecular testing performed on preceding/subsequent biopsies, and the results across multiple samples were compared. In two patients, the NGS-based fluid analysis identified clinically actionable alterations that were not detected by other hotspot testing. In four patients treated with tyrosine kinase inhibitors, malignant fluid sequencing confirmed driver alterations from prior testing and revealed new resistance mechanisms. Hence, given adequate DNA input and tumor cellularity, comprehensive genomic profiling of malignant effusions may be used to establish mutational status at diagnosis and inform treatment resistance during targeted therapy.

    View details for PubMedID 29269277

  • Identification of a Novel MPL Loss of Function Mutation in a Patient with Cyclic Thrombocytopenia and Characterization of This Syndrome Zhang, H., Hou, Y., Brar, R. S., Zhang, B., Chen, Z., Abidi, P., Jin, J., Gotlib, J. R., Zehnder, J. L. AMER SOC HEMATOLOGY. 2016

    View details for Web of Science ID 000394452300114

  • Methodologic Considerations in the Application of Next-Generation Sequencing of Human TRB Repertoires for Clinical Use. journal of molecular diagnostics Xu, L., You, X., Zheng, P., Zhang, B. M., Gupta, P. K., Lavori, P., Meyer, E., Zehnder, J. L. 2016

    Abstract

    Next-generation sequencing (NGS) of immune receptors has become a standard tool to assess minimal residual disease (MRD) in patients treated for lymphoid malignancy, and it is being used to study the T-cell repertoire in many clinical settings. To better understanding the potential clinical utility and limitations of this application outside of MRD, we developed a BIOMED-2 primer-based NGS method and characterized its performance in controls and patients with graft-versus-host disease (GVHD) after allogeneic hematopoietic transplant. For controls and patients with GVHD, replicate sequencing of the same T-cell receptor β (TRB) libraries was highly reproducible. Higher variability was observed in sequencing of different TRB libraries made from the same DNA stock. Variability was increased in patients with GVHD compared with controls; patients with GVHD also had lower diversity than controls. In the T-cell repertoire of a healthy person, approximately 99.6% of the CDR3 clones were in low abundance, with frequency <10(-3). A single library could identify >93% of the clones with frequency ≥10(-3) in the repertoire. Sequencing in duplicate increased the average detection rate to >97%. This work demonstrates that NGS reliably and robustly characterizes TRB populations in healthy individuals and patients with GVHD with frequency ≥10(-3) and provides a methodologic framework for applying NGS immune repertoire methods to clinical testing applications beyond MRD.

    View details for DOI 10.1016/j.jmoldx.2016.07.009

    View details for PubMedID 27815002

  • Development of a Cloud-Based Comprehensive Cancer Transcriptome Profiling Pipeline for Clinical Diagnostics Costa, H. A., Kunder, C., Zehnder, J. L., Bustamante, C. D. ELSEVIER SCIENCE INC. 2016: 989

    View details for Web of Science ID 000387201000232

  • Targeted Next-Generation Sequencing-Based Mutation Profiling of Fine Needle Aspiration Samples: Utility and Technical Adequacy Lin, C., Yang, S., Costa, H. A., Weiel, J., Lo, A., Zhang, M., Kunder, C., Long, S., Zehnder, J. ELSEVIER SCIENCE INC. 2016: 999

    View details for Web of Science ID 000387201000275

  • Detection of POLE Exonuclease Domain Mutations by Targeted Genomic Profiling in Hypermutated, Microsatellite Stable Colorectal Carcinoma Steiner, D. F., Lo, A., Jones, C., Gojenola, L., Kunder, C., Zehnder, J., Suarez, C. J. ELSEVIER SCIENCE INC. 2016: 1006

    View details for Web of Science ID 000387201000301

  • Value of NGS-Based T-Cell Clonality Analysis in Cases of Mycosis Fungoides with Discrepant Fragment Size-Based Tissue and Blood Results Zhang, B. M., Xu, L., You, X., Suarez, C. J., Kim, J., Zehnder, J. L. ELSEVIER SCIENCE INC. 2016: 951

    View details for Web of Science ID 000387201000079

  • A study of the mutational landscape of pediatric-type follicular lymphoma and pediatric nodal marginal zone lymphoma. Modern pathology Ozawa, M. G., Bhaduri, A., Chisholm, K. M., Baker, S. A., Ma, L., Zehnder, J. L., Luna-Fineman, S., Link, M. P., Merker, J. D., Arber, D. A., Ohgami, R. S. 2016; 29 (10): 1212-1220

    Abstract

    Pediatric-type follicular lymphoma and pediatric marginal zone lymphoma are two of the rarest B-cell lymphomas. These lymphomas occur predominantly in the pediatric population and show features distinct from their more common counterparts in adults: adult-type follicular lymphoma and adult-type nodal marginal zone lymphoma. Here we report a detailed whole-exome deep sequencing analysis of a cohort of pediatric-type follicular lymphomas and pediatric marginal zone lymphomas. This analysis revealed a recurrent somatic variant encoding p.Lys66Arg in the transcription factor interferon regulatory factor 8 (IRF8) in 3 of 6 cases (50%) of pediatric-type follicular lymphoma. This specific point mutation was not detected in pediatric marginal zone lymphoma or in adult-type follicular lymphoma. Additional somatic point mutations in pediatric-type follicular lymphoma were observed in genes involved in transcription, intracellular signaling, and cell proliferation. In pediatric marginal zone lymphoma, no recurrent mutation was identified; however, somatic point mutations were observed in genes involved in cellular adhesion, cytokine regulatory elements, and cellular proliferation. A somatic variant in AMOTL1, a recurrently mutated gene in splenic marginal zone lymphoma, was also identified in a case of pediatric marginal zone lymphoma. The overall non-synonymous mutational burden was low in both pediatric-type follicular lymphoma and pediatric marginal zone lymphoma (4.6 mutations per exome). Altogether, these findings support a distinctive genetic basis for pediatric-type follicular lymphoma and pediatric marginal zone lymphoma when compared with adult subtypes and to one another. Moreover, identification of a recurrent point mutation in IRF8 provides insight into a potential driver mutation in the pathogenesis of pediatric-type follicular lymphoma with implications for novel diagnostic or therapeutic strategies.Modern Pathology advance online publication, 24 June 2016; doi:10.1038/modpathol.2016.102.

    View details for DOI 10.1038/modpathol.2016.102

    View details for PubMedID 27338637

  • Population-specific single-nucleotide polymorphism confers increased risk of venous thromboembolism in African Americans. Molecular genetics & genomic medicine Daneshjou, R., Cavallari, L. H., Weeke, P. E., Karczewski, K. J., Drozda, K., Perera, M. A., Johnson, J. A., Klein, T. E., Bustamante, C. D., Roden, D. M., Shaffer, C., Denny, J. C., Zehnder, J. L., Altman, R. B. 2016; 4 (5): 513-520

    Abstract

    African Americans have a higher incidence of venous thromboembolism (VTE) than European descent individuals. However, the typical genetic risk factors in populations of European descent are nearly absent in African Americans, and population-specific genetic factors influencing the higher VTE rate are not well characterized.We performed a candidate gene analysis on an exome-sequenced African American family with recurrent VTE and identified a variant in Protein S (PROS1) V510M (rs138925964). We assessed the population impact of PROS1 V510M using a multicenter African American cohort of 306 cases with VTE compared to 370 controls. Additionally, we compared our case cohort to a background population cohort of 2203 African Americans in the NHLBI GO Exome Sequencing Project (ESP).In the African American family with recurrent VTE, we found prior laboratories for our cases indicating low free Protein S levels, providing functional support for PROS1 V510M as the causative mutation. Additionally, this variant was significantly enriched in the VTE cases of our multicenter case-control study (Fisher's Exact Test, P = 0.0041, OR = 4.62, 95% CI: 1.51-15.20; allele frequencies - cases: 2.45%, controls: 0.54%). Similarly, PROS1 V510M was also enriched in our VTE case cohort compared to African Americans in the ESP cohort (Fisher's Exact Test, P = 0.010, OR = 2.28, 95% CI: 1.26-4.10).We found a variant, PROS1 V510M, in an African American family with VTE and clinical laboratory abnormalities in Protein S. Additionally, we found that this variant conferred increased risk of VTE in a case-control study of African Americans. In the ESP cohort, the variant is nearly absent in ESP European descent subjects (n = 3, allele frequency: 0.03%). Additionally, in 1000 Genomes Phase 3 data, the variant only appears in African descent populations. Thus, PROS1 V510M is a population-specific genetic risk factor for VTE in African Americans.

    View details for DOI 10.1002/mgg3.226

    View details for PubMedID 27652279

  • Gender and duration of disease differentiate responses to rituximab-dexamethasone therapy in adults with immune thrombocytopenia AMERICAN JOURNAL OF HEMATOLOGY Chapin, J., Lee, C. S., Zhang, H., Zehnder, J. L., Bussel, J. B. 2016; 91 (9): 907–11

    Abstract

    Adults often develop chronic immune thrombocytopenia (ITP) for which treatment order is uncertain. Rituximab and three cycles of dexamethasone (4R + 3Dex) improve treatment responses and short-term disease control but long-term outcome is not known. In adults with ITP treated with 4R + 3D, we sought long-term outcome and associated prognostic variables. Forty-nine adults treated at Weill-Cornell received 4R + 3Dex. Their clinical characteristics were reviewed. Duration was median time to treatment failure; Kaplan-Meier estimates were developed. Vbeta Tcell receptor (VBTCR) repertoire was obtained after treatment in 36 patients. Patients were adults with ITP 18-64 years old, median age 37. The 27 females were twice as likely to have an ongoing response to 4R + 3Dex (44.1%) as males (19.6%; P = 0.009). For ITP duration <12 months, 52.7% of patients had continuing responses to 4R + 3Dex compared to 15.3% of patients with diagnosis >12 months (P = 0.02). Females with ITP duration of <12 months had continuing responses in 78.6%, compared to males with <12 months duration of ITP (21.2%). For patients with disease duration <12 months, 67% of females had continuing responses, compared to 31% of males (P = 0.004). Post-treatment polyclonal VBTCR was seen in 9/10 continuing responders (six female, three male) but only 13/26 relapsers/nonresponders (P = 0.068). Durable remissions after treatment with 4R + 3Dex were more frequent in female patients with <12 months of ITP duration and those with polyclonal VBTCR after treatment, emphasizing the roles of duration of disease, gender and T cells in chronic ITP. Differences in pathophysiology of ITP by gender and by duration of ITP require further study. Am. J. Hematol. 91:907-911, 2016. © 2016 Wiley Periodicals, Inc.

    View details for PubMedID 27220625

  • DESCRIPTION AND LABORATORY VALIDATION OF A NOVEL ALGORITHM FOR CALCULATION OF TEG PLATELET MAPPING (TM) WHEN ATYPICAL ACTIVATOR F TRACING" OCCURS Jin, J., Zehnder, J. WILEY-BLACKWELL. 2016: E428

    View details for Web of Science ID 000385237100145

  • Germ line variants predispose to both JAK2 V617F clonal hematopoiesis and myeloproliferative neoplasms. Blood Hinds, D. A., Barnholt, K. E., Mesa, R. A., Kiefer, A. K., Do, C. B., Eriksson, N., Mountain, J. L., Francke, U., Tung, J. Y., Nguyen, H. M., Zhang, H., Gojenola, L., Zehnder, J. L., Gotlib, J. 2016; 128 (8): 1121-1128

    Abstract

    We conducted a genome-wide association study (GWAS) to identify novel predisposition alleles associated with Philadelphia chromosome-negative myeloproliferative neoplasms (MPNs) and JAK2 V617F clonal hematopoiesis in the general population. We recruited a web-based cohort of 726 individuals with polycythemia vera, essential thrombocythemia, and myelofibrosis and 252 637 population controls unselected for hematologic phenotypes. Using a single-nucleotide polymorphism (SNP) array platform with custom probes for the JAK2 V617F mutation (V617F), we identified 497 individuals (0.2%) among the population controls who were V617F carriers. We performed a combined GWAS of the MPN cases plus V617F carriers in the control population (n = 1223) vs the remaining controls who were noncarriers for V617F (n = 252 140). For these MPN cases plus V617F carriers, we replicated the germ line JAK2 46/1 haplotype (rs59384377: odds ratio [OR] = 2.4, P = 6.6 × 10(-89)), previously associated with V617F-positive MPN. We also identified genome-wide significant associations in the TERT gene (rs7705526: OR = 1.8, P = 1.1 × 10(-32)), in SH2B3 (rs7310615: OR = 1.4, P = 3.1 × 10(-14)), and upstream of TET2 (rs1548483: OR = 2.0, P = 2.0 × 10(-9)). These associations were confirmed in a separate replication cohort of 446 V617F carriers vs 169 021 noncarriers. In a joint analysis of the combined GWAS and replication results, we identified additional genome-wide significant predisposition alleles associated with CHEK2, ATM, PINT, and GFI1B All SNP ORs were similar for MPN patients and controls who were V617F carriers. These data indicate that the same germ line variants endow individuals with a predisposition not only to MPN, but also to JAK2 V617F clonal hematopoiesis, a more common phenomenon that may foreshadow the development of an overt neoplasm.

    View details for DOI 10.1182/blood-2015-06-652941

    View details for PubMedID 27365426

  • ROS: novel regulators of thrombopoiesis BLOOD Zhang, B., Zehnder, J. L. 2016; 128 (5): 613-?

    View details for DOI 10.1182/blood-2016-06-718544

    View details for Web of Science ID 000383843000004

    View details for PubMedID 27492312

  • Prozone Effect in the Diagnosis of Lupus Anticoagulant for the Lupus Anticoagulant-Hypoprothrombinemia Syndrome. American journal of clinical pathology Jin, J., Zehnder, J. L. 2016; 146 (2): 262-267

    Abstract

    The main clinical sequela of a lupus anticoagulant is increased thrombosis risk. However, bleeding due to lupus anticoagulant-hypoprothrombinemia syndrome is a rare but well-described manifestation of antiphospholipid syndrome. The association of acute acquired hypoprothrombinemia is caused by a lupus anticoagulant's specificity to prothrombin, which results in clearance of prothrombin and bleeding due to hypoprothrombinemia (usually <10% of normal). Severe life-threatening bleeding is most frequently reported in children with systemic lupus erythematosus or in healthy children after viral infection. In such cases, steroid therapy is usually effective in controlling the bleeding problems and improving prothrombin levels.We report one pediatric patient with a lupus anticoagulant who had acute hemorrhagic diathesis.The diagnosis in this case was complicated by the presence of a prozone effect in lupus anticoagulant testing. The prozone effect (also known as hook effect) refers to situations where very high concentrations of antibody mask detection, typically in antigen-antibody reactions, which depend on visualization of agglutination. Decreasing the antibody/antigen ratio results in detectable antigen-antibody complexes.We report for the first time a variation on this theme in a patient with a lupus anticoagulant-type antiphospholipid antibody and hypoprothrombinemia, which corrected with immunosuppression and restoration of normal prothrombin levels.

    View details for DOI 10.1093/ajcp/aqw106

    View details for PubMedID 27473743

  • Template for Reporting Results of Monitoring Tests for Patients With Chronic Myelogenous Leukemia (BCR-ABL1(+)). Archives of pathology & laboratory medicine Kelley, T. W., Arber, D. A., Gibson, C., Jones, D., Khoury, J. D., Medeiros, B. C., O'Malley, D. P., Patel, K. P., Pilichowska, M., Vasef, M. A., Wallentine, J., Zehnder, J. L. 2016; 140 (7): 672-674

    View details for DOI 10.5858/arpa.2015-0399-CP

    View details for PubMedID 26653363

  • Template for Reporting Results of Biomarker Testing of Specimens From Patients With Myeloproliferative Neoplasms. Archives of pathology & laboratory medicine Kelley, T. W., Arber, D. A., Gibson, C., Jones, D., Khoury, J. D., Medeiros, B. C., O'Malley, D. P., Patel, K. P., Pilichowska, M., Vasef, M. A., Wallentine, J., Zehnder, J. L. 2016; 140 (7): 675-677

    View details for DOI 10.5858/arpa.2015-0400-CP

    View details for PubMedID 26653364

  • LABORATORY ADHERENCE TO ISTH GUIDELINES ON PLATELET AGGREGATION TESTING. INITIAL REVIEW OF RESULTS FROM AN INTERNATIONAL SURVEY. Moffat, K. A., Zehnder, J., Meijer, P., Plumhoff, E., Hayward, C. M. WILEY-BLACKWELL. 2016: 74–75

    View details for Web of Science ID 000383521500175

  • Clinical activity of ponatinib in a patient with FGFR1-rearranged mixed-phenotype acute leukemia. Leukemia Khodadoust, M. S., Luo, B., Medeiros, B. C., Johnson, R. C., Ewalt, M. D., Schalkwyk, A. S., Bangs, C. D., Cherry, A. M., Arai, S., Arber, D. A., Zehnder, J. L., Gotlib, J. 2016; 30 (4): 947-950

    View details for DOI 10.1038/leu.2015.136

    View details for PubMedID 26055304

  • Measuring the Effect of Dipyridamole and Clopidogrel Using Thromboelastography/Platelet Mapping (R): Is There Evidence of a Meaningful Dose-Response Relationship in Children Supported with the Berlin Heart EXCOR Ventricular Assist Device? May, L. J., Lo, C., Tesoro, T. M., Zehnder, J. L., Yang, L., Chen, S., Lee, J., McElhinney, D. B., Rosenthal, D. N., Maeda, K., Almond, C. S. ELSEVIER SCIENCE INC. 2016: S47

    View details for DOI 10.1016/j.healun.2016.01.127

    View details for Web of Science ID 000374718100104

  • Isolated Follicles Enriched for Centroblasts and Lacking t(14;18)/BCL2 in Lymphoid Tissue: Diagnostic and Clinical Implications PLOS ONE Nybakken, G. E., Bala, R., Gratzinger, D., Jones, C. D., Zehnder, J. L., Bangs, C. D., Cherry, A., Warnke, R. A., Natkunam, Y. 2016; 11 (3)

    Abstract

    We sought to address the significance of isolated follicles that exhibit atypical morphologic features that may be mistaken for lymphoma in a background of reactive lymphoid tissue. Seven cases that demonstrated centroblast-predominant isolated follicles and absent BCL2 staining in otherwise-normal lymph nodes were studied. Four of seven cases showed clonal B-cell proliferations amid a polyclonal B cell background; all cases lacked the IGH-BCL2 translocation and BCL2 protein expression. Although three patients had invasive breast carcinoma at other sites, none were associated with systemic lymphoma up to 44 months after diagnosis. The immunoarchitectural features of these highly unusual cases raise the question of whether a predominance of centroblasts and/or absence of BCL2 expression could represent a precursor lesion or atypical reactive phenomenon. Differentiating such cases from follicular lymphoma or another mimic is critical, lest patients with indolent proliferations be exposed to unnecessarily aggressive treatment.

    View details for DOI 10.1371/journal.pone.0151735

    View details for Web of Science ID 000372582800093

    View details for PubMedCentralID PMC4798531

  • Clonal architecture of CXCR4 WHIM-like mutations in Waldenstrom Macroglobulinaemia BRITISH JOURNAL OF HAEMATOLOGY Xu, L., Hunter, Z. R., Tsakmaklis, N., Cao, Y., Yang, G., Chen, J., Liu, X., Kanan, S., Castillo, J. J., Tai, Y., Zehnder, J. L., Brown, J. R., Carrasco, R. D., Advani, R., Sabile, J. M., Argyropoulos, K., Palomba, M. L., Morra, E., Trojani, A., Greco, A., Tedeschi, A., Varettoni, M., Arcaini, L., Munshi, N. M., Anderson, K. C., Treon, S. P. 2016; 172 (5): 735-744

    Abstract

    CXCR4(WHIM) somatic mutations are distinctive to Waldenström Macroglobulinaemia (WM), and impact disease presentation and treatment outcome. The clonal architecture of CXCR4(WHIM) mutations remains to be delineated. We developed highly sensitive allele-specific polymerase chain reaction (AS-PCR) assays for detecting the most common CXCR4(WHIM) mutations (CXCR4(S338X C>A and C>G) ) in WM. The AS-PCR assays detected CXCR4(S338X) mutations in WM and IgM monoclonal gammopathy of unknown significance (MGUS) patients not revealed by Sanger sequencing. By combined AS-PCR and Sanger sequencing, CXCR4(WHIM) mutations were identified in 44/102 (43%), 21/62 (34%), 2/12 (17%) and 1/20 (5%) untreated WM, previously treated WM, IgM MGUS and marginal zone lymphoma patients, respectively, but no chronic lymphocytic leukaemia, multiple myeloma, non-IgM MGUS patients or healthy donors. Cancer cell fraction analysis in WM and IgM MGUS patients showed CXCR4(S338X) mutations were primarily subclonal, with highly variable clonal distribution (median 35·1%, range 1·2-97·5%). Combined AS-PCR and Sanger sequencing revealed multiple CXCR4(WHIM) mutations in many individual WM patients, including homozygous and compound heterozygous mutations validated by deep RNA sequencing. The findings show that CXCR4(WHIM) mutations are more common in WM than previously revealed, and are primarily subclonal, supporting their acquisition after MYD88(L265P) in WM oncogenesis. The presence of multiple CXCR4(WHIM) mutations within individual WM patients may be indicative of targeted CXCR4 genomic instability.

    View details for DOI 10.1111/bjh.13897

    View details for PubMedID 26659815

  • Somatic Mutational Landscape of Inflammatory Bowel Disease-Associated Signet Ring Cell Colorectal Carcinoma Lo, A. A., Costa, H. A., Zehnder, J., Rao, S., Bustamante, C. D., Yang, G., Beaubier, N. NATURE PUBLISHING GROUP. 2016: 456A

    View details for Web of Science ID 000370302503280

  • Comparative Study of Isocitrate Dehydrogenase 1 Mutations in Intraheptatic Cholangiocarcinoma by Immunohistochemistry and Pyrosequencing Lo, A. A., Costa, H. A., Zehnder, J., Dittman, D., Rao, S., Yang, G., Beaubier, N. NATURE PUBLISHING GROUP. 2016: 443A–444A

    View details for Web of Science ID 000369270702496

  • Somatic Mutational Landscape of Inflammatory Bowel Disease-Associated Signet Ring Cell Colorectal Carcinoma Lo, A. A., Costa, H. A., Zehnder, J., Rao, S., Bustamante, C. D., Yang, G., Beaubier, N. NATURE PUBLISHING GROUP. 2016: 456A

    View details for Web of Science ID 000369270702545

  • Comparative Study of Isocitrate Dehydrogenase 1 Mutations in Intraheptatic Cholangiocarcinoma by Immunohistochemistry and Pyrosequencing Lo, A. A., Costa, H. A., Zehnder, J., Dittman, D., Rao, S., Yang, G., Beaubier, N. NATURE PUBLISHING GROUP. 2016: 443A–444A

    View details for Web of Science ID 000370302503231

  • Isolated Follicles Enriched for Centroblasts and Lacking t(14;18)/BCL2 in Lymphoid Tissue: Diagnostic and Clinical Implications. PloS one Nybakken, G. E., Bala, R., Gratzinger, D., Jones, C. D., Zehnder, J. L., Bangs, C. D., Cherry, A., Warnke, R. A., Natkunam, Y. 2016; 11 (3)

    Abstract

    We sought to address the significance of isolated follicles that exhibit atypical morphologic features that may be mistaken for lymphoma in a background of reactive lymphoid tissue. Seven cases that demonstrated centroblast-predominant isolated follicles and absent BCL2 staining in otherwise-normal lymph nodes were studied. Four of seven cases showed clonal B-cell proliferations amid a polyclonal B cell background; all cases lacked the IGH-BCL2 translocation and BCL2 protein expression. Although three patients had invasive breast carcinoma at other sites, none were associated with systemic lymphoma up to 44 months after diagnosis. The immunoarchitectural features of these highly unusual cases raise the question of whether a predominance of centroblasts and/or absence of BCL2 expression could represent a precursor lesion or atypical reactive phenomenon. Differentiating such cases from follicular lymphoma or another mimic is critical, lest patients with indolent proliferations be exposed to unnecessarily aggressive treatment.

    View details for DOI 10.1371/journal.pone.0151735

    View details for PubMedID 26991267

    View details for PubMedCentralID PMC4798531

  • Factor V Leiden AMERICAN JOURNAL OF HEMATOLOGY Van Cott, E. M., Khor, B., Zehnder, J. L. 2016; 91 (1): 46–49

    Abstract

    Factor V Leiden (FVLeiden ) is a common hereditary thrombophilia that causes activated protein C (APC) resistance. This review describes many of the most fascinating features of FVLeiden , including background features, mechanisms of hypercoagulability, the founder mutation concept, the "FVLeiden paradox," synergistic interaction with other thrombotic risk factors, the intertwined relationship between FVLeiden and APC resistance testing, and other, uncommon mutations implicated in causing APC resistance. In addition, there are several conditions where laboratory tests for APC resistance and FVLeiden are or can be discrepant, including lupus anticoagulants, anticoagulants such as direct thrombin inhibitors (dabigatran, argatroban, and bivalirudin) and rivaroxaban, as well as pseudohomozygous, pseudo-wildtype, liver transplant, and bone marrow transplant patients. The laboratory test error rate for FVLeiden is also presented.

    View details for PubMedID 26492443

  • The Clonal Architecture of CXCR4mutations in Waldenstrom's Macroglobulinemia Shows Highly Variable Subclonal Distribution, and Multiple Mutations within Individual Patients Indicative of Targeted Genomic Instability Xu, L., Hunter, Z., Tsakmaklis, N., Cao, Y., Yang, G., Chen, J., Liu, X., Kanan, S., Castillo, J. J., Tai, Y., Zehnder, J. L., Brown, J. R., Carrasco, R. D., Advani, R. H., Argyropoulos, K. V., Palomba, M., Morra, E., Trojani, A., Greco, A., Tedeschi, A., Varettoni, M., Arcaini, L., Munshi, N. C., Anderson, K. C., Treon, S. P. AMER SOC HEMATOLOGY. 2015

    View details for Web of Science ID 000368019004286

  • The VarScan2's SNVs-Near-Indel Filter: Is It Necessary? Suarez, C. J., Stehr, H., Fung, E., Kunder, C. A., Ewalt, M. D., Lal, A., Alizadeh, A., Diehn, M., Schrijver, I., Zehnder, J. ELSEVIER SCIENCE INC. 2015: 801

    View details for Web of Science ID 000363830000216

  • Next-Generation Sequencing for Detection of Clonal TRG Gene Rearrangements Shows Improved Specificity and Positive Predictive Value Compared to Fragment Analysis Using BIOMED-2 Primers and Capillary Electrophoresis Ewalt, M. D., Klass, M., Panganiban, J., Huang, Y., Stenzel, T., Hutt, K., Ma, L., Arber, D. A., Merker, J. D., Schrijver, I., Zehnder, J. L. ELSEVIER SCIENCE INC. 2015: 770

    View details for Web of Science ID 000363830000086

  • Potential Pitfalls with TruSeq Amplicon: Cancer Panel Variant Detection Lai, A., Kunder, C. A., Ewalt, M. D., Schrijver, I., Zehnder, J., Suarez, C. J. ELSEVIER SCIENCE INC. 2015: 852

    View details for Web of Science ID 000363830000423

  • IBRUTINIB IN PREVIOUSLY TREATED PATIENTS WITH WALDENSTROM'S MACROGLOBULINEMIA IS HIGHLY ACTIVE, PRODUCES DURABLE RESPONSES, AND IS IMPACTED BY MYD88 AND CXCR4 MUTATION STATUS Treon, S., Tripsas, C., Meid, K., Warren, D., Varma, G., Green, R., Argyropoulos, K., Yang, G., Cao, Y., Xu, L., Patterson, C., Rodig, S., Zehnder, J., Aster, J., Harris, N. L., Kanan, S., Ghobrial, I., Laubach, J., Hunter, Z., Salman, Z., Cheng, M., Li, J., Clow, F., Graef, T., Castillo, J., Palomba, M. L., Advani, R. FERRATA STORTI FOUNDATION. 2015: 311

    View details for Web of Science ID 000361204902289

  • Next-generation sequencing of acute myeloid leukemia identifies the significance of TP53, U2AF1, ASXL1, and TET2 mutations MODERN PATHOLOGY Ohgami, R. S., Ma, L., Merker, J. D., Gotlib, J. R., Schrijver, I., Zehnder, J. L., Arber, D. A. 2015; 28 (5): 706-714

    Abstract

    We assessed the frequency and clinicopathologic significance of 19 genes currently identified as significantly mutated in myeloid neoplasms, RUNX1, ASXL1, TET2, CEBPA, IDH1, IDH2, DNMT3A, FLT3, NPM1, TP53, NRAS, EZH2, CBL, U2AF1, SF3B1, SRSF2, JAK2, CSF3R, and SETBP1, across 93 cases of acute myeloid leukemia (AML) using capture target enrichment and next-generation sequencing. Of these cases, 79% showed at least one nonsynonymous mutation, and cases of AML with recurrent genetic abnormalities showed a lower frequency of mutations versus AML with myelodysplasia-related changes (P<0.001). Mutational analysis further demonstrated that TP53 mutations are associated with complex karyotype AML, whereas ASXL1 and U2AF1 mutations are associated with AML with myelodysplasia-related changes. Furthermore, U2AF1 mutations were specifically associated with trilineage morphologic dysplasia. Univariate analysis demonstrated that U2AF1 and TP53 mutations are associated with absence of clinical remission, poor overall survival (OS), and poor disease-free survival (DFS; P<0.0001), whereas TET2 and ASXL1 mutations are associated with poor OS (P<0.03). In multivariate analysis, U2AF1 and TP53 mutations retained independent prognostic significance in OS and DFS, respectively. Our results demonstrate unique relationships between mutations in AML, clinicopathologic prognosis, subtype categorization, and morphologic dysplasia.Modern Pathology advance online publication, 21 November 2014; doi:10.1038/modpathol.2014.160.

    View details for DOI 10.1038/modpathol.2014.160

    View details for Web of Science ID 000353774200010

    View details for PubMedID 25412851

  • Ibrutinib in Previously Treated Waldenstrom's Macroglobulinemia NEW ENGLAND JOURNAL OF MEDICINE Treon, S. P., Tripsas, C. K., Meid, K., Warren, D., Varma, G., Green, R., Argyropoulos, K. V., Yang, G., Cao, Y., Xu, L., Patterson, C. J., Rodig, S., Zehnder, J. L., Aster, J. C., Harris, N. L., Kanan, S., Ghobrial, I., Castillo, J. J., Laubach, J. P., Hunter, Z. R., Salman, Z., Li, J., Cheng, M., Clow, F., Graef, T., Palomba, M. L., Advani, R. H. 2015; 372 (15): 1430-1440

    Abstract

    MYD88(L265P) and CXCR4(WHIM) mutations are highly prevalent in Waldenström's macroglobulinemia. MYD88(L265P) triggers tumor-cell growth through Bruton's tyrosine kinase, a target of ibrutinib. CXCR4(WHIM) mutations confer in vitro resistance to ibrutinib.We performed a prospective study of ibrutinib in 63 symptomatic patients with Waldenström's macroglobulinemia who had received at least one previous treatment, and we investigated the effect of MYD88 and CXCR4 mutations on outcomes. Ibrutinib at a daily dose of 420 mg was administered orally until disease progression or the development of unacceptable toxic effects.After the patients received ibrutinib, median serum IgM levels decreased from 3520 mg per deciliter to 880 mg per deciliter, median hemoglobin levels increased from 10.5 g per deciliter to 13.8 g per deciliter, and bone marrow involvement decreased from 60% to 25% (P<0.01 for all comparisons). The median time to at least a minor response was 4 weeks. The overall response rate was 90.5%, and the major response rate was 73.0%; these rates were highest among patients with MYD88(L265P)CXCR4(WT) (with WT indicating wild-type) (100% overall response rate and 91.2% major response rate), followed by patients with MYD88(L265P)CXCR4(WHIM) (85.7% and 61.9%, respectively) and patients with MYD88(WT)CXCR4(WT) (71.4% and 28.6%). The estimated 2-year progression-free and overall survival rates among all patients were 69.1% and 95.2%, respectively. Treatment-related toxic effects of grade 2 or higher included neutropenia (in 22% of the patients) and thrombocytopenia (in 14%), which were more common in heavily pretreated patients; postprocedural bleeding (in 3%); epistaxis associated with the use of fish-oil supplements (in 3%); and atrial fibrillation associated with a history of arrhythmia (5%).Ibrutinib was highly active, associated with durable responses, and safe in pretreated patients with Waldenström's macroglobulinemia. MYD88 and CXCR4 mutation status affected responses to this drug. (Funded by Pharmacyclics and others; ClinicalTrials.gov number, NCT01614821.).

    View details for DOI 10.1056/NEJMoa1501548

    View details for PubMedID 25853747

  • Mast Cells in Systemic Mastocytosis Have Distinctly Brighter CD45 Expression by Flow Cytometry AMERICAN JOURNAL OF CLINICAL PATHOLOGY Chisholm, K. M., Merker, J. D., Gotlib, J. R., Gitana, G., Lefterova, M., Zehnder, J. L., George, T. I., Arber, D. A., Ohgami, R. S. 2015; 143 (4): 527-534

    Abstract

    We sought to determine the significance of bright CD45 expression on mast cells in cases of systemic mastocytosis vs mast cells in bone marrows uninvolved by systemic mastocytosis and compare this CD45 expression with CD25 and CD2 expression on mast cells.Multiparameter flow cytometry was performed on 31 cases of systemic mastocytosis and 70 bone marrow cases that were not involved by systemic mastocytosis. Bright expression of CD45 was defined as more than 20% of CD117+ mast cells showing brighter CD45 expression than the average expression level of lymphocytes.Mast cells with bright CD45 expression were seen in 26 systemic mastocytosis cases and three bone marrows uninvolved by systemic mastocytosis (sensitivity, 84%; specificity, 96%). CD25 alone had a greater sensitivity (100%) but lower specificity (93%) compared with bright CD45 for identifying abnormal mast cells, while CD2 alone had lower sensitivity but higher specificity. To reach a specificity of 100%, CD25 together with bright CD45 on mast cells was the optimal combination to detect cases of systemic mastocytosis.A combination of bright CD45 and CD25 appears to specifically identify abnormal mast cells in cases of systemic mastocytosis. Further studies will be necessary to confirm these results.

    View details for DOI 10.1309/AJCPZ3J4GEEYIRRA

    View details for PubMedID 25780004

  • Thromboelastography/Platelet Mapping (R) and Aspirin: Is There Evidence of a Meaningful Dose-Response Relationship in Children Supported With the Berlin Heart EXCOR Ventricular Assist Device? May, L. J., Lo, C., Tesoro, T. M., Zehnder, J. L., Chen, S., Lee, J., Desai, M., McElhinney, D. B., Rosenthal, D. N., Maeda, K., Almond, C. S. ELSEVIER SCIENCE INC. 2015: S77

    View details for DOI 10.1016/j.healun.2015.01.202

    View details for Web of Science ID 000353251500186

  • The utility of IgM, CD21, HGAL and LMO2 in the diagnosis of pediatric follicular lymphoma HUMAN PATHOLOGY Karnik, T., Ozawa, M. G., Lefterova, M., Luna-Fineman, S., Alvarez, E., Link, M., Zehnder, J. L., Arber, D. A., Ohgami, R. S. 2015; 46 (4): 629-633

    Abstract

    Pediatric follicular lymphoma (pFL) is a rare neoplasm with features differing from follicular lymphoma arising in adults. Here, we describe a rare case of pFL that showed morphologic features partially overlapping with progressive transformation of germinal centers and reactive follicular hyperplasia. As typical of pFL, neoplastic B cells within follicles did not express B-cell leukemia/lymphoma 2 (BCL2). However, this case showed additional distinctive abnormal findings, which contributed to the diagnosis: (1) diffuse and uniform staining of immunoglobulin M (IgM) on cells within and outside of follicles, (2) abnormally dim expression of CD21 on follicular dendritic cells, and (3) expression of human germinal center-associated lymphoma (HGAL) and LIM domain only 2 (LMO2) on B cells in interfollicular and follicular areas. This case demonstrates the utility of these abnormal features, which can be seen in adult- or usual-type follicular lymphoma, in the diagnosis of pFL. Further studies are necessary to evaluate the significance of these findings in other cases of pFL.

    View details for DOI 10.1016/j.humpath.2014.12.016

    View details for Web of Science ID 000352117000020

    View details for PubMedID 25701230

  • Molecular Immunoglobulin/T-Cell Receptor Clonality Analysis by Bidirectional Amplicon Sequencing on MiSeq with a Phasing Primer Design Strategy Zhang, B. M., You, X., Xu, L., Schrijver, I., Zehnder, J. L. ELSEVIER SCIENCE INC. 2014: 714–15

    View details for Web of Science ID 000343794200083

  • Next-Generation Molecular Diagnostic Testing of Cystic Fibrosis Using Complementary Long Padlock Probes Odegaard, J. I., Lefterova, M. I., Shen, P., Fung, E., Peng, G., Broady, K. M., Gu, W., Zehnder, J., Davis, R. W., Wang, W., Schrijver, I., Scharfe, C. ELSEVIER SCIENCE INC. 2014: 712

    View details for Web of Science ID 000343794200071

  • Standardization of BCR-ABL1 Quantitation: Successes and Challenges, the Stanford Experience Ewalt, M. D., Gojenola, L., Zehnder, J. L., Schrijver, I. ELSEVIER SCIENCE INC. 2014: 714

    View details for Web of Science ID 000343794200080

  • An Unusual Case of Classical Hodgkin Lymphoma Expressing Multiple T-Cell Antigens: Contributions of Molecular Studies to Diagnosis in Hematopathology Ewalt, M. D., Natkunam, Y., Zehnder, J. L. ELSEVIER SCIENCE INC. 2014: 721

    View details for Web of Science ID 000343794200110

  • 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

  • Immunoglobulin and T cell receptor gene high-throughput sequencing quantifies minimal residual disease in acute lymphoblastic leukemia and predicts post-transplantation relapse and survival. Biology of blood and marrow transplantation Logan, A. C., Vashi, N., Faham, M., Carlton, V., Kong, K., Buño, I., Zheng, J., Moorhead, M., Klinger, M., Zhang, B., Waqar, A., Zehnder, J. L., Miklos, D. B. 2014; 20 (9): 1307-1313

    Abstract

    Minimal residual disease (MRD) quantification is an important predictor of outcome after treatment for acute lymphoblastic leukemia (ALL). Bone marrow ALL burden ≥ 10(-4) after induction predicts subsequent relapse. Likewise, MRD ≥ 10(-4) in bone marrow before initiation of conditioning for allogeneic (allo) hematopoietic cell transplantation (HCT) predicts transplantation failure. Current methods for MRD quantification in ALL are not sufficiently sensitive for use with peripheral blood specimens and have not been broadly implemented in the management of adults with ALL. Consensus-primed immunoglobulin (Ig), T cell receptor (TCR) amplification and high-throughput sequencing (HTS) permit use of a standardized algorithm for all patients and can detect leukemia at 10(-6) or lower. We applied the LymphoSIGHT HTS platform (Sequenta Inc., South San Francisco, CA) to quantification of MRD in 237 samples from 29 adult B cell ALL patients before and after allo-HCT. Using primers for the IGH-VDJ, IGH-DJ, IGK, TCRB, TCRD, and TCRG loci, MRD could be quantified in 93% of patients. Leukemia-associated clonotypes at these loci were identified in 52%, 28%, 10%, 35%, 28%, and 41% of patients, respectively. MRD ≥ 10(-4) before HCT conditioning predicted post-HCT relapse (hazard ratio [HR], 7.7; 95% confidence interval [CI], 2.0 to 30; P = .003). In post-HCT blood samples, MRD ≥10(-6) had 100% positive predictive value for relapse with median lead time of 89 days (HR, 14; 95% CI, 4.7 to 44, P < .0001). The use of HTS-based MRD quantification in adults with ALL offers a standardized approach with sufficient sensitivity to quantify leukemia MRD in peripheral blood. Use of this approach may identify a window for clinical intervention before overt relapse.

    View details for DOI 10.1016/j.bbmt.2014.04.018

    View details for PubMedID 24769317

  • 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

  • STAT3 mutations are present in aggressive B-cell lymphomas including a subset of diffuse large B-cell lymphomas with CD30 expression. Haematologica Ohgami, R. S., Ma, L., Monabati, A., Zehnder, J. L., Arber, D. A. 2014; 99 (7): e105-7

    Abstract

    -

    View details for DOI 10.3324/haematol.2013.101543

    View details for PubMedID 24837465

  • 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

  • High sensitivity and specificity of a new functional flow cytometry assay for clinically significant heparin-induced thrombocytopenia antibodies INTERNATIONAL JOURNAL OF LABORATORY HEMATOLOGY Garritsen, H. S., Probst-Kepper, M., Legath, N., Eberl, W., Samaniego, S., Woudenberg, J., Schuitemaker, J. H., Kroll, H., Gurney, D. A., Moore, G. W., Zehnder, J. L. 2014; 36 (2): 135-143

    Abstract

    Heparin-induced thrombocytopenia (HIT) is a life-threatening condition, in which the anticoagulant heparin, platelet factor 4 (PF4), and platelet-activating antibodies form complexes with prothrombotic properties. Laboratory tests to support clinical diagnosis are subdivided into functional, platelet activation assays, which lack standardization, or immunological assays, which have moderate specificity toward HIT. In this study, clinical performance of HITAlert, a novel in vitro diagnostic (IVD) registered platelet activation assay, was tested in a large cohort of HIT-suspected patients and compared with immunological assays.From 346 HIT-suspected patients (single center), clinical data including 4T pretest probability results, citrated platelet-poor plasmas, and sera were collected, allowing direct comparison of clinical observations with HITAlert results. HITAlert performance was compared with PF4 IgG ELISA (246 patients, three centers) and PF4 PaGIA (298 patients, single center).HITAlert showed high sensitivity (88.2%) and specificity (99.1%) when compared with clinical diagnosis. Agreement of HITAlert with PF4 ELISA- and PF4 PaGIA-positive patients is low (52.7 and 23.2%, respectively), while agreement with PF4 IgG ELISA- and PF4 PaGIA-negative patients is very high (98.1 and 99.1%, respectively).HITAlert performance is excellent when compared with clinical HIT diagnosis, making it a suitable assay for rapid testing of platelet activation due to anticoagulant therapy.

    View details for DOI 10.1111/ijlh.12136

    View details for Web of Science ID 000332777000004

    View details for PubMedID 23981347

  • 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

  • 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

  • A Balanced Look at the Implications of Genomic (and Other "Omics") Testing for Disease Diagnosis and Clinical Care. Genes Boyd, S. D., Galli, S. J., Schrijver, I., Zehnder, J. L., Ashley, E. A., Merker, J. D. 2014; 5 (3): 748-766

    Abstract

    The tremendous increase in DNA sequencing capacity arising from the commercialization of "next generation" instruments has opened the door to innumerable routes of investigation in basic and translational medical science. It enables very large data sets to be gathered, whose interpretation and conversion into useful knowledge is only beginning. A challenge for modern healthcare systems and academic medical centers is to apply these new methods for the diagnosis of disease and the management of patient care without unnecessary delay, but also with appropriate evaluation of the quality of data and interpretation, as well as the clinical value of the insights gained. Most critically, the standards applied for evaluating these new laboratory data and ensuring that the results and their significance are clearly communicated to patients and their caregivers should be at least as rigorous as those applied to other kinds of medical tests. Here, we present an overview of conceptual and practical issues to be considered in planning for the integration of genomic methods or, in principle, any other type of "omics" testing into clinical care.

    View details for DOI 10.3390/genes5030748

    View details for PubMedID 25257203

  • STAT3 mutations are frequent in CD30+ T-cell lymphomas and T-cell large granular lymphocytic leukemia. Leukemia Ohgami, R. S., Ma, L., Merker, J. D., Martinez, B., Zehnder, J. L., Arber, D. A. 2013; 27 (11): 2244-2247

    View details for DOI 10.1038/leu.2013.104

    View details for PubMedID 23563237

  • VIETNAMESE NON-SMALL CELL LUNG CANCER PATIENTS IN CALIFORNIA: MOLECULAR PROFILES AND CLINICAL CHARACTERISTICS Nguyen, K. H., Das, M., Ramchandran, K., Shrager, J., Merritt, R. E., Hoang, C., Burt, B., Tisch, A., Pagtama, J., Zehnder, J., Berry, G., Wakelee, H. A., Anh-Hoa Nguyen, Neal, J. W. LIPPINCOTT WILLIAMS & WILKINS. 2013: S208–S209

    View details for Web of Science ID 000339624901009

  • Comprehensive whole-genome sequencing of an early-stage primary myelofibrosis patient defines low mutational burden and non-recurrent candidate genes. Haematologica Merker, J. D., Roskin, K. M., Ng, D., Pan, C., Fisk, D. G., King, J. J., Hoh, R., Stadler, M., Okumoto, L. M., Abidi, P., Hewitt, R., Jones, C. D., Gojenola, L., Clark, M. J., Zhang, B., Cherry, A. M., George, T. I., Snyder, M., Boyd, S. D., Zehnder, J. L., Fire, A. Z., Gotlib, J. 2013; 98 (11): 1689-1696

    Abstract

    In order to identify novel somatic mutations associated with classic BCR/ABL1-negative myeloproliferative neoplasms, we performed high-coverage genome sequencing of DNA from peripheral blood granulocytes and cultured skin fibroblasts from a patient with MPL W515K-positive primary myelofibrosis. The primary myelofibrosis genome had a low somatic mutation rate, consistent with that observed in similar hematopoietic tumor genomes. Interfacing of whole-genome DNA sequence data with RNA expression data identified three somatic mutations of potential functional significance: a nonsense mutation in CARD6, implicated in modulation of NF-kappaB activation; a 19-base pair deletion involving a potential regulatory region in the 5'-untranslated region of BRD2, implicated in transcriptional regulation and cell cycle control; and a non-synonymous point mutation in KIAA0355, an uncharacterized protein. Additional mutations in three genes (CAP2, SOX30, and MFRP) were also evident, albeit with no support for expression at the RNA level. Re-sequencing of these six genes in 178 patients with polycythemia vera, essential thrombocythemia, and myelofibrosis did not identify recurrent somatic mutations in these genes. Finally, we describe methods for reducing false-positive variant calls in the analysis of hematologic malignancies with a low somatic mutation rate. This trial is registered with ClinicalTrials.gov (NCT01108159).

    View details for DOI 10.3324/haematol.2013.092379

    View details for PubMedID 23872309

  • Comprehensive whole-genome sequencing of an early-stage primary myelofibrosis patient defines low mutational burden and non-recurrent candidate genes. Haematologica Merker, J. D., Roskin, K. M., Ng, D., Pan, C., Fisk, D. G., King, J. J., Hoh, R., Stadler, M., Okumoto, L. M., Abidi, P., Hewitt, R., Jones, C. D., Gojenola, L., Clark, M. J., Zhang, B., Cherry, A. M., George, T. I., Snyder, M., Boyd, S. D., Zehnder, J. L., Fire, A. Z., Gotlib, J. 2013; 98 (11): 1689-1696

    Abstract

    In order to identify novel somatic mutations associated with classic BCR/ABL1-negative myeloproliferative neoplasms, we performed high-coverage genome sequencing of DNA from peripheral blood granulocytes and cultured skin fibroblasts from a patient with MPL W515K-positive primary myelofibrosis. The primary myelofibrosis genome had a low somatic mutation rate, consistent with that observed in similar hematopoietic tumor genomes. Interfacing of whole-genome DNA sequence data with RNA expression data identified three somatic mutations of potential functional significance: a nonsense mutation in CARD6, implicated in modulation of NF-kappaB activation; a 19-base pair deletion involving a potential regulatory region in the 5'-untranslated region of BRD2, implicated in transcriptional regulation and cell cycle control; and a non-synonymous point mutation in KIAA0355, an uncharacterized protein. Additional mutations in three genes (CAP2, SOX30, and MFRP) were also evident, albeit with no support for expression at the RNA level. Re-sequencing of these six genes in 178 patients with polycythemia vera, essential thrombocythemia, and myelofibrosis did not identify recurrent somatic mutations in these genes. Finally, we describe methods for reducing false-positive variant calls in the analysis of hematologic malignancies with a low somatic mutation rate. This trial is registered with ClinicalTrials.gov (NCT01108159).

    View details for DOI 10.3324/haematol.2013.092379

    View details for PubMedID 23872309

  • Minimal residual disease quantification using consensus primers and high- throughput IGH sequencing predicts post-transplant relapse in chronic lymphocytic leukemia LEUKEMIA Logan, A. C., Zhang, B., Narasimhan, B., Carlton, V., Zheng, J., Moorhead, M., Krampf, M. R., Jones, C. D., Waqar, A. N., Faham, M., Zehnder, J. L., Miklos, D. B. 2013; 27 (8): 1659-1665

    Abstract

    Quantification of minimal residual disease (MRD) following allogeneic hematopoietic cell transplantation (allo-HCT) predicts post-transplant relapse in patients with chronic lymphocytic leukemia (CLL). We utilized an MRD-quantification method that amplifies immunoglobulin heavy chain (IGH) loci using consensus V and J segment primers followed by high-throughput sequencing (HTS), enabling quantification with a detection limit of one CLL cell per million mononuclear cells. Using this IGH-HTS approach, we analyzed MRD patterns in over 400 samples from 40 CLL patients who underwent reduced-intensity allo-HCT. Nine patients relapsed within 12 months post-HCT. Of the 31 patients in remission at 12 months post-HCT, disease-free survival was 86% in patients with MRD <10(-4) and 20% in those with MRD 10(-4) (relapse hazard ratio (HR) 9.0; 95% confidence interval (CI) 2.5-32; P<0.0001), with median follow-up of 36 months. Additionally, MRD predicted relapse at other time points, including 9, 18 and 24 months post-HCT. MRD doubling time <12 months with disease burden 10(-5) was associated with relapse within 12 months of MRD assessment in 50% of patients, and within 24 months in 90% of patients. This IGH-HTS method may facilitate routine MRD quantification in clinical trials.Leukemia advance online publication, 12 March 2013; doi:10.1038/leu.2013.52.

    View details for DOI 10.1038/leu.2013.52

    View details for Web of Science ID 000322823200006

    View details for PubMedID 23419792

  • Oxidative stress and immune thrombocytopenia. Seminars in hematology Zhang, B., Zehnder, J. L. 2013; 50 (3): e1-4

    Abstract

    Immune thrombocytopenia (ITP) is an autoimmune disorder characterized by increased platelet destruction or decreased platelet production. The mechanism of the disease has been extensively studied so that we now have a much improved understanding of the pathophysiology; however, the trigger of the autoimmunity remains unclear. More recently, oxidative stress was identified to be involved in the pathogenesis of ITP and provides a new hypothesis for the initiation of autoimmunity in patients with ITP. In this review, oxidative stress and its impact on autoimmunity, particularly ITP, will be covered.

    View details for DOI 10.1053/j.seminhematol.2013.06.011

    View details for PubMedID 23953344

  • A distinct evolution of the T-cell repertoire categorizes treatment refractory gastrointestinal acute graft-versus-host disease. Blood Meyer, E. H., Hsu, A. R., Liliental, J., Löhr, A., Florek, M., Zehnder, J. L., Strober, S., Lavori, P., Miklos, D. B., Johnson, D. S., Negrin, R. S. 2013; 121 (24): 4955-4962

    Abstract

    Steroid refractory gastrointestinal (GI) acute graft versus host disease (aGVHD) is a major cause of mortality in hematopoietic stem cell transplantation (HCT) without immune markers to establish a diagnosis or guide therapy. We found that T cell receptor β (TCRβ) CDR3 repertoire sequencing reveals patterns that could eventually serve as a disease biomarker of T cell alloreactivity in aGVHD. We identified T cell clones in GI biopsies in a heterogeneous group of 15 allogeneic HCT patients with GI aGVHD symptoms. Seven steroid-refractory aGVHD patients showed a more conserved TCRβ clonal structure between different biopsy sites in the GI tract than eight primary-therapy responsive patients. Tracking GI clones identified at endoscopy longitudinally in the blood also revealed an increased clonal expansion in patients with steroid-refractory disease. Immune repertoire sequencing-based methods could enable a novel personalized way to guide diagnosis and therapy in diseases where T cell activity is a major determinant.

    View details for DOI 10.1182/blood-2013-03-489757

    View details for PubMedID 23652802

  • A distinct evolution of the T-cell repertoire categorizes treatment refractory gastrointestinal acute graft-versus-host disease BLOOD Meyer, E. H., Hsu, A. R., Liliental, J., Loehr, A., Florek, M., Zehnder, J. L., Strober, S., Lavori, P., Miklos, D. B., Johnson, D. S., Negrin, R. S. 2013; 121 (24): 4955-4962

    Abstract

    Steroid refractory gastrointestinal (GI) acute graft versus host disease (aGVHD) is a major cause of mortality in hematopoietic stem cell transplantation (HCT) without immune markers to establish a diagnosis or guide therapy. We found that T cell receptor β (TCRβ) CDR3 repertoire sequencing reveals patterns that could eventually serve as a disease biomarker of T cell alloreactivity in aGVHD. We identified T cell clones in GI biopsies in a heterogeneous group of 15 allogeneic HCT patients with GI aGVHD symptoms. Seven steroid-refractory aGVHD patients showed a more conserved TCRβ clonal structure between different biopsy sites in the GI tract than eight primary-therapy responsive patients. Tracking GI clones identified at endoscopy longitudinally in the blood also revealed an increased clonal expansion in patients with steroid-refractory disease. Immune repertoire sequencing-based methods could enable a novel personalized way to guide diagnosis and therapy in diseases where T cell activity is a major determinant.

    View details for DOI 10.1182/blood-2013-03-489757

    View details for Web of Science ID 000321896300024

  • 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

  • Indolent T-Lymphoblastic Proliferation (iT-LBP): A Review of Clinical and Pathologic Features and Distinction from Malignant T-Lymphoblastic Lymphoma ADVANCES IN ANATOMIC PATHOLOGY Ohgami, R. S., Arber, D. A., Zehnder, J. L., Natkunam, Y., Warnke, R. A. 2013; 20 (3): 137-140

    Abstract

    In recent years, a new pathologic entity has emerged: indolent T-lymphoblastic proliferation (iT-LBP). iT-LBPs share immunophenotypic similarities with T-lymphoblastic lymphoma; however, T-lymphoblastic proliferations are clinically indolent, and unlike the malignant counterpart, these expansions of nonclonal terminal deoxynucleotidyl transferase (TdT)+ T cells do not require treatment. Here we review the clinical and pathologic features, which are required for an accurate diagnosis of an iT-LBP. We demonstrate specific criteria can be used to accurately diagnose iT-LBP, notably: (1) confluent groups of TdT+ T cells in a biopsy specimen, (2) relative preservation of surrounding normal lymphoid architecture, (3) TdT+ T cells without morphologic atypia, (4) absence of thymic epithelium, (5) nonclonal TdT+ T cells, (6) immunophenotype of developmentally normal immature thymic T cells, and (7) clinical evidence of indolence (follow-up >6 mo without progression).

    View details for DOI 10.1097/PAP.0b013e31828d17ec

    View details for Web of Science ID 000317588700001

    View details for PubMedID 23574769

  • CLINICAL UTILITY OF ENDOGENOUS THROMBIN POTENTIAL IN MONITORING PATIENTS TREATED WITH UNFRACTIONATED HEPARIN Jin, J., Zehnder, J. WILEY-BLACKWELL. 2013: 69–69

    View details for Web of Science ID 000318286300136

  • Sequential azacitidine plus lenalidomide combination for elderly patients with untreated acute myeloid leukemia. Haematologica Pollyea, D. A., Zehnder, J., Coutre, S., Gotlib, J. R., Gallegos, L., Abdel-Wahab, O., Greenberg, P., Zhang, B., Liedtke, M., Berube, C., Levine, R., Mitchell, B. S., Medeiros, B. C. 2013; 98 (4): 591-596

    Abstract

    There are limited treatment options for older patients with acute myeloid leukemia and prognosis of these patients remains poor, thereby warranting development of novel therapies. We evaluated the efficacy and safety of azacitidine in combination with lenalidomide as front-line therapy for older patients with acute myeloid leukemia. Patients ≥ 60 years of age with untreated acute myeloid leukemia received azacitidine 75 mg/m2 for 7 days followed by escalating doses of lenalidomide daily for 21 days starting on day 8 of each cycle every 6 weeks. Patients received continued therapy until disease progression, unacceptable toxicity, or completion of 12 cycles. Forty-two patients (median age, 74 years) were enrolled with equal distribution according to European LeukemiaNet risk. The overall response rate was 40% (rate of complete remission with or without complete recovery of blood counts = 28%). The median time to complete remission with or without complete recovery of blood counts was 12 weeks, and duration of this status was 28 weeks (range, 4 - >104 weeks). Therapy-related acute myeloid leukemia and a high score on the Hematopoietic Cell Transplantation Comorbidity Index were negative predictors of response. Early death was noted in 17% of patients. Grades ≥ 3 toxicities were uncommon and most adverse events were gastrointestinal, fatigue and myelosuppression. In conclusion, a sequential combination of azacitidine plus lenalidomide has clinical activity in older patients with acute myeloid leukemia, and further studies of this combination are underway.

    View details for DOI 10.3324/haematol.2012.076414

    View details for PubMedID 23242596

  • Impaired B Cell Clonotype Diversification After Allogeneic Hematopoietic Cell Transplantation Predicts Graft-Versus-Host Disease BMT Tandem Meetings Logan, A., Sahaf, B., Zhang, B., Arai, S., Carlton, V., Zheng, J., Moorhead, M., Krampf, M. R., Jones, C. D., Waqar, A. N., Faham, M., Shizuru, J. A., Zehnder, J. L., Miklos, D. B. ELSEVIER SCIENCE INC. 2013: S148–S149

    View details for Web of Science ID 000314441900072

  • 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

  • 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

  • The Expansion of Gastrointestinal-Associated alpha beta T Cell Clones in Peripheral Blood Associates with Severe Steroid Refractory GVHD BMT Tandem Meetings Meyer, E. H., Liliental, J. A., Florek, M., Lohr, A., Hsu, A., Johnson, D., Lavori, P., Zehnder, J. L., Miklos, D. B., Strober, S., Negrin, R. ELSEVIER SCIENCE INC. 2013: S334–S335

    View details for Web of Science ID 000314441900449

  • 2-Hydroxyglutarate in IDH mutant acute myeloid leukemia: predicting patient responses, minimal residual disease and correlations with methylcytosine and hydroxymethylcytosine levels LEUKEMIA & LYMPHOMA Pollyea, D. A., Kohrt, H. E., Zhang, B., Zehnder, J., Schenkein, D., Fantin, V., Straley, K., Vasanthakumar, A., Abdel-Wahab, O., Levine, R., Godley, L. A., Medeiros, B. C. 2013; 54 (2): 408-410

    View details for DOI 10.3109/10428194.2012.701009

    View details for Web of Science ID 000313285400034

    View details for PubMedID 22680765

  • Discordant aPTT and anti-Xa values and outcomes in hospitalized patients treated with intravenous unfractionated heparin. Annals of pharmacotherapy Price, E. A., Jin, J., Nguyen, H. M., Krishnan, G., Bowen, R., Zehnder, J. L. 2013; 47 (2): 151-158

    Abstract

    Both the activated partial thromboplastin time (aPTT) and anti-Xa assay can be used to monitor unfractionated heparin (UFH). Following implementation of an anti-Xa method for heparin dosing protocols in our hospital, we became aware of many patients with discordant aPTT and anti-Xa values.To determine the frequency of discordant aPTT and anti-Xa values in a large cohort of hospitalized patients treated with UFH, as well as the demographics, coagulation status, indication for UFH, and clinical outcomes in this population.All aPTT and anti-Xa values from adults hospitalized between February and August 2009 at Stanford Hospital who were treated with UFH were analyzed. All samples were drawn simultaneously. A polynomial fit correlating aPTT and anti-Xa with a 99% confidence limit was designed. Paired aPTT/anti-Xa values were grouped according to whether the paired values fell within or outside of the concordant area. Patients were placed into groups based on concordance status, and clinical outcomes were assessed.A total of 2321 paired values from 539 patients were studied; 42% of data pairs had a high aPTT value relative to the anti-Xa value. Patients with elevated baseline prothrombin time/international normalized ratio or aPTT frequently demonstrated disproportionate relative prolongation of the aPTT. Patients with at least 2 consecutive high aPTT to anti-Xa values had increased 21-day major bleeding (9% vs 3%; p = 0.0316) and 30-day mortality (14% dead vs 5% dead at 30 days; p = 0.0202) compared with patients with consistently concordant values.aPTT and anti-Xa values are frequently discordant when used to measure UFH in hospitalized patients. A disproportionate prolongation of the aPTT relative to the anti-Xa was the most common discordant pattern in our study. Patients with relatively high aPTT to anti-Xa values appear to be at increased risk of adverse outcomes. Monitoring both aPTT and Xa values may have utility in managing such patients.

    View details for DOI 10.1345/aph.1R635

    View details for PubMedID 23386070

  • The Expansion of Gastrointestinal-associated alpha beta T Cell Clones in Peripheral Blood Over Time Is a Disease Feature of Severe Acute Graft-Versus-Host Disease 54th Annual Meeting and Exposition of the American-Society-of-Hematology (ASH) Meyer, E. H., Liliental, J., Florek, M., Loehr, A., Hsu, A., Johnson, D. S., Miklos, D. B., Zehnder, J. L., Negrin, R. S. AMER SOC HEMATOLOGY. 2012

    View details for Web of Science ID 000313838901092

  • Azacitidine Plus Lenalidomide for Untreated AML Patients Ineligible for Conventional Chemotherapy Pollyea, D. A., Zehnder, J. L., Coutre, S., Gotlib, J., Gallegos, L., Greenberg, P., Zhang, B., Liedtke, M., Levine, R. L., Medeiros, B. C. AMER SOC HEMATOLOGY. 2012

    View details for Web of Science ID 000313838906082

  • The Role of Oxidative Stress in Pediatric Immune Thrombocytopenia Lo, C., Zhang, B., Cusmano-Ozog, K., Wong, W., Jeng, M., Cowan, T., Zehnder, J. L. AMER SOC HEMATOLOGY. 2012

    View details for Web of Science ID 000314049602365

  • A Germline Variant in the TERT Gene Is a Novel Predisposition Allele Associated with Myeloproliferative Neoplasms Hinds, D. A., Barnholt, K. E., Zehnder, J. L., Kiefer, A. K., Do, C. B., Eriksson, N., Mountain, J. L., Francke, U., Tung, J. A., Huong (Marie) Nguyen, Levine, R. L., Mesa, R. A., Gotlib, J. AMER SOC HEMATOLOGY. 2012

    View details for Web of Science ID 000313838900250

  • Correlation of Symptom Assessment with Genotyping Analysis of Saliva Samples in a Large Cohort of Myeloproliferative Neoplasm Patients 54th Annual Meeting and Exposition of the American-Society-of-Hematology (ASH) Nguyen, H. (., Hinds, D. A., Barnholt, K. E., Kiefer, A. K., Do, C. B., Eriksson, N., Mountain, J. L., Francke, U., Tung, J. A., Levine, R. L., Zehnder, J. L., Gotlib, J., Mesa, R. A. AMER SOC HEMATOLOGY. 2012

    View details for Web of Science ID 000314049602145

  • Estimation of JAK2 V617F Prevalence by Detection of the Mutation in Saliva Samples From Online MPN and General Population Cohorts 54th Annual Meeting and Exposition of the American-Society-of-Hematology (ASH) Barnholt, K. E., Hinds, D. A., Kiefer, A. K., Do, C. B., Eriksson, N., Mountain, J. L., Francke, U., Tung, J. A., Nguyen, H. (., Levine, R. L., Mesa, R. A., Gotlib, J., Zehnder, J. L. AMER SOC HEMATOLOGY. 2012

    View details for Web of Science ID 000314049602140

  • Whole Genome Sequence Analysis of Primary Myelofibrosis. 54th Annual Meeting and Exposition of the American-Society-of-Hematology (ASH) Merker, J. D., Roskin, K., Ng, D., Pan, C., Fisk, D. G., Jones, C. D., Gojenola, L., Clark, M. J., Zhang, B., Cherry, M., Snyder, M., Boyd, S. D., Zehnder, J. L., Fire, A. Z., Gotlib, J. AMER SOC HEMATOLOGY. 2012

    View details for Web of Science ID 000313838905376

  • 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

  • USING A FUNCTIONAL FLOW CYTOMETRIC ASSAY (HITALERT) TO SUPPORT CLINICAL HEPARIN-INDUCED THROMBOCYTOPENIA (HIT) DIAGNOSIS Garritsen, H., Probst-Kepper, M., Legath, N., Eberl, W., Samianego, S., Schuitemaker, J., de Boef, E., Woudenberg, J., Guerney, D., Moore, G. W., Zehnder, J. L. WILEY-BLACKWELL. 2012: 394

    View details for Web of Science ID 000310386300054

  • Prophylactic rituximab after allogeneic transplantation decreases B-cell alloimmunity with low chronic GVHD incidence BLOOD Arai, S., Sahaf, B., Narasimhan, B., Chen, G. L., Jones, C. D., Lowsky, R., Shizuru, J. A., Johnston, L. J., Laport, G. G., Weng, W., Benjamin, J. E., Schaenman, J., Brown, J., Ramirez, J., Zehnder, J. L., Negrin, R. S., Miklos, D. B. 2012; 119 (25): 6145-6154

    Abstract

    B cells are involved in the pathogenesis of chronic GVHD (cGVHD). We hypothesized that prophylactic anti-B-cell therapy delivered 2 months after transplantation would decrease allogeneic donor B-cell immunity and possibly the incidence of cGVHD. Therefore, in the present study, patients with high-risk chronic lymphocytic leukemia (n = 22) and mantle-cell lymphoma (n = 13) received a total lymphoid irradiation of 80 cGy for 10 days and antithymocyte globulin 1.5 mg/kg/d for 5 days. Rituximab (375 mg/m(2)) was infused weekly on days 56, 63, 70, and 77 after transplantation. The incidence of acute GVHD was 6%. The cumulative incidence of cGVHD was 20%. Nonrelapse mortality was 3%. Rituximab treatment after allogeneic transplantation significantly reduced B-cell allogeneic immunity, with complete prevention of alloreactive H-Y Ab development in male patients with female donors (P = .01). Overall survival and freedom from progression at 4 years for chronic lymphocytic leukemia patients were 73% and 47%, respectively; for mantle-cell lymphoma patients, they were 69% and 53%, respectively.

    View details for DOI 10.1182/blood-2011-12-395970

    View details for PubMedID 22563089

  • Aggressive EBV-associated Lymphoproliferative Disorder: A Prodrome to Diffuse Large B-cell Lymphoma? APPLIED IMMUNOHISTOCHEMISTRY & MOLECULAR MORPHOLOGY Batra, R., Medeiros, B. C., Zehnder, J. L., Warnke, R. A., Natkunam, Y. 2012; 20 (3): 325-330

    Abstract

    A 19-year-old male patient presented with intermittent high fever and left cervical lymphadenopathy. The lymph node biopsy findings were interpreted as "Epstein-Barr virus (EBV)-associated lymphoproliferative disorder consistent with infectious mononucleosis." No molecular studies were performed at that time. The patient was followed without treatment. Five months later, the patient again presented with fever, lymphadenopathy, and splenomegaly. The lymph node biopsy showed features of a diffuse large B-cell lymphoma. Molecular studies on this lymph node biopsy showed a clonal EBV population, although polymerase chain reaction studies failed to reveal a clonal B-cell or T-cell population. A concurrent bone marrow biopsy showed features consistent with hemophagocytic syndrome. He had elevated ferritin, soluble interleukin-2 receptors and persistent EBV viremia. The patient responded to Rituxan for a short period with undetectable EBV levels. Subsequent right cervical lymph node, liver, and jejunal biopsies showed involvement by diffuse large B-cell lymphoma and the patient expired soon thereafter.

    View details for Web of Science ID 000303140100012

    View details for PubMedID 22505014

  • The Risk of Using a Patient's Medical Record to Determine Platelet Function Inhibition by Aspirin Therapy Urwyler, N., Stave, C., Theiler, L., Zehnder, J. WILEY-BLACKWELL. 2012: S183–S184

    View details for Web of Science ID 000302999500124

  • Comparison of Aptt and anti-Xa Activity with Patient Outcomes in a Large Cohort of Hospitalized Patients Treated with Unfractionated Heparin Jin, J., Price, E. A., Houng (Marie) Nguyen, H., Krishnan, G., Balise, R., Bowen, R., Zehnder, J. WILEY-BLACKWELL. 2012: S174–S174

    View details for Web of Science ID 000302999500099

  • Controversies in Heparin Monitoring AMERICAN JOURNAL OF HEMATOLOGY Zehnder, J., Price, E., Jin, J. 2012; 87: S137-S140

    View details for DOI 10.1002/ajh.23210

    View details for Web of Science ID 000302999500024

    View details for PubMedID 22495972

  • Safety, efficacy and biological predictors of response to sequential azacitidine and lenalidomide for elderly patients with acute myeloid leukemia LEUKEMIA Pollyea, D. A., Kohrt, H. E., Gallegos, L., Figueroa, M. E., Abdel-Wahab, O., Zhang, B., Bhattacharya, S., Zehnder, J., Liedtke, M., Gotlib, J. R., Coutre, S., Berube, C., Melnick, A., Levine, R., Mitchell, B. S., Medeiros, B. C. 2012; 26 (5): 893-901

    Abstract

    Acute myeloid leukemia (AML) is a disease of the elderly. Poor outcomes with standard therapies necessitate novel approaches. Outpatient regimens sufficiently potent and well tolerated to induce remissions and enable continuation therapy may be beneficial. In this phase-1 study, we determined the maximum tolerated dose (MTD) and the efficacy for sequential azacitidine and lenalidomide as remission induction and continuation therapy in elderly, previously untreated patients. We investigated the impact on global DNA methylation and bone marrow cytokines, and sought biological predictors of response. Eighteen patients were enrolled. The MTD was not reached. Median follow-up was 8.2 months (10.3 months for survivors). Common adverse events included fatigue, injection site reactions, constipation, nausea, pruritus and febrile neutropenia. Ten patients responded (56%), and the rate of complete remissions (CRs) or CRs with incomplete recovery of blood counts for evaluable patients was 44% (7/16). The median response duration was 6.2 months. DNA demethylation and changes in bone marrow cytokines were observed; responders had a unique cytokine profile and a trend towards lower methylation levels. Sequential azacitidine and lenalidomide was well tolerated with encouraging clinical and biological activity in previously untreated elderly AML patients. This trial is registered at ClinicalTrials.gov (NCT00890929).

    View details for DOI 10.1038/leu.2011.294

    View details for Web of Science ID 000303883500005

    View details for PubMedID 22033493

  • UNIVERSAL MINIMAL RESIDUAL DISEASE QUANTIFICATION USING CONSENSUS PRIMERS AND HIGH-THROUGHPUT IGH SEQUENCING PREDICTS POST-TRANSPLANT CLL RELAPSE BETTER THAN PATIENT-SPECIFIC PCR BMT Tandem Meeting Logan, A. C., Zhang, M. B., Faham, M., Carlton, V., Zheng, J., Moorhead, M., Willis, T., Jones, C. D., Wagar, A. N., Zehnder, J. L., Miklos, D. B. ELSEVIER SCIENCE INC. 2012: S288–S288

    View details for Web of Science ID 000299398600228

  • Tailored temozolomide therapy according to MGMT methylation status for elderly patients with acute myeloid leukemia AMERICAN JOURNAL OF HEMATOLOGY Medeiros, B. C., Kohrt, H. E., Gotlib, J., Coutre, S. E., Zhang, B., Arber, D. A., Zehnder, J. L. 2012; 87 (1): 45-50

    Abstract

    Temozolomide sensitivity is determined by methylation of the O(6)-methylguanine-DNA methyltransferase (MGMT) promoter. This study assessed whether the temozolomide dose can be tailored by MGMT promoter status and whether protracted, low-dose temozolomide can "prime" blasts in patients with unmethylated MGMT (unMGMT). Elderly patients with high-risk AML were stratified by MGMT methylation. Patients with methylated MGMT (mMGMT) received temozolomide 200 mg/m(2) orally for 7 days every 4 weeks, while patients with unMGMT received temozolomide 100 mg/m(2) orally for 14 days followed by 200 mg/m(2) orally for 7 days every 6weeks. Of 36 patients (median age, 75 years), 31 (86%) had an unMGMT promoter. Overall response rate for the entire cohort was 36%. Patients with mMGMT and unMGMT had similar response rates (40% vs. 29%). Median duration of response and overall survival (OS) among responders were 29 and 35 weeks, respectively. Induction deaths (ID) occurred in 25% of patients, mostly caused by disease progression. Hematological toxicities were the most common adverse event. Toxicities were similar between patients on conventional versus protracted schedules. High HCT-CI scores were predictive of lower CR rate, higher ID, and shorter OS, while bone marrow blast count <50% at screening predicted for improved responses. Temozolomide, dosed according to MGMT methylation status, demonstrated modest clinical activity in elderly patients with AML, especially in those presenting with fewer comorbidities and low disease burden. The trial was registered on www.ClinicalTrials.gov as #NCT00611247.

    View details for DOI 10.1002/ajh.22191

    View details for Web of Science ID 000298257700010

    View details for PubMedID 22052619

  • High-throughput VDJ sequencing for quantification of minimal residual disease in chronic lymphocytic leukemia and immune reconstitution assessment PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Logan, A. C., Gao, H., Wang, C., Sahaf, B., Jones, C. D., Marshall, E. L., Buno, I., Armstrong, R., Fire, A. Z., Weinberg, K. I., Mindrinos, M., Zehnder, J. L., Boyd, S. D., Xiao, W., Davis, R. W., Miklos, D. B. 2011; 108 (52): 21194-21199

    Abstract

    The primary cause of poor outcome following allogeneic hematopoietic cell transplantation (HCT) for chronic lymphocytic leukemia (CLL) is disease recurrence. Detection of increasing minimal residual disease (MRD) following HCT may permit early intervention to prevent clinical relapse; however, MRD quantification remains an uncommon diagnostic test because of logistical and financial barriers to widespread use. Here we describe a method for quantifying CLL MRD using widely available consensus primers for amplification of all Ig heavy chain (IGH) genes in a mixture of peripheral blood mononuclear cells, followed by high-throughput sequencing (HTS) for disease-specific IGH sequence quantification. To achieve accurate MRD quantification, we developed a systematic bioinformatic methodology to aggregate cancer clone sequence variants arising from systematic and random artifacts occurring during IGH-HTS. We then compared the sensitivity of IGH-HTS, flow cytometry, and allele-specific oligonucleotide PCR for MRD quantification in 28 samples collected from 6 CLL patients following allogeneic HCT. Using amplimer libraries generated with consensus primers from patient blood samples, we demonstrate the sensitivity of IGH-HTS with 454 pyrosequencing to be 10(-5), with a high correlation between quantification by allele-specific oligonucleotide PCR and IGH-HTS (r = 0.85). From the same dataset used to quantify MRD, IGH-HTS also allowed us to profile IGH repertoire reconstitution after HCT-information not provided by the other MRD methods. IGH-HTS using consensus primers will broaden the availability of MRD quantification in CLL and other B cell malignancies, and this approach has potential for quantitative evaluation of immune diversification following transplant and nontransplant therapies.

    View details for DOI 10.1073/pnas.1118357109

    View details for Web of Science ID 000298479900065

    View details for PubMedID 22160699

    View details for PubMedCentralID PMC3248502

  • A novel splice donor mutation in the thrombopoietin gene leads to exon 2 skipping in a Filipino family with hereditary thrombocythemia. Blood Zhang, B., Ng, D., Jones, C., Oh, S. T., Nolan, G. P., Salehi, S., Wong, W., Zehnder, J. L., Gotlib, J. 2011; 118 (26): 6988-6990

    View details for DOI 10.1182/blood-2011-10-386177

    View details for PubMedID 22194398

  • A novel splice donor mutation in the thrombopoietin gene leads to exon 2 skipping in a Filipino family with hereditary thrombocythemia BLOOD Ng, D., Jones, C., Oh, S. T., Nolan, G. P., Salehi, S., Wong, W., Zehnder, J. L., Gotlib, J. 2011; 118 (26): 6988-?

    View details for DOI 10.1182/blood-2011-10-386177

    View details for Web of Science ID 000298401000038

  • Clonally identical Hodgkin's disease develops after allogeneic hematopoietic cell transplant for CLL BONE MARROW TRANSPLANTATION Tseng, D., Jones, C. D., Anderson, M., Warnke, R., Zehnder, J. L., Miklos, D. B. 2011; 46 (12): 1576-1578

    View details for DOI 10.1038/bmt.2010.340

    View details for Web of Science ID 000298326500015

    View details for PubMedID 21258419

  • Immature T-Cell Populations in Lymph Nodes of Castleman Disease and Angioimmunoblastic T-Cell Lymphoma Suggest Alternate Sites of T-Cell Development 53rd Annual Meeting and Exposition of the American-Society-of-Hematology (ASH) Ohgami, R. S., Chun, S., Ohgami, J., Zehnder, J. L., Van de Rijn, M., Arber, D. A., Natkunam, Y., Warnke, R. AMER SOC HEMATOLOGY. 2011: 1395–96

    View details for Web of Science ID 000299597104588

  • 2-Hydroxyglutarate in IDH mutant AML: Predicting Patient Responses, Minimal Residual Disease and Correlations with Methylcytosine and Hydroxymethylcytosine Levels Pollyea, D. A., Kohrt, H. E., Zhang, B., Zehnder, J. L., Schenkein, D. P., Fantin, V., Straley, K., Vasanthakumar, A., Abdel-Wahab, O., Levine, R. L., Godley, L., Medeiros, B. C. AMER SOC HEMATOLOGY. 2011: 1073–74

    View details for Web of Science ID 000299597103375

  • Massively Parallel Immunoglobulin Gene Sequencing Provides Ultra-Sensitive Minimal Residual Disease Detection and Predicts Post-Transplant Relapse in Acute Lymphoblastic Leukemia by Three to Six Months Logan, A. C., Vashi, N., Faham, M., Carlton, V., Buno, I., Zheng, J., Moorhead, M., Klinger, M., Willis, T., Zhang, M. B., Waqar, A., Zehnder, J. L., Miklos, D. B. AMER SOC HEMATOLOGY. 2011: 1752–53

    View details for Web of Science ID 000299597106272

  • Prolonged aPTT Relative to Anti-Xa Is Associated with Increased 30-Day Mortality in Hospitalized Patients Treated with Unfractionated Heparin 53rd Annual Meeting and Exposition of the American-Society-of-Hematology (ASH) Jin, J., Price, E., Huong Nguyen, H., Krishnan, G., Balise, R., Bowen, R. A., Zehnder, J. L. AMER SOC HEMATOLOGY. 2011: 559–59

    View details for Web of Science ID 000299597101517

  • High-Throughput Immunoglobulin Gene Sequencing Quantifies Minimal Residual Disease in CLL with 10e-6 Sensitivity and Strongly Predicts Relapse After Allogeneic Hematopoietic Cell Transplantation 53rd Annual Meeting and Exposition of the American-Society-of-Hematology (ASH) Logan, A. C., Faham, M., Carlton, V., Zheng, J., Moorhead, M., Willis, T., Jones, C. D., Zhang, M. B., Waqar, A. N., Zehnder, J. L., Miklos, D. B. AMER SOC HEMATOLOGY. 2011: 1090–91

    View details for Web of Science ID 000299597103408

  • Refining the diagnosis of T-cell large granular lymphocytic leukemia by combining distinct patterns of antigen expression with T-cell clonality studies LEUKEMIA Ohgami, R. S., Ohgami, J. K., Pereira, I. T., Gitana, G., Zehnder, J. L., Arber, D. A. 2011; 25 (9): 1439-1443

    Abstract

    T-cell large granular lymphocytic (LGL) leukemia is a complex diagnosis, requiring persistent clonal expansions of LGLs, and cytopenias. Often the diagnosis is unclear as non-clonal expansions of LGLs commonly occur in reactive conditions. To better understand T-LGL leukemia, we performed a comprehensive clinicopathologic analysis of 85 patients with LGL expansions. Interestingly, distinct CD8+(dim)/CD57+ populations, seen by flow cytometry, were significantly associated with clonal T-LGL leukemia (P < 0.001) as well as neutropenia (median absolute neutrophil count (ANC) 1.45 vs 3.19 × 10(9)/l; P = 0.0017). Furthermore, cases with distinct CD8+(dim)/CD57+ populations and monoclonal T cells had even lower ANCs (median ANC 1.41 × 10(9)/l; P = 0.001) compared with cases without these dual criteria. Additionally, complete or partial loss of CD5 expression was independently associated with clonal T-LGL leukemia (P<0.001) and neutropenia (median ANC 1.41 vs 2.70 × 10(9)/l; P = 0.002). This study describes specific immunophenotypic parameters to better define clonal cases of T-LGL leukemia associated with significant neutropenia.

    View details for DOI 10.1038/leu.2011.107

    View details for Web of Science ID 000294665400008

    View details for PubMedID 21617700

  • Evaluation of a Gene Expression Microarray-based Assay to Determine Tissue Type of Origin on a Diverse Set of 49 Malignancies AMERICAN JOURNAL OF SURGICAL PATHOLOGY Beck, A. H., Rodriguez-Paris, J., Zehnder, J., Schrijver, I. 2011; 35 (7): 1030-1037

    Abstract

    The Tissue of Origin Frozen (TOO-FRZ) assay from Pathwork Diagnostics has been cleared by the Food and Drug Administration as a diagnostic study for malignancies of unknown primary. The goal of this study was to evaluate the performance of TOO-FRZ on a diverse collection of malignancies. We collected a diverse set of 49 malignancies. We classified each case into 1 of 4 groups: common morphology from a tissue type included in the TOO-FRZ assay (n=29), uncommon morphology from a tissue type included in the TOO-FRZ assay (n=10), tumor from a tissue type not included in the TOO-FRZ assay (n=3), and malignancies of unknown primary (n=7). We found strong diagnostic performance for common morphologies from tissue types on the TOO-FRZ [overall accuracy=26 of 29 (90%, 95% CI, 73% to 97%)], with perfect performance in all tissue types except gastric (0 of 2) and pancreatic (1 of 2) tissues. There was a significant decline in performance for uncommon morphologies from tissue types included in the TOO-FRZ assay [6 of 10 (60%) cases with an indeterminate result, 1 of 10 (10%) cases with an incorrect prediction, and 3 of 10 (30%) with a correct prediction] and for tumors from tissue types not included in the assay (incorrect prediction in 2 of 3 cases). For the 7 malignancies of unknown primary in our study set, the TOO-FRZ provided a likely clinically useful result in only 2 of 7 cases. These results provide an insight into the strengths and limitations of this molecular assay for the surgical pathologist, and our findings suggest future directions for research in this area.

    View details for DOI 10.1097/PAS.0b013e3182178b59

    View details for Web of Science ID 000291676200011

    View details for PubMedID 21602661

  • Sequential azacitidine and lenalidomide in elderly acute myeloid leukemia: completed results of the phase I study Pollyea, D. A., Kohrt, H. E., Gallegos, L., Zhang, B., Figueroa, M., Melnick, A., Berube, C., Coutre, S. E., Gotlib, J. R., Zehnder, J. L., Liedtke, M., Mitchell, B. S., Medeiros, B. C. AMER SOC CLINICAL ONCOLOGY. 2011

    View details for DOI 10.1200/jco.2011.29.15_suppl.6505

    View details for Web of Science ID 000208880302012

  • Mutagenesis of Varicella-Zoster Virus Glycoprotein I (gI) Identifies a Cysteine Residue Critical for gE/gI Heterodimer Formation, gI Structure, and Virulence in Skin Cells JOURNAL OF VIROLOGY Oliver, S. L., Sommer, M. H., Reichelt, M., Rajamani, J., Vlaycheva-Beisheim, L., Stamatis, S., Cheng, J., Jones, C., Zehnder, J., Arvin, A. M. 2011; 85 (9): 4095-4110

    Abstract

    Varicella-zoster virus (VZV) is the alphaherpesvirus that causes chicken pox (varicella) and shingles (zoster). The two VZV glycoproteins gE and gI form a heterodimer that mediates efficient cell-to-cell spread. Deletion of gI yields a small-plaque-phenotype virus, ΔgI virus, which is avirulent in human skin using the xenograft model of VZV pathogenesis. In the present study, 10 mutant viruses were generated to determine which residues were required for the typical function of gI. Three phosphorylation sites in the cytoplasmic domain of gI were not required for VZV virulence in vivo. Two deletion mutants mapped a gE binding region in gI to residues 105 to 125. A glycosylation site, N116, in this region did not affect virulence. Substitution of four cysteine residues highly conserved in the Alphaherpesvirinae established that C95 is required for gE/gI heterodimer formation. The C95A and Δ105-125 (with residues 105 to 125 deleted) viruses had small-plaque phenotypes with reduced replication kinetics in vitro similar to those of the ΔgI virus. The Δ105-125 virus was avirulent for human skin in vivo. In contrast, the C95A mutant replicated in vivo but with significantly reduced kinetics compared to those of the wild-type virus. In addition to abolished gE/gI heterodimer formation, gI from the C95A or the Δ105-125 mutant was not recognized by monoclonal antibodies that detect the canonical conformation of gI, demonstrating structural disruption of gI in these viruses. This alteration prevented gI incorporation into virus particles. Thus, residues C95 and 105 to 125 are critical for gI structure required for gE/gI heterodimer formation, virion incorporation, and ultimately, effective viral spread in human skin.

    View details for DOI 10.1128/JVI.02596-10

    View details for Web of Science ID 000289618600005

    View details for PubMedID 21345964

    View details for PubMedCentralID PMC3126246

  • The role of vanin-1 and oxidative stress-related pathways in distinguishing acute and chronic pediatric ITP BLOOD Zhang, B., Lo, C., Shen, L., Sood, R., Jones, C., Cusmano-Ozog, K., Park-Snyder, S., Wong, W., Jeng, M., Cowan, T., Engleman, E. G., Zehnder, J. L. 2011; 117 (17): 4569-4579

    Abstract

    Pediatric immune thrombocytopenia (ITP) is usually self-limited. However, approximately 20% of children develop chronic ITP, which can be associated with significant morbidity because of long-term immunosuppression and splenectomy in refractory cases. To explore the molecular mechanism of chronic ITP compared with acute ITP, we studied 63 pediatric patients with ITP. Gene expression analysis of whole blood revealed distinct signatures for acute and chronic ITP. Oxidative stress-related pathways were among the most significant chronic ITP-associated pathways. Overexpression of VNN1, an oxidative stress sensor in epithelial cells, was most strongly associated with progression to chronic ITP. Studies of normal persons demonstrated VNN1 expression in a variety of blood cells. Exposure of blood mononuclear cells to oxidative stress inducers elicited dramatic up-regulation of VNN1 and down-regulation of PPARγ, indicating a role for VNN1 as a peripheral blood oxidative stress sensor. Assessment of redox state by tandem mass spectrometry demonstrated statistically significant lower glutathione ratios in patients with ITP versus healthy controls; lower glutathione ratios were also seen in untreated patients with ITP compared with recently treated patients. Our work demonstrates distinct patterns of gene expression in acute and chronic ITP and implicates oxidative stress pathways in the pathogenesis of chronic pediatric ITP.

    View details for DOI 10.1182/blood-2010-09-304931

    View details for PubMedID 21325602

  • Distinct Populations of CD8+/CD57+ Large Granular Lymphocytes Are Associated with T-Cell Monoclonality and Define a Subset of Clinically and Pathologically Significant Patients: A Case-Control Study of 70 Patients with Large Granular Lymphocytic Proliferations Ohgami, R. S., Pereira, I. T., Zehnder, J. L., Arber, D. A. NATURE PUBLISHING GROUP. 2011: 312A

    View details for Web of Science ID 000291285000768

  • Distinct Populations of CD8+/CD57+ Large Granular Lymphocytes Are Associated with T-Cell Monoclonality and Define a Subset of Clinically and Pathologically Significant Patients: A Case-Control Study of 70 Patients with Large Granular Lymphocytic Proliferations. Ohgami, R. S., Pereira, I. T., Zehnder, J. L., Arber, D. A. NATURE PUBLISHING GROUP. 2011: 312A

    View details for Web of Science ID 000287282301483

  • Varicella-Zoster Virus Glycoprotein E Is a Critical Determinant of Virulence in the SCID Mouse-Human Model of Neuropathogenesis JOURNAL OF VIROLOGY Zerboni, L., Berarducci, B., Rajamani, J., Jones, C. D., Zehnder, J. L., Arvin, A. 2011; 85 (1): 98-111

    Abstract

    Varicella-zoster virus (VZV) is a neurotropic alphaherpesvirus. VZV infection of human dorsal root ganglion (DRG) xenografts in immunodeficient mice models the infection of sensory ganglia. We examined DRG infection with recombinant VZV (recombinant Oka [rOka]) and the following gE mutants: gEΔ27-90, gEΔCys, gE-AYRV, and gE-SSTT. gEΔ27-90, which lacks the gE domain that interacts with a putative receptor insulin-degrading enzyme (IDE), replicated as extensively as rOka, producing infectious virions and significant cytopathic effects within 14 days of inoculation. Since neural cells express IDE, the gE/IDE interaction was dispensable for VZV neurotropism. In contrast, gEΔCys, which lacks gE/gI heterodimer formation, was significantly impaired at early times postinfection; viral genome copy numbers increased slowly, and infectious virus production was not detected until day 28. Delayed replication was associated with impaired cell-cell spread in ganglia, similar to the phenotype of a gI deletion mutant (rOkaΔgI). However, at later time points, infection of satellite cells and other supportive nonneuronal cells resulted in extensive DRG tissue damage and cell loss such that cytopathic changes observed at day 70 were more severe than those for rOka-infected DRG. The replication of gE-AYRV, which is impaired for trans-Golgi network (TGN) localization, and the replication of gE-SSTT, which contains mutations in an acidic cluster, were equivalent to that of rOka, causing significant cytopathic effects and infectious virus production by day 14; genome copy numbers were equivalent to those of rOka. These experiments suggest that the gE interaction with cellular IDE, gE targeting to TGN sites of virion envelopment, and phosphorylation at SSTT are dispensable for VZV DRG infection, whereas the gE/gI interaction is critical for VZV neurovirulence.

    View details for DOI 10.1128/JVI.01902-10

    View details for Web of Science ID 000285095800008

    View details for PubMedID 20962081

    View details for PubMedCentralID PMC3014186

  • Development of Antibodies to Human Thrombin and Factor V in a Patient Exposed to Topical Bovine Thrombin PEDIATRIC BLOOD & CANCER Lo, C. Y., Jones, C., Glader, B., Zehnder, J. L. 2010; 55 (6): 1195-1197

    Abstract

    Bovine topical thrombin is commonly used for local hemostasis in pediatric surgery. Acquired inhibitors to coagulation factors, particularly to factor V and bovine thrombin, have been infrequently reported in the pediatric population. We report a 3-year-old male who developed a coagulopathy and clinical bleeding after cardiothoracic surgery, during which bovine topical thrombin was used for local hemostasis. Laboratory tests revealed elevated prothrombin, partial thromboplastin, and thrombin times, and a low factor V activity level. He was found to have both human-thrombin and factor V inhibitors, among the first reported cases of these combined inhibitors secondary to bovine topical thrombin. He was treated with intravenous immunoglobulin and steroids with a rapid and durable response.

    View details for DOI 10.1002/pbc.22699

    View details for PubMedID 20979176

  • Development of North American Consensus Guidelines for Medical Laboratories That Perform and Interpret Platelet Function Testing Using Light Transmission Aggregometry AMERICAN JOURNAL OF CLINICAL PATHOLOGY Hayward, C. P., Moffat, K. A., Raby, A., Israels, S., Plumhoff, E., Flynn, G., Zehnder, J. L. 2010; 134 (6): 955-963

    Abstract

    Platelet function testing is important for the diagnostic evaluation of common and rare bleeding disorders. Our study goals were to promote best practices and reduce unnecessary testing variances by developing North American guidelines on platelet function testing. Guidelines were developed by consensus for expert recommendations (minimum level for approval, 70%) that included recommendations on the evaluation and interpretation of light transmission platelet aggregometry (LTA). To assess consensus, medical opinions on recommendations were gathered from diagnostic laboratories that perform LTA, in collaboration with the Quality Management Program-Laboratory Services (QMP-LS) in Ontario, Canada (10 laboratories), and the North American Specialized Coagulation Laboratory Association (NASCOLA; 47 laboratories, 5 overlapping the QMP-LS group). Adequate consensus was achieved for all and 89% of recommendations for the QMP-LS and NASCOLA groups, respectively. The recommendations adopted provide North American laboratories with additional guidance on platelet function testing, including how to interpret LTA abnormalities.

    View details for DOI 10.1309/AJCP9V3RRVNZMKDS

    View details for Web of Science ID 000284440100013

    View details for PubMedID 21088160

  • Temozolomide In Acute Myeloid Leukemia: A MGMT Promoter Methylation Status-Based Treatment Stratification 52nd Annual Meeting and Exposition of the American-Society-of-Hematology (ASH) Medeiros, B. C., Kohrt, H. E., Rajwanshi, R., Gotlib, J., Coutre, S., Liedtke, M., Berube, C., Zhang, M., Arber, D. A., Zehnder, J. L. AMER SOC HEMATOLOGY. 2010: 1357–58

    View details for Web of Science ID 000289662203644

  • Distinct CD8(+)/CD57(+) Large Granular Lymphocytic Populations Seen by Flow Cytometry Are Associated with Monoclonal T-Cell Populations, a Retrospective Case-Control Study of Large Granular Lymphocytic Leukemia Ohgami, R. S., Zehnder, J. L., Arber, D. A. AMER SOC HEMATOLOGY. 2010: 1116

    View details for Web of Science ID 000289662203039

  • Identification of Novel LNK Mutations In Patients with Chronic Myeloproliferative Neoplasms and Related Disorders 52nd Annual Meeting and Exposition of the American-Society-of-Hematology (ASH) Oh, S. T., Zahn, J. M., Jones, C. D., Zhang, B., Loh, M. L., Kantarjian, H., Simonds, E. F., Bruggner, R. V., Abidi, P., Natsoulis, G., Bell, J., Buenrostro, J., Nolan, G. P., Zehnder, J. L., Ji, H. P., Gotlib, J. AMER SOC HEMATOLOGY. 2010: 143–44

    View details for Web of Science ID 000289662200316

  • Identification of a Novel Splice Donor Mutation In the Thrombopoietin Gene In a Philippine Family with Hereditary Thrombocythemia 52nd Annual Meeting and Exposition of the American-Society-of-Hematology (ASH) Gotlib, J., Zhang, B., Jones, C. D., Riess, J., Wong, W. B., Simonds, E. F., Hale, M. B., Abidi, P., McClung, J., Nolan, G. P., Oh, S. T., Zehnder, J. L. AMER SOC HEMATOLOGY. 2010: 1272–72

    View details for Web of Science ID 000289662203418

  • High-Throughput VDJ Sequencing Is Superior to Quantitative PCR and Flow Cytometry for the Quantification of Minimal Residual Disease In Chronic Lymphocytic Leukemia After Hematopoietic Cell Transplantation. 52nd Annual Meeting and Exposition of the American-Society-of-Hematology (ASH) Logan, A. C., Wang, C., Sahaf, B., Jones, C. D., Marshall, E. L., Buno, I., Fire, A. Z., Mindrinos, M., Zehnder, J. L., Boyd, S. D., Miklos, D. B. AMER SOC HEMATOLOGY. 2010: 552–52

    View details for Web of Science ID 000289662201393

  • Complete donor T-cell engraftment 30 days after allogeneic transplantation predicts molecular remission in high-risk chronic lymphocytic leukaemia BRITISH JOURNAL OF HAEMATOLOGY Jones, C. D., Arai, S., Lowsky, R., Tyan, D. B., Zehnder, J. L., Miklos, D. B. 2010; 150 (5): 637-639

    View details for DOI 10.1111/j.1365-2141.2010.08252.x

    View details for Web of Science ID 000281060700020

    View details for PubMedID 20528878

    View details for PubMedCentralID PMC2935248

  • Heparin-induced thrombocytopenia: Current status and diagnostic challenges AMERICAN JOURNAL OF HEMATOLOGY Otis, S. A., Zehnder, J. L. 2010; 85 (9): 700-706

    Abstract

    Heparin-induced thrombocytopenia (HIT) is a fairly common and potentially catastrophic complication of heparin therapy. Diagnosing HIT remains a challenge, as the patients at risk often have other reasons for thrombocytopenia and/or thrombosis. HIT is considered a clinicopathologic disorder whose diagnosis is generally made on the basis of both clinical criteria and the presence of "HIT antibodies" in the patient's serum or plasma. There are two basic laboratory approaches to detect HIT antibodies. The immunoassays detect antibodies based on their binding properties, whereas the functional assays detect antibodies based on their platelet-activating properties. Prompt and accurate diagnosis of HIT is imperative, as overdiagnosis exposes patients to alternative anticoagulants and their associated bleeding risks, whereas under- or delayed diagnosis leaves patients vulnerable to the thromboembolic sequelae of HIT, which can be life threatening. A critical interpretation of laboratory results by the clinician is an essential component of diagnosing HIT. This requires a keen understanding of the current concepts in the pathophysiologic mechanisms of the disease, and the application of these concepts when interpreting the results of both the functional and immunoassays. Equally important is an awareness of the strengths and weaknesses, as well as the current lack of standardization and proficiency testing, of these assays.

    View details for DOI 10.1002/ajh.21770

    View details for Web of Science ID 000281601900012

    View details for PubMedID 20665476

  • Novel mutations in the inhibitory adaptor protein LNK drive JAK-STAT signaling in patients with myeloproliferative neoplasms BLOOD Oh, S. T., Simonds, E. F., Jones, C., Hale, M. B., Goltsev, Y., Gibbs, K. D., Merker, J. D., Zehnder, J. L., Nolan, G. P., Gotlib, J. 2010; 116 (6): 988-992

    Abstract

    Dysregulated Janus kinase-signal transducer and activator of transcription (JAK-STAT) signaling due to activation of tyrosine kinases is a common feature of myeloid malignancies. Here we report the first human disease-related mutations in the adaptor protein LNK, a negative regulator of JAK-STAT signaling, in 2 patients with JAK2 V617F-negative myeloproliferative neoplasms (MPNs). One patient exhibited a 5 base-pair deletion and missense mutation leading to a premature stop codon and loss of the pleckstrin homology (PH) and Src homology 2 (SH2) domains. A second patient had a missense mutation (E208Q) in the PH domain. BaF3-MPL cells transduced with these LNK mutants displayed augmented and sustained thrombopoietin-dependent growth and signaling. Primary samples from MPN patients bearing LNK mutations exhibited aberrant JAK-STAT activation, and cytokine-responsive CD34(+) early progenitors were abnormally abundant in both patients. These findings indicate that JAK-STAT activation due to loss of LNK negative feedback regulation is a novel mechanism of MPN pathogenesis.

    View details for DOI 10.1182/blood-2010-02-270108

    View details for Web of Science ID 000280881700021

    View details for PubMedID 20404132

    View details for PubMedCentralID PMC2924231

  • Hereditary diffuse gastric cancer due to a previously undescribed CDH1 splice site mutation HUMAN PATHOLOGY Matsukuma, K. E., Mullins, F. M., Dietz, L., Zehnder, J. L., Ford, J. M., Chun, N. M., Schrijver, I. 2010; 41 (8): 1200-1203

    Abstract

    Our patient was a 52-year-old man who was diagnosed with signet ring cell gastric adenocarcinoma. An extensive family history of gastric cancer raised suspicion for hereditary diffuse gastric cancer. Sequencing of the patient's CDH1 gene revealed a novel point mutation in a strictly conserved splice site within intron 6, c.833-2 A > G. This mutation was predicted to result in loss of function due to defective RNA splicing. To characterize the pathogenic mechanism of this mutation, we amplified the patient's CDH1 gene products by reverse transcriptase polymerase chain reaction. Primers flanking the region of the mutation detected 3 distinct transcripts. In addition to the wild-type product, a larger product consistent with activation of a cryptic splice site within intron 6 and a smaller product shown to result from exon 7 skipping were detected. In summary, we have identified a novel CDH1 mutation in a large hereditary diffuse gastric cancer kindred and identified its pathogenic mechanism.

    View details for DOI 10.1016/j.humpath.2010.01.022

    View details for PubMedID 20624523

  • Future research in ITP: an ICIS consensus 3rd Intercontinental-Cooperative-ITP-Study-Group Expert Meeting Zehnder, J. L., Semple, J. W., Imbach, P., Neufeld, E. J., Buchanan, G. R., Cines, D. B. SPRINGER. 2010: S19–S23

    Abstract

    While much has been learned about the basic immunology and clinical characteristics of immune thrombocytopenia, many important questions remain with regard to pathogenesis, disease progression, identification of novel therapeutic targets and approaches, and clinical trials that rationalize and optimize use of existing therapies. The answers to these questions are likely to impact our understanding of the pathogenesis and therapeutic targets of autoimmune disease in general.

    View details for DOI 10.1007/s00277-010-0917-1

    View details for Web of Science ID 000279682100005

  • Future research in ITP: an ICIS consensus. Annals of hematology Zehnder, J. L., Semple, J. W., Imbach, P., Neufeld, E. J., Buchanan, G. R., Cines, D. B. 2010; 89: 19-23

    Abstract

    While much has been learned about the basic immunology and clinical characteristics of immune thrombocytopenia, many important questions remain with regard to pathogenesis, disease progression, identification of novel therapeutic targets and approaches, and clinical trials that rationalize and optimize use of existing therapies. The answers to these questions are likely to impact our understanding of the pathogenesis and therapeutic targets of autoimmune disease in general.

    View details for PubMedID 20309690

  • Individual Variation in the Germline Ig Gene Repertoire Inferred from Variable Region Gene Rearrangements JOURNAL OF IMMUNOLOGY Boyd, S. D., Gaeta, B. A., Jackson, K. J., Fire, A. Z., Marshall, E. L., Merker, J. D., Maniar, J. M., Zhang, L. N., Sahaf, B., Jones, C. D., Simen, B. B., Hanczaruk, B., Nguyen, K. D., Nadeau, K. C., Egholm, M., Miklos, D. B., Zehnder, J. L., Collins, A. M. 2010; 184 (12): 6986-6992

    Abstract

    Individual variation in the Ig germline gene repertoire leads to individual differences in the combinatorial diversity of the Ab repertoire, but the study of such variation has been problematic. The application of high-throughput DNA sequencing to the study of rearranged Ig genes now makes this possible. The sequencing of thousands of VDJ rearrangements from an individual, either from genomic DNA or expressed mRNA, should allow their germline IGHV, IGHD, and IGHJ repertoires to be inferred. In addition, where previously mere glimpses of diversity could be gained from sequencing studies, new large data sets should allow the rearrangement frequency of different genes and alleles to be seen with clarity. We analyzed the DNA of 108,210 human IgH chain rearrangements from 12 individuals and determined their individual IGH genotypes. The number of reportedly functional IGHV genes and allelic variants ranged from 45 to 60, principally because of variable levels of gene heterozygosity, and included 14 previously unreported IGHV polymorphisms. New polymorphisms of the IGHD3-16 and IGHJ6 genes were also seen. At heterozygous loci, remarkably different rearrangement frequencies were seen for the various IGHV alleles, and these frequencies were consistent between individuals. The specific alleles that make up an individual's Ig genotype may therefore be critical in shaping the combinatorial repertoire. The extent of genotypic variation between individuals is highlighted by an individual with aplastic anemia who appears to lack six contiguous IGHD genes on both chromosomes. These deletions significantly alter the potential expressed IGH repertoire, and possibly immune function, in this individual.

    View details for DOI 10.4049/jimmunol.1000445

    View details for Web of Science ID 000278516700047

    View details for PubMedID 20495067

  • Comprehensive and Efficient HBB Mutation Analysis for Detection of beta-Hemoglobinopathies in a Pan-Ethnic Population AMERICAN JOURNAL OF CLINICAL PATHOLOGY Chan, O. T., Westover, K. D., Dietz, L., Zehnder, J. L., Schrijver, I. 2010; 133 (5): 700-707

    Abstract

    Current methods that assay hemoglobin beta-globin chain variants can have limited clinical sensitivity when applied techniques identify only a predefined panel of mutations. Even sequence-based assays may be limited depending on which gene regions are investigated. We sought to develop a clinically practical yet inclusive molecular assay to identify beta-globin mutations in multicultural populations. We highlight the beta-globin mutation detection assay (beta-GMDA), an extensive gene sequencing assay. The polymerase chain reaction (PCR) primers are located to encompass virtually all hemoglobin beta locus (HBB) mutations. In addition, this assay is able to detect, by gap PCR, a common large deletion (Delta619 base pair), which would be missed by sequencing alone. We describe our 5-year experience with the beta-GMDA and indicate its capability for detecting homozygous, heterozygous, and compound heterozygous sequence changes, including previously unknown HBB variants. The beta-GMDA offers superior sensitivity and ease of use with comprehensive detection of HBB mutations that result in beta-globin chain variants.

    View details for DOI 10.1309/AJCP7HQ2KWGHECIO

    View details for Web of Science ID 000277476500004

    View details for PubMedID 20395516

  • Combined Use of PCR-Based TCRG and TCRB Clonality Tests on Paraffin-Embedded Skin Tissue in the Differential Diagnosis of Mycosis Fungoides and Inflammatory Dermatoses JOURNAL OF MOLECULAR DIAGNOSTICS Zhang, B., Beck, A. H., Taube, J. M., Kohler, S., Seo, K., Zwerner, J., Viakhereva, N., Sundram, U., Kim, Y. H., Schrijver, I., Arber, D. A., Zehnder, J. L. 2010; 12 (3): 320-327

    Abstract

    The distinction between mycosis fungoides (MF) and inflammatory dermatoses (ID) by clinicopathologic criteria can be challenging. There is limited information regarding the performance characteristics and utility of TCRG and TCRB clonality assays in diagnosis of MF and ID from paraffin-embedded tissue sections. In this study, PCR tests were performed with both TCRG and TCRB BIOMED-2 clonality methods followed by capillary electrophoresis and Genescan analysis using DNA samples from 35 MF and 96 ID patients with 69 and 133 paraffin-embedded specimens, respectively. Performance characteristics were determined for each test individually and in combination. TCRG and TCRB tests demonstrated identical sensitivity (64%) and specificity (84%) when analyzed as individual assays. The positive predictive value, negative predictive value, and change of posttest MF probability over a range of MF pretest probabilities were obtained. These data were used to construct an algorithm for sequential use of TCRG and TCRB. As single tests, commercially available BIOMED-2 PCR-based TCRG and TCRB clonality tests on paraffin-embedded tissue have no significant difference in terms of sensitivity and specificity. Combined use of the two tests in patients with intermediate pretest probabilities as proposed in the algorithm could improve test utility.

    View details for DOI 10.2353/jmoldx.2010.090123

    View details for Web of Science ID 000277531700009

    View details for PubMedID 20203005

    View details for PubMedCentralID PMC2860468

  • Presentation of Extranodal Natural Killer T-Cell Lymphoma, Nasal Type, With Poorly Circumscribed Erythematous Patches JOURNAL OF CLINICAL ONCOLOGY Pincus, L. B., Zehnder, J. L., Neuhaus, I. M., Andreadis, C., McCalmont, T. H. 2010; 28 (6): E94-E95

    View details for DOI 10.1200/JCO.2009.24.3428

    View details for Web of Science ID 000274653200030

    View details for PubMedID 19933911

  • Acute Myeloid Leukemia (AML) with Monosomal Karyotype Is Characterized by Absence of NPM1 and FLT3 Mutations, Worse Clinical Outcome and Usually Falls within AML with Myelodysplasia-Related Changes (MRC) Weinberg, O. K., Seetharam, M., Ren, L., Ma, L., Seo, K., Zehnder, J., Gotlib, J., Arber, D. A. NATURE PUBLISHING GROUP. 2010: 328A

    View details for Web of Science ID 000274582502144

  • Acute Myeloid Leukemia (AML) with Monosomal Karyotype Is Characterized by Absence of NPM1 and FLT3 Mutations, Worse Clinical Outcome and Usually Falls within AML with Myelodysplasia-Related Changes (MRC) Weinberg, O. K., Seetharam, M., Ren, L., Ma, L., Seo, L., Zehnder, J., Gotlib, J., Arber, D. A. NATURE PUBLISHING GROUP. 2010: 328A

    View details for Web of Science ID 000274337301479

  • Design and Evaluation of a Real-Time PCR Assay for Quantification of JAK2 V617F and Wild-Type JAK2 Transcript Levels in the Clinical Laboratory JOURNAL OF MOLECULAR DIAGNOSTICS Merker, J. D., Jones, C. D., Oh, S. T., Schrijver, I., Gotlib, J., Zehnder, J. L. 2010; 12 (1): 58-64

    Abstract

    The somatic mutation JAK2 V617F is associated with BCR-ABL1-negative myeloproliferative neoplasms. Detection of this mutation aids diagnosis of these neoplasms, and quantification of JAK2 V617F may provide a method to monitor response to therapy. For these reasons, we designed a clinical assay that uses allele-specific PCR and real-time detection with hydrolysis probes for the quantification of JAK2 V617F, wild-type JAK2, and GAPDH transcripts. Mutant and wild-type JAK2 were quantified by using external plasmid standards that contain the relevant JAK2 V617F or JAK2 sequence, respectively. We tested 55 peripheral blood specimens from patients with suspected myeloproliferative neoplasms and 55 peripheral blood specimens from patients not known to have myeloproliferative neoplasms. Low-level, nonspecific amplification was detected in reactions containing a high copy number of plasmid standards and in specimens from patients not known to have myeloproliferative neoplasms, necessitating the use of a laboratory-established mutant to wild-type cutoff. The limit of detection established by using cell line dilutions is 0.1%, and this method identified three JAK2 V617F-positive patients who were not detected by a less sensitive method. The assay characteristics and our initial evaluation indicate this method can be used for the detection and quantification of JAK2 V617F, which should be useful for diagnosis of myeloproliferative neoplasms and potentially for monitoring minimal residual disease in future trials of therapies targeted to myeloproliferative neoplasms.

    View details for DOI 10.2353/jmoldx.2010.090068

    View details for Web of Science ID 000273664100009

    View details for PubMedID 19959796

    View details for PubMedCentralID PMC2797719

  • Anti-Glycoprotein H Antibody Impairs the Pathogenicity of Varicella-Zoster Virus in Skin Xenografts in the SCID Mouse Model JOURNAL OF VIROLOGY Vleck, S. E., Oliver, S. L., Reichelt, M., Rajamani, J., Zerboni, L., Jones, C., Zehnder, J., Grose, C., Arvin, A. M. 2010; 84 (1): 141-152

    Abstract

    Varicella-zoster virus (VZV) infection is usually mild in healthy individuals but can cause severe disease in immunocompromised patients. Prophylaxis with varicella-zoster immunoglobulin can reduce the severity of VZV if given shortly after exposure. Glycoprotein H (gH) is a highly conserved herpesvirus protein with functions in virus entry and cell-cell spread and is a target of neutralizing antibodies. The anti-gH monoclonal antibody (MAb) 206 neutralizes VZV in vitro. To determine the requirement for gH in VZV pathogenesis in vivo, MAb 206 was administered to SCID mice with human skin xenografts inoculated with VZV. Anti-gH antibody given at 6 h postinfection significantly reduced the frequency of skin xenograft infection by 42%. Virus titers, genome copies, and lesion size were decreased in xenografts that became infected. In contrast, administering anti-gH antibody at 4 days postinfection suppressed VZV replication but did not reduce the frequency of infection. The neutralizing anti-gH MAb 206 blocked virus entry, cell fusion, or both in skin in vivo. In vitro, MAb 206 bound to plasma membranes and to surface virus particles. Antibody was internalized into vacuoles within infected cells, associated with intracellular virus particles, and colocalized with markers for early endosomes and multivesicular bodies but not the trans-Golgi network. MAb 206 blocked spread, altered intracellular trafficking of gH, and bound to surface VZV particles, which might facilitate their uptake and targeting for degradation. As a consequence, antibody interference with gH function would likely prevent or significantly reduce VZV replication in skin during primary or recurrent infection.

    View details for DOI 10.1128/JVI.01338-09

    View details for Web of Science ID 000272564300013

    View details for PubMedID 19828615

    View details for PubMedCentralID PMC2798403

  • Measurement and Clinical Monitoring of Human Lymphocyte Clonality by Massively Parallel V-D-J Pyrosequencing SCIENCE TRANSLATIONAL MEDICINE Boyd, S. D., Marshall, E. L., Merker, J. D., Maniar, J. M., Zhang, L. N., Sahaf, B., Jones, C. D., Simen, B. B., Hanczaruk, B., Nguyen, K. D., Nadeau, K. C., Egholm, M., Miklos, D. B., Zehnder, J. L., Fire, A. Z. 2009; 1 (12)

    Abstract

    The complex repertoire of immune receptors generated by B and T cells enables recognition of diverse threats to the host organism. In this work, we show that massively parallel DNA sequencing of rearranged immune receptor loci can provide direct detection and tracking of immune diversity and expanded clonal lymphocyte populations in physiological and pathological contexts. DNA was isolated from blood and tissue samples, a series of redundant primers was used to amplify diverse DNA rearrangements, and the resulting mixtures of barcoded amplicons were sequenced using long-read ultra deep sequencing. Individual DNA molecules were then characterized on the basis of DNA segments that had been joined to make a functional (or nonfunctional) immune effector. Current experimental designs can accommodate up to 150 samples in a single sequence run, with the depth of sequencing sufficient to identify stable and dynamic aspects of the immune repertoire in both normal and diseased circumstances. These data provide a high-resolution picture of immune spectra in normal individuals and in patients with hematological malignancies, illuminating, in the latter case, both the initial behavior of clonal tumor populations and the later suppression or re-emergence of such populations after treatment.

    View details for DOI 10.1126/scitranslmed.3000540

    View details for Web of Science ID 000277263200001

    View details for PubMedID 20161664

    View details for PubMedCentralID PMC2819115

  • Increased VNN1/PPARG Gene Expression Ratio Is Correlated with Developing Chronic ITP and Oxidative Stress Exposure to PBMC in Vitro 51st Annual Meeting and Exposition of the American-Society-of-Hematology Zhang, B., Shen, L., Jeng, M., Jones, C., Wong, W., Engleman, E. E., Zehnder, J. L. AMER SOC HEMATOLOGY. 2009: 368–68

    View details for Web of Science ID 000272725801073

  • Critical Values in the Coagulation Laboratory: Results of a Survey of the North American Specialized Coagulation Laboratory Association Pai, M., Moffat, K. A., Plumhoff, E., Adcock, D. M., Zehnder, J. L., Hayward, C. M. AMER SOC HEMATOLOGY. 2009: 576

    View details for Web of Science ID 000272725801595

  • AML Patients with Monosomal Karyotype Are Characterized by Absence of NPM1 and FLT3 Mutations and Worse Clinical Outcome. Seetharam, M., Weinberg, O. K., Ren, L., Ma, L., Seo, K., Zehnder, J. L., Gotlib, J. R., Arber, D. A. AMER SOC HEMATOLOGY. 2009: 1035

    View details for Web of Science ID 000272725803193

  • Clinical Assay Design for Detection of CEBPA Mutations in Patients with Acute Myeloid Leukemia Deucher, A., Chiang, T., Seo, K., Arber, D. A., Zehnder, J., Schrijver, I. AMER SOC INVESTIGATIVE PATHOLOGY, INC. 2009: 627

    View details for Web of Science ID 000271681400060

  • Interim results of protracted low doses of temozolomide in high-risk acute myeloid leukemia 45th Annual Meeting of the American-Society-of-Clinical-Oncology (ASCO) Medeiros, B. C., Gotlib, J. R., Coutre, S. E., Jones, C., Khan, S. A., Rajwanshi, R., Rajwanshi, R., Zehnder, J., Zehnder, J. AMER SOC CLINICAL ONCOLOGY. 2009

    View details for Web of Science ID 000276606605042

  • Baseline status of O (6)-methylguanine-DNA-methyltransferase (MGMT) promoter methylation and effect of protracted doses of temozolomide on MGMT in acute leukemia. Medeiros, B., Khan, S., Rajwanshi, R., Jones, C., Zehnder, J. AMER ASSOC CANCER RESEARCH. 2009

    View details for Web of Science ID 000209702702111

  • Glycogen synthase kinase 3 beta missplicing contributes to leukemia stem cell generation PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Abrahamsson, A. E., Geron, I., Gotlib, J., Dao, K. T., Barroga, C. F., Newton, I. G., Giles, F. J., Durocher, J., Creusot, R. S., Karimi, M., Jones, C., Zehnder, J. L., Keating, A., Negrin, R. S., Weissman, I. L., Jamieson, C. H. 2009; 106 (10): 3925-3929

    Abstract

    Recent evidence suggests that a rare population of self-renewing cancer stem cells (CSC) is responsible for cancer progression and therapeutic resistance. Chronic myeloid leukemia (CML) represents an important paradigm for understanding the genetic and epigenetic events involved in CSC production. CML progresses from a chronic phase (CP) in hematopoietic stem cells (HSC) that harbor the BCR-ABL translocation, to blast crisis (BC), characterized by aberrant activation of beta-catenin within granulocyte-macrophage progenitors (GMP). A major barrier to predicting and inhibiting blast crisis transformation has been the identification of mechanisms driving beta-catenin activation. Here we show that BC CML myeloid progenitors, in particular GMP, serially transplant leukemia in immunocompromised mice and thus are enriched for leukemia stem cells (LSC). Notably, cDNA sequencing of Wnt/beta-catenin pathway regulatory genes, including adenomatous polyposis coli, GSK3beta, axin 1, beta-catenin, lymphoid enhancer factor-1, cyclin D1, and c-myc, revealed a novel in-frame splice deletion of the GSK3beta kinase domain in the GMP of BC samples that was not detectable by sequencing in blasts or normal progenitors. Moreover, BC CML progenitors with misspliced GSK3beta have enhanced beta-catenin expression as well as serial engraftment potential while reintroduction of full-length GSK3beta reduces both in vitro replating and leukemic engraftment. We propose that CP CML is initiated by BCR-ABL expression in an HSC clone but that progression to BC may include missplicing of GSK3beta in GMP LSC, enabling unphosphorylated beta-catenin to participate in LSC self-renewal. Missplicing of GSK3beta represents a unique mechanism for the emergence of BC CML LSC and might provide a novel diagnostic and therapeutic target.

    View details for DOI 10.1073/pnas.0900189106

    View details for Web of Science ID 000264036900051

    View details for PubMedID 19237556

    View details for PubMedCentralID PMC2646624

  • Clinical characterization of acute myeloid leukemia with myelodysplasia-related changes as defined by the 2008 WHO classification system 50th Annual Meeting of the American-Society-of-Hematology/ASH/ASCO Joint Symposium Weinberg, O. K., Seetharam, M., Ren, L., Seo, K., Ma, L., Merker, J. D., Gotlib, J., Zehnder, J. L., Arber, D. A. AMER SOC HEMATOLOGY. 2009: 1906–8

    Abstract

    Although some studies have validated the 2001 World Health Organization (WHO) classification of acute myeloid leukemia (AML), including the importance of multilineage dysplasia, others have suggested that multilineage dysplasia correlates with unfavorable cytogenetics but has no independent impact on prognosis. In 2008, the revised WHO classification has expanded this category into "AML with myelodysplasia-related changes" (AML-MRC). We evaluated the clinical, pathologic, cytogenetic, and molecular features of 100 AML patients using the 2008 WHO criteria. Patients underwent genetic screening for NPM1, FLT3-ITD, FLT3-D835, and CEBPA mutations. Compared with patients with AML, not otherwise specified, patients with AML-MRC were significantly older (P= .014), presented with a lower hemoglobin (P= .044), more frequently expressed CD14 (P= .048), and exhibited a decreased frequency of CEBPA mutations (P= .001). Multivariate analysis indicated that patients with AML-MRC had a significantly worse overall survival, progression-free survival, and complete response compared with AML-not otherwise specified (all P< .001). These data support the clinical, morphologic, and cytogenetic criteria for this 2008 WHO AML category.

    View details for DOI 10.1182/blood-2008-10-182782

    View details for Web of Science ID 000263723700007

    View details for PubMedID 19131546

  • Laboratory Practice Guidelines for Detecting and Reporting BCR-ABL Drug Resistance Mutations in Chronic Myelogenous Leukemia and Acute Lymphoblastic Leukemia A Report of the Association for Molecular Pathology JOURNAL OF MOLECULAR DIAGNOSTICS Jones, D., Kamel-Reid, S., Bahler, D., Dong, H., Elenitoba-Johnson, K., Press, R., Quigley, N., Rothberg, P., Sabath, D., Viswanatha, D., Weck, K., Zehnder, J. 2009; 11 (1): 4-11

    Abstract

    The BCR-ABL tyrosine kinase produced by the t(9;22)(q34;q11) translocation, also known as the Philadelphia chromosome, is the initiating event in chronic myeloid leukemia (CML) and Ph+ acute lymphoblastic leukemia (ALL). Targeting of BCR-ABL with tyrosine kinase inhibitors (TKIs) has resulted in rapid clinical responses in the vast majority of patients with CML and Philadelphia chromosome+ ALL. However, long-term use of TKIs occasionally results in emergence of therapy resistance, in part through the selection of clones with mutations in the BCR-ABL kinase domain. We present here an overview of the current practice in monitoring for such mutations, including the methods used, the clinical and laboratory criteria for triggering mutational analysis, and the guidelines for reporting BCR-ABL mutations. We also present a proposal for a public database for correlating mutational status with in vitro and in vivo responses to different TKIs to aid in the interpretation of mutation studies.

    View details for DOI 10.2353/jmoldx.2009.080095

    View details for PubMedID 19095773

  • High-Throughput Immune Receptor Sequencing for Diagnosis, Prognosis and Monitoring of Lymphoid Malignancies Boyd, S. D., Merker, J. D., Zehnder, J. L., Fire, A. Z. NATURE PUBLISHING GROUP. 2009: 368A

    View details for Web of Science ID 000262486301670

  • High-Throughput Immune Receptor Sequencing for Diagnosis, Prognosis and Monitoring of Lymphoid Malignancies Boyd, S. D., Merker, J. D., Zehnder, J. L., Fire, A. Z. NATURE PUBLISHING GROUP. 2009: 368A

    View details for Web of Science ID 000262371501669

  • Acute Myeloid Leukemia with Myelodysplasia-Related Changes as Defined by the 2008 WHO Classification System Weinberg, O. K., Seetharam, M., Ren, L., Ma, L., Seo, K., Merker, J., Gotlib, J., Zehnder, J., Arber, D. A. NATURE PUBLISHING GROUP. 2009: 291A

    View details for Web of Science ID 000262371501315

  • Acute Myeloid Leukemia with Myelodysplasia-Related Changes as Defined by the 2008 WHO Classification System 98th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Weinberg, O. K., Seetharam, M., Ren, L., Ma, L., Seo, K., Merker, J., Gotlib, J., Zehnder, J., Arber, D. A. NATURE PUBLISHING GROUP. 2009: 291A–291A

    View details for Web of Science ID 000262486301315

  • Molecular stratification of patients with normal karyotype acute myeloid leukemia based on initial assessment of FLT3-internal tandem duplication status at first complete remission LEUKEMIA & LYMPHOMA Medeiros, B. C., Gotlib, J., Zehnder, J. 2009; 50 (5): 851-853

    View details for DOI 10.1080/10428190902838400

    View details for Web of Science ID 000266201800029

    View details for PubMedID 19452323

  • Clinical Characterization of Acute Myeloid Leukemia with Myelodysplasia-Related Changes as Defined by the 2008 WHO Classification System 50th Annual Meeting of the American-Society-of-Hematology/ASH/ASCO Joint Symposium Weinberg, O. K., Seetharam, M., Ren, L., Ma, L., Seo, K., Merker, J. D., Gotlib, J. R., Zehnder, J. L., Arber, D. A. AMER SOC HEMATOLOGY. 2008: 341–42

    View details for Web of Science ID 000262104701146

  • Complete Donor Chimerism Predicts Molecular Remission in High Risk CLL Following Nonmyeloablative Transplantation Jones, C., Letzinger, R., Zehnder, J. L., Miklos, D. AMER SOC HEMATOLOGY. 2008: 1127–28

    View details for Web of Science ID 000262104703719

  • Tailored Temozolomide Therapy for Elderly Patients with Acute Myeloid Leukemia. Medeiros, B. C., Gotlib, J. R., Coutre, S. E., Mignea, A., Zehnder, J. AMER SOC HEMATOLOGY. 2008: 351

    View details for Web of Science ID 000262104701178

  • Elevated Vanin 1 and Advillin Expression Is Associated with Progression to Chronic ITP in Children 50th Annual Meeting of the American-Society-of-Hematology/ASH/ASCO Joint Symposium Zhang, B., Sood, R., Jones, C., Wong, W., Jeng, M., Zehnder, J. L. AMER SOC HEMATOLOGY. 2008: 152–53

    View details for Web of Science ID 000262104700398

  • High-Throughput Sequencing for Diagnosis, Prognosis and Monitoring of Lymphoid Malignancies. 50th Annual Meeting of the American-Society-of-Hematology/ASH/ASCO Joint Symposium Boyd, S. D., Merker, J. D., Zehnder, J. L., Fire, A. Z. AMER SOC HEMATOLOGY. 2008: 1294–94

    View details for Web of Science ID 000262104704397

  • Design and Validation of a Real-Time PCR Assay for Quantification of JAK2 V617F and Wild-type JAK2 Transcript Levels 14th Annual Meeting of the Association-for-Molecular-Pathology Merker, J. D., Jones, C. D., Oh, S. T., Khan, S., Schrijver, I., Gotlib, J., Zehnder, J. L. ELSEVIER SCIENCE INC. 2008: 581–81

    View details for Web of Science ID 000260533600084

  • Superficial venous thrombosis associated with congenital absence of the inferior vena cava and previous episode of deep venous thrombosis AMERICAN JOURNAL OF HEMATOLOGY Evanchuk, D. M., Von Gehr, A., Zehnder, J. L. 2008; 83 (3): 250-252

    Abstract

    Congenital malformations of the inferior vena cava (IVC) are uncommon and may be associated with an increased risk of venous thrombosis. We report the case of a man with congenital absence of the IVC and remote history of deep venous thrombosis who now presents with severe abdominal wall superficial thrombophlebitis. To our knowledge, this is the first report of a patient with IVC absence who has developed both deep and superficial venous thromboses.

    View details for DOI 10.1002/ajh.21089

    View details for Web of Science ID 000253559700018

    View details for PubMedID 17918250

  • Interlaboratory performance of a microarray-based gene expression test to determine tissue of origin in poorly differentiated and undifferentiated cancers JOURNAL OF MOLECULAR DIAGNOSTICS Dumur, C. I., Lyons-Weiler, M., Sciulli, C., Garrett, C. T., Schrijver, I., Holley, T. K., Rodriguez-Paris, J., Pollack, J. R., Zehnder, J. L., Price, M., Hagenkord, J. M., Rigl, C. T., Buturovic, L. J., Anderson, G. G., Monzon, F. A. 2008; 10 (1): 67-77

    Abstract

    Clinical workup of metastatic malignancies of unknown origin is often arduous and expensive and is reported to be unsuccessful in 30 to 60% of cases. Accurate classification of uncertain primary cancers may improve with microarray-based gene expression testing. We evaluated the analytical performance characteristics of the Pathwork tissue of origin test, which uses expression signals from 1668 probe sets in a gene expression microarray, to quantify the similarity of tumor specimens to 15 known tissues of origin. Sixty archived tissue specimens from poorly and undifferentiated tumors (metastatic and primary) were analyzed at four laboratories representing a wide range of preanalytical conditions (eg, personnel, reagents, instrumentation, and protocols). Cross-laboratory comparisons showed highly reproducible results between laboratories, with correlation coefficients between 0.95 to 0.97 for measurements of similarity scores, and an average 93.8% overall concordance between laboratories in terms of final tissue calls. Bland-Altman plots (mean coefficients of reproducibility of 32.48+/-3.97) and kappa statistics (kappa >0.86) also indicated a high level of agreement between laboratories. We conclude that the Pathwork tissue of origin test is a robust assay that produces consistent results in diverse laboratory conditions reflecting the preanalytical variations found in the everyday clinical practice of molecular diagnostics laboratories.

    View details for DOI 10.2353/jmoldx.2008.070099

    View details for Web of Science ID 000252521200009

    View details for PubMedID 18083688

    View details for PubMedCentralID PMC2175545

  • Significance of NPM1 and FLT3 mutations in acute myeloid leukemia with multilineage dysplasia: Does NPM1 identify a lower risk group? Weinberg, O., Merker, J., Beck, A., Seetharam, M., Gotlib, J., Zehnder, J., Arber, D. A. NATURE PUBLISHING GROUP. 2008: 281A

    View details for Web of Science ID 000252181101411

  • Significance of NPM1 and FLT3 mutations in acute myeloid leukemia with multilineage dysplasia: Does NPM1 identify a lower risk group? 97th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Weinberg, O., Merker, J., Beck, A., Seetharam, M., Gotlib, J., Zehnder, J., Arber, D. A. NATURE PUBLISHING GROUP. 2008: 281A–281A

    View details for Web of Science ID 000252180201412

  • Gene expression and pathway analysis of immune thrombocytopenic purpura BRITISH JOURNAL OF HAEMATOLOGY Sood, R., Wong, W., Gotlib, J., Jeng, M., Zehnder, J. L. 2008; 140 (1): 99-103

    Abstract

    A global expression profile of peripheral blood from patients with immune thrombocytopenic purpura (ITP) was performed that identified an ITP-specific signature, which also included interferon (IFN)-induced genes. Several genes correlated with ITP have been shown to be associated with expression signatures in systemic lupus erythematosis and rheumatoid arthritis, indicating an overlap with other autoimmune disorders. Pathway analysis demonstrated that IFN signalling, death receptor and protein ubiquitination pathways were associated with ITP. These results provide the first glimpse of the genes and pathways consistently aberrant in ITP, identifying new targets for investigations of pathogenesis and treatment of ITP.

    View details for DOI 10.1111/j.1365-2141.2007.06881.x

    View details for PubMedID 18005267

  • Laboratory testing for heparin-induced thrombocytopenia is inconsistent in North America: A survey of North American specialized coagulation laboratories THROMBOSIS AND HAEMOSTASIS Price, E. A., Hayward, C. R., Moffat, K. A., Moore, J. C., Warkentin, T. E., Zehnder, J. L. 2007; 98 (6): 1357-1361

    Abstract

    Heparin-induced thrombocytopenia (HIT) is a serious complication of heparin therapy. As HIT is considered a clinico-pathologic entity, laboratory practices have an important role in diagnosing or excluding HIT. It was the objective of this study to assess the current status of laboratory testing for HIT in North America. An online survey consisting of 67 questions related to laboratory testing for HIT was developed by the North American Specialized Coagulation Laboratory Association (NASCOLA), and distributed to its 59 members. The survey included queries about HIT test ordering practices, HIT immunoassay and activation assays performed, and reporting practices. Data was collected from the 44 NASCOLA laboratories who responded. Of these sites, 88% performed immunoassays for HIT, commonly using commercial assays. However, sites varied in practices related to use of controls, immunoglobulin class of antibody detected, and in result interpretation and reporting. Platelet activation assays for HIT were performed by 36% of sites, commonly using assays of serotonin release (50%) or heparin-induced platelet aggregation (43%). Sites varied in the use of washed platelets versus platelet-rich plasma, controls, and heparin concentrations. This survey is the first comprehensive assessment of patterns of practice in HIT testing among diagnostic coagulation laboratories in North America. We observed site-specific variability of testing methods encompassing all stages of testing, including pre-analytical handling, testing methodologies, and result interpretation and reporting. The variability in HIT platelet activation assay methods among institutions indicates a need for proficiency testing to assess assay performance, and for consensus guidelines on HIT laboratory testing.

    View details for DOI 10.1160/TH07-06-0401

    View details for Web of Science ID 000251687400031

    View details for PubMedID 18064336

  • Effect of Ginkgo biloba (EGb 761) aggregation and platelet and aspirin on platelet analysis among older adults at risk of cardiovascular disease: a randomized clinical trial BLOOD COAGULATION & FIBRINOLYSIS Gardner, C. D., Zehnder, J. L., Rigby, A. J., Nicholus, J. R., Farquhar, J. W. 2007; 18 (8): 787-793

    Abstract

    Several case reports have implicated Ginkgo biloba in clinically adverse bleeding disorders. Ginkgo biloba has been reported to increase pain-free walking distance among patients with peripheral artery disease (PAD). Standard PAD therapy includes 325 mg/day aspirin. The objective of this study was to examine potential adverse effects of concomitant aspirin and Ginkgo biloba on platelet function. Ginkgo biloba (EGb 761, 300 mg/day) was compared with placebo for effects on measures of platelet aggregation among adults consuming 325 mg/day aspirin in a randomized, double-blind, placebo-controlled, parallel design trial of 4-week duration. Participants were adults, age 69 +/- 10 years, with PAD or risk factors for cardiovascular disease. Outcome measures included platelet function analysis (PFA-100 analyzer) using ADP as an agonist (n = 26 placebo; n = 29 ginkgo), and platelet aggregation using ADP, epinephrine, collagen and ristocetin as agonists (n = 21 placebo; n = 23 ginkgo). Participants kept daily logs of bleeding or bruising episodes. There were no clinically or statistically significant differences between treatment groups for any agonists, for either PFA-100 analysis or platelet aggregation. Reports of bleeding or bruising were infrequent and similar for both study groups. In conclusion, in older adults with PAD or cardiovascular disease risk, a relatively high dose of Ginkgo biloba combined with 325 mg/day daily aspirin did not have a clinically or statistically detectable impact on indices of coagulation examined over 4 weeks, compared with the effect of aspirin alone. No adverse bleeding events were observed, although the trial was limited to a small sample size.

    View details for Web of Science ID 000251271200012

    View details for PubMedID 17982321

  • Missplicing of glycogen synthase kinase 3 beta: A potential mechanism of blast crisis chronic myeloid leukemia stem cell generation 49th Annual Meeting of the American-Society-of-Hematology Abrahamsson, A., Geron, I., Gotlib, J., Dao, K., Giles, F., Newton, I., Kavaterchik, E., Durocher, J., Creusot, R., Karimi, M., Jones, C., Zehnder, J., Keating, A., Negrin, R., Weissman, I. L., Jamieson, C. H. AMER SOC HEMATOLOGY. 2007: 238A–239A

    View details for Web of Science ID 000251100801044

  • Large granular lymphocyte leukemia: Clonality reconsidered. Witteles, W., Zhang, B., Schrijver, I., Arber, D., Gotlib, J., Zehnder, J. AMER SOC HEMATOLOGY. 2007: 912A

    View details for Web of Science ID 000251100804139

  • T-cell clonality analysis in biopsy specimens from two different skin sites shows high specificity in the diagnosis of patients with suggested mycosis fungoides 43rd Annual Meeting of the American-Society-of-Dermatopathology Thurber, S. E., Zhang, B., Kim, Y. H., Schrijver, I., Zehnder, J., Kohler, S. MOSBY-ELSEVIER. 2007: 782–90

    Abstract

    The diagnosis of mycosis fungoides (MF) is often difficult because of significant clinical and histopathologic overlap with inflammatory dermatoses. T-cell receptor (TCR)gamma chain rearrangement by polymerase chain reaction (PCR) (TCR-PCR) is a helpful adjuvant tool in this setting, but several of the inflammatory dermatoses in the differential diagnosis of MF may contain a clonal T-cell proliferation.We examined whether analysis for T-cell clonality and comparison of the clones with the standardized BIOMED-2 PCR multiplex primers for the TCRgamma chain from two anatomically distinct skin sites improves diagnostic accuracy.We examined two biopsy specimens each from 10 patients with unequivocal MF, from 18 patients with inflammatory dermatoses, and from 18 patients who could initially not be definitively given a diagnosis based on clinical and histopathologic criteria.Eight of 10 patients with unequivocal MF had an identical clone in both biopsy specimens. Two of 18 patients with inflammatory dermatoses were found to have a clone in one of the biopsy specimens. On further follow-up of the 18 patients with morphologically nondiagnostic biopsy specimens, 13 of 18 were later confirmed to have MF and 5 of 18 had inflammatory dermatoses. Eleven of 13 patients with MF had an identical clone in both biopsy specimens; two of 13 had a polyclonal amplification pattern in both biopsy specimens. Four of 5 patients with inflammatory dermatoses had no clone in either biopsy specimen. One patient with an inflammatory dermatosis had an identical clone in both specimens. The sensitivity of TCR-PCR analysis to evaluate for an identical clone at different anatomic skin sites (dual TCR-PCR) is 82.6% and the specificity is 95.7%.The number of patients in the study group was limited.These data suggest that dual TCR-PCR is a very promising technique with high specificity in distinguishing MF from inflammatory dermatoses.

    View details for DOI 10.1016/j.jaad.2007.06.004

    View details for Web of Science ID 000250387100004

    View details for PubMedID 17646032

  • Analysis of dystrophin gene alterations in Duchenne and Becker muscular dystrophy patients using a custom comparative genomic hybridization array Howell, W. D., Scheffer-Wong, A., Tsang, P., Zehnder, J. L., Schrijver, I. AMER SOC INVESTIGATIVE PATHOLOGY, INC. 2007: 650

    View details for Web of Science ID 000250849300017

  • Identification of an intronic SNP leading to allele dropout during validation of a CDH1 sequencing assay: Implications for designing PCR-based assays Mullins, F. M., Dietz, L., Lay, M., Zehnder, J. L., Ford, J., Chun, N., Schrijver, I. AMER SOC INVESTIGATIVE PATHOLOGY, INC. 2007: 656–57

    View details for Web of Science ID 000250849300044

  • Analytical performance of a microarray-based gene expression test to determine tissue of origin in uncertain primary cancers Dumur, C. I., Lyons-Weiler, M., Garrett, C. T., Holley, T. K., Schrijver, I., Rodriquez-Paris, J., Pollack, J., Zehnder, J. L., Sciulli, C. M., Hagenkord, J. M., Price, M., Rigl, C. T., Buturovic, L. J., Anderson, G., Monzon-Bordonaba, F. A. AMER SOC INVESTIGATIVE PATHOLOGY, INC. 2007: 685

    View details for Web of Science ID 000250849300166

  • Identification of an intronic single nucleotide polymorphism leading to allele dropout during validation of a CDH1 sequencing assay: implications for designing polymerase chain reaction-based assays GENETICS IN MEDICINE Mullins, F. M., Dietz, L., Lay, M., Zehnder, J. L., Ford, J., Chun, N., Schrijver, I. 2007; 9 (11): 752-760

    Abstract

    The CDH1 gene encodes the cell adhesion protein E-cadherin, and CDH1 germline mutations are associated with hereditary diffuse gastric cancer. Identification of individuals at high risk of developing diffuse gastric cancer affords the opportunity for endoscopic screening or elective prophylactic gastrectomy. We set out to develop a CDH1 sequencing assay for clinical use.All exons of the CDH1 gene were amplified and sequenced with published and modified primers.While validating the assay, we encountered a case in which a single nucleotide polymorphism located in intron 15 led to allele dropout and therefore to a false-negative result. The polymorphism leading to allele dropout was located within a primer-binding sequence, five bases away from the 3' end of the primer. A frameshift mutation in exon 15 was detected by an alternative primer that binds away from the polymorphic site. A search of the University of California Santa Cruz single nucleotide polymorphism database revealed other polymorphisms located within primer-binding sites. A total of 12 primers in nine primer sets were modified to minimize allele dropout risk.The approach of designing primers to avoid known single nucleotide polymorphisms can be generalized to the design of any polymerase chain reaction-based assay and should be employed whenever possible.

    View details for DOI 10.1097/GIM.0b013e318159a369

    View details for PubMedID 18007144

  • Antithrombotic therapy and pregnancy: consensus report and recommendations for prevention and treatment of venous thromboembolism and adverse pregnancy outcomes AMERICAN JOURNAL OF OBSTETRICS AND GYNECOLOGY Duhl, A. J., Paidas, M. J., Ural, S. H., Branch, W., Casele, H., Cox-Gill, J., Hamersley, S. L., Hyers, T. M., Katz, V., Kuhlmann, R., Nutescu, E. A., Thorp, J. A., Zehnder, J. L. 2007; 197 (5)

    Abstract

    Venous thromboembolism and adverse pregnancy outcomes are potential complications of pregnancy. Numerous studies have evaluated both the risk factors for and the prevention and management of these outcomes in pregnant patients. This consensus group was convened to provide concise recommendations, based on the currently available literature, regarding the use of antithrombotic therapy in pregnant patients at risk for venous thromboembolic events and adverse pregnancy outcomes.

    View details for DOI 10.1016/j.ajog.2007.04.022

    View details for Web of Science ID 000250915500004

    View details for PubMedID 17980177

  • Aberrant infection and persistence of varicella-zoster virus in human dorsal root ganglia in vivo absence of glycoprotein I PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Zerboni, L., Reichelt, M., Jones, C. D., Zehnder, J. L., Ito, H., Arvin, A. M. 2007; 104 (35): 14086-14091

    Abstract

    Varicella-zoster virus (VZV) causes varicella, establishes latency in sensory ganglia, and reactivates as herpes zoster. Human dorsal root ganglia (DRGs) xenografts in immunodeficient mice provide a model for evaluating VZV neuropathogenesis. Our investigation of the role of glycoprotein I (gI), which is dispensable in vitro, examines the functions of a VZV gene product during infection of human neural cells in vivo. Whereas intact recombinant Oka (rOka) initiated a short replicative phase followed by persistence in DRGs, the gI deletion mutant, rOkaDeltagI, showed prolonged replication with no transition to persistence up to 70 days after infection. Only a few varicella-zoster nucleocapsids and cytoplasmic virions were observed in neurons, and the major VZV glycoprotein, gE, was retained in the rough endoplasmic reticulum in the absence of gI. VZV neurotropism was not disrupted when DRG xenografts were infected with rOka mutants lacking gI promoter elements that bind cellular transactivators, specificity factor 1 (Sp1) and upstream stimulatory factor (USF). Because gI is essential and Sp1 and USF contribute to VZV pathogenesis in skin and T cells in vivo, these DRG experiments indicate that the genetic requirements for VZV infection are less stringent in neural cells in vivo. The observations demonstrate that gI is important for VZV neurotropism and suggest that a strategy to reduce neurovirulence by deleting gI could prolong active infection in human DRGs.

    View details for DOI 10.1073/pnas.0706023104

    View details for Web of Science ID 000249187500042

    View details for PubMedID 17709745

    View details for PubMedCentralID PMC1955823

  • Testing for maternal cell contamination in prenatal samples - A comprehensive survey of current diagnostic practices in 35 molecular diagnostic laboratories JOURNAL OF MOLECULAR DIAGNOSTICS Schrijver, I., Cherny, S. C., Zehnder, J. L. 2007; 9 (3): 394-400

    Abstract

    The potential presence of maternal cell contamination (MCC) in chorionic villus or amniotic fluid samples poses a serious preanalytical risk for prenatal misdiagnosis. The aim of this study was to identify current diagnostic practices in the absence of comprehensive practice guidelines. Thirty-five clinical molecular laboratories that conduct prenatal testing agreed to participate in a clinical practice survey. The survey included questions about sample requirements, test indications, assay type, test performance and limitations, criteria and management of uninformative test results, reporting, and billing. Sixty percent of participating laboratories performed testing on direct and cultured amniotic fluid, whereas forty percent tested cultured cells only. Most also accepted chorionic villus samples. Although MCC testing of fetal samples is recommended in guidelines by the American College of Medical Genetics, only 60% of surveyed laboratories performed it without exception. Commercially available assays were used by 75% of participating laboratories, and at least five identity markers were evaluated at 87% of the laboratories. The reported lower limit of MCC detection ranged from 1 to 20% but was not determined in all laboratories. MCC testing was performed in the majority of molecular diagnostic laboratories, but guidelines for standardization are needed to ensure optimal and accurate prenatal patient care.

    View details for DOI 10.2353/jmoldx.2007.070017

    View details for Web of Science ID 000247691200015

    View details for PubMedID 17591939

    View details for PubMedCentralID PMC1899411

  • Recalcitrant cutaneous langerhans cell histiocytosis in a child with T-cell acute lymphoblastic leukemia Khuu, P., Bruckner, A., Wei, M., Lacavo, N., George, T. J., Zehnder, J., Jeng, M. WILEY-LISS. 2007: 757–58

    View details for Web of Science ID 000246239900056

  • International standardized approach for flow cytometric residual disease monitoring in chronic lymphocytic leukaemia LEUKEMIA Rawstron, A. C., Villamor, N., Ritgen, M., Boettcher, S., Ghia, P., Zehnder, J. L., Lozanski, G., Colomer, D., Moreno, C., Geuna, M., Evans, P. A., Natkunam, Y., Coutre, S. E., Avery, E. D., Rassenti, L. Z., Kipps, T. J., Caligaris-Cappio, F., Kneba, M., Byrd, J. C., Hallek, M. J., Montserrat, E., Hillmen, P. 2007; 21 (5): 956-964

    Abstract

    The eradication of minimal residual disease (MRD) in chronic lymphocytic leukaemia (CLL) predicts for improved outcome. However, the wide variety of MRD techniques makes it difficult to interpret and compare different clinical trials. Our aim was to develop a standardized flow cytometric CLL-MRD assay and compare it to real-time quantitative allele-specific oligonucleotide (RQ-ASO) Immunoglobulin heavy chain gene (IgH) polymerase chain reaction (PCR). Analysis of 728 paired blood and marrow samples demonstrated high concordance (87%) for patients off-therapy. Blood analysis was equally or more sensitive than marrow in 92% of samples but marrow analysis was necessary to detect MRD within 3 months of alemtuzumab therapy. Assessment of 50 CLL-specific antibody combinations identified three (CD5/CD19 with CD20/CD38, CD81/CD22 and CD79b/CD43) with low inter-laboratory variation and false-detection rates. Experienced operators demonstrated an accuracy of 95.7% (specificity 98.8%, sensitivity 91.1%) in 141 samples with 0.01-0.1% CLL. There was close correlation and 95% concordance with RQ-ASO IgH-PCR for detection of CLL above 0.01%. The proposed flow cytometry approach is applicable to all sample types and therapeutic regimes, and sufficiently rapid and sensitive to guide therapy to an MRD-negativity in real time. These techniques may be used as a tool for assessing response and comparing the efficacy of different therapeutic approaches.

    View details for DOI 10.1038/sj.leu.2404584

    View details for Web of Science ID 000245999900014

    View details for PubMedID 17361231

  • Clinical evaluation of a novel oncologic tissue of origin assay based on gene expression microarray. Schrijver, I., Rodriguez-Paris, J., Zehnder, J. L., Pollack, J. R. ASSOC CLINICAL SCIENTISTS. 2007: 197–97

    View details for Web of Science ID 000246264700023

  • Severe coagulation factor V deficiency associated with an interstitial deletion of chromosome 1q JOURNAL OF THROMBOSIS AND HAEMOSTASIS Caudill, J. S., Sood, R., Zehnder, J. L., Pruthi, R. K., Steensma, D. P. 2007; 5 (3): 626-628

    View details for Web of Science ID 000244277800034

    View details for PubMedID 17166249

  • Safety and efficacy of a ginkgo biloba-containing dietary supplement on cognitive function, quality of life, and platelet function in healthy, cognitively intact older adults JOURNAL OF THE AMERICAN DIETETIC ASSOCIATION Carlson, J. J., Farquhar, J. W., Dinucci, E., Ausserer, L., Zehnder, J., Miller, D., Berra, K., Hagerty, L., Haskell, W. L. 2007; 107 (3): 422-432

    Abstract

    To determine if a ginkgo biloba-containing supplement improves cognitive function and quality of life, alters primary hemostasis, and is safe in healthy, cognitively intact older adults.Four-month, randomized, double-blind, placebo-controlled parallel design.Ninety men and women (age range 65 to 84 years) were recruited to a university clinic. Eligibility included those without dementia or depression, not taking psychoactive medications or medications or supplements that alter hemostasis.Ninety subjects were randomly assigned to placebo or a ginkgo biloba-based supplement containing 160 mg ginkgo biloba, 68 mg gotu kola, and 180 mg decosahexaenoic acid per day for 4 months.Assessments included: six standardized cognitive function tests, the SF-36 Quality of Life questionnaire, the Platelet Function Analyzer-100 (Dade Behring, Eschbom, Germany), and the monitoring of adverse events.Baseline characteristics and study hypotheses were tested using analysis of covariance. Tests were two-tailed with a 0.05 significance level.Seventy-eight subjects (87%) completed both baseline and 4-month testing (n=36 in placebo group, n=42 in ginkgo biloba group). At baseline, the participants' cognitive function was above average. One of six cognitive tests indicated significant protocol differences at 4 months (P=0.03), favoring the placebo. There were no significant differences in quality of life, platelet function, or adverse events.These finding do not support the use of a ginkgo biloba-containing supplement for improving cognitive function or quality of life in cognitively intact, older, healthy adults. However, high baseline scores may have contributed to the null findings. The ginkgo biloba product seems safe and did not alter platelet function, though additional studies are needed to evaluate the interaction of varying doses of ginkgo biloba and ginkgo biloba-containing supplements with medications and supplements that alter hemostasis.

    View details for DOI 10.1016/j.jada.2006.12.011

    View details for Web of Science ID 000244551100016

    View details for PubMedID 17324660

  • Molecular characterization and subcellular localization of Tyr478del: a pathogenic in-frame deletion in coagulation factor V JOURNAL OF THROMBOSIS AND HAEMOSTASIS Jones, C. D., Yeung, C., Negro, F., Zehnder, J. L. 2007; 5 (2): 431-433

    View details for Web of Science ID 000243563000038

    View details for PubMedID 17269939

  • Rituximab infusion two months after total lymphoid irradiation-antithymocyte globulin (TLI-ATG) nonmyeloablative transplantation maintains B-cell disease control with minimal GVHD Tandem BMT Meeting 2007 Arai, S., Sahaf, B., Jones, C., Zehnder, J., Lowsky, R., Strober, S., Shizuru, J., Negrin, R., Johnston, L., Laport, G., Goldstein, K., Brown, J., Elder, L., Tierney, K., Lavori, P., Miklos, D. ELSEVIER SCIENCE INC. 2007: 103–

    View details for Web of Science ID 000244028300284

  • T-cell clonality analysis in biopsies from two different sites improves specificity of diagnosis in patients with suspected mycosis fungoides Thurber, S., Zehnder, J., Kohler, S., Bessoni, J., Schrijver, I., Kim, Y. BLACKWELL PUBLISHING. 2007: 82

    View details for Web of Science ID 000243233600026

  • Gene expression profile of idiopathic thrombocytopenic purpura (ITP) reveals elevated expression of interferon regulated genes. 48th Annual Meeting of the American-Society-of-Hematology Sood, R., Wong, W., Gotlib, J., Jeng, M., Zehnder, J. L. AMER SOC HEMATOLOGY. 2006: 211A–211A

    View details for Web of Science ID 000242440000703

  • Severe factor V deficiency associated with chromosome 1q deletion. Caudill, J. S., Sood, R., Zehnder, J. L., Thorland, E. C., Rodriguez, V., Pruthi, R. K., Steensma, D. P. AMER SOC HEMATOLOGY. 2006: 310A

    View details for Web of Science ID 000242440001302

  • Rituximab infusion two months after nonmyeloablative transplantation maintains B-cell disease control with minimal GVHD. 48th Annual Meeting of the American-Society-of-Hematology Arai, S., Sahaf, B., Jones, C., Zehnder, J., Lowsky, R., Strober, S., Shizuru, J., Negrin, R., Johnston, L., Laport, G., Goldstein, K., Brown, J. (., Miklos, D. AMER SOC HEMATOLOGY. 2006: 823A–823A

    View details for Web of Science ID 000242440003696

  • Microsatellite instability testing: Comparison of the Promega MSI Analysis System, the Bethesda panel, and immunohistochemistry Putcha, G. V., Schrijver, I., Longacre, T., Zehnder, J. AMER SOC INVESTIGATIVE PATHOLOGY, INC. 2006: 665

    View details for Web of Science ID 000241811800199

  • Testing for maternal cell contamination in prenatal samples: a comprehensive survey of current diagnostic practices in 35 molecular diagnostic laboratories in the US Cherny, S. C., Zehnder, J. L., Schrijver, I. AMER SOC INVESTIGATIVE PATHOLOGY, INC. 2006: 622–23

    View details for Web of Science ID 000241811800019

  • Gene expression profile of idiopathic thrombocytopenic purpura (ITP) 2nd Expert Meeting of the Intercontinental-Childhood-ITP-Study-Group (ICIS) Sood, R., Wong, W., Jeng, M., Zehnder, J. L. WILEY PERIODICALS, INC. 2006: 675–77

    Abstract

    To search for novel mechanisms that contribute to the pathophysiology of idiopathic thrombocytopenic purpura (ITP), we determined the whole blood gene expression profile in five ITP patients and five control samples. Using DNA microarrays that contained 24,473 unique putative genes, we found 176 cDNAs that were strongly correlated with ITP. These included a cluster of interferon-regulated genes and TLR7, as well many less-well characterized genes which are candidates for further study. We believe this approach is likely to yield new insights into our understanding of the molecular pathophysiology of ITP.

    View details for DOI 10.1002/pbc.20981

    View details for Web of Science ID 000240405600010

    View details for PubMedID 16933260

  • Maintenance rituximab following induction chemoimmunotherapy may prolong progression-free survival in mantle cell lymphoma: a pilot study from the Wisconsin Oncology Network ANNALS OF ONCOLOGY Kahl, B. S., Longo, W. L., Eickhoff, J. C., Zehnder, J., Jones, C., Blank, J., McFarland, T., Bottner, W., Rezazedeh, H., Werndli, J., Bailey, H. H. 2006; 17 (9): 1418-1423

    Abstract

    There is no standard first line treatment for mantle cell lymphoma.This was a multicenter phase II pilot study of rituximab and modified hyper-fractionated cyclophosphamide, vincristine doxorubicin, dexamethasone (modified R-hyperCVAD) administered every 28 days for four to six cycles followed by rituximab maintenance therapy consisting of four weekly doses every 6 months for 2 years. Unlike traditional hyperCVAD regimens, no methotrexate or cytarabine was administered.Of 22 patients, the overall response rate was 77% and the complete response rate was 64%. With a median follow-up time of 37 months in surviving patients, the median PFS was 37 months and the median OS was not reached. The achievement of a molecular remission did not correlate with improved outcome. The major toxicity was expected myelosuppression. Two patients died during induction treatment. There were no major adverse effects during maintenance therapy.In a multicenter trial, modified R-hyperCVAD was tolerable and effective induction therapy for untreated MCL. Maintenance rituximab appeared to prolong PFS without increasing toxicity.

    View details for DOI 10.1093/annonc/mdl127

    View details for Web of Science ID 000240587900012

    View details for PubMedID 16766582

  • Detection of the JAK2 V617F mutation by LightCycler PCR and probe dissociation analysis JOURNAL OF MOLECULAR DIAGNOSTICS Lay, M., Mariappan, R., Gotlib, J., Dietz, L., Sebastian, S., Schrijver, I., Zehnder, J. L. 2006; 8 (3): 330-334

    Abstract

    A point mutation in the JAK2 gene, a member of the tyrosine kinase family, was recently identified and shown to be associated with several myeloproliferative disorders. Several studies identified the same JAK2 point mutation (1,849G>T), resulting in the substitution of a valine to phenylalanine at codon 617 (V617F). We developed a simple and sensitive method to detect this mutation via polymerase chain reaction and probe dissociation analysis using the LightCycler platform, and we compared this method to existing restriction fragment-length polymorphism, direct sequencing, and amplification refractory mutation system methods. We found that the LightCycler method offered advantages of speed, reliability, and more straightforward interpretation over the restriction fragment-length polymorphism and sequencing approaches.

    View details for DOI 10.2353/jmoldx.2006.050130

    View details for Web of Science ID 000239106800006

    View details for PubMedID 16825505

    View details for PubMedCentralID PMC1867612

  • First molecular characterization of a patient with combined factor V and factor VII deficiency THROMBOSIS AND HAEMOSTASIS Traynis, I., Jones, C. D., Gibb, C. B., Acharya, S. S., Zehnder, J. L. 2006; 95 (6): 1031-1032

    View details for DOI 10.1160/TH06-03-0177

    View details for Web of Science ID 000238557200019

    View details for PubMedID 16732384

  • Mast cells can promote the development of multiple features of chronic asthma in mice JOURNAL OF CLINICAL INVESTIGATION Yu, M., Tsai, M., Tam, S., Jones, C., Zehnder, J., Galli, S. J. 2006; 116 (6): 1633-1641

    Abstract

    Bronchial asthma, the most prevalent cause of significant respiratory morbidity in the developed world, typically is a chronic disorder associated with long-term changes in the airways. We developed a mouse model of chronic asthma that results in markedly increased numbers of airway mast cells, enhanced airway responses to methacholine or antigen, chronic inflammation including infiltration with eosinophils and lymphocytes, airway epithelial goblet cell hyperplasia, enhanced expression of the mucin genes Muc5ac and Muc5b, and increased levels of lung collagen. Using mast cell-deficient (Kit(W-sh/W-sh) and/or Kit(W/W-v)) mice engrafted with FcRgamma+/+ or FcRgamma-/- mast cells, we found that mast cells were required for the full development of each of these features of the model. However, some features also were expressed, although usually at less than wild-type levels, in mice whose mast cells lacked FcRgamma and therefore could not be activated by either antigen- and IgE-dependent aggregation of Fc epsilonRI or the binding of antigen-IgG1 immune complexes to Fc gammaRIII. These findings demonstrate that mast cells can contribute to the development of multiple features of chronic asthma in mice and identify both Fc Rgamma-dependent and Fc Rgamma-independent pathways of mast cell activation as important for the expression of key features of this asthma model.

    View details for DOI 10.1172/JCI25702

    View details for Web of Science ID 000237979700025

    View details for PubMedID 16710480

    View details for PubMedCentralID PMC1462940

  • Successful transduction of liver in hemophilia by AAV-Factor IX and limitations imposed by the host immune response (vol 12, pg 342, 2006) NATURE MEDICINE Manno, C. S., Arruda, V. R., Pierce, G. F., Glader, B., Ragni, M., Rasko, J., Ozelo, M. C., Hoots, K., Blatt, P., Konkle, B., Dake, M., Kaye, R., Razavi, M., Zajko, A., Zehnder, J., Nakai, H., Chew, A., Leonard, D., Wright, J. F., Lessard, R. R., Sommer, J. M., TIGGES, M., Sabatino, D., Luk, A., Jiang, H., Mingozzi, F., Couto, L., Ertl, H. C., High, K. A., Kay, M. A. 2006; 12 (5): 592-592

    View details for Web of Science ID 000238149100046

  • The JAK2 V617F mutation occurs in hematopoietic stem cells in polycythemia vera and predisposes toward erythroid differentiation PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Jamieson, C. H., Gotlib, J., Durocher, J. A., Chao, M. P., Mariappan, M. R., Lay, M., Jones, C., Zehnder, J. L., Lilleberg, S. L., Weissman, I. L. 2006; 103 (16): 6224-6229

    Abstract

    Although a large proportion of patients with polycythemia vera (PV) harbor a valine-to-phenylalanine mutation at amino acid 617 (V617F) in the JAK2 signaling molecule, the stage of hematopoiesis at which the mutation arises is unknown. Here we isolated and characterized hematopoietic stem cells (HSC) and myeloid progenitors from 16 PV patient samples and 14 normal individuals, testing whether the JAK2 mutation could be found at the level of stem or progenitor cells and whether the JAK2 V617F-positive cells had altered differentiation potential. In all PV samples analyzed, there were increased numbers of cells with a HSC phenotype (CD34+CD38-CD90+Lin-) compared with normal samples. Hematopoietic progenitor assays demonstrated that the differentiation potential of PV was already skewed toward the erythroid lineage at the HSC level. The JAK2 V617F mutation was detectable within HSC and their progeny in PV. Moreover, the aberrant erythroid potential of PV HSC was potently inhibited with a JAK2 inhibitor, AG490.

    View details for DOI 10.1073/pnas.0601462103

    View details for Web of Science ID 000236999000031

    View details for PubMedID 16603627

    View details for PubMedCentralID PMC1434515

  • Gene expression patterns in human placenta PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Sood, R., Zehnder, J. L., Druzin, M. L., Brown, P. O. 2006; 103 (14): 5478-5483

    Abstract

    The placenta is the principal metabolic, respiratory, excretory, and endocrine organ for the first 9 months of fetal life. Its role in fetal and maternal physiology is remarkably diverse. Because of the central role that the placenta has in fetal and maternal physiology and development, the possibility that variation in placental gene expression patterns might be linked to important abnormalities in maternal or fetal health, or even variations in later life, warrants investigation. As an initial step, we used DNA microarrays to analyze gene expression patterns in 72 samples of amnion, chorion, umbilical cord, and sections of villus parenchyma from 19 human placentas from successful full-term pregnancies. The umbilical cord, chorion, amnion, and villus parenchyma samples were readily distinguished by differences in their global gene-expression patterns, many of which seemed to be related to physiology and histology. Differentially expressed genes have roles that include placental trophoblast secretion, signal transduction, metabolism, immune regulation, cell adhesion, and structure. We found interindividual differences in expression patterns in villus parenchyma and systematic differences between the maternal, fetal, and intermediate layers. A group of genes that was expressed in both the maternal and fetal villus parenchyma sections of placenta included genes that may be associated with preeclampsia. We identified sets of genes whose expression in placenta was significantly correlated with the sex of the fetus. This study provides a rich and diverse picture of the molecular variation in the placenta from healthy pregnancies.

    View details for DOI 10.1073/pnas.0508035103

    View details for Web of Science ID 000236636400044

    View details for PubMedID 16567644

    View details for PubMedCentralID PMC1414632

  • Successful transduction of liver in hemophilia by AAV-factor IX and limitations imposed by the host immune response NATURE MEDICINE Manno, C. S., Arruda, V. R., Pierce, G. F., Glader, B., Ragni, M., Rasko, J., Ozelo, M. C., Hoots, K., Blatt, P., Konkle, B., Dake, M., Kaye, R., Razavi, M., Zajko, A., Zehnder, J., Nakai, H., Chew, A., Leonard, D., Wright, J. F., Lessard, R. R., Sommer, J. M., TIGGES, M., Sabatino, D., Luk, A., Jiang, H. Y., Mingozzi, F., Couto, L., Ertl, H. C., High, K. A., Kay, M. A. 2006; 12 (3): 342-347

    Abstract

    We have previously shown that a single portal vein infusion of a recombinant adeno-associated viral vector (rAAV) expressing canine Factor IX (F.IX) resulted in long-term expression of therapeutic levels of F.IX in dogs with severe hemophilia B. We carried out a phase 1/2 dose-escalation clinical study to extend this approach to humans with severe hemophilia B. rAAV-2 vector expressing human F.IX was infused through the hepatic artery into seven subjects. The data show that: (i) vector infusion at doses up to 2 x 10(12) vg/kg was not associated with acute or long-lasting toxicity; (ii) therapeutic levels of F.IX were achieved at the highest dose tested; (iii) duration of expression at therapeutic levels was limited to a period of approximately 8 weeks; (iv) a gradual decline in F.IX was accompanied by a transient asymptomatic elevation of liver transaminases that resolved without treatment. Further studies suggested that destruction of transduced hepatocytes by cell-mediated immunity targeting antigens of the AAV capsid caused both the decline in F.IX and the transient transaminitis. We conclude that rAAV-2 vectors can transduce human hepatocytes in vivo to result in therapeutically relevant levels of F.IX, but that future studies in humans may require immunomodulation to achieve long-term expression.

    View details for DOI 10.1038/nm1358

    View details for Web of Science ID 000235802900035

    View details for PubMedID 16474400

  • Microsatellite instability in ovarian cancer subtypes and in synchronous ovarian and endometrial cancers: The Stanford experience Jensen, K. C., Putcha, G. V., Otani, I. M., Schrijver, Zehnder, J., Longacre, T. A. NATURE PUBLISHING GROUP. 2006: 182A

    View details for Web of Science ID 000234207601277

  • Microsatellite instability in ovarian cancer subtypes and in synchronous ovarian and endometrial cancers: The Stanford experience Jensen, K. C., Putcha, G. V., Otani, I. M., Schrijver, Zehnder, J. NATURE PUBLISHING GROUP. 2006: 182A

    View details for Web of Science ID 000234094501282

  • Bioluminescent imaging of human leukemic stem cell engraftment. 47th Annual Meeting of the American-Society-of-Hematology Jamieson, C., Karimi, M., Creusot, R., Negrin, R., Gotlib, J., Chao, M., Jones, C., Keating, A., Fathman, C. G., Zehnder, J., Weissman, I. L. AMER SOC HEMATOLOGY. 2005: 205A–205A

    View details for Web of Science ID 000233426001165

  • Gene expression profiling of multiple patients with cyclic thrombocytopenia reveals consistent profile. Sood, R., Gourley, E., Schrier, S. L., Go, R., Zehnder, J. L. AMER SOC HEMATOLOGY. 2005: 67A

    View details for Web of Science ID 000233426000220

  • First molecular characterization of a family with combined FV and FVII deficiency. Traynis, Jones, C. D., Gibb, C. B., Acharya, S., Zehnder, J. L. AMER SOC HEMATOLOGY. 2005: 508A

    View details for Web of Science ID 000233426003148

  • Molecular progenitor profiling in human myeloproliferative disorders. 47th Annual Meeting of the American-Society-of-Hematology Jamieson, C. H., Gotlib, J., Chao, M., Mariappan, M. R., LayRaj, M., Jones, C., Zehnder, J., Durocher, J., Lilleberg, S., Coutre, S., Weissman, I. L. AMER SOC HEMATOLOGY. 2005: 38A–39A

    View details for Web of Science ID 000233426000119

  • Characterization of a novel prothrombin variant, Prothrombin C20209T, as a modifier of thrombotic risk among African-Americans JOURNAL OF THROMBOSIS AND HAEMOSTASIS Itakura, H., Telen, M. J., Hoppe, C. C., White, D. A., Zehnder, J. L. 2005; 3 (10): 2357-2359

    View details for Web of Science ID 000232443200036

    View details for PubMedID 16194213

  • A comparison study of different PCR assays in measuring circulating plasma Epstein-Barr virus DNA levels in patients with nasopharyngeal carcinoma CLINICAL CANCER RESEARCH Le, Q. T., Jones, C. D., Yau, T. K., Shirazi, H. A., Wong, P. H., Thomas, E. N., Patterson, B. K., Lee, A. W., Zehnder, J. L. 2005; 11 (16): 5700-5707

    Abstract

    To compare the performance of three PCR assays in measuring circulating Epstein-Barr virus (EBV). DNA levels in nasopharyngeal carcinoma patients and to confirm its prognostic significance.Plasma from 58 newly diagnosed nasopharyngeal carcinoma patients were collected before, during, and every 3 to 6 months after radiotherapy. EBV DNA levels were determined by real-time quantitative PCR using primer/probe sets for polymerase-1 (Pol-1), latent membrane protein 2 (Lmp2), and BamHI-W. Pretreatment levels from the three assays were correlated with each other and serial measurements from the Pol-1 assay were correlated with clinical variables.Pol-1 was more accurate than BamHI-W in predicting EBV DNA concentrations in cell lines. Of the three assays, BamHI-W yielded the highest concentrations followed by Pol-1 in plasmas (n = 23). The correlation coefficient was 0.99 (P < 0.0001) for Pol-1 and Lmp2, 0.66 (P < 0.0001) for Pol-1 and BamHI-W, and 0.55 (P < 0.0001) for BamHI-W and Lmp2. Elevated pretreatment DNA levels as detected by Pol-1 were correlated with advanced nodal stage (P = 0.04) and overall stage (P = 0.028). There was no correlation between pretreatment EBV DNA levels and freedom-from-relapse or overall survival; however, there was a significant correlation between posttreatment levels and these variables. The 2-year freedom-from-relapse and overall survival rates were 92% and 94% for patients with undetectable, and 37% and 55% for those with detectable, posttreatment levels (P < 0.0001 and P < 0.002).The three PCR assays yielded similar results in detecting EBV DNA in plasmas. The Pol-1-detected posttreatment EBV DNA level was the strongest predictor for treatment outcomes.

    View details for DOI 10.1158/1078-0432.CCR-05-0648

    View details for PubMedID 16115906

  • Identification of mislabeled specimen by molecular methods: Case report and review INTERNATIONAL JOURNAL OF SURGICAL PATHOLOGY Mariappan, M. R., Zehnder, J., Arber, D. A., Lay, M., Fadare, O., Schrijver, R. 2005; 13 (3): 253-258

    Abstract

    Specimen misidentification is a common cause of errors in surgical pathology. We report a case where bone-marrow biopsies from patients of different genders were mislabeled and molecular methods were applied to resolve the identity. A short tandem repeat (STR)-polymerase chain reaction-based assay, commonly used in paternity testing, was employed in an attempt to assign the correct identity to the specimens. However, the specimens had been processed by decalcification and the DNA yield was poor. One of the markers in the assay is the non-STR amelogenin locus that distinguishes the X and Y chromosomes. This amelogenin marker results in a product of low molecular weight, enabling unequivocal resolution of identity despite a poor DNA yield. The prevalence of errors in pathology due to specimen misidentifications is reviewed.

    View details for Web of Science ID 000231185700004

    View details for PubMedID 16086080

  • Varicella-zoster virus infection of human dorsal root ganglia in vivo PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Zerboni, L., Ku, C. C., Jones, C. D., Zehnder, J. L., Arvin, A. M. 2005; 102 (18): 6490-6495

    Abstract

    Varicella-zoster virus (VZV) causes varicella and establishes latency in sensory ganglia. VZV reactivation results in herpes zoster. We developed a model using human dorsal root ganglion (DRG) xenografts in severe combined immunodeficient (SCID) mice to investigate VZV infection of differentiated neurons and satellite cells in vivo. DRG engrafted under the kidney capsule and contained neurons and satellite cells within a typical DRG architecture. VZV clinical isolates infected the neurons within DRG. At 14 days postinfection, VZ virions were detected by electron microscopy in neuronal cell nuclei and cytoplasm but not in satellite cells. The VZV genome copy number was 7.1 x 10(7) to 8.0 x 10(8) copies per 10(5) cells, and infectious virus was recovered. This initial phase of viral replication was followed within 4-8 weeks by a transition to VZV latency, characterized by the absence of infectious virus release, the cessation of virion assembly, and a reduction in VZV genome copies to 3.7 x 10(5) to 4.7 x 10(6) per 10(5) cells. VZV persistence in DRG was achieved without any requirement for VZV-specific adaptive immunity and was associated with continued transcription of the ORF63 regulatory gene. The live attenuated varicella vaccine virus exhibited the same pattern of short-term replication, persistence of viral DNA, and prominent ORF63 transcription as the clinical isolates. VZV-infected T cells transferred virus from the circulation into DRG, suggesting that VZV lymphotropism facilitates its neurotropism. DRG xenografts may be useful for investigating neuropathogenic mechanisms of other human viruses.

    View details for DOI 10.1073/pnas.0501045102

    View details for Web of Science ID 000228918400045

    View details for PubMedID 15851670

    View details for PubMedCentralID PMC1088374

  • High frequency of premature termination mutations in the factor V gene: Three factor V deficiency case reports and a mutation review THROMBOSIS AND HAEMOSTASIS Schrijver, I., Hong, D. W., Mandle, L., Jones, C. D., DiMichele, D., Monahan, P. E., Zehnder, J. L. 2005; 93 (3): 610-611

    View details for Web of Science ID 000227808200034

    View details for PubMedID 15735818

  • Identification of a novel factor V A2 domain tyrosine deletion mutation in a patient with Factor V deficiency and bleeding. Jones, C. D., Negro, F., Darnell, K., Zehnder, J. L. AMER SOC HEMATOLOGY. 2004: 94B

    View details for Web of Science ID 000225127700362

  • Gene expression patterns in placenta from normal and preeclamptic pregnancies Sood, R., Zehnder, J., Druzin, M., Brown, P. O. W B SAUNDERS CO LTD. 2004: A21

    View details for Web of Science ID 000223898300085

  • Granulocyte-macrophage progenitors as candidate leukemic stem cells in blast-crisis CML NEW ENGLAND JOURNAL OF MEDICINE Jamieson, C. H., Ailles, L. E., Dylla, S. J., Muijtjens, M., Jones, C., Zehnder, J. L., Gotlib, J., Li, K., Manz, M. G., Keating, A., Sawyers, C. L., Weissman, I. L. 2004; 351 (7): 657-667

    Abstract

    The progression of chronic myelogenous leukemia (CML) to blast crisis is supported by self-renewing leukemic stem cells. In normal mouse hematopoietic stem cells, the process of self-renewal involves the beta-catenin-signaling pathway. We investigated whether leukemic stem cells in CML also use the beta-catenin pathway for self-renewal.We used fluorescence-activated cell sorting to isolate hematopoietic stem cells, common myeloid progenitors, granulocyte-macrophage progenitors, and megakaryocyte-erythroid progenitors from marrow during several phases of CML and from normal marrow. BCR-ABL, beta-catenin, and LEF-1 transcripts were compared by means of a quantitative reverse-transcriptase-polymerase-chain-reaction assay in normal and CML hematopoietic stem cells and granulocyte-macrophage progenitors. Confocal fluorescence microscopy and a lymphoid enhancer factor/T-cell factor reporter assay were used to detect nuclear beta-catenin in these cells. In vitro replating assays were used to identify self-renewing cells as candidate leukemic stem cells, and the dependence of self-renewal on beta-catenin activation was tested by lentiviral transduction of hematopoietic progenitors with axin, an inhibitor of the beta-catenin pathway.The granulocyte-macrophage progenitor pool from patients with CML in blast crisis and imatinib-resistant CML was expanded, expressed BCR-ABL, and had elevated levels of nuclear beta-catenin as compared with the levels in progenitors from normal marrow. Unlike normal granulocyte-macrophage progenitors, CML granulocyte-macrophage progenitors formed self-renewing, replatable myeloid colonies, and in vitro self-renewal capacity was reduced by enforced expression of axin.Activation of beta-catenin in CML granulocyte-macrophage progenitors appears to enhance the self-renewal activity and leukemic potential of these cells.

    View details for PubMedID 15306667

  • Rapid combined genotyping assay for four achondroplasia and hypochondroplasia mutations by real-time PCR with multiple detection probes GENETIC TESTING Schrijver, I., Lay, M. J., Zehnder, J. L. 2004; 8 (2): 185-189

    Abstract

    Achondroplasia (ACH) and hypochondroplasia (HYCH) are the most prevalent genetic short-stature syndromes. Whereas the diagnosis of ACH can be established on clinical and radiologic grounds alone in the majority of cases, HYCH is more difficult to confirm. Molecular genetic analysis of both skeletal dysplasias can be especially helpful for the purpose of prenatal diagnosis, in early childhood to differentiate definitively between the largely overlapping phenotypes, and in atypical presentations. The two most prevalent mutations for each syndrome cause substitution of a single respective nucleotide. These mutations can be identified by a variety of molecular methods, including PCR with restriction enzyme digestion or direct DNA sequencing. We have developed a single-step, real-time PCR assay in which two detection probes are applied in combination with a single anchor probe at each mutation position. Because the two most prevalent mutations for each syndrome cause substitution of a single respective nucleotide, this approach guarantees optimal differentiation during probe dissociation analysis after amplification. This assay, which is performed on the LightCycler thermocycler, enables the rapid and reliable detection of the two most common FGFR3 mutations associated with ACH (1138G --> A and 1138G --> C; G380R) and HYCH (1620C --> A and 1620 C --> G; N540K) in a single test.

    View details for Web of Science ID 000223513900019

    View details for PubMedID 15345118

  • CyclinD1/CyclinD3 ratio by real-time PCR improves specificity for the diagnosis of mantle cell lymphoma JOURNAL OF MOLECULAR DIAGNOSTICS Jones, C. D., Darnell, K. H., Warnke, R. A., Zehnder, J. L. 2004; 6 (2): 84-89

    Abstract

    We developed a real-time, quantitative, reverse transcription PCR assay for cyclin D1 (CCND1) expression to aid in the diagnosis of mantle cell lymphoma (MCL). The diagnosis of MCL can be problematic, and existing CCND1 expression assays show a lack of specificity, with elevated expression also detected in other lymphoproliferative disorders. We postulated that evaluating CCND1 expression relative to CCND3 expression by quantitative PCR could offer an improved specificity over an evaluation of CCND1 alone. This method quantitates both CCND1 and CCND3, each normalized to a housekeeping gene (GADPH), using the 5'-exonuclease technique. We analyzed 107 clinical specimens: MCL (17), chronic lymphocytic leukemias (CLL) (10), other non-MCL hematolymphoid disorders (41), non-malignant tissues with an epithelial component (7) and other normal samples (32). This method correctly identified 16 of 17 MCLs, and there were no false positives among any of the other diagnostic groups tested including CLL. CLL presents the major diagnostic dilemma at this institution when diagnosing MCL. Sensitivity studies showed that this method could detect an elevated CCND1/CCND3 ratio when the tumor infiltrate is at least 10% of the cells. We compared the specificity of CCND1 expression alone against the CCND1/CCND3 ratio to demonstrate the increased specificity for the latter. We conclude that the CCND1/CCND3 ratio is a sensitive and specific test for the diagnosis of MCL.

    View details for Web of Science ID 000221002500002

    View details for PubMedID 15096562

  • Molecular diagnosis of hypercoagulable states LABORATORY MEDICINE Nagy, P. L., Schrijver, I., Zehnder, J. L. 2004; 35 (4): 214-221

    View details for Web of Science ID 000220469400016

  • Randomized trial of folic acid for prevention of cardiovascular events in end-stage renal disease JOURNAL OF THE AMERICAN SOCIETY OF NEPHROLOGY Wrone, E. M., Hornberger, J. M., Zehnder, J. L., McCann, L. M., Coplon, N. S., Fortmann, S. P. 2004; 15 (2): 420-426

    Abstract

    High serum total homocysteine (tHcy) is gaining scrutiny as a risk factor for cardiovascular disease in the general population. The relationship between tHcy and mortality and cardiovascular events in patients with end-stage renal disease (ESRD) is unsettled. This randomized trial evaluates the efficacy of high-dose folic acid in preventing events in ESRD. A total of 510 patients on chronic dialysis were randomized to 1, 5, or 15 mg of folic acid contained in a renal multivitamin with a median follow-up of 24 mo. Mortality, cardiovascular events, and homocysteine levels were assessed. There were 189 deaths, and 121 patients experienced at least one cardiovascular event. Composite rates of mortality and cardiovascular events among the folic acid groups did not differ (at 24 mo: 43.7% in 1 mg group, 38.6% in 5 mg group, 47.1% in 15 mg group; log-rank P = 0.47). Unexpectedly, high baseline tHcy was associated with lower event rates. From lowest to highest quartile, event rates at 24 mo were 54.5% for Q1, 41.8% for Q2, 41.2% for Q3, and 34.7% for Q4 (log-rank P = 0.033). In contrast to some studies describing tHcy as a risk factor for mortality and cardiovascular events, this study found a reverse relationship between tHcy and events in ESRD patients. Administration of high-dose folic acid did not affect event rates.

    View details for DOI 10.1097/01.ASN.0000110181.64655.6C

    View details for Web of Science ID 000188604600020

    View details for PubMedID 14747389

  • Lymphomatoid papulosis associated with mycosis fungoides: A study of 21 patients including analyses for clonality. (vol 49, pg 620, 2003) JOURNAL OF THE AMERICAN ACADEMY OF DERMATOLOGY Zackheim, H. S., Jones, C., LeBoit, P. E., Kashani-Sabet, M., McCalmont, T. H., Zehnder, J. 2004; 50 (2): 202

    View details for Web of Science ID 000188710100004

  • Congenital and acquired thrombocytopenia. Hematology / the Education Program of the American Society of Hematology. American Society of Hematology. Education Program Cines, D. B., Bussel, J. B., McMillan, R. B., Zehnder, J. L. 2004: 390-406

    Abstract

    The diagnosis and management of thrombocytopenia is a growing component in the practice of hematology. The frequency with which hematologists are called in consultation for thrombocytopenia continues to increase with the advent of routine automated platelet determinations and the introduction of new medications. For most patients, such as those with inherited and auto-immune thrombocytopenia, emphasis is focused on efforts to treat or forestall bleeding without excess drug-induced toxicity or burden to the patient. However, in disorders such as heparin-induced thrombocytopenia (HIT), avoidance of thrombotic complications is the key to management. In this chapter, we provide the pediatric and adult hematologist with new insights into the pathogenesis and recognition of congenital inherited thrombocytopenias (CTP), a hitherto difficult to comprehend constellation of clinical entities. We also highlight new approaches to the diagnosis and treatment of two of the more common thrombocytopenic conditions encountered in practice, autoimmune or idiopathic thrombocytopenic purpura (ITP) and HIT. In Section I, Dr. James Bussel discusses CTPs and their distinction from childhood ITP. He emphasizes the clinical features that enable the pediatrician and hematologist to suspect the diagnosis of CTP and those that are of use to subcategorize the various entities, where possible. He also emphasizes newer molecular markers that afford definitive diagnosis in some cases and provide insight into platelet production. This section highlights the characteristic associated findings and differences in the natural history and approaches to management of the various entities. In Section II, Dr. Robert McMillan discusses adult chronic ITP. He revisits the utility of platelet antibody determination in diagnosis and review new insights into pathogenesis. The role of Helicobacter pylori infection and the timing of splenectomy in the management of acute and emergent ITP are examined. New insights into the natural history of ITP post-splenectomy and management strategies for patients with severe, chronic, refractory ITP are discussed. In Section III, Dr. James Zehnder updates us on HIT. He emphasizes new insights into the clinical presentation and pathogenesis of this condition. He critically reviews the utility of laboratory testing for heparin-dependent antibodies. Recent studies on the use of direct thrombin inhibitors are examined and the management of cardiopulmonary bypass surgery in patients with HIT is discussed.

    View details for PubMedID 15561694

  • BCR/ABL expression by highly purified chronic myelogenous leukemic hematopoietic stem cells and myeloid progenitors pre and post-imatinib therapy. 45th Annual Meeting and Exhibition of the American-Society-of-Hematology Jamieson, C. H., Jones, C., Zehnder, J., Gotlib, J., Manz, M., Weissman, I. L. AMER SOC HEMATOLOGY. 2003: 418A–418A

    View details for Web of Science ID 000186536701519

  • Gene expression profiling of cyclical thrombocytopenia. Sood, R., Gourley, E. J., Schrier, S. L., Brown, P. O., Zehnder, J. L. AMER SOC HEMATOLOGY. 2003: 789A

    View details for Web of Science ID 000186536702916

  • The role of beta-catenin in chronic myelogenous leukemic progenitor expansion. 45th Annual Meeting and Exhibition of the American-Society-of-Hematology Jamieson, C. H., Ailles, L. E., Muijtjens, M., Jones, C., Zehnder, J., Gotlib, J., Dylla, S., Li, K., Weissman, I. L. AMER SOC HEMATOLOGY. 2003: 570A–570A

    View details for Web of Science ID 000186536702096

  • Prothrombin gene variants in non-Caucasians with fetal loss and intrauterine growth retardation JOURNAL OF MOLECULAR DIAGNOSTICS Schrijver, I., Lenzi, T. J., Jones, C. D., Lay, M. J., Druzin, M. L., Zehnder, J. L. 2003; 5 (4): 250-253

    Abstract

    Thrombotic predisposition may affect pregnancy outcome, but in non-Caucasians the contributing genetic factors are poorly characterized. Two recently identified prothrombin gene mutations (20209C>T and 20221C>T) have been observed in non-Caucasian patients with thrombosis. The mutations are located near the commonly identified variant 20210G>A and have not been reported in Caucasian patients. The authors report a novel connection with pregnancy complications. The identification of sequence variants other than 20210G>A in the 3'-untranslated region of the prothrombin gene suggests that additional nucleotide substitutions may contribute to the development of thrombotic events and adverse pregnancy outcomes, especially in less well-characterized populations.

    View details for Web of Science ID 000186292700009

    View details for PubMedID 14573785

  • Lymphomatoid papulosis associated with mycosis fungoides: A study of 21 patients including analyses for clonality JOURNAL OF THE AMERICAN ACADEMY OF DERMATOLOGY Zackheim, H. S., Jones, C., LeBoit, P. E., Kashani-Sabet, M., McCalmont, T. H., Zehnder, J. 2003; 49 (4): 620-623

    Abstract

    Although an association of lymphomatoid papulosis (LyP) with mycosis fungoides (MF) is recognized, our understanding of this relation is limited.We sought to document the clinical experience at the University of California, San Francisco, in 21 patients who had both LyP and MF and to do clonality studies in 7 of those patients in whom this was possible.We conducted chart review of the 21 patients and analysis for T-cell receptor-gamma gene rearrangements by the polymerase chain reaction.Of 54 patients, 21 (39%) with LyP had associated MF. LyP preceded MF in 14 (67%), MF preceded LyP in 4 (19%), and there was concurrent appearance in 3 (14%). Of the 21 patients, 20 (95%) were type A and only 1 (5%) was type B. An identical clone was found in lesions of both LyP and MF in all 7 patients in whom analysis was possible.Findings of this study strengthen the idea that LyP and MF are related T-cell lymphoproliferative disorders.

    View details for DOI 10.1067/S0190-9622(03)01577-9

    View details for Web of Science ID 000185606500006

    View details for PubMedID 14512906

  • Heterozygous prothrombin G20210A gene mutation in a patient with livedoid vasculitis ARCHIVES OF DERMATOLOGY Gotlib, J., Kohler, S., Reicherter, P., Oro, A. E., Zehnder, J. L. 2003; 139 (8): 1081-1083

    View details for Web of Science ID 000184692700021

    View details for PubMedID 12925402

  • Comprehensive validation of a real-time quantitative bcr-abl assay for clinical laboratory use AMERICAN JOURNAL OF CLINICAL PATHOLOGY Jones, C. D., Yeung, C., Zehnder, J. L. 2003; 120 (1): 42-48

    Abstract

    We developed and extensively validated a real-time PCR assay for the quantitation of bcr-abl to determine residual disease in patients with chronic myelogenous leukemia. This method quantitates the p210 and the p190 bcr-abl RNA fusion transcripts with results normalized to a housekeeping gene, using the 5'-exonuclease technique and the ABI PRISM 7700 Sequence Detection System (Applied Biosystems, Foster City, CA). We parallel tested 372 clinical specimens and 50 peripheral blood samples from patients not known to have any myeloproliferative disorders. The results were 100% specific. Sensitivity studies showed that this method can detect bcr-abl in cell lines diluted to 0.0001% and can detect a single bcr-abl plasmid spiked into negative RNA. The between-run reproducibility showed a coefficient of variance (CV) of 12.3%, and within-run reproducibility showed a CV of 13.8%. This method can be used to reliably monitor the disease load in patients with bcr-abl-positive diseases.

    View details for DOI 10.1309/60A9C8WGEGHRNXEE

    View details for Web of Science ID 000183730300005

    View details for PubMedID 12866371

  • Diagnostic single nucleotide polymorphism analysis of factor V Leiden and prothrombin 20210G > A - A comparison of the nanogen electronic microarray with restriction enzyme digestion and the Roche LightCycler AMERICAN JOURNAL OF CLINICAL PATHOLOGY Schrijver, I., Lay, M. J., Zehnder, J. L. 2003; 119 (4): 490-496

    Abstract

    Genetic thrombosis risk factors include a sequence variant in the prothrombin gene (20210G > A) and factor V Leiden (1691G > A). These single nucleotide polymorphisms can be diagnosed with restriction fragment length polymorphism (RFLP) analysis, fluorescent genotyping on the LightCycler (Roche Diagnostics, Indianapolis, IN), and microarray-based testing on the novel NanoChip electronic microarray (NanoChip Molecular Biology Workstation, Nanogen, San Diego, CA). We compared these methods for accuracy, time to results, throughput, and interpretation. Results from 789 of 800 individual amplicons analyzed on the NanoChip were in complete agreement with the other assays. Eleven were "no calls" (uninterpreted by the NanoChip system) resulting from failed polymerase chain reaction amplifications. Although the NanoChip System, when used in a low-throughput setting, requires more overall time than the LightCycler, it is nearly equivalent per genotyping call. Owing to minimal sample handling, assay results are more reliable on the NanoChip platform and on the LightCycler than with RFLP. The NanoChip assay is reliable and may be especially valuable to laboratories with a large volume of thrombophilia test requests.

    View details for DOI 10.1309/3VTR7TL2X7TXL0XY

    View details for Web of Science ID 000181872500002

    View details for PubMedID 12710121

  • Lack of human herpesvirus 8 and Epstein-Barr virus in Kikuchi's histiocytic necrotizing lymphadenitis HUMAN PATHOLOGY George, T. I., Jones, C. D., Zehnder, J. L., Warnke, R. A., Dorfman, R. F. 2003; 34 (2): 130-135

    Abstract

    Kikuchi's histiocytic necrotizing lymphadenitis is a self-limited disorder that typically involves the cervical lymph nodes of young women. Although a viral etiology has been postulated, a definitive viral agent has not been identified. Recent reports have suggested that human herpesvirus 8 (HHV 8) or Epstein-Barr virus (EBV) may play an etiologic role. We investigated the presence of HHV 8 and EBV in archival tissue from 34 cases of Kikuchi's histiocytic necrotizing lymphadenitis. We examined 29 cases for HHV 8 using a nested polymerase chain reaction (PCR) on paraffin-embedded or frozen tissue, and 24 cases for EBV RNA using in situ hybridization (ISH) for EBER1. Controls included reactive lymph nodes from 8 adult women presenting with cervical or axillary lymphadenopathy. The study patients included 7 men and 27 women with a mean age of 28 years. All patients were previously healthy without evidence of immunocompromise and presented with cervical, axillary, or inguinal lymphadenopathy. Two cases exhibited EBV RNA by ISH; this was confirmed by PCR for EBV DNA. HHV 8 DNA was not amplified by nested PCR in any of the cases of Kikuchi's histiocytic necrotizing lymphadenitis or reactive lymph nodes; control PCR demonstrated the presence of amplifiable DNA in all cases. These findings suggest that HHV 8 and EBV do not play causative roles in Kikuchi's histiocytic necrotizing lymphadenitis.

    View details for DOI 10.1053/hupa.2003.11

    View details for PubMedID 12612880

  • Psoriasiform mycosis fungoides with fatal outcome after treatment with cyclosporine JOURNAL OF THE AMERICAN ACADEMY OF DERMATOLOGY Zackheim, H. S., Koo, J., LeBoit, P. E., McCalmont, T. H., Bowman, P. H., Kashani-Sabet, M., Jones, C., Zehnder, J. 2002; 47 (1): 155-157

    View details for DOI 10.1067/mjd.2002.120571

    View details for Web of Science ID 000176560400023

    View details for PubMedID 12077599

  • Homozygous factor V splice site mutation associated with severe factor V deficiency BLOOD Schrijver, I., Koerper, M. A., Jones, C. D., Zehnder, J. L. 2002; 99 (8): 3063-3065

    Abstract

    We investigated a family whose proband has a severe bleeding disorder and factor V antigenic and functional levels of 8% and less than 1% of control values, respectively. Molecular analysis of the factor V gene revealed a novel homozygous mutation in the last nucleotide of exon 10. 1701G>T causes activation of a cryptic exonic splice site in exon 10, which encodes part of the factor V heavy chain (A2 domain). This leads to the deletion of 35 nucleotides and results in a frameshift with a premature stop codon at amino acid position 498. The G1701 and corresponding Gln509 are conserved in murine, bovine, and porcine factor V and in human factor VIII. Few factor V deficiency mutations have been identified as yet. Several are present in the heterozygous form in combination with factor V Leiden (Arg506Gln). This is the first reported homozygous splice site mutation in a patient with factor V deficiency.

    View details for Web of Science ID 000174866500055

    View details for PubMedID 11929802

  • Randomized trial of folic acid for homocysteine reduction in end-stage renal disease. Wrone, E., Hornberger, J., Zehnder, J., McCann, L., Coplon, N., Fortmann, S. W B SAUNDERS CO. 2002: A34

    View details for Web of Science ID 000174835000126

  • The effect of a supplement containing Ginkgo biloba (Gb), fish oil (DHA) and Gotu kola (Gk) on platelet function in older healthy adults Carlson, J. J., Farquhar, J. W., Zehnder, J., Haskell, W., Miller, D. FEDERATION AMER SOC EXP BIOL. 2002: A646

    View details for Web of Science ID 000174533603589

  • Novel factor VC2-domain mutation (R2074H) in two families with factor V deficiency and bleeding THROMBOSIS AND HAEMOSTASIS Schrijver, I., Houissa-Kastally, R., Jones, C. D., Garcia, K. C., Zehnder, J. L. 2002; 87 (2): 294-299

    Abstract

    The molecular basis of Factor V deficiency has been defined in few patients only. We report a homozygous nucleotide change (G6395A) in two Tunisian probands with Factor V deficiency and bleeding episodes. This substitution results in the replacement of an arginine (R) by a histidine (H) in amino acid position 2074, located in the Factor V C2-domain. Mutations in this protein domain have not previously been described. Several lines of evidence support that this sequence variant is indeed disease causing: 1) Crystal structures of Factor V and molecular C2-domain modeling studies of H2074 suggest that the conserved R2074 is required for correct folding; 2) Structure-function studies of selective Factor V mutants (R2074A) demonstrate the importance of R2074 for structural stability of the Factor V C2-domain and for cofactor activity (1); 3) In Factor VIII, point mutations in codon 2209, which corresponds to position 2074 in Factor V, cause hemophilia A.

    View details for Web of Science ID 000173869300020

    View details for PubMedID 11858490

  • Immunological consequences of topical bovine thrombin AMERICAN JOURNAL OF PATHOLOGY Zehnder, J. L., Leung, L. L. 2001; 159 (6): 2371-2371

    View details for Web of Science ID 000172457400040

    View details for PubMedID 11733385

    View details for PubMedCentralID PMC1850577

  • Real-time PCR assay for quantitation of BCR-ABL mRNA in patients with chronic phase CML treated with the tyrosine kinase inhibitor GLEEVEC (TM) (STI-571). Gotlib, J., Jones, C. D., Zheng, K. H., Yeung, C., Dugan, K., Fechter, L., Falkow, R., Zehnder, J. L., Coutre, S. AMER SOC HEMATOLOGY. 2001: 258B–259B

    View details for Web of Science ID 000172134201177

  • A polymorphism in the BCL-6 gene is associated with follicle center lymphoma LEUKEMIA & LYMPHOMA Lossos, I. S., Jones, C. D., Zehnder, J. L., Levy, R. 2001; 42 (6): 1343-1350

    Abstract

    Follicle center lymphoma (FCL) accounts for approximately 40% of all non-Hodgkin's lymphomas (NHL). The genetic-environmental interactions involved in the etiology and pathogenesis of this disease are unknown. In our previous study a single nucleotide polymorphism (SNP) (397C) in the regulatory untranslated first intron region of the BCL-6 gene was found in four of the eight FCL patients but in none of the 10 healthy controls. To further evaluate the potential association between the 397C allele of the BCL-6 gene and FCL, we performed a case-control study. Genomic DNA was isolated from 85 FCL patients, from 98 control cases without a previous history of malignancy, treated at Stanford University Medical Center for non-malignant disorders and from 90 samples from the DNA Polymorphism Discovery Resource. The 397G and the 397C polymorphic alleles were identified by a PCR-RFLP method. To evaluate the possible effect of this polymorphism on gene expression, BCL-6 mRNA levels in nine FCL tumors with the 397G-G genotype and in nine FCL tumors with the 397G-C genotype were measured by quantitative real-time RT-PCR. The 397C polymorphic allele was found in 32 FCL cases (37.6%), in 20 controls (20.4%) and in 17 (18.9%) samples from the DNA Polymorphism Discovery Resource. The prevalence of the 397G-C and 397C-C genotypes was significantly higher in FCL cases than in control group (p = 0.01). No difference in BCL-6 gene expression was observed between FCL cases with 397G-G and 397G-C genotypes. The present study demonstrates a possible association between the 397C allele of the BCL-6 proto-oncogene and FCL. The similar levels of BCL-6 mRNA expression in 397G-G and in 397G-C FCL cases suggests that any possible oncogenic effect of the polymorphic allele would not simply be related to a direct effect on BCL-6 gene expression and suggests the existence of other FCL susceptibility genes that are in linkage disequilibrium with the 397C allele of the BCL-6 gene.

    View details for Web of Science ID 000173558700022

    View details for PubMedID 11911418

  • An MTHFR variant, homocysteine, and cardiovascular comorbidity in renal disease 32nd Annual Meeting of the American-Society-of-Nephrology Wrone, E. M., Zehnder, J. L., Hornberger, J. M., McCann, L. M., Coplon, N. S., Fortmann, S. P. NATURE PUBLISHING GROUP. 2001: 1106–13

    Abstract

    It is unclear whether total serum homocysteine (tHcy) and the C677T mutation of methylenetetrahydrofolate reductase (MTHFR) are associated with cardiovascular disease (CVD) in patients with end-stage renal disease (ESRD).A cross-sectional sample of 459 patients with ESRD on chronic dialysis was assessed to determine whether tHcy and the C677T mutation are associated with CVD prevalence in multiple logistic regression. As CVD mortality is high, we examined the relationship between homozygosity and duration of dialysis.Mean tHcy was higher in patients without a history of CVD (35.2 micromol/L vs. 30.4 micromol/L, P = 0.02). In multivariate models, CVD was negatively associated with tHcy and positively associated with TT genotype, male gender, and body mass index. Mean tHcy levels were higher among those with the TT genotype compared with those with the CC genotype when adjusted for age, folate, creatinine, and albumin (37.9 micromol/L vs. 31.9 micromol/L, P = 0.005). Among whites, the prevalence of the TT genotype was higher in those having undergone less than one year of dialysis (P = 0.002).The C677T genotype of MTHFR is associated with CVD in ESRD and may be a more meaningful marker than tHcy for abnormal homocysteine metabolism in ESRD. Prospective data from ongoing clinical trials are needed to improve our understanding of these findings. Screening for this polymorphism may help guide prevention measures.

    View details for Web of Science ID 000170668100029

    View details for PubMedID 11532106

  • Expression of a single gene, BCL-6, strongly predicts survival in patients with diffuse large B-cell lymphoma BLOOD Lossos, I. S., Jones, C. D., Warnke, R., Natkunam, Y., Kaizer, H., Zehnder, J. L., Tibshirani, R., Levy, R. 2001; 98 (4): 945-951

    Abstract

    Diffuse large B-cell lymphoma (DLBCL) is characterized by a marked degree of morphologic and clinical heterogeneity. Establishment of parameters that can predict outcome could help to identify patients who may benefit from risk-adjusted therapies. BCL-6 is a proto-oncogene commonly implicated in DLBCL pathogenesis. A real-time reverse transcription-polymerase chain reaction assay was established for accurate and reproducible determination of BCL-6 mRNA expression. The method was applied to evaluate the prognostic significance of BCL-6 expression in DLBCL. BCL-6 mRNA expression was assessed in tumor specimens obtained at the time of diagnosis from 22 patients with primary DLBCL. All patients were subsequently treated with anthracycline-based chemotherapy regimens. These patients could be divided into 2 DLBCL subgroups, one with high BCL-6 gene expression whose median overall survival (OS) time was 171 months and the other with low BCL-6 gene expression whose median OS was 24 months (P =.007). BCL-6 gene expression also predicted OS in an independent validation set of 39 patients with primary DLBCL (P =.01). BCL-6 protein expression, assessed by immunohistochemistry, also predicted longer OS in patients with DLBCL. BCL-6 gene expression was an independent survival predicting factor in multivariate analysis together with the elements of the International Prognostic Index (IPI) (P =.038). By contrast, the aggregate IPI score did not add further prognostic information to the patients' stratification by BCL-6 gene expression. High BCL-6 mRNA expression should be considered a new favorable prognostic factor in DLBCL and should be used in the stratification and the design of risk-adjusted therapies for patients with DLBCL. (Blood. 2001;98:945-951)

    View details for Web of Science ID 000170364100008

    View details for PubMedID 11493437

  • The transcriptional program in human mast cells stimulated via the Fc[epsilon]RI: New insights into the immunological functions of mast cells in allergic inflammation.epsilon Tsai, M., Sayama, K., Diehn, M., Zehnder, J., Jones, C., Botstein, D., Brown, P. O., GALLI, S. J. FEDERATION AMER SOC EXP BIOL. 2001: A1020–A1020

    View details for Web of Science ID 000167454201772

  • CD31 mismatching affects marrow transplantation outcome BIOLOGY OF BLOOD AND MARROW TRANSPLANTATION GRUMET, F. C., Hiraki, D. D., Brown, B. W., Zehnder, J. L., Zacks, E. S., Draksharapu, A., Parnes, J., Negrin, R. S. 2001; 7 (9): 503-512

    Abstract

    Graft-versus-host disease (GVHD) complicating allogeneic bone marrow transplantation (BMT) is often attributed to mismatched minor histocompatibility antigens (mHags), which are poorly defined in humans. CD31 is a candidate human mHag relevant to acute GVHD, but reports disagree about its level of significance, the role of HLA restriction, and the relative importance of different polymorphic codons within the molecule. We therefore examined in greater detail the impact of CD31-matching on BMT outcome in a prospective study from a single institution. Samples of recipient and donor DNA were collected pretransplantation for all patients receiving unmanipulated bone marrow from an HLA-identical sibling over a 45-month period at our institution. CD31 DNA typing of alleles at the 3 polymorphic codons 125 (L or V), 563 (N or S), and 670 (R or G) was performed for 118 patient-donor pairs plus 2 additional pairs who had codon 125 typing only. Donor-recipient CD31 nonidentity was tested for correlation with BMT clinical outcome measures of severe acute GVHD, chronic GVHD, relapse, and survival. Gene frequencies of approximately 0.5 for each allele at all 3 codons were comparable to previous reports. Because complete association was seen for 563N with 670G and for 563S with 670R, nonidentity for those codons was analyzed as a single genetic marker designated codon 563/670. Donor-recipient CD31 nonidentity was a significant risk factor for overall survival, both at codon 563/670 (hazard ratio [hr] = 2.58, P = .005) and at codon 125 (hr = 1.07, P = .036). Similar results held for disease-free survival. Nonidentity at codon 563/670 was also a significant risk factor (odds ratio [OR] = 11.15, P = .011) for severe (grades III, IV) versus no (grade 0) acute GVHD. Nonidentity at codon 125 posed less but still significant risk (OR = 9.30, P = .030). When the comparison group without severe acute GVHD was expanded to include grade I as well as grade 0 patients, the risk from CD31 nonidentity increased for both codon 563/670 (OR = 12.31, P = .010) and codon 125 (OR = 11.24, P = .011). CD31 nonidentity remained a significant independent risk factor for survival and for severe acute GVHD when tested in multivariate analysis with the covariates of adulthood, recipient-donor sex difference, ethnic group, disease, pretransplantation risk category, HLA-A2 type, B44-like types, and GVHD prophylactic regimen. CD31 nonidentity showed a trend but failed to achieve statistical significance as a risk factor for relapse and for chronic GVHD. In conclusion, donor-recipient CD31 nonidentity is a significant risk factor for survival and for severe acute GVHD in HLA-identical sibling BMT. The stronger associations with codon 563/670 suggest that polymorphism may be more important than the linked polymorphism at codon 125.

    View details for Web of Science ID 000171449200004

    View details for PubMedID 11669217

  • Lack of human herpesvirus 8 & Epstein-Barr virus in Kikuchi's histiocytic necrotizing lymphadenitis George, T. I., Jones, C. D., Zehnder, J., Warnke, R. A., Dorfman, R. F. NATURE PUBLISHING GROUP. 2001: 164A

    View details for Web of Science ID 000166634900973

  • CD10 and BCL-6 expression in high-grade marginal zone B-cell lymphoma Higgins, J. P., Jones, C. D., Zehnder, J. L., Warnke, R. A. NATURE PUBLISHING GROUP. 2001: 166A

    View details for Web of Science ID 000166634900987

  • Lack of human herpesvirus 8 & Epstein-Barr virus in Kikuchi's histiocytic necrotizing lymphadenitis George, T. I., Jones, C. D., Zehnder, J., Warnke, R. A., Dorfman, R. F. NATURE PUBLISHING GROUP. 2001: 164A

    View details for Web of Science ID 000166622400969

  • Sensitive detection of clonal immunoglobulin rearrangements in frozen and paraffin embedded tissues by polymerase chain reaction heteroduplex analysis DIAGNOSTIC MOLECULAR PATHOLOGY Ranheim, E. A., Jones, C. D., Zehnder, J. L. 2000; 9 (4): 177-183

    Abstract

    Molecular detection of a clonal population of B or T cells through analysis of rearranged antigen receptor genes is an essential adjunct to the morphologic, flow cytometric, and immunohistochemical evaluation of tissue specimens for the presence of leukemia or lymphoma. Combining polymerase chain reaction (PCR) with heteroduplex annealing and polyacrylamide gel electrophoresis (PAGE) has been used to detect clonal T-cell receptor rearrangements, particularly in skin biopsy specimens. The authors have developed a similar PCR heteroduplex assay for detection of clonal VDJ immunoglobulin gene rearrangements using two sets of primers based on relatively conserved consensus regions in the J(H) and framework I and 2 regions of the immunoglobulin heavy chain V region gene. This method is able to detect a clonal rearrangement when the clone comprises as little as 1% of the population in a polyclonal B-cell background. It may be used on fresh, frozen, or paraffin-embedded tissue and detects a clonal population in a majority of lymphoma subtypes. Compared with conventional PCR analysis, this method requires only a short additional cycle of denaturation and slow renaturation before PAGE. Interpretation is simplified as the clonal PCR product migrates away from the polyclonal background products.

    View details for Web of Science ID 000165556700001

    View details for PubMedID 11129440

  • BCL-6 proto-oncogene polymorphism and susceptibility to follicle center lymphoma (FCL). Lossos, I. S., Jones, C. D., Zehnder, J. L., Levy, R. AMER SOC HEMATOLOGY. 2000: 574A–575A

    View details for Web of Science ID 000165256102468

  • Real-time PCR assay for quantitation of BCL-6 mRNA. Lossos, I. S., Jones, C. D., Zehnder, J. L., Warnke, R., Levy, R. AMER SOC HEMATOLOGY. 2000: 166B

    View details for Web of Science ID 000165256200763

  • Identification of a novel factor VC2-domain mutation (R2074H) in two separate families with factor V deficiency and bleeding. Schrijver, Houissa-Kastally, R., Jones, C. D., Zehnder, J. L. AMER SOC HEMATOLOGY. 2000: 533A

    View details for Web of Science ID 000165256102292

  • Argatroban reexposure in patients with heparin-induced thrombocytopenia (HIT). Lewis, B. E., Wallis, D. E., Zehnder, J. L., Barton, J. C. AMER SOC HEMATOLOGY. 2000: 52A–52A

    View details for Web of Science ID 000165256100217

  • Search for ATM gene mutations in patients with follicle center lymphoma (FCL), in the FCL tumors and in tumors following transformation to a higher-grade. Lossos, I. S., Thorstenson, Y., Oefner, P. J., Chu, G., Levy, R. AMER SOC HEMATOLOGY. 2000: 574A-+

    View details for Web of Science ID 000165256102466

  • Peripheral T-cell lymphoma complicated by a proliferation of large B cells 88th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Higgins, J. P., van de Rijn, M., Jones, C. D., Zehnder, J. L., Warnke, R. A. AMER SOC CLINICAL PATHOLOGY. 2000: 236–47

    Abstract

    We studied 14 cases that showed a morphologic appearance of peripheral T-cell lymphoma and contained substantial numbers of CD20+ large B cells. In all but 2 cases, the CD20+ large cells showed a mix of kappa and lambda light chain expression. Two cases showed a focal predominance of kappa expression. In situ hybridization using the EBER1 probe for detection of Epstein-Barr virus (EBV) RNA was performed on every case. EBV RNA was present in 10 cases. Of 8 cases with EBV RNA stained by immunohistochemistry for the latent membrane protein of EBV, 6 were positive. Double-labeling immunohistochemistry and in situ hybridization confirmed that EBV was present in the large B cells. Polymerase chain reaction (PCR) analysis showed a clonal rearrangement of the T-cell receptor (TCR)-gamma chain gene in 12 of 13 cases tested. One additional case showed a clonal rearrangement of the TCR-beta chain gene by Southern blot hybridization. PCR analysis showed a clonal immunoglobulin gene rearrangement in 5 cases, a suggestion of a clonal rearrangement in 1, an oligoclonal pattern in 4, and a polyclonal pattern in 4. The finding of large B and T cells may result in a misdiagnosis of a reactive process or of T-cell-rich B-cell lymphoma. The presence of EBV in some cases could cause further confusion with the reactive T- and B-immunoblastic proliferation of infectious mononucleosis.

    View details for Web of Science ID 000088460700010

    View details for PubMedID 10941339

  • PCR-heteroduplex analysis of T-cell receptor gamma gene rearrangement in paraffin-embedded skin biopsies AMERICAN JOURNAL OF DERMATOPATHOLOGY Kohler, S., Jones, C. D., Warnke, R. A., Zehnder, J. L. 2000; 22 (4): 321-327

    Abstract

    We developed a rapid, simple, and sensitive method for the detection of T-cell receptor-gamma (TCRgamma) gene rearrangements in paraffin-embedded skin biopsies. Available techniques often require either fresh tissue, several primer pairs, nested amplifications, or specialized electrophoresis steps such as denaturing gradient gel electrophoresis. Our method is based on heteroduplex analysis of polymerase chain reaction (PCR) products of the TCRgamma in a nondenaturing modified polyacrylamide gel using a single pair of primers and is adapted for paraffin-embedded tissue. When tested against Southern blot analysis, the PCR results correlated in 8 of 9 cases. Six mature cutaneous B-cell lymphomas and 29 inflammatory skin disorders all resulted in a polyclonal amplification pattern. When analyzing 3-mm or 4-mm punch biopsies of 51 cases of cutaneous T-cell lymphoma, 37 (72.5%) showed a clonal rearrangement with this technique. For 7 cases of patch stage mycosis fungoides, frozen tissue and formalin-fixed and paraffin-embedded tissue was available, and in 5 of 7 cases (71%), the results in frozen and paraffin-embedded tissue were concordant. One case showed a clonal pattern in frozen tissue but not in paraffin-embedded tissue, and one case was polyclonal in frozen tissue but monoclonal in paraffin-embedded tissue. Using serial dilutions of DNA from a T-cell ALL in a polyclonal background (tonsil), we established a sensitivity of 0.5%. Heteroduplex PCR of the TCRgamma is a rapid, sensitive, and inexpensive screening procedure as well as a useful adjunct to histologic analysis and immunophenotyping of cutaneous T-cell proliferations.

    View details for Web of Science ID 000088565400005

    View details for PubMedID 10949457

  • Blastic/blastoid transformation of follicular lymphoma - Immunohistologic and molecular analyses of five cases AMERICAN JOURNAL OF SURGICAL PATHOLOGY Natkunam, Y., Warnke, R. A., Zehnder, J. L., Jones, C. D., Milatovich-Cherry, A., Cornbleet, P. J. 2000; 24 (4): 525-534

    Abstract

    Progression of follicular lymphoma to a higher-grade malignancy frequently heralds a poor prognosis. Clinical transformation is variably accompanied by a spectrum of histologic changes characterized by alteration in growth and cytology. Although several cytogenetic events and potential oncogenes have been documented in this progression, the underlying molecular mechanisms are largely unknown. We present five patients with an unusual histologic transformation of follicular lymphoma manifested by blastic/blastoid morphology. This transformation is histologically distinct from other types of transformation of follicular lymphoma. All five cases exhibited the t(14;18) translocation and expressed the BCL-2 protein. In addition, two of the five patients showed increased levels of the p53 protein within neoplastic cells implicating a possible role for this oncogene in blastic/blastoid transformation. The lack of BCL-1 and myeloid antigens by immunohistochemistry and flow cytometry studies served to distinguish blastic/blastoid transformation of follicular lymphoma from its morphologic mimics. This distinction is clinically important because lymphoblastic and myeloid leukemias require significantly different therapeutic modalities and show better prognosis. Moreover, the lack of Epstein-Barr virus-specific mRNA suggests that this virus is unlikely to participate in blastic/blastoid transformation of follicular lymphoma.

    View details for Web of Science ID 000086211700006

    View details for PubMedID 10757399

  • Antibodies against the first Ig-like domain of human platelet endothelial cell adhesion molecule-1 (PECAM-1) that inhibit PECAM-1-dependent homophilic adhesion block in vivo neutrophil recruitment JOURNAL OF IMMUNOLOGY Nakada, M. T., Amin, K., Christofidou-Solomidou, M., O'Brien, C. D., Sun, J., Gurubhagavatula, I., Heavner, G. A., Taylor, A. H., Paddock, C., Sun, Q. H., Zehnder, J. L., Newman, P. J., Albelda, S. M., DeLisser, H. M. 2000; 164 (1): 452-462

    Abstract

    Platelet endothelial cell adhesion molecule (PECAM-1), a member of the Ig superfamily, is found on endothelial cells and neutrophils and has been shown to be involved in the migration of leukocytes across the endothelium. Adhesion is mediated, at least in part, through binding interactions involving its first N-terminal Ig-like domain, but it is still unclear which sequences in this domain are required for in vivo function. Therefore, to identify functionally important regions of the first Ig-like domain of PECAM-1 that are required for the participation of PECAM-1 in in vivo neutrophil recruitment, a panel of mAbs against this region of PECAM-1 was generated and characterized in in vitro adhesion assays and in an in vivo model of cutaneous inflammation. It was observed that mAbs that disrupted PECAM-1-dependent homophilic adhesion in an L cell aggregation assay also blocked TNF-alpha-induced intradermal accumulation of neutrophils in a transmigration model using human skin transplanted onto SCID mice. Localization of the epitopes of these Abs indicated that these function-blocking Abs mapped to specific regions on either face of domain 1. This suggests that these regions of the first Ig-like domain may contain or be close to binding sites involved in PECAM-1-dependent homophilic adhesion, and thus may represent potential targets for the development of antiinflammatory reagents.

    View details for Web of Science ID 000084321200060

    View details for PubMedID 10605042

  • Blastic/blastoid transformation of follicular lymphoma: Clinicopathologic, immunophenotypic cytogenetic and molecular analyses of five cases. Natkunam, Y., Warnke, R. A., Zehnder, J. L., Milatovich-Cherry, A., Cornbleet, P. J. AMER SOC HEMATOLOGY. 1999: 251B

    View details for Web of Science ID 000083790701168

  • Familial coagulation factor V deficiency due to multiple discrete base pair substitutions. Jones, C. D., Parks, S. P., Koerper, M. A., Zehnder, J. L. AMER SOC HEMATOLOGY. 1999: 232A–233A

    View details for Web of Science ID 000083790301077

  • Kikuchi's necrotizing lymphadenitis and Kaposi's sarcoma-associated herpes virus. George, T. I., Dorfman, R. F., Jones, C. D., Zehnder, J. L. AMER SOC HEMATOLOGY. 1999: 52B

    View details for Web of Science ID 000083790700230

  • Real-time t(11;14) and t(14;18) PCR assays provide sensitive and quantitative assessments of minimal residual disease (MRD) LEUKEMIA Olsson, K., Gerard, C. J., Zehnder, J., Jones, C., Ramanathan, R., Reading, C., Hanania, E. G. 1999; 13 (11): 1833-1842

    Abstract

    Non-Hodgkin's lymphoma (NHL) arises as a clonal transformation of normal B and T cell differentiation and is often characterized by a higher incidence of specific chromosomal translocations. We have developed real-time TaqMan PCR assays directed toward two of these tumor-associated DNA markers, the t(14;18)(q32;q21.3) at the major breakpoint region of the bcl-2 gene and the t(11;14)(q13;q32) at the bcl-1 major translocation cluster. During analysis of serial dilutions of t(14;18)-positive DNA, the t(14;18) real-time PCR was at least as sensitive as nested PCR and demonstrated enhanced quantitative potential. Moreover, in a blinded comparison of the t(14;18) real-time PCR and a clinically validated nested PCR protocol using 134 cell line and patient DNA samples, the real-time PCR detected the translocation in 30.0% more cases than nested PCR. Both the t(14;18) and t(11;14) real-time PCR assays were used to quantitate minimal residual disease (MRD) in an NHL clinical trial assessing the safety and efficacy of a tumor-purging protocol in autologous stem cell transplantation. The assays were also used to evaluate disease depletion in an ex vivo tumor spiking model in which normal peripheral blood was spiked with tumor cell lines and processed according to the clinical purging method. PCR data from both the clinical trial and the ex vivo model demonstrated a 4 to 6 log reduction in tumor cells during CD34+ and CD34+ Thy-1+ enrichment. Because the t(14;18) and t(11;14) real-time PCR assays are very sensitive, quantitative, rapid, and require no post-PCR manipulation, they may serve as practical alternatives to nested PCR.

    View details for Web of Science ID 000083664000023

    View details for PubMedID 10557059

  • Familial coagulation factor V deficiency caused by a novel 4 base pair insertion in the factor V gene: factor V Stanford [erratum]. Thrombosis and haemostasis Zehnder, J. L., Hiraki, D. D., Jones, C. D., Gross, N., GRUMET, F. C. 1999; 82 (5): XII-?

    View details for PubMedID 10681265

  • Variability of polymerase chain reaction detection of the bcl-2-IgH translocation in an international multicentre study ANNALS OF ONCOLOGY Johnson, P. W., Swinbank, K., MacLennan, S., Colomer, D., Debuire, B., Diss, T., Gabert, J., Gupta, R. K., Haynes, A., Kneba, M., Lee, M. S., MacIntyre, E., Mensink, E., Moos, M., MORGAN, G. J., NERI, A., Johnson, A., Reato, G., Salles, G., van 't Veer, M. B., Zehnder, J. L., Zucca, E., Selby, P. J., Cotter, F. E. 1999; 10 (11): 1349-1354

    Abstract

    The capacity of the polymerase chain reaction (PCR) to detect very low numbers of cells bearing a t(14;18) translocation has led to its application in assessment of the results of treatment for follicular lymphoma, and suggestions that therapy might be guided by molecular studies. To test the reliability of PCR a collaborative study was undertaken to compare results from different laboratories in Europe and North America.Twenty laboratories with records of publication in molecular diagnostics were sent blood from normal donors with varying numbers of t(14;18)-bearing cells added from a cell line with a translocation in the major breakpoint region (MBR) of the bcl-2 gene. Samples contained 1000, 100, 10, 1 or 0 cells per ml of whole blood and were sent blinded in duplicate. PCR methodology varied widely, with the total number of amplification cycles between 30 and 70, and 13 different primers used for the MBR region. Twelve laboratories used nested PCR and eight single round amplification.The sensitivity of nested and single round PCR was similar at 100 cells/ml but below this the nested method proved significantly more sensitive. The false positive rate was 28%, with 11 samples from 9 laboratories reported as positive when no t(14;18) cells were added. PCR product size and sequence analysis showed that false positives were due to contamination from cell-line DNA rather than background translocations in the donors. There was no significant difference in false positive rates between nested and single round techniques.The polymerase chain reaction to detect bcl-2-IgH rearrangements is presently carried out with widely disparate results. Further effort is required to bring forward a standard PCR protocol which can be re-tested in different laboratories to improve accuracy and reproducibility. The application of quantitative techniques such as real-time PCR may resolve many of the problems presently encountered.

    View details for Web of Science ID 000084979300018

    View details for PubMedID 10631464

  • Familial coagulation factor V deficiency caused by a novel 4 base pair insertion in the factor V gene: Factor v Stanford THROMBOSIS AND HAEMOSTASIS Zehnder, J. L., Hiraki, D. D., Jones, C. D., Gross, N., GRUMET, F. C. 1999; 82 (3): 1097-1099

    Abstract

    An index patient with pseudohomozygosity for factor V Leiden was identified. Each of his two children inherited a different paternal factor V allele; a daughter was heterozygous for factor V Leiden, with 100% factor V activity, and a son was heterozygous for factor V deficiency, with 50% factor V activity. Genomic DNA was obtained from family members, and the 25 factor V exons and flanking intronic regions were sequenced in the proband and confirmed in the children. Within exon 13 of factor V, a 4 base insertion was found at NT 2856 in the proband and son. but not the daughter. This mutation, here designated factor V Stanford, results in a frameshift with loss of a thrombin activation site (R1545V) and premature termination of translation at amino acid 1560.

    View details for Web of Science ID 000082421100018

    View details for PubMedID 10494770

  • Mast-cell heparin demystified. Nature Zehnder, J. L., GALLI, S. J. 1999; 400 (6746): 714-715

    View details for PubMedID 10466718

  • Use of the polymerase chain reaction in the evaluation of cutaneous T-cell infiltrates DERMATOLOGIC CLINICS Kohler, S., Zehnder, J. L. 1999; 17 (3): 657-?

    Abstract

    Histologic evaluation of suspected cutaneous T-cell neoplasia is challenging. There is significant overlap with features of benign condition, and neoplastic cells often occur in a reactive background. Recently, molecular techniques using paraffin-embedded tissue have been applied to the diagnosis of cutaneous T-cell infiltrates. These methods are useful for determining whether a clonal population of T-cells is present. The advantages and limitation of molecular diagnostic methods in the diagnosis of cutaneous T-cell infiltrates are discussed.

    View details for Web of Science ID 000081318700015

    View details for PubMedID 10410865

  • Aggressive natural killer-like T-cell malignancy with leukemic presentation following solid organ transplantation AMERICAN JOURNAL OF CLINICAL PATHOLOGY Natkunam, Y., Warnke, R. A., Zehnder, J. L., Cornbleet, P. J. 1999; 111 (5): 663-671

    Abstract

    NK-like T-cell malignancies are part of a spectrum of lymphoproliferative diseases that complicate immunosuppression associated with solid organ transplantation. We describe 2 patients with long-standing immunosuppression following solid organ transplantation. Both patients had systemic symptoms that included fever, myalgia, and weight loss. Organ involvement and lymphadenopathy were not initially observed. Unique to these 2 cases are the initial leukemic symptoms, which led to further characterization and identification of NK-like T-cell malignancies. Both patients exhibited an anomalous T/NK phenotype, CD56 positivity, and atypical blastic architecture of the large granular lymphocytes. Clonal rearrangement of T-cell receptor genes was detected in both patients. In 1 patient, a cytogenetic abnormality involving 8q24 was demonstrated. The disease course in both patients was aggressive, with involvement of multiple sites and rapid demise. This study emphasizes the importance of including NK-like T-cell malignancies in the differential diagnosis of lymphoproliferative disorders associated with immunosuppression and recognizing that an aggressive clinical course may follow leukemic presentation of disease.

    View details for Web of Science ID 000079920500011

    View details for PubMedID 10230357

  • Aggressive cutaneous NK and NK-like T-cell lymphomas - Clinicopathologic, immunohistochemical, and molecular analyses of 12 cases AMERICAN JOURNAL OF SURGICAL PATHOLOGY Natkunam, Y., SMOLLER, B. R., Zehnder, J. L., Dorfman, R. F., Warnke, R. A. 1999; 23 (5): 571-581

    Abstract

    Natural killer (NK) and NK-like T-cell lymphomas are rare hematolymphoid malignancies that predominate in the upper aerodigestive system. They also involve other extranodal sites, including the skin. Primary cutaneous manifestations of NK and NK-like T-cell lymphomas are uncommon, and the clinicopathologic features are poorly understood. We have studied 12 patients of varied ethnic backgrounds with CD56-positive lymphomas in the skin. Six patients subsequently progressed to disseminated disease. These lymphomas showed the following immunophenotype: CD56+, CD43+, TCRb-, CD3-/+, CD20-, CD30-/+, CD4-, and CD8-. Two cases exhibited T-cell receptor gene rearrangements supporting a T-cell origin for these lymphomas, whereas the remaining 10 cases were likely derived from NK cells. Our results show inconsistent association of these lymphomas with Epstein-Barr virus (EBV), the multidrug resistance phenotype, and expression of P53. In addition, we found a previously unreported correlation between lymphomas harboring EBV mRNA and the expression of the multidrug resistance phenotype. These lymphomas were aggressive and were associated with rapid clinical progression, treatment failure, multiple relapses, and an average survival of 15 months from the time of diagnosis. Our results indicate the importance of recognizing this disease as a distinct subset of aggressive cutaneous lymphomas that may be diagnosed on the basis of morphology, immunophenotype, and gene rearrangement studies.

    View details for Web of Science ID 000080178200012

    View details for PubMedID 10328090

  • The international normalized ratio during concurrent warfarin and argatroban anticoagulation: Differential contributions of each agent and effects of the choice of thromboplastin used CLINICAL CHEMISTRY Hursting, M. J., Zehnder, J. L., Joffrion, J. L., Becker, J. C., Knappenberger, G. D., Schwarz, R. P. 1999; 45 (3): 409-412

    View details for Web of Science ID 000078979100015

    View details for PubMedID 10053045

  • Blinded comparison demonstrates that BCL-2 real-time PCR assay provides more sensitive and quantitative assessment of residual disease than nested PCR. Olsson, K., Zehnder, J., Jones, C., Cherry, A., Ramanathan, R., Reading, C., Hanania, E. G. W B SAUNDERS CO. 1998: 238A

    View details for Web of Science ID 000077121300966

  • Familial coagulation factor V deficiency caused by a novel 4 base pair insertion in the factor V gene. Zehnder, J. L., Hiraki, D., Jones, C., Gross, N., Grumet, F. C. W B SAUNDERS CO. 1998: 354A–355A

    View details for Web of Science ID 000077121301451

  • Sensitive detection of clonal immunoglobulin rearrangements in frozen and paraffin embedded tissues by PCR heteroduplex analysis. Ranheim, E. A., Jones, C. D., Zehnder, J. L. W B SAUNDERS CO. 1998: 229A

    View details for Web of Science ID 000077121300936

  • Performance of the Avocet(PT) prothrombin time (PT)/INR system. Ansell, J. E., Zweig, S., Meyer, B., Zehnder, J., Lewis, J. AMER SOC HEMATOLOGY. 1998: 112B–112B

    View details for Web of Science ID 000077121400477

  • Mesenteric artery thrombosis: A case report of combined protein S and protein C deficiency AMERICAN JOURNAL OF HEMATOLOGY Atkins, K. A., Zehnder, J. L. 1998; 58 (3): 246-247

    Abstract

    Individuals with more than one defect in natural coagulant/anticoagulant systems have been postulated to be at an increased risk for thrombotic events. We report a case of combined protein S and C deficiency in a young woman, which resulted in fatal arterial mesenteric thrombosis. The role of coagulation defects in arterial thrombosis is discussed.

    View details for Web of Science ID 000074510000017

    View details for PubMedID 9662280

  • Factor V Leiden and the risk of proximal venous thrombosis after total hip arthroplasty JOURNAL OF ARTHROPLASTY Woolson, S. T., Zehnder, J. L., Maloney, W. J. 1998; 13 (2): 207-210

    Abstract

    Deep vein thrombosis (DVT) remains a major cause of morbidity in patients undergoing total hip arthroplasty (THA). Despite postoperative DVT prophylaxis, 20-50% of THA patients still develop DVT. Currently, there is no accurate way of predicting which patients will develop DVT despite standard prophylaxis. The presence of factor V Leiden is the most common cause of inherited DVT risk. It has been postulated that patients who have factor V Leiden and are subjected to thrombogenic stressors such as THA would have an increased risk of thrombosis. The factor V Leiden genotype of 36 patients who developed proximal DVT after surgery and 45 control patients who had THA but did not develop DVT was determined. All patients had had prophylaxis against thrombosis using intermittent pneumatic compression alone or in combination with warfarin or aspirin. Surveillance for proximal DVT was performed on all patients prior to discharge by duplex ultrasound. The 2 groups were similar in age, sex, and type of operation. Three of 36 study patients who had developed DVT (8%) and 2 of 45 control patients who had not developed DVT (4%) were heterozygotes for factor V Leiden; these prevalences were not statistically different. Heterozygosity for factor V Leiden is not associated with DVT prophylaxis failure in patients undergoing THA.

    View details for Web of Science ID 000072498700014

    View details for PubMedID 9526216

  • A microplate allele-specific oligonucleotide hybridization assay for detection of factor V Leiden DIAGNOSTIC MOLECULAR PATHOLOGY Zehnder, J. L., Benson, R. C., Cheng, S. 1997; 6 (6): 347-352

    Abstract

    Factor V Leiden is the most common genetic risk factor for thrombosis. Currently, the determination of factor V Leiden genotype is limited to laboratories with expertise in molecular methods to develop "home brew" assays using polymerase chain reaction (PCR) to amplify genomic DNA, followed by analysis of Mnl I restriction fragments. These methods are not standardized, are labor intensive, and have low throughput. We describe a method for determination of factor V Leiden genotype using allele-specific oligonucleotide capture probes coated onto 96 well plates, requiring only a thermal cycler and a microplate spectrophotometer to perform. With an automated strip washer and plate reader, genotypes could be determined in 80 min from completion of PCR. Within-run and between-run coefficients of variation for the assay were < 10%. In all 160 cases studied, the microplate assay correctly identified the factor V genotype. The microplate oligonucleotide hybridization assay is a simple and reliable system for determination of factor V Leiden genotype. The assay offers an automatable, high-throughput alternative to current testing methodologies.

    View details for Web of Science ID 000072819400007

    View details for PubMedID 9559295

  • Cross-linking hybridization assay for direct detection of factor V Leiden mutation 29th Annual Oak Ridge Conference Zehnder, J., VANATTA, R., Jones, C., Sussman, H., Wood, M. AMER ASSOC CLINICAL CHEMISTRY. 1997: 1703–8

    Abstract

    A nucleic acid photocross-linking technology was used in the development of a direct assay for factor V Leiden, a point mutation in the factor V gene (G1691A) that is the most common inherited risk factor for thrombosis. This cross-linking hybridization assay included two allele-specific capture probes and six signal-generating reporter probes; all were modified with a photoactivated cross-linking compound. By using two different capture probes complementary to a 16-base sequence at the factor V Leiden mutation site, but differing in the nucleotide opposite the mutation site (C vs T), wild-type and factor V Leiden alleles were differentiated in purified DNA specimens. The assay was also successfully applied to genomic DNA in leukocytes isolated from whole blood; the factor V status of 122 patients as determined by this method was in complete concordance with a standard PCR-based assay and clearly discriminated between healthy individuals and factor V Leiden heterozygotes.

    View details for Web of Science ID A1997XV89700033

    View details for PubMedID 9299963

  • Heparin-induced platelet aggregation vs platelet factor 4 enzyme-linked immunosorbent assay in the diagnosis of heparin-induced thrombocytopenia-thrombosis AMERICAN JOURNAL OF CLINICAL PATHOLOGY Look, K. A., Sahud, M., Flaherty, S., Zehnder, J. L. 1997; 108 (1): 78-82

    Abstract

    Thrombosis occurs in an unpredictable subset of patients with heparin-induced thrombocytopenia (HIT). The diagnosis of HIT requires clinical suspicion and laboratory confirmation. Although the "gold-standard" diagnostic test is considered to be the serotonin release assay (SRA), most laboratories use heparin-induced platelet aggregation (HIPA), which is highly specific but reported to be less sensitive than the SRA. Recently, the heparin-platelet factor 4 (PF4) enzyme-linked immunosorbent assay (ELISA) has been reported to have comparable sensitivity to the SRA. We compared the HIPA and PF4 ELISA in serum samples from 146 patients examined for HIT and assessed whether either test predicted thrombotic risk. Results for 81 patients were positive for HIPA, PF4 ELISA, or both. Of these, 91% were HIPA-positive, while only 60% were PF4 ELISA-positive. Clinical information was available on 63 patients, 17 of whom had thrombotic events (10 venous, 6 arterial, and 1 both). Neither the HIPA nor the PF4 ELISA predicted thrombotic risk, but the HIPA proved to be a more sensitive test for laboratory confirmation.

    View details for Web of Science ID A1997XG01600013

    View details for PubMedID 9208982

  • Administration of a CD31-derived peptide delays the onset and significantly increases survival from lethal graft-versus-host disease BLOOD Chen, Y. F., Schlegel, P. G., Tran, N. P., Thompson, D., Zehnder, J. L., Chao, N. J. 1997; 89 (4): 1452-1459

    Abstract

    The CD31 monoclonal antibody, LYP21, binds to the CD31 domain 6 and inhibits the human mixed-lymphocyte reaction (MLR) in a specific and dose-dependent fashion. A synthetic CD31 peptide based on human CD31 epitope (amino acids 551 to 574) recognized by LYP21 is equally effective in inhibiting the MLR. In this study, we used the murine homolog of CD31 peptide 551 to 574 and a control peptide to study the role of CD31 molecule on T-cell activation. In vitro, CD31 peptide inhibited the MLR across several major and minor histocompatibility differences in a specific and dose-dependent fashion, similar to the results observed in the human system. Maximal inhibition was achieved at a dose of 200 microg/mL. In the cytotoxic T-lymphocyte (CTL) assay, CD31 peptide inhibited CTL responses by 97%. To study the in vivo effect of this peptide, graft-versus-host disease (GVHD) across minor histocompatibility barriers was induced in the B10.D2 (H-2d) --> BALB/c (H-2d) model. BALB/c recipients received CD31 peptide (100 microg/d), or phosphate-buffered saline (PBS), or control peptide (100 microg/d) intraperitoneally (IP) for the first 5 weeks. CD31 peptide delayed onset of graft-versus-host disease and significantly increased long-term survival. Twelve of 14 mice receiving CD31 peptide survived more than 100 days after transplantation, as compared with none of 10 mice receiving PBS and none of five mice receiving control peptide (P = .0001). Long-term engraftment of allogeneic bone marrow was documented in all transplanted mice by polymerase chain reaction (PCR) analysis of microsatellite region in the interleukin (IL)-1beta gene. Our data suggest that the CD31 molecule has an important functional role in T-cell activation in vitro and in vivo.

    View details for Web of Science ID A1997WG79700038

    View details for PubMedID 9028970

  • Sensitivity and specificity of the APC resistance assay in detection of individuals with factor V Leiden AMERICAN JOURNAL OF CLINICAL PATHOLOGY Zehnder, J. L., Benson, R. C. 1996; 106 (1): 107-111

    Abstract

    Resistance to activated protein C (APC) is the most common cause of familial thrombophilia. The partial thromboplastin time (PTT)-based test for resistance to APC has been widely employed as a screening test for this disorder. However, the utility of this test for screening is not well characterized. More than 90% of patients with resistance to APC have the G1691A mutation in factor V (factor V Leiden). The authors studied the ability of a commercial APC resistance assay to correctly identify the factor V Leiden genotype in 130 individuals. At the recommended assay cut-off value of 2, the sensitivity of the APC resistance assay was 50%, with a specificity of 98%. Increasing the cut-off value increased the sensitivity but decreased the specificity of the test. Receiver operating characteristic (ROC) curve analysis indicated that the test was of intermediate utility. There was considerable overlap in APC ratios in the range of 2 to 3 between subjects with a normal factor V genotype and heterozygotes for factor V Leiden. The authors conclude that the APC resistance assay in its present form is not a useful screening test for factor V Leiden heterozygotes. Until the performance of this assay is improved, patients should have molecular diagnostic testing performed to determine their factor V Leiden status.

    View details for Web of Science ID A1996UW50500018

    View details for PubMedID 8701918

  • Recurrent thrombosis due to compound heterozygosity for factor V Leiden and factor V deficiency BLOOD COAGULATION & FIBRINOLYSIS Zehnder, J. L., Jain, M. 1996; 7 (3): 361-362

    Abstract

    A point mutation in the factor V gene (factor V Leiden) is the most common cause of familial thrombophilia. Patients with factor V Leiden have an increased risk of thrombosis, particularly those homozygous for the mutation. However, the phenotype in individuals with the mutation is variable, suggesting that other factors influence thrombotic risk. We describe for the first time a family in which two independent defects in factor V co-exist: heterozygosity for factor V Leiden and factor V deficiency. Compound heterozygosity for these two defects results in a phenotype similar to a homozygous factor V Leiden state with profound resistance to APC and recurrent thrombosis.

    View details for Web of Science ID A1996UL37900012

    View details for PubMedID 8735145

  • Polymorphism of adhesion molecule CD31 and its role in acute graft-versus-host disease NEW ENGLAND JOURNAL OF MEDICINE Behar, E., Chao, N. J., Hiraki, D. D., Krishnaswamy, S., Brown, B. W., Zehnder, J. L., GRUMET, F. C. 1996; 334 (5): 286-291

    Abstract

    Graft-versus-host disease (GVHD) caused by poorly defined minor (i.e., other than HLA) histocompatibility antigens remains a serious problem in recipients of bone marrow transplants. We sought to determine whether the CD31 adhesion molecule is a minor alloantigen.We directly sequenced samples of complementary DNA (cDNA) encoding CD31 molecules from 21 unrelated normal subjects. Sequence-specific primers were then designed to amplify alleles by the polymerase chain reaction, thereby permitting CD31 typing of genomic DNA from additional normal subjects. To assess the relevance of CD31 matching to bone marrow transplantation, we performed CD31 typing of 46 recipients of bone marrow (32 without GVHD and 14 with severe [grade III or IV] acute GVHD) and their HLA-identical sibling donors. The immunoreactivity of CD31 phenotypes with anti-CD31 monoclonal antibodies was compared by flow cytometry.Direct sequencing of cDNA for CD31 from the 21 normal subjects identified a single polymorphism, CTG-->GTG (Leu-->Val), at codon 125; we designated the resulting alleles CD31.L and CD31.V, respectively. The CD31 genotypes of these and 142 other unrelated subjects were of the expected frequencies. Among the transplant recipients, 71 percent of those with acute GVHD had CD31 genotypes that were not identical to the donor's genotype, as compared with 22 percent of the recipients without GVHD (P = 0.004). The binding of anti-CD31 monoclonal antibodies as measured by fluorescence-activated cell sorting correlated with the CD31 types of homozygous cell lines.The adhesion molecule CD31 is polymorphic. When donor and recipient genotypes are not identical, the risk of GVHD increases. Prospective CD31 typing may reduce the risk of acute GVHD.

    View details for Web of Science ID A1996TV69500002

    View details for PubMedID 8532023

  • Administration of a CD31-derived peptide reduces lethal graft-versus-host disease across the minor histocompatibility complex barrier in mice Chen, Y., Schlegel, P. G., Tran, N., Thompson, D., Zehnder, J. L., Chao, N. J. AMER SOC HEMATOLOGY. 1995: 2286–86

    View details for Web of Science ID A1995TH91002287

  • PECAM-1 (CD31) is a minor histocompatibility determinant in acute GVHD. Chao, N. J., Behar, E., Hiraki, D., Krishnaswamy, S., Brown, B. W., Zehnder, J., Grumet, F. C. W B SAUNDERS CO. 1995: 2475

    View details for Web of Science ID A1995TH91002476

  • HISTIDINE-RICH GLYCOPROTEIN - IS THERE A ROLE IN HEMOSTASIS OR IMMUNE FUNCTION JOURNAL OF LABORATORY AND CLINICAL MEDICINE Zehnder, J. L., Leung, L. L. 1995; 125 (6): 682-683

    View details for Web of Science ID A1995RB44500004

    View details for PubMedID 7769360

  • LAMININ MEDIATES AGGREGATION OF CD31-TRANSFECTED CELLS Zehnder, J. L., BUTCHER, E. C., Leung, L. L. SCHATTAUER GMBH-VERLAG MEDIZIN NATURWISSENSCHAFTEN. 1995: 1153–53

    View details for Web of Science ID A1995RP38500966

  • INVOLVEMENT OF CD31 IN LYMPHOCYTE-MEDIATED IMMUNE-RESPONSES - IMPORTANCE OF THE MEMBRANE-PROXIMAL IMMUNOGLOBULIN DOMAIN AND IDENTIFICATION OF AN INHIBITING CD31 PEPTIDE BLOOD Zehnder, J. L., Shatsky, M., Leung, L. L., BUTCHER, E. C., McGregor, J. L., Levitt, L. J. 1995; 85 (5): 1282-1288

    Abstract

    CD31 (PECAM-1) is an immunoglobulin gene superfamily cell adhesion molecule found on vascular endothelium, platelets, and leukocytes. Lymphocyte expression of CD31 is most closely associated with the CD45RA+CD8+ naive T phenotype. CD31 has recently been shown to play a role in leukocyte egress to inflammatory sites. The mechanism of CD31 adhesion remains under investigation. Several investigators have reported evidence for a heterotypic ligand. We have previously shown that CD31 is phosphorylated with cell activation, which suggests a possible role for CD31 in cell activation events. We therefore studied the effects of CD31 antibodies on in vitro assays of lymphocyte activation. One CD31 antibody, LYP21, inhibited the mixed lymphocyte reaction (MLR) in a specific and dose-dependent fashion. An LYP21 epitope was localized to the sixth Ig domain of CD31. This peptide and a scrambled control peptide were synthesized and used to study effects of this epitope on lymphocyte activation. The CD31 peptide strongly inhibited the MLR. Because CD31 is expressed on both stimulator and responder populations, stimulator peripheral blood leukocytes and responder lymphocyte populations were separately incubated with CD31 peptide or control peptide and then washed before mixing. The CD31 peptide inhibited the MLR equally when either stimulator or responder cells were preincubated with the CD31 peptide. We further sorted responder cells into CD31-high and CD31-low populations and separately incubated these subsets with peptides. The CD31 peptide strongly inhibited MLRs, regardless of level of responder-cell CD31 expression. Examination of MLR reactions involving the CD31 peptide showed dispersed small aggregates of cells, rather than the single large aggregate observed in control MLRs. The CD31 peptide did not affect activation of lymphocytes by phorbol myristate acetate (PMA) and ionomycin. These results suggest that a surface CD31-ligand interaction may have a functional role in alloimmune lymphocyte activation and identify a functionally important domain of CD31.

    View details for Web of Science ID A1995QJ43500018

    View details for PubMedID 7858258

  • ACQUIRED ACTIVATED PROTEIN-C RESISTANCE CAUSED BY FACTOR-VA ANTIBODY - A POSSIBLE MECHANISM OF INCREASED THROMBOSIS IN THE ANTIPHOSPHOLIPID ANTIBODY SYNDROME LIN, R. Z., ZEHNDER, J. L. W B SAUNDERS CO. 1994: A83

    View details for Web of Science ID A1994PR75400319

  • ADHESION MECHANISM OF CD31 (PECAM-1) ZEHNDER, J. L., BUTCHER, E. C. SLACK INC. 1994: A129

    View details for Web of Science ID A1994NF02000103

  • THE CELL-ADHESION MOLECULE CD31 IS PHOSPHORYLATED AFTER CELL ACTIVATION - DOWN-REGULATION OF CD31 IN ACTIVATED LYMPHOCYTES-T JOURNAL OF BIOLOGICAL CHEMISTRY Zehnder, J. L., Hirai, K., Shatsky, M., McGregor, J. L., Levitt, L. J., Leung, L. L. 1992; 267 (8): 5243-5249

    Abstract

    We report the independent cloning of the cDNA for CD31, a recently described cell adhesion molecule of the immunoglobulin gene superfamily present on platelets, granulocytes, monocytes, lymphocytes, and endothelial cells. Northern analysis revealed three major mRNA transcripts in Jurkat (a human T cell line) and K562 and HEL (leukemia cell lines) cells with an additional 5.3-kilobase transcript seen in cultured human umbilical vein endothelial cells. Following T cell activation, CD31 mRNA was down-regulated by Northern analysis, and decreased CD31 protein expression was confirmed by immunoblots. The down-regulation of CD31 was partially mediated by decreased transcription as demonstrated by nuclear run-on studies. CD31 became rapidly phosphorylated in platelets, Jurkat cells, and endothelial cells after cell activation. We were unable to demonstrate the presence of a phosphotyrosine in CD31 using monoclonal and polyclonal phosphotyrosine antibodies. In addition, CD31 phosphorylation in platelets was induced by phorbol ester and was blocked by staurosporin, a protein kinase C inhibitor, suggesting that CD31 phosphorylation is mediated by protein kinase C and involves serine and/or threonine residues. The phosphorylation of CD31 following cell activation may modulate its cellular adhesiveness, and the down-regulation of its expression may serve to impart target specificity and to localize effector lymphocytes to areas of inflammation.

    View details for Web of Science ID A1992HH74700040

    View details for PubMedID 1544907

  • DEVELOPMENT OF ANTIBODIES TO THROMBIN AND FACTOR-V WITH RECURRENT BLEEDING IN A PATIENT EXPOSED TO TOPICAL BOVINE THROMBIN BLOOD Zehnder, J. L., Leung, L. L. 1990; 76 (10): 2011-2016

    Abstract

    A 65 year old patient who was exposed to topical bovine thrombin during cardiac surgery developed markedly prolonged clotting times and a severe bleeding diathesis. Mixing studies with normal plasma failed to correct the clotting times. Platelet transfusions, immunosuppressive and immunomodulatory therapies were ineffective, but plasmapheresis was effective in decreasing clotting times and in the resolution of clinical bleeding events. The patient's purified IgG reacted with bovine thrombin by immunoblotting and enzyme-linked immunosorbent assay (ELISA). However, the IgG reacted minimally with human thrombin. In view of the severe bleeding, a coexisting inhibitor was sought. The patient's factor V activity was 1% of normal and was not corrected by mixing with normal plasma, demonstrating the presence of an inhibitor against factor V. The patient's IgG reacted with both bovine and human factor V. Immunoblotting localized the site of antibody binding to the light chain of activated bovine factor V. Detectable amounts of bovine factor V were found in commercial bovine thrombin preparations by ELISA. The data suggest that patients exposed to topical bovine thrombin may develop antibodies to thrombin and factor V. Anti-thrombin antibodies may mask coexisting factor V inhibitors responsible for clinical bleeding.

    View details for Web of Science ID A1990EJ39100016

    View details for PubMedID 2242423

  • DEVELOPMENT OF ANTIBODIES TO THROMBIN AND FACTOR-V WITH RECURRENT BLEEDING IN A PATIENT EXPOSED TO TOPICAL BOVINE THROMBIN Zehnder, J. L., Leung, L. L. SLACK INC. 1990: A427–A427

    View details for Web of Science ID A1990CZ24401154

  • A PLATELET MEMBRANE GLYCOPROTEIN, GP128, IS SYNTHESIZED BY HUMAN VASCULAR ENDOTHELIAL-CELLS AND SECRETED INTO THE MEDIA Zehnder, J. L., Parmentier, S., McGregor, J. L., Leung, L. L. SCHATTAUER GMBH-VERLAG MEDIZIN NATURWISSENSCHAFTEN. 1989: 44–44

    View details for Web of Science ID A1989AR58400138

  • TORSADES-DE-POINTES PRECIPITATED BY A CHINESE HERBAL REMEDY AMERICAN JOURNAL OF CARDIOLOGY Bryerash, M., Zehnder, J., ANGELCHIK, P., Maisel, A. 1987; 60 (14): 1186-1187

    View details for Web of Science ID A1987K936300016

    View details for PubMedID 3687752

  • REINVESTIGATION OF THE REACTION OF CHYMOTRYPSIN WITH N-FURYLACRYLOYLTRYPTOPHAN DERIVATIVES AT ACIDIC PH BIOCHEMICAL JOURNAL Fink, A. L., Feldman, R., Zehnder, J. 1979; 181 (3): 733-736

    Abstract

    The reaction of alpha-chymotrypsin with N alpha-3-(2-furyl)acryloyl-L-tryptophan methyl ester (FA-Trp-OMe) and amide has been investigated in aqueous and dimethylsulphoxide cryosolvent solutions from pH2 to 7 and over a wide temperature range. Previous reports have suggested that an intermediate preceding the acyl-enzyme can be detected spectrophotometrically in the reaction with methyl esters of FA-Trp and FA-Tyr at low pH [Yu & Viswanatha (1969) Eur. J. Biochem. 11, 347--352), and that this intermediate is an oxazolinone [Coletti-Previero et al. (1970) FEBS Lett. 11, 213--217]. We show that the previous interpretations of the time-dependent spectral changes were incorrect, and that the only detected intermediate is the acyl-enzyme. This may be isolated by gel filtration at pH less than 2.5, 1 degree C, owing to its relative stability. The pH-dependence of the rates of acylation and deacylation from pH 8.5 to 2.0 are consistent with a single ionization of pK congruent to 7.0 in both aqueous and cryosolvent solutions.

    View details for Web of Science ID A1979HL98500027

    View details for PubMedID 42387