Mark Pegram

All Publications

• First-in-Human, First-in-Class Phase I Trial of the Anti-CD47 Antibody Hu5F9-G4 in Patients With Advanced Cancers JOURNAL OF CLINICAL ONCOLOGY Sikic, B., Lakhani, N., Patnaik, A., Shah, S. A., Chandana, S. R., Rasco, D., Colevas, A., O'Rourke, T., Narayanan, S., Papadopoulos, K., Fisher, G. A., Villalobos, V., Prohaska, S. S., Howard, M., Beeram, M., Chao, M. P., Agoram, B., Chen, J. Y., Huang, J., Axt, M., Liu, J., Volkmer, J., Majeti, R., Weissman, I. L., Takimoto, C. H., Supan, D., Wakelee, H. A., Aoki, R., Pegram, M. D., Padda, S. K. 2019; 37 (12): 946-+

  View details for DOI 10.1200/JCO.18.02018

  View details for Web of Science ID 000466717700002

• Clinical validation of an immunohistochemistry-based CanAssist-Breast test for distant recurrence prediction in hormone receptor-positive breast cancer patients. Cancer medicine Bakre, M. M., Ramkumar, C., Attuluri, A. K., Basavaraj, C., Prakash, C., Buturovic, L., Madhav, L., Naidu, N., R, P., Somashekhar, S. P., Gupta, S., Doval, D. C., Pegram, M. D. 2019

  Abstract

  CanAssist-Breast (CAB) is an immunohistochemistry (IHC)-based prognostic test for early-stage Hormone Receptor (HR+)-positive breast cancer patients. CAB uses a Support Vector Machine (SVM) trained algorithm which utilizes expression levels of five biomarkers (CD44, ABCC4, ABCC11, N-Cadherin, and Pan-Cadherin) and three clinical parameters such as tumor size, grade, and node status as inputs to generate a risk score and categorizes patients as low- or high-risk for distant recurrence within 5years of diagnosis. In this study, we present clinical validation of CAB. CAB was validated using a retrospective cohort of 857 patients. All patients were treated either with endocrine therapy or chemoendocrine therapy. Risk categorization by CAB was analyzed by calculating Distant Metastasis-Free Survival (DMFS) and recurrence rates using Kaplan-Meier survival curves. Multivariate analysis was performed to calculate Hazard ratios (HR) for CAB high-risk vs low-risk patients. The results showed that Distant Metastasis-Free Survival (DMFS) was significantly different (P-0.002) between low- (DMFS: 95%) and high-risk (DMFS: 80%) categories in the endocrine therapy treated alone subgroup (n=195) as well as in the total cohort (n=857, low-risk DMFS: 95%, high-risk DMFS: 84%, P<0.0001). In addition, the segregation of the risk categories was significant (P=0.0005) in node-positive patients, with a difference in DMFS of 12%. In multivariate analysis, CAB risk score was the most significant predictor of distant recurrence with hazard ratio of 3.2048 (P<0.0001). CAB stratified patients into discrete risk categories with high statistical significance compared to Ki-67 and IHC4 score-based stratification. CAB stratified a higher percentage of the cohort (82%) as low-risk than IHC4 score (41.6%) and could re-stratify >74% of high Ki-67 and IHC4 score intermediate-risk zone patients into low-risk category. Overall the data suggest that CAB can effectively predict risk of distant recurrence with clear dichotomous high- or low-risk categorization.

  View details for PubMedID 30848103

• First-in-Human, First-in-Class Phase I Trial of the Anti-CD47 Antibody Hu5F9-G4 in Patients With Advanced Cancers. Journal of clinical oncology : official journal of the American Society of Clinical Oncology Sikic, B. I., Lakhani, N., Patnaik, A., Shah, S. A., Chandana, S. R., Rasco, D., Colevas, A. D., O'Rourke, T., Narayanan, S., Papadopoulos, K., Fisher, G. A., Villalobos, V., Prohaska, S. S., Howard, M., Beeram, M., Chao, M. P., Agoram, B., Chen, J. Y., Huang, J., Axt, M., Liu, J., Volkmer, J., Majeti, R., Weissman, I. L., Takimoto, C. H., Supan, D., Wakelee, H. A., Aoki, R., Pegram, M. D., Padda, S. K. 2019: JCO1802018

  Abstract

  PURPOSE: To evaluate the safety, pharmacokinetics, and pharmacodynamics of Hu5F9-G4 (5F9), a humanized IgG4 antibody that targets CD47 to enable phagocytosis.PATIENTS AND METHODS: Adult patients with solid tumors were treated in four cohorts: part A, to determine a priming dose; part B, to determine a weekly maintenance dose; part C, to study a loading dose in week 2; and a tumor biopsy cohort.RESULTS: Sixty-two patients were treated: 11 in part A, 14 in B, 22 in C, and 15 in the biopsy cohort. Part A used doses that ranged from 0.1 to 3 mg/kg. On the basis of tolerability and receptor occupancy studies that showed 100% CD47 saturation on RBCs, 1 mg/kg was selected as the priming dose. In subsequent groups, patients were treated with maintenance doses that ranged from 3 to 45 mg/kg, and most toxicities were mild to moderate. These included transient anemia (57% of patients), hemagglutination on peripheral blood smear (36%), fatigue (64%), headaches (50%), fever (45%), chills (45%), hyperbilirubinemia (34%), lymphopenia (34%), infusion-related reactions (34%), and arthralgias (18%). No maximum tolerated dose was reached with maintenance doses up to 45 mg/kg. At doses of 10 mg/kg or more, the CD47 antigen sink was saturated by 5F9, and a 5F9 half-life of approximately 13 days was observed. Strong antibody staining of tumor tissue was observed in a patient at 30 mg/kg. Two patients with ovarian/fallopian tube cancers had partial remissions for 5.2 and 9.2 months.CONCLUSION: 5F9 is well tolerated using a priming dose at 1 mg/kg on day 1 followed by maintenance doses of up to 45 mg/kg weekly.

  View details for PubMedID 30811285

• Induced Pluripotent Stem Cells as a Novel Cancer Vaccine. Expert opinion on biological therapy Wang, L., Pegram, M. D., Wu, J. C. 2019

  Abstract

  Introduction: Although many current cancer therapies are effective, the mortality rate globally is unacceptably high. Cancer remains the second leading cause of death worldwide after heart disease and has caused nearly 10 million deaths in 2018. Additionally, current preventive therapies for cancer are underdeveloped, undermining the quality of life of high-risk individuals. Therefore, new treatment options for targeting cancer are urgently needed. In a recent study, researchers adopted an autologous iPSC-based vaccine to present a broad spectrum of tumor antigens to the immune system and succeeded in orchestrating a strong prophylactic immunity towards multiple types of cancer in mice. Areas Covered: In this review, we provide an overview of how cancer develops, the role of immune surveillance in cancer progression, the current status and challenges of cancer immunotherapy as well as the genetic overlap between pluripotent stem cells and cancer cells. Finally, we discuss the rationale for an autologous iPSC-based vaccine and its applications in murine cancer models. Expert Opinion: The autologous iPSC-based vaccine is a promising preventive and therapeutic strategy for fighting various types of cancers. Continuing efforts and clinical/translational follow-up studies may bring an autologous iPSC-based cancer vaccination approach from bench to bedside.

  View details for DOI 10.1080/14712598.2019.1650909

  View details for PubMedID 31364894

• PF-05280014 (a trastuzumab biosimilar) plus paclitaxel compared with reference trastuzumab plus paclitaxel for HER2-positive metastatic breast cancer: a randomised, double-blind study. British journal of cancer Pegram, M. D., Bondarenko, I., Zorzetto, M. M., Hingmire, S., Iwase, H., Krivorotko, P. V., Lee, K. S., Li, R. K., Pikiel, J., Aggarwal, R., Ewesuedo, R., Freyman, A., Li, R., Vana, A., Yin, D., Zacharchuk, C., Tan-Chiu, E. 2018

  Abstract

  BACKGROUND: This randomised, double-blind study compared PF-05280014 (a trastuzumab biosimilar) with reference trastuzumab (Herceptin) sourced from the European Union (trastuzumab-EU), when each was given with paclitaxel as first-line treatment for HER2-positive metastatic breast cancer.METHODS: Between 4 April 2014 and 22 January 2016, 707 participants were randomised 1:1 to receive intravenous PF-05280014 plus paclitaxel (PF-05280014 group; n=352) or trastuzumab-EU plus paclitaxel (trastuzumab-EU group; n=355). PF-05280014 or trastuzumab-EU was administered weekly (first dose 4mg/kg, subsequent doses 2mg/kg), with the option to change to a 3-weekly regimen (6mg/kg) from Week 33. Treatment with PF-05280014 or trastuzumab-EU could continue until disease progression. Paclitaxel (starting dose 80mg/m2) was administered on Days 1, 8 and 15 of 28-day cycles for at least six cycles or until maximal benefit of response. The primary endpoint was objective response rate (ORR), evaluating responses achieved by Week 25 and confirmed by Week 33, based on blinded central radiology review.RESULTS: The risk ratio for ORR was 0.940 (95% CI: 0.842-1.049). The 95% CI fell within the pre-specified equivalence margin of 0.80-1.25. ORR was 62.5% (95% CI: 57.2-67.6%) in the PF-05280014 group and 66.5% (95% CI: 61.3-71.4%) in the trastuzumab-EU group. As of data cut-off on 11 January 2017 (using data up to 378 days post-randomisation), there were no notable differences between groups in progression-free survival (median: 12.16 months in the PF-05280014 group vs. 12.06 months in the trastuzumab-EU group; 1-year rate: 54% vs. 51%) or overall survival (median: not reached in either group; 1-year rate: 89.31% vs. 87.36%). Safety outcomes and immunogenicity were similar between the treatment groups.CONCLUSION: When given as first-line treatment for HER2-positive metastatic breast cancer, PF-05280014 plus paclitaxel demonstrated equivalence to trastuzumab-EU plus paclitaxel in terms of ORR.CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov, NCT01989676.

  View details for PubMedID 30568294

• Use of cyclin-dependent kinase (CDK) 4/6 inhibitors for hormone receptor-positive, human epidermal growth factor receptor 2-negative, metastatic breast cancer: a roundtable discussion by The Breast Cancer Therapy Expert Group (BCTEG) BREAST CANCER RESEARCH AND TREATMENT Abraham, J., Coleman, R., Elias, A., Holmes, F., Kalinsky, K., Kittaneh, M., Lower, E., Mahtani, R., Mamounas, E., Pegram, M., Vogel, C., Breast Canc Therapy Expert Grp B 2018; 171 (1): 11?20

  Abstract

  To provide an overview of clinical data supporting the use of cyclin-dependent kinases 4 and 6 (CDK 4/6) inhibitors in the treatment of hormone receptor-positive (HR+), human epidermal growth factor receptor 2-negative (HER2-), metastatic breast cancer (mBC), from the perspective of the practicing oncologist community.A recent roundtable discussion was convened by The Breast Cancer Therapy Expert Group (BCTEG) to review existing data on this topic and its impact on their current practice.Level 1 evidence now supports use of a CDK 4/6 inhibitor in combination with endocrine therapy for patients with HR+, HER2-, mBC. Currently, there are no biomarkers that reliably define patients who will, or will not, benefit from the addition of a CDK 4/6 inhibitor to their endocrine therapy. Additional research is needed to identify the optimal sequencing of CDK 4/6 inhibitors in relation to other therapies as well as the optimal duration of therapy; at present, evidence suggests that use in the upfront setting is better than waiting for a later line of therapy, or adding after endocrine therapy has started.Thus far, three CDK 4/6 inhibitors-palbociclib, ribociclib, and more recently, abemaciclib-have been approved for use in the setting of HR+, HER2-, mBC.  The degrees to which these agents differ in terms of CDK4/6 affinity, side-effect profiles, dosing, degree of central nervous system (CNS) penetration, optimal use in combination with antiestrogen therapy, and across other subsets of breast cancer, remain an active area of investigation.

  View details for PubMedID 29725889

• High caveolin-1 expression in African American women with early-stage triple-negative breast cancer Baumbach-Reardon, L. L., Getz, J., Ahearn, M. E., Carpten, J., Pegram, M. AMER ASSOC CANCER RESEARCH. 2018

  View details for DOI 10.1158/1538-7445.AM2018-4352

  View details for Web of Science ID 000468819503016

• Generation of HER2-specific antibody immunity during trastuzumab adjuvant therapy associates with reduced relapse in resected HER2 breast cancer BREAST CANCER RESEARCH Norton, N., Fox, N., McCarl, C., Tenner, K. S., Ballman, K., Erskine, C. L., Necela, B. M., Northfelt, D., Tan, W. W., Calfa, C., Pegram, M., Colon-Otero, G., Perez, E. A., Clynes, R., Knutson, K. L. 2018; 20: 52

  Abstract

  Resected HER2 breast cancer patients treated with adjuvant trastuzumab and chemotherapy have superior survival compared to patients treated with chemotherapy alone. We previously showed that trastuzumab and chemotherapy induce HER2-specific antibodies which correlate with improved survival in HER2 metastatic breast cancer patients. It remains unclear whether the generation of immunity required trastuzumab and whether endogenous antibody immunity is associated with improved disease-free survival in the adjuvant setting. In this study, we addressed this question by analyzing serum anti-HER2 antibodies from a subset of patients enrolled in the NCCTG trial N9831, which includes an arm (Arm A) in which trastuzumab was not used. Arms B and C received trastuzumab sequentially or concurrently to chemotherapy, respectively.Pre-and post-treatment initiation sera were obtained from 50 women enrolled in N9831. Lambda IgG antibodies (to avoid detection of trastuzumab) to HER2 were measured and compared between arms and with disease-free survival.Prior to therapy, across all three arms, N9831 patients had similar mean anti-HER2 IgG levels. Following treatment, the mean levels of antibodies increased in the trastuzumab arms but not the chemotherapy-only arm. The proportion of patients who demonstrated antibodies increased by 4% in Arm A and by 43% in the Arms B and C combined (p?=?0.003). Cox modeling demonstrated that larger increases in antibodies were associated with improved disease-free survival in all patients (HR?=?0.23; p?=?0.04).These results show that the increased endogenous antibody immunity observed in adjuvant patients treated with combination trastuzumab and chemotherapy is clinically significant, in view of its correlation with improved disease-free survival. The findings may have important implications for predicting treatment outcomes in patients treated with trastuzumab in the adjuvant setting.ClinicalTrials.gov, NCT00005970 . Registered on July 5, 2000.

  View details for PubMedID 29898752

  View details for PubMedCentralID PMC6000975

• Innovative Strategies: Targeting Subtypes in Metastatic Breast Cancer. American Society of Clinical Oncology educational book. American Society of Clinical Oncology. Annual Meeting Pegram, M. D., Zong, Y., Yam, C., Goetz, M. P., Moulder, S. L. 2018: 65?77

  Abstract

  Metastatic breast cancer continues to be a life-threatening diagnosis that impacts hundreds of thousands of patients around the world. Targeted therapies are usually associated with less toxicity compared with cytotoxic chemotherapies and often induce response or durable disease control in estrogen receptor (ER) and/or HER2+ breast cancers. Drugs that target CDK 4/6 either alone or in combination with endocrine therapy have demonstrated substantial improvements in progression-free survival (PFS) compared with endocrine monotherapy. Most recently, PARP inhibitors have shown longer PFS compared with physician's choice of chemotherapy in BRCA-associated cancers, leading to the first U.S. Food and Drug Administration (FDA) approval of a targeted therapy with the potential to benefit a subgroup of patients with triple-negative breast cancer (TNBC). Finally, newer drug delivery strategies using antibody drug conjugates have also allowed a "targeted approach" to deliver moderate to extremely potent cytotoxins directly to sites of metastatic disease, with less toxicity.

  View details for PubMedID 30231328

• Endocrine therapy and related issues in hormone receptor-positive early breast cancer: a roundtable discussion by the breast cancer therapy expert group (BCTEG) BREAST CANCER RESEARCH AND TREATMENT Abraham, J., Caldera, H., Coleman, R., Elias, A., Goetz, M. P., Kittaneh, M., Lower, E., Mahtani, R., Mamounas, E., Pegram, M., Rugo, H., Schwartzberg, L., Traina, T., Vogel, C. 2018; 169 (1): 1?7

  Abstract

  Management of breast cancer is a rapidly evolving field, and, although evidence-based guidelines are available for clinicians to provide direction on critical issues in patient care, clinicians often left to address these issues in the context of community practice situations with their patients. These include the patient's comorbid conditions, actual versus perceived benefit of treatments, patient's compliance as well as financial/reimbursement issues, and long-term tolerability of therapy.A meeting of global oncology experts was convened in January 2017 with the belief that there is a gap in clinical practice guidance on several fundamental issues in breast cancer care, particularly in the community setting, where oncologists may encounter multiple tumor types. The goal was to discuss some of the most important questions in this area and provide some guidance for practicing oncologists.Topics addressed included risk of contralateral breast cancer recurrence in patients with estrogen receptor-positive early breast cancer who have undergone 5 years of adjuvant endocrine therapy, adverse events associated with endocrine therapy and their management, emergent data on adjuvant bisphosphonate therapy and its apparent benefit in reducing breast cancer recurrence, recent findings of extended adjuvant endocrine therapy trials, and the use of currently available genomic biomarker tests as a means of further informing treatment decisions.A summary of the discussion on these topics and several 'expert opinion statements' are provided herein in an effort to convey the collective insights of the panel as it relates to current standard practice.

  View details for PubMedID 29352368

  View details for PubMedCentralID PMC5882621

• Electrophoretic cytopathology resolves ERBB2 forms with single-cell resolution NPJ PRECISION ONCOLOGY Kang, C., Ward, T. M., Bockhorn, J., Schiffman, C., Huang, H., Pegram, M. D., Herr, A. E. 2018; 2: 10

  Abstract

  In addition to canonical oncoproteins, truncated isoforms and proteolysis products are implicated in both drug resistance and disease progression. In HER2-positive breast tumors, expression of truncated HER2 isoforms resulting from alternative translation and/or carboxy-terminal fragments (CTFs) resulting from proteolysis (collectively, t-erbB2) have been associated with shortened progression-free survival of patients. Thus, to advance clinical pathology and inform treatment decisions, we developed a high-selectivity cytopathology assay capable of distinguishing t-erbB2 from full-length HER2 expression without the need for isoform-specific antibodies. Our microfluidic, single-cell western blot, employs electrophoretic separations to resolve full-length HER2 from the smaller t-erbB2 in each ~28?pL single-cell lysate. Subsequently, a pan-HER2 antibody detects all resolved HER2 protein forms via immunoprobing. In analysis of eight breast tumor biopsies, we identified two tumors comprised of 15% and 40% t-erbB2-expressing cells. By single-cell western blotting of the t-erbB2-expressing cells, we observed statistically different ratios of t-erbB2 proteins to full-length HER2 expression. Further, target multiplexing and clustering analyses scrutinized signaling, including ribosomal S6, within the t-erbB2-expressing cell subpopulation. Taken together, cytometric assays that report both protein isoform profiles and signaling state offer cancer classification taxonomies with unique relevance to precisely describing drug resistance mechanisms in which oncoprotein isoforms/fragments are implicated.

  View details for PubMedID 29872719

• Anti-HER2 scFv-directed extracellular vesicle-mediated mRNA-based gene delivery inhibits growth of HER2-positive human breast tumor xenografts by prodrug activation. Molecular cancer therapeutics Wang, J., Forterre, A. V., Zhao, J., Frimannsson, D. O., Delcayre, A., Antes, T. J., Efron, B., Jeffrey, S. S., Pegram, M. D., Matin, A. C. 2018

  Abstract

  This paper deals with specific targeting of the prodrug/enzyme regimen, CNOB/HChrR6, to treat a serious disease namely HER2+ve human breast cancer with minimal off-target toxicity. HChrR6 is an improved bacterial enzyme that converts CNOB into the cytotoxic drug MCHB. Extracellular vesicles (EVs) were used for mRNA-based HchrR6 gene delivery: EVs may cause minimal immune rejection, and mRNA may be superior to DNA for gene delivery. To confine HChrR6 generation and CNOB activation to the cancer, the EVHB chimeric protein was constructed. It contains high affinity anti-HER2 scFv antibody (ML39) and is capable of latching on to EV surface. Cells transfected with EVHB-encoding plasmid generated EVs displaying this protein ("directed EVs"). Transfection of a separate batch of cells with the new plasmid, XPort/HChrR6, generated EVs containing HChrR6 mRNA; incubation with pure EVHB enabled these to target the HER2 receptor, generating "EXO-DEPT" EVs. EXO-DEPT treatment specifically enabled HER2-overexpressing BT474 cells to convert CNOB into MCHB in actinomycin D independent manner, showing successful and specific delivery of HCHrR6 mRNA. EXO-DEPTs --but not undirected EVs-- plus CNOB caused near-complete growth-arrest of orthotopic BT474 xenografts in vivo, demonstrating for the first time EV-mediated delivery of functional exogenous mRNA to tumors. EXO-DEPTs may be generated from patient's own dendritic cells to evade immune rejection, and without plasmids and their potentially harmful genetic material, raising the prospect of clinical use of this regimen. This approach can be employed to treat any disease overexpressing a specific marker.

  View details for PubMedID 29483213

• Targeted disruption of transcriptional effector GLI2 attenuates breast tumor growth and metastasis Beachy, P., Cai, S., Ma, Y., Hatakeyama, J., Zhao, C., Stoffels, M., Verhaegh, W., van de Stolpe, A., Pegram, M. AMER ASSOC CANCER RESEARCH. 2018

  View details for Web of Science ID 000425489401310

• Neratinib Efficacy and Circulating Tumor DNA Detection of HER2 Mutations in HER2 Non-amplified Metastatic Breast Cancer. Clinical cancer research : an official journal of the American Association for Cancer Research Ma, C. X., Bose, R., Gao, F., Freedman, R. A., Telli, M. L., Kimmick, G., Winer, E. P., Naughton, M. J., Goetz, M. P., Russell, C., Tripathy, D., Cobleigh, M., Forero, A., Pluard, T. J., Anders, C. K., Niravath, P., Thomas, S., Anderson, J., Bumb, C., Banks, K. C., Lanman, R. B., Bryce, R., Lalani, A. S., Pfeifer, J. D., Hayes, D. F., Pegram, M. D., Blackwell, K., Bedard, P. L., Al-Kateb, H., Ellis, M. J. 2017

  Abstract

  Based on promising preclinical data, we conducted a single arm phase II trial to assess the clinical benefit rate (CBR) of neratinib, defined  as complete/partial response (CR/PR) or stable disease (SD) ?24 weeks, in HER2(mut) non-amplified metastatic breast cancer (MBC).  Secondary endpoints included progression-free survival (PFS), toxicity, and circulating tumor DNA (ctDNA) HER2(mut) detection. 

  Experimental Methods:  Tumor tissue positive for HER2(mut) was required for eligibility.  Neratinib was administered 240mg daily with prophylactic loperamide.  ctDNA sequencing was performed retrospectively for 54 patients (14 positive and 40 negative for tumor HER2(mut)). 

  Results:  2.4% (9/381) tumors sequenced centrally harbored HER2(mut) (lobular 7.8% vs ductal 1.6%; p=0.026).  Thirteen additional HER2(mut) cases were identified locally.  21 of these 22 HER2(mut) cases were estrogen receptor positive.  Sixteen patients (median age 58 [31-74] years and 3 [2-10] prior metastatic regimens) received neratinib.  The CBR was 31% (90%CI 13-55%), including 1 CR, 1 PR and 3 SD ?24 weeks).  Median PFS was 16 (90%CI: 8-31) weeks. Diarrhea (grade 2 44%, grade 3 25%) was the most common adverse event.  Baseline ctDNA sequencing identified the same HER2(mut) in 11 of 14 tumor positive cases (sensitivity 79%, 90%CI: 53-94%) and correctly assigned 32 of 32 informative negative cases (specificity 100%, 90%CI: 91-100%).  Additionally ctDNA HER2(mut) variant allele frequency decreased in 9 of 11 paired samples at week four, followed by an increase upon progression.

  Conclusions: Neratinib is active in HER2(mut), non-amplified MBC.  ctDNA sequencing offers a non-invasive strategy to identify patients with HER2(mut) cancers for clinical trial participation.

  View details for PubMedID 28679771

• Relationship between Tumor Biomarkers and Efficacy in EMILIA, a Phase III Study of Trastuzumab Emtansine in HER2-Positive Metastatic Breast Cancer. Clinical cancer research Baselga, J., Lewis Phillips, G. D., Verma, S., Ro, J., Huober, J., Guardino, A. E., Samant, M. K., Olsen, S., de Haas, S. L., Pegram, M. D. 2016; 22 (15): 3755-3763

  Abstract

  HER2-positive breast cancer is heterogeneous. Some tumors express mutations, like activating PIK3CA mutations or reduced PTEN expression, that negatively correlate with response to HER2-targeted therapies. In this exploratory analysis, we investigated whether the efficacy of trastuzumab emtansine (T-DM1), an antibody-drug conjugate comprised of the cytotoxic agent DM1 linked to the HER2-targeted antibody trastuzumab, was correlated with the expression of specific biomarkers in the phase III EMILIA study.Tumors were evaluated for HER2 (n = 866), EGFR (n = 832), and HER3 (n = 860) mRNA expression by quantitative reverse transcriptase PCR; for PTEN protein expression (n = 271) by IHC; and for PIK3CA mutations (n = 259) using a mutation detection kit. Survival outcomes were analyzed by biomarker subgroups. T-DM1 was also tested on cell lines and in breast cancer xenograft models containing PIK3CA mutations.Longer progression-free survival (PFS) and overall survival (OS) were observed with T-DM1 compared with capecitabine plus lapatinib in all biomarker subgroups. PIK3CA mutations were associated with shorter median PFS (mutant vs. wild type: 4.3 vs. 6.4 months) and OS (17.3 vs. 27.8 months) in capecitabine plus lapatinib-treated patients, but not in T-DM1-treated patients (PFS, 10.9 vs. 9.8 months; OS, not reached in mutant or wild type). T-DM1 showed potent activity in cell lines and xenograft models with PIK3CA mutations.Although other standard HER2-directed therapies are less effective in tumors with PI3KCA mutations, T-DM1 appears to be effective in both PI3KCA-mutated and wild-type tumors. Clin Cancer Res; 22(15); 3755-63. ©2016 AACR.

  View details for DOI 10.1158/1078-0432.CCR-15-2499

  View details for PubMedID 26920887

• Combined niclosamide with cisplatin inhibits epithelial-mesenchymal transition and tumor growth in cisplatin-resistant triple-negative breast cancer TUMOR BIOLOGY Liu, J., Chen, X., Ward, T., Pegram, M., Shen, K. 2016; 37 (7): 9825-9835

  Abstract

  Women with triple-negative breast cancer have worse prognosis compared to other breast cancer subtypes. Acquired drug resistance remains to be an important reason influencing triple-negative breast cancer treatment efficacy. A prevailing theory postulates that the cancer resistance and recurrence results from a subpopulation of tumor cells with stemness program, which are often insensitive to cytotoxic drugs such as cisplatin. Recent studies suggested that niclosamide, an anti-helminthic drug, has potential therapeutic activities against breast cancer stem cells, which prompts us to determine its roles on eliminating cisplatin-resistant cancer cells. Hence, we established a stable cisplatin-resistant MDA-MB-231 cell line (231-CR) through continuously exposure to increasing concentrations of cisplatin (5-20 ?mol/l). Interestingly, 231-CR exhibited properties associated to epithelial-mesenchymal transition with enhanced invasion, preserved proliferation, increased mammosphere formation, and reduced apoptosis compared to naive MDA-MB-231 sensitive cells (231-CS). Importantly, niclosamide or combination with cisplatin inhibited both 231-CS and 231-CR cell proliferation in vitro. In addition, niclosamide reversed the EMT phenotype of 231-CR by downregulation of snail and vimentin. Mechanistically, niclosamide treatment in combination with or without cisplatin significantly inhibited Akt, ERK, and Src signaling pathways. In vivo study showed that niclosamide or combination with cisplatin could repress the growth of xenografts originated from either 231-CS or 231-CR cells, with prominent suppression of Ki67 expression. These findings suggested that niclosamide might serve as a novel therapeutic strategy, either alone or in combination with cisplatin, for triple-negative breast cancer treatment, especially those resistant to cisplatin.

  View details for DOI 10.1007/s13277-015-4650-1

  View details for Web of Science ID 000382174500134

  View details for PubMedID 26810188

• Improved Survival of HER2(+) Breast Cancer Patients Treated with Trastuzumab and Chemotherapy Is Associated with Host Antibody Immunity against the HER2 Intracellular Domain CANCER RESEARCH Knutson, K. L., Clynes, R., Shreeder, B., Yeramian, P., Kemp, K. P., Ballman, K., Tenner, K. S., Erskine, C. L., Norton, N., Northfelt, D., Tan, W., Calfa, C., Pegram, M., Mittendorf, E. A., Perez, E. A. 2016; 76 (13): 3702-3710

  Abstract

  The addition of trastuzumab to chemotherapy extends survival among patients with HER2(+) breast cancer. Prior work showed that trastuzumab and chemotherapy augments HER2 extracellular domain (ECD)-specific antibodies. The current study investigated whether combination therapy induced immune responses beyond HER2-ECD and, importantly, whether those immune responses were associated with survival. Pretreatment and posttreatment sera were obtained from 48 women with metastatic HER2(+) breast cancer on NCCTG (now Alliance for Clinical Trials in Oncology) studies, N0337 and N983252. IgG to HER2 intracellular domain (ICD), HER2-ECD, p53, IGFBP2, CEA, and tetanus toxoid were examined. Sera from 25 age-matched controls and 26 surgically resected HER2(+) patients were also examined. Prior to therapy, some patients with metastatic disease had elevated antibodies to IGFBP2, p53, HER2-ICD, HER2-ECD, and CEA, but not to tetanus toxin, relative to controls and surgically resected patients. Treatment augmented antibody responses to HER2-ICD in 69% of metastatic patients, which was highly associated with improved progression-free survival (PFS; HR = 0.5, P = 0.0042) and overall survival (OS; HR = 0.7, P = 0.038). Augmented antibody responses to HER2-ICD also correlated (P = 0.03) with increased antibody responses to CEA, IGFBP2, and p53, indicating that treatment induces epitope spreading. Paradoxically, patients who already had high preexisting immunity to HER2-ICD did not respond to therapy with increased antibodies to HER2-ICD and demonstrated poorer PFS (HR = 1.6, P < 0.0001) and OS (HR = 1.4, P = 0.0006). Overall, the findings further demonstrate the importance of the adaptive immune system in the efficacy of trastuzumab-containing regimens. Cancer Res; 76(13); 3702-10. ©2016 AACR.

  View details for DOI 10.1158/0008-5472.CAN-15-3091

  View details for Web of Science ID 000380071400008

  View details for PubMedID 27197192

• Classification of large circulating tumor cells isolated with ultra-high throughput microfluidic Vortex technology ONCOTARGET Che, J., Yu, V., Dhar, M., Renier, C., Matsumoto, M., Heirich, K., Garon, E. B., Goldman, J., Rao, J., Sledge, G. W., Pegram, M. D., Sheth, S., Jeffrey, S. S., Kulkarni, R. P., Sollier, E., Di Carlo, D. 2016; 7 (11): 12748-12760

  Abstract

  Circulating tumor cells (CTCs) are emerging as rare but clinically significant non-invasive cellular biomarkers for cancer patient prognosis, treatment selection, and treatment monitoring. Current CTC isolation approaches, such as immunoaffinity, filtration, or size-based techniques, are often limited by throughput, purity, large output volumes, or inability to obtain viable cells for downstream analysis. For all technologies, traditional immunofluorescent staining alone has been employed to distinguish and confirm the presence of isolated CTCs among contaminating blood cells, although cells isolated by size may express vastly different phenotypes. Consequently, CTC definitions have been non-trivial, researcher-dependent, and evolving. Here we describe a complete set of objective criteria, leveraging well-established cytomorphological features of malignancy, by which we identify large CTCs. We apply the criteria to CTCs enriched from stage IV lung and breast cancer patient blood samples using the High Throughput Vortex Chip (Vortex HT), an improved microfluidic technology for the label-free, size-based enrichment and concentration of rare cells. We achieve improved capture efficiency (up to 83%), high speed of processing (8 mL/min of 10x diluted blood, or 800 ?L/min of whole blood), and high purity (avg. background of 28.8±23.6 white blood cells per mL of whole blood). We show markedly improved performance of CTC capture (84% positive test rate) in comparison to previous Vortex designs and the current FDA-approved gold standard CellSearch assay. The results demonstrate the ability to quickly collect viable and pure populations of abnormal large circulating cells unbiased by molecular characteristics, which helps uncover further heterogeneity in these cells.

  View details for PubMedID 26863573

• Phase 1b/2a study of trastuzumab emtansine (T-DM1), paclitaxel, and pertuzumab in HER2-positive metastatic breast cancer BREAST CANCER RESEARCH Krop, I. E., Modi, S., LoRusso, P. M., Pegram, M., Guardino, E., Althaus, B., Lu, D., Strasak, A., Elias, A. 2016; 18

  View details for DOI 10.1186/s13058-016-0691-7

  View details for Web of Science ID 000372492000002

• Niclosamide inhibits epithelial-mesenchymal transition and tumor growth in lapatinib-resistant human epidermal growth factor receptor 2-positive breast cancer. international journal of biochemistry & cell biology Liu, J., Chen, X., Ward, T., Mao, Y., Bockhorn, J., Liu, X., Wang, G., Pegram, M., Shen, K. 2016; 71: 12-23

  Abstract

  Acquired resistance to lapatinib, a human epidermal growth factor receptor 2 kinase inhibitor, remains a clinical problem for women with human epidermal growth factor receptor 2-positive advanced breast cancer, as metastasis is commonly observed in these patients. Niclosamide, an anti-helminthic agent, has recently been shown to exhibit cytotoxicity to tumor cells with stem-like characteristics. This study was designed to identify the mechanisms underlying lapatinib resistance and to determine whether niclosamide inhibits lapatinib resistance by reversing epithelial-mesenchymal transition. Here, two human epidermal growth factor receptor 2-positive breast cancer cell lines, SKBR3 and BT474, were exposed to increasing concentrations of lapatinib to establish lapatinib-resistant cultures. Lapatinib-resistant SKBR3 and BT474 cells exhibited up-regulation of the phenotypic epithelial-mesenchymal transition markers Snail, vimentin and ?-smooth muscle actin, accompanied by activation of nuclear factor-?B and Src and a concomitant increase in stem cell marker expression (CD44(high)/CD24(low)), compared to naive lapatinib-sensitive SKBR3 and BT474 cells, respectively. Interestingly, niclosamide reversed epithelial-mesenchymal transition, induced apoptosis and inhibited cell growth by perturbing aberrant signaling pathway activation in lapatinib-resistant human epidermal growth factor receptor 2-positive cells. The ability of niclosamide to alleviate stem-like phenotype development and invasion was confirmed. Collectively, our results demonstrate that lapatinib resistance correlates with epithelial-mesenchymal transition and that niclosamide inhibits lapatinib-resistant cell viability and epithelial-mesenchymal transition. These findings suggest a role of niclosamide or derivatives optimized for more favorable bioavailability not only in reversing lapatinib resistance but also in reducing metastatic potential during the treatment of human epidermal growth factor receptor 2-positive breast cancer.

  View details for DOI 10.1016/j.biocel.2015.11.014

  View details for PubMedID 26643609

• Hydrogel Pore-Size Modulation for Enhanced Single-Cell Western Blotting ADVANCED MATERIALS Duncombe, T. A., Kang, C., Maity, S., Ward, T. M., Pegram, M. D., Murthy, N., Herr, A. E. 2016; 28 (2): 327-334

  View details for DOI 10.1002/adma.201503939

  View details for Web of Science ID 000367842400015

• Phase 1b/2a study of trastuzumab emtansine (T-DM1), paclitaxel, and pertuzumab in HER2-positive metastatic breast cancer. Breast cancer research Krop, I. E., Modi, S., LoRusso, P. M., Pegram, M., Guardino, E., Althaus, B., Lu, D., Strasak, A., Elias, A. 2016; 18 (1): 34-?

  Abstract

  In pre-clinical studies, the anti-tumor activity of T-DM1 was enhanced when combined with taxanes or pertuzumab. This phase 1b/2a study evaluated the safety/tolerability of T-DM1?+?paclitaxel?±?pertuzumab in HER2-positive advanced breast cancer.In phase 1b (n?=?60), a 3?+?3 dose-escalation approach was used to determine the maximum tolerated dose (MTD) of T-DM1?+?paclitaxel?±?pertuzumab. The primary objective of phase 2a was feasibility, with 44 patients randomized to T-DM1?+?paclitaxel?±?pertuzumab at the MTD identified in phase 1b.The MTD was T-DM1 3.6 mg/kg every three weeks (q3w) or 2.4 mg/kg weekly?+?paclitaxel 80 mg/m(2) weekly?±?pertuzumab 840 mg loading dose followed by 420 mg q3w. Phase 2a patients had received a median of 5.0 (range: 0-10) prior therapies for advanced cancer. In phase 2a, 51.2 % received ?12 paclitaxel doses within 15 weeks, and 14.0 % received 12 paclitaxel doses by week 12. Common all-grade adverse events (AEs) were peripheral neuropathy (90.9 %) and fatigue (79.5 %). A total of 77.3 % experienced grade ?3 AEs, most commonly neutropenia (25.0 %) and peripheral neuropathy (18.2 %). Among the 42 phase 2a patients with measurable disease, the objective response rate (ORR) was 50.0 % (95 % confidence interval (CI) 34.6-65.4); the clinical benefit rate (CBR) was 56.8 % (95 % CI 41.6-71.0). No pharmacokinetic interactions were observed between T-DM1 and paclitaxel.This regimen showed clinical activity. Although there is potential for paclitaxel to be added to T-DM1?±?pertuzumab, peripheral neuropathy was common in this heavily pretreated population.ClinicalTrials.gov NCT00951665 . Registered August 3, 2009.

  View details for DOI 10.1186/s13058-016-0691-7

  View details for PubMedID 26979312

  View details for PubMedCentralID PMC4791863

• Hydrogel Pore-Size Modulation for Enhanced Single-Cell Western Blotting. Advanced materials (Deerfield Beach, Fla.) Duncombe, T. A., Kang, C. C., Maity, S., Ward, T. M., Pegram, M. D., Murthy, N., Herr, A. E. 2016; 28 (2): 327?34

  Abstract

  Pore-gradient microgel arrays enable thousands of parallel high-resolution single-cell protein electrophoresis separations for targets accross a wide molecular mass (25-289 kDa), yet within 1 mm separation distances. Dual crosslinked hydrogels facilitate gel-pore expansion after electrophoresis for efficient and uniform immunoprobing. The photopatterned, light-activated, and acid-expandable hydrogel underpins single-cell protein analysis, here for oncoprotein-related signaling in human breast biopsy.

  View details for PubMedID 26567472

  View details for PubMedCentralID PMC4708057

• "Vertical" Inhibition of HER2 Yields Horizontal Gains in the Clinic. Clinical cancer research Sledge, G. W., Pegram, M. D. 2015; 21 (12): 2663-2665

  Abstract

  HER2-targeted therapy has moved beyond trastuzumab to include other monoclonals targeting the cell surface, receptor tyrosine kinase inhibitors of HER2, and antibody-drug conjugates. Afatinib, a small molecule receptor tyrosine kinase inhibitor, now joins the ranks of HER2-targeting agents in combination with trastuzumab. The combination brings new opportunities and challenges.

  View details for DOI 10.1158/1078-0432.CCR-14-3183

  View details for PubMedID 26078429

• Everything old is neu again: cellular senescence in HER2-positive breast cancer. Journal of the National Cancer Institute Sledge, G. W., Pegram, M. D. 2015; 107 (5)

  View details for DOI 10.1093/jnci/djv091

  View details for PubMedID 25972602

• Personalization of loco-regional care for primary breast cancer patients (part 1) FUTURE ONCOLOGY Toi, M., Winer, E. P., Benson, J. R., Inamoto, T., Forbes, J. F., von Minckwitz, G., Robertson, J. F., Grobmyer, S. R., Jatoi, I., Sasano, H., Kunkler, I., Ho, A. Y., Yamauchi, C., Chow, L. W., Huang, C., Han, W., Noguchi, S., Pegram, M. D., Yamauchi, H., Lee, E., Larionov, A. A., Bevilacqua, J. L., Yoshimura, M., Sugie, T., Yamauchi, A., Krop, I. E., Noh, D. Y., Klimberg, V. S. 2015; 11 (9): 1297-1300

  Abstract

  ABSTRACT? Kyoto Breast Cancer Consensus Conference, Kyoto, Japan, 18-20 February 2014 The loco-regional management of breast cancer is increasingly complex with application of primary systemic therapies, oncoplastic techniques and genetic testing for breast cancer susceptibility. Personalization of loco-regional treatment is integral to optimization of breast cancer care. Clinical and pathological tumor stage, biological features and host factors influence loco-regional treatment strategies and extent of surgical procedures. Key issues including axillary staging, axillary treatment, radiation therapy, primary systemic therapy (PST), preoperative hormonal therapy and genetic predisposition were identified and discussed at the Kyoto Breast Cancer Consensus Conference (KBCCC2014). In the first of a two part conference scene, consensus recommendations for axillary management are presented and focus on the following topics: indications for completion axillary lymph node dissection in primary surgical patients with ?2 macrometastases or any sentinel nodal deposits after PST; the timing of sentinel lymph node biopsy in the context of PST; use of axillary irradiation as a component of primary treatment plans and the role of intraoperative node assessment in the post-Z0011 era.

  View details for DOI 10.2217/fon.15.65

  View details for Web of Science ID 000354390500002

  View details for PubMedID 25952777

• Personalization of loco-regional care for primary breast cancer patients (part 2) FUTURE ONCOLOGY Toi, M., Winer, E. P., Benson, J. R., Inamoto, T., Forbes, J. F., von Minckwitz, G., Robertson, J. F., Grobmyer, S. R., Jatoi, I., Sasano, H., Kunkler, I., Ho, A. Y., Yamauchi, C., Chow, L. W., Huang, C., Han, W., Noguchi, S., Pegram, M. D., Yamauchi, H., Lee, E., Larionov, A. A., Bevilacqua, J. L., Yoshimura, M., Sugie, T., Yamauchi, A., Krop, I. E., Noh, D. Y., Klimberg, V. S. 2015; 11 (9): 1301-1305

  Abstract

  Kyoto Breast Cancer Consensus Conference, Kyoto, Japan, 18-20 February 2014 The loco-regional management of breast cancer is increasingly complex with application of primary systemic therapies, oncoplastic techniques and genetic testing for breast cancer susceptibility. Personalization of loco-regional treatment is integral to optimization of breast cancer care. Clinical and pathological tumor stage, biological features and host factors influence loco-regional treatment strategies and extent of surgical procedures. Key issues including axillary staging, axillary treatment, radiation therapy, primary systemic therapy (PST), preoperative hormonal therapy and genetic predisposition were identified and discussed at the Kyoto Breast Cancer Consensus Conference (KBCCC2014). In the second of a two part conference scene, consensus recommendations for radiation treatment, primary systemic therapies and management of genetic predisposition are reported and focus on the following topics: influence of both clinical response to PST and stage at presentation on recommendations for postmastectomy radiotherapy; use of regional nodal irradiation in selected node-positive patients and those with adverse pathological factors; extent of surgical resection following downstaging of tumors with PST; use of preoperative hormonal therapy in premenopausal women with larger, node-negative luminal A-like tumors and managing increasing demands for contralateral prophylactic mastectomy in patients with a unilateral sporadic breast cancer.

  View details for DOI 10.2217/fon.15.66

  View details for Web of Science ID 000354390500003

  View details for PubMedID 25952778

• Association Studies of Fc gamma Receptor Polymorphisms with Outcome in HER2(+) Breast Cancer Patients Treated with Trastuzumab in NCCTG (Alliance) Trial N9831 CANCER IMMUNOLOGY RESEARCH Norton, N., Olson, R. M., Pegram, M., Tenner, K., Ballman, K. V., Clynes, R., Knutson, K. L., Perez, E. A. 2014; 2 (10): 962-969

  Abstract

  Patients with HER2+ breast cancer treated with trastuzumab and chemotherapy have superior survival compared with patients treated with chemotherapy alone. Polymorphisms within FCGR2A and FCGR3A are associated with binding affinity of natural killer cells to the IgG1 portion of trastuzumab, and a polymorphism in FCGR2B (I232T) is associated with impaired regulatory activity. The association of these polymorphisms with clinical response among trastuzumab-treated patients is equivocal, with both positive and negative associations. We performed genotyping analysis on the FCGR3A V158F, FCGR2A R131H, and FCGR2B I232T polymorphisms in 1,325 patients from the N9831 clinical trial. Patients in arm A (N = 419) received chemotherapy only. Patients in arms B (N = 469) and C (N = 437) were treated with chemotherapy and trastuzumab (sequentially in arm B and concurrently in arm C). Using log-rank test and Cox proportional hazard models, we compared disease-free survival (DFS) among genotypic groups within pooled arms B/C. We found no differences in DFS between trastuzumab-treated patients who had the FCGR3A 158 V/V and/or FCGR2A 131 H/H high-affinity genotypes and patients without those genotypes. Furthermore, there was no significant interaction between FCGR3A and FCGR2A and treatment. However, there was a difference in DFS for FCGR2B I232T, with I/I patients deriving benefit from trastuzumab (P < 0.001), compared with the T carriers who did not (P = 0.81). The interaction between FCGR2B genotype and treatment was statistically significant (P = 0.03). Our analysis did not reveal an association between Fc?R high-affinity genotypes and outcomes. However, it seems that the FCGR2B inhibitory gene may be predictive of adjuvant trastuzumab benefit.

  View details for DOI 10.1158/2326-6066.CIR-14-0059

  View details for Web of Science ID 000346134100006

  View details for PubMedID 24989892

  View details for PubMedCentralID PMC4215796

• Assessing the Discordance Rate Between Local and Central HER2 Testing in Women With Locally Determined HER2-Negative Breast Cancer CANCER Kaufman, P. A., Bloom, K. J., Burris, H., Gralow, J. R., Mayer, M., Pegram, M., Rugo, H. S., Swain, S. M., Yardley, D. A., Chau, M., Lalla, D., Yoo, B., Brammer, M. G., Vogel, C. L. 2014; 120 (17): 2657-2664

  Abstract

  The importance of human epidermal growth factor receptor 2 (HER2) as a prognostic and predictive marker in invasive breast cancer is well established. Accurate assessment of HER2 status is essential to determine optimal treatment options.Breast cancer tumor tissue samples from the VIRGO observational cohort tissue substudy that were locally HER2-negative were retested centrally with both US Food and Drug Administration (FDA)-approved immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) assays, using FDA-approved assay cutoffs; results were compared.Of the 552 unique patient samples centrally retested with local HER2-negative results recorded, tumor samples from 22 (4.0%) patients were determined to be HER2-positive (95% confidence interval [CI]?=?2.5%-5.7%). Of these, 18 had been tested locally by only one testing methodology; 15 of 18 were HER2-positive after the central retesting, based on the testing methodology not performed locally. Compared with the 530 patients with centrally confirmed HER2-negative tumors, the 22 patients with centrally determined HER2-positive tumors were younger (median age 56.5 versus 60.0 years) and more likely to have ER/PR-negative tumors (27.3% versus 22.3%). These patients also had shorter median progression-free survival (6.4 months [95% CI?=?3.8-15.9 months] versus 9.1 months [95% CI?=?8.3-10.3 months]) and overall survival (25.9 months [95% CI?=?13.8-not estimable] versus 27.9 months [95% CI?=?25.0-32.9 months]).This study highlights the limitations of employing just one HER2 testing methodology in current clinical practice. It identifies a cohort of patients who did not receive potentially efficacious therapy because their tumor HER2-positivity was not determined by the test initially used. Because of inherent limitations in testing methodologies, it is inadvisable to rely on a single test to rule out potential benefit from HER2-targeted therapy.

  View details for DOI 10.1002/cncr.28710

  View details for Web of Science ID 000340559000006

  View details for PubMedID 24930388

• Breast Cancer Version 3.2014 Clinical Practice Guidelines in Oncology JOURNAL OF THE NATIONAL COMPREHENSIVE CANCER NETWORK Gradishar, W. J., Anderson, B. O., Blair, S. L., Burstein, H. J., Cyr, A., Elias, A. D., Farrar, W. B., Forero, A., Giordano, S. H., Goldstein, L. J., Hayes, D. F., Hudis, C. A., Isakoff, S. J., Ljung, B. E., Marcom, P. K., Mayer, I. A., McCormick, B., Miller, R. S., Pegram, M., Pierce, L. J., Reed, E. C., Salerno, K. E., Schwartzberg, L. S., Smith, M. L., Soliman, H., Somlo, G., Ward, J. H., Wolff, A. C., Zellars, R., Shead, D. A., Kumar, R. 2014; 12 (4): 542-590

  Abstract

  Breast cancer is the most common malignancy in women in the United States and is second only to lung cancer as a cause of cancer death. The overall management of breast cancer includes the treatment of local disease with surgery, radiation therapy, or both, and the treatment of systemic disease with cytotoxic chemotherapy, endocrine therapy, biologic therapy, or combinations of these. The NCCN Guidelines specific to management of large clinical stage II and III tumors are discussed in this article. These guidelines are the work of the members of the NCCN Breast Cancer Panel. Expert medical clinical judgment is required to apply these guidelines in the context of an individual patient to provide optimal care. Although not stated at every decision point of the guidelines, patient participation in prospective clinical trials is the preferred option of treatment for all stages of breast cancer.

  View details for Web of Science ID 000334014900011

• Breast cancer version 3.2014. Journal of the National Comprehensive Cancer Network Gradishar, W. J., Anderson, B. O., Blair, S. L., Burstein, H. J., Cyr, A., Elias, A. D., Farrar, W. B., Forero, A., Giordano, S. H., Goldstein, L. J., Hayes, D. F., Hudis, C. A., Isakoff, S. J., Ljung, B. E., Marcom, P. K., Mayer, I. A., McCormick, B., Miller, R. S., Pegram, M., Pierce, L. J., Reed, E. C., Salerno, K. E., Schwartzberg, L. S., Smith, M. L., Soliman, H., Somlo, G., Ward, J. H., Wolff, A. C., Zellars, R., Shead, D. A., Kumar, R. 2014; 12 (4): 542-590

  Abstract

  Breast cancer is the most common malignancy in women in the United States and is second only to lung cancer as a cause of cancer death. The overall management of breast cancer includes the treatment of local disease with surgery, radiation therapy, or both, and the treatment of systemic disease with cytotoxic chemotherapy, endocrine therapy, biologic therapy, or combinations of these. The NCCN Guidelines specific to management of large clinical stage II and III tumors are discussed in this article. These guidelines are the work of the members of the NCCN Breast Cancer Panel. Expert medical clinical judgment is required to apply these guidelines in the context of an individual patient to provide optimal care. Although not stated at every decision point of the guidelines, patient participation in prospective clinical trials is the preferred option of treatment for all stages of breast cancer.

  View details for PubMedID 24717572

• Treating the HER2 Pathway in Early and Advanced Breast Cancer HEMATOLOGY-ONCOLOGY CLINICS OF NORTH AMERICA Pegram, M. D. 2013; 27 (4): 751-?

  Abstract

  ERBB2 gene amplification occurs in ?20% of human breast cancers (BC) and is associated with an adverse clinical prognosis, indicating that it may be playing a critical role in disease pathogenesis. Therapeutic strategies targeting pathologic ERBB2 overexpression have revolutionized the diagnosis and treatment of BC. Indeed, humanized anti-ERBB2 antibodies, small molecule ERBB2 kinase inhibitors and ERBB2-targeting antibody-drug conjugates have proven safety and efficacy based upon evidence from randomized phase III clinical trials. Recent progress in targeting ERBB2 alteration will be reviewed, with focus on data that has informed changes in clinical practice for the treatment of BC.

  View details for DOI 10.1016/j.hoc.2013.05.007

  View details for Web of Science ID 000323798000008

  View details for PubMedID 23915743

• Truncated p110 ERBB2 induces mammary epithelial cell migration, invasion and orthotopic xenograft formation, and is associated with loss of phosphorylated STAT5. Oncogene Ward, T. M., IORNS, E., Liu, X., Hoe, N., Kim, P., Singh, S., Dean, S., Jegg, A., Gallas, M., Rodriguez, C., Lippman, M., Landgraf, R., Pegram, M. D. 2013; 32 (19): 2463-2474

  Abstract

  Truncated-ERBB2 isoforms (t-ERBB2s), resulting from receptor proteolysis or alternative translation of the ERBB2 mRNA, exist in a subset of human breast tumors. t-ERBB2s lack the receptor extracellular domain targeted by therapeutic anti-ERBB2 antibodies and antibody-drug conjugates, including trastuzumab, trastuzumab-DM1 and pertuzumab. In clinical studies, expression of t-ERBB2 in breast tumors correlates with metastasis as well as trastuzumab resistance. By using a novel immuno-microarray method, we detect a significant t-ERBB2 fraction in 18 of 31 (58%) of immunohistochemistry (IHC)3+ ERBB2+ human tumor specimens, and further show that t-ERBB2 isoforms are phosphorylated in a subset of IHC3+ samples (10 of 31, 32%). We investigated t-ERBB2 biological activity via engineered expression of full-length and truncated ERBB2 isoforms in human mammary epithelial cells (HMECs), including HMEC and MCF10A cells. Expression of p110 t-ERBB2, but not p95m (m=membrane, also 648CTF) or intracellular ERBB2s, significantly enhanced cell migration and invasion in multiple cell types. In addition, only expression of the p110 isoform led to human breast epithelial cell (HMLE) xenograft formation in vivo. Expression of t-ERBB2s did not result in hyperactivation of the phosphoinositide kinase-3/AKT or mitogen-activated protein kinase signaling pathways in these cells; rather, phosphoproteomic array profiling revealed attenuation of phosphorylated signal transducer and activator of transcription 5 (STAT5) in p110-t-ERBB2-expressing cells compared to controls. Short hairpin-mediated silencing of STAT5 phenocopied p110-t-ERBB2-driven cell migration and invasion, while expression of constitutively active STAT5 reversed these effects. Thus, we provide novel evidence that (1) expression of p110 t-ERBB2 is sufficient for full transformation of HMEC, yielding in vivo xenograft formation, and (2) truncated p110 t-ERBB2 expression is associated with decreased phosphorylation of STAT5.

  View details for DOI 10.1038/onc.2012.256

  View details for PubMedID 22751112

• The use of neoadjuvant platinum-based chemotherapy in locally advanced breast cancer that is triple negative: retrospective analysis of 144 patients. Breast cancer research and treatment Hurley, J., Reis, I. M., Rodgers, S. E., Gomez-Fernandez, C., Wright, J., Leone, J. P., Larrieu, R., Pegram, M. D. 2013; 138 (3): 783-794

  Abstract

  Triple-negative breast cancers comprise about 20 % of breast cancers. They have poor prognosis and have no standard therapy. The aim of this study was to evaluate pathologic complete response (pCR), progression-free survival (PFS), and overall survival (OS) in patients with TNBC treated with neoadjuvant platinum-based chemotherapy. This is a retrospective study of one hundred and forty-four women with TNBC treated with neoadjuvant platinum-containing chemotherapy for locally advanced breast cancer at the University of Miami between January 1, 1999, and January 1, 2011. The medical record was reviewed to obtain data on clinical characteristics, including ethnicity, race, age, clinical stage, treatment regimen, and vital status. This study was approved by the University of Miami IRB. All patients had locally advanced breast cancer with at least one of the following features at presentation: T3, T4, N2, and N3. The mean tumor size by palpation was 9.4 cm. The clinical T-stage at presentation was 1.4 % T1, 8.3 % T2, 52.8 % T3, and 37.5 % T4 (19.4 % T4d). The nodal status by physical exam at presentation was 23 % N0, 37.5 % N1, 34 % N2, and 5.5 % N3. pCR in breast and axilla was seen in 31 %. PFS and OS were 55 and 59 %, respectively, at 7 years. Cisplatin offered a survival advantage over carboplatin in both PFS (P = 0.007) and OS (P = 0.018). Node positivity was the most important predictor of survival. Cisplatin/docetaxel neoadjuvant therapy was well tolerated and an effective therapy in locally advanced TNB.

  View details for DOI 10.1007/s10549-013-2497-y

  View details for PubMedID 23542956

• PI3K independent activation of mTORC1 as a target in lapatinib-resistant ERBB2+breast cancer cells BREAST CANCER RESEARCH AND TREATMENT Jegg, A., Ward, T. M., Iorns, E., Hoe, N., Zhou, J., Liu, X., Singh, S., Landgraf, R., Pegram, M. D. 2012; 136 (3): 683-692

  Abstract

  Therapies targeting the ERBB2 receptor, including the kinase inhibitor lapatinib (Tykerb, GlaxoSmithKline), have improved clinical outcome for women with ERBB2-amplified breast cancer. However, acquired resistance to lapatinib remains a significant clinical problem, and the mechanisms governing resistance remain poorly understood. We sought to define molecular alterations that confer an acquired lapatinib resistance phenotype in ER-/ERBB2+ human breast cancer cells. ERBB2-amplified SKBR3 breast cancer cells were rendered resistant to lapatinib via culture in increasing concentrations of the drug, and molecular changes associated with a resistant phenotype were interrogated using a collaborative enzyme-enhanced immunoassay platform and immunoblotting techniques for detection of phosphorylated signaling cascade proteins. Interestingly, despite apparent inactivation of the PI3K/AKT signaling pathway, resistant cells exhibited constitutive activation of mammalian target of rapamycin complex 1 (mTORC1) and were highly sensitive to mTOR inhibition with rapamycin and the dual PI3K/mTOR inhibitor NVP-BEZ235. These data demonstrate a role for downstream activation of mTORC1 in the absence of molecular alterations leading to PI3K/AKT hyperactivation as a potential mechanism of lapatinib resistance in this model of ERBB2+ breast cancer and support the rationale of combination or sequential therapy using ERBB2 and mTOR-targeting molecules to prevent or target resistance to lapatinib. Moreover, our data suggest that assessment of mTOR substrate phosphorylation (i.e., S6) may serve as a more robust biomarker to predict sensitivity to mTOR inhibitors in the context of lapatinib resistance than PI3K mutations, loss of PTEN and p-AKT levels.

  View details for DOI 10.1007/s10549-012-2252-9

  View details for Web of Science ID 000312071000006

  View details for PubMedID 23089982

• Preoperative systemic therapy in locoregional management of early breast cancer: highlights from the Kyoto Breast Cancer Consensus Conference BREAST CANCER RESEARCH AND TREATMENT Toi, M., Benson, J. R., Winer, E. P., Forbes, J. F., von Minckwitz, G., Golshan, M., Robertson, J. F., Sasano, H., Cole, B. F., Chow, L. W., Pegram, M. D., Han, W., Huang, C., Ikeda, T., Kanao, S., Lee, E., Noguchi, S., Ohno, S., Partridge, A. H., Rouzier, R., Tozaki, M., Sugie, T., Yamauchi, A., Inamoto, T. 2012; 136 (3): 919-926

  Abstract

  Data reviewed at the Kyoto Breast Cancer Consensus Conference (KBCCC) showed that preoperative systemic therapy (PST) could optimize surgery through the utilization of information relating to pre- and post-PST tumor stage, therapeutic sensitivity, and treatment-induced changes in the biological characteristics of the tumor. As such, it was noted that the biological characteristics of the tumor, such as hormone receptors, human epidermal growth factor receptor-2, histological grade, cell proliferative activity, mainly defined by the Ki67 labeling index, and the tumor's multi-gene signature, should be considered in the planning of both systemic and local therapy. Furthermore, the timing of axillary sentinel lymph node diagnosis (i.e., before or after the PST) was also noted to be critical in that it may influence the likelihood of axillary preservation, even in node positive cases. In addition, axillary diagnosis with ultrasound and concomitant fine needle aspiration cytology or core needle biopsy (CNB) was reported to contribute to the construction of a treatment algorithm for patient-specific or individualized axillary surgery. Following PST, planning for breast surgery should therefore be based on tumor subtype, tumor volume and extent, therapeutic response to PST, and patient preference. Nomograms for predicting nodal status and drug sensitivity were also recognized as a tool to support decision-making in the selection of surgical treatment. Overall, review of data at the KBCCC showed that PST increases the likelihood of patients receiving localized surgery and individualized treatment regimens.

  View details for DOI 10.1007/s10549-012-2333-9

  View details for Web of Science ID 000312071000030

  View details for PubMedID 23143284

• Trastuzumab Emtansine for HER2-Positive Advanced Breast Cancer NEW ENGLAND JOURNAL OF MEDICINE Verma, S., Miles, D., Gianni, L., Krop, I. E., Welslau, M., Baselga, J., Pegram, M., Oh, D., Dieras, V., Guardino, E., Fang, L., Lu, M. W., Olsen, S., Blackwell, K. 2012; 367 (19): 1783-1791

  Abstract

  Trastuzumab emtansine (T-DM1) is an antibody-drug conjugate incorporating the human epidermal growth factor receptor 2 (HER2)-targeted antitumor properties of trastuzumab with the cytotoxic activity of the microtubule-inhibitory agent DM1. The antibody and the cytotoxic agent are conjugated by means of a stable linker.We randomly assigned patients with HER2-positive advanced breast cancer, who had previously been treated with trastuzumab and a taxane, to T-DM1 or lapatinib plus capecitabine. The primary end points were progression-free survival (as assessed by independent review), overall survival, and safety. Secondary end points included progression-free survival (investigator-assessed), the objective response rate, and the time to symptom progression. Two interim analyses of overall survival were conducted.Among 991 randomly assigned patients, median progression-free survival as assessed by independent review was 9.6 months with T-DM1 versus 6.4 months with lapatinib plus capecitabine (hazard ratio for progression or death from any cause, 0.65; 95% confidence interval [CI], 0.55 to 0.77; P<0.001), and median overall survival at the second interim analysis crossed the stopping boundary for efficacy (30.9 months vs. 25.1 months; hazard ratio for death from any cause, 0.68; 95% CI, 0.55 to 0.85; P<0.001). The objective response rate was higher with T-DM1 (43.6%, vs. 30.8% with lapatinib plus capecitabine; P<0.001); results for all additional secondary end points favored T-DM1. Rates of grade 3 or 4 adverse events were higher with lapatinib plus capecitabine than with T-DM1 (57% vs. 41%). The incidences of thrombocytopenia and increased serum aminotransferase levels were higher with T-DM1, whereas the incidences of diarrhea, nausea, vomiting, and palmar-plantar erythrodysesthesia were higher with lapatinib plus capecitabine.T-DM1 significantly prolonged progression-free and overall survival with less toxicity than lapatinib plus capecitabine in patients with HER2-positive advanced breast cancer previously treated with trastuzumab and a taxane. (Funded by F. Hoffmann-La Roche/Genentech; EMILIA ClinicalTrials.gov number, NCT00829166.).

  View details for DOI 10.1056/NEJMoa1209124

  View details for Web of Science ID 000310773200005

  View details for PubMedID 23020162

• Comprehensive molecular portraits of human breast tumours NATURE Koboldt, D. C., Fulton, R. S., McLellan, M. D., Schmidt, H., Kalicki-Veizer, J., McMichael, J. F., Fulton, L. L., Dooling, D. J., Ding, L., Mardis, E. R., Wilson, R. K., Ally, A., Balasundaram, M., Butterfield, Y. S., Carlsen, R., Carter, C., Chu, A., Chuah, E., Chun, H. E., Coope, R. J., Dhalla, N., Guin, R., Hirst, C., Hirst, M., Holt, R. A., Lee, D., Li, H. I., Mayo, M., Moore, R. A., Mungall, A. J., Pleasance, E., Robertson, A. G., Schein, J. E., Shafiei, A., Sipahimalani, P., Slobodan, J. R., Stoll, D., Tam, A., Thiessen, N., Varhol, R. J., Wye, N., Zeng, T., Zhao, Y., Birol, I., Jones, S. J., Marra, M. A., Cherniack, A. D., Saksena, G., Onofrio, R. C., Pho, N. H., Carter, S. L., Schumacher, S. E., Tabak, B., Hernandez, B., Gentry, J., Huy Nguyen, H., Crenshaw, A., Ardlie, K., Beroukhim, R., Winckler, W., Getz, G., Gabriel, S. B., Meyerson, M., Chin, L., Park, P. J., Kucherlapati, R., Hoadley, K. A., Auman, J. T., Fan, C., Turman, Y. J., Shi, Y., Li, L., Topal, M. D., He, X., Chao, H., Prat, A., Silva, G. O., Iglesia, M. D., Zhao, W., Usary, J., Berg, J. S., Adams, M., Booker, J., Wu, J., Gulabani, A., Bodenheimer, T., Hoyle, A. P., Simons, J. V., Soloway, M. G., Mose, L. E., Jefferys, S. R., Balu, S., Parker, J. S., Hayes, D. N., Perou, C. M., Malik, S., Mahurkar, S., Shen, H., Weisenberger, D. J., Triche, T., Lai, P. H., Bootwalla, M. S., Maglinte, D. T., Berman, B. P., Van den Berg, D. J., Baylin, S. B., Laird, P. W., Creighton, C. J., Donehower, L. A., Getz, G., Noble, M., Voet, D., Saksena, G., Gehlenborg, N., DiCara, D., Zhang, J., Zhang, H., Wu, C., Liu, S. Y., Lawrence, M. S., Zou, L., Sivachenko, A., Lin, P., Stojanov, P., Jing, R., Cho, J., Sinha, R., Park, R. W., Nazaire, M., Robinson, J., Thorvaldsdottir, H., Mesirov, J., Park, P. J., Chin, L., Reynolds, S., Kreisberg, R. B., Bernard, B., Bressler, R., Erkkila, T., Lin, J., Thorsson, V., Zhang, W., Shmulevich, I., Ciriello, G., Weinhold, N., Schultz, N., Gao, J., Cerami, E., Gross, B., Jacobsen, A., Sinha, R., Aksoy, B. A., Antipin, Y., Reva, B., Shen, R., Taylor, B. S., Ladanyi, M., Sander, C., Anur, P., Spellman, P. T., Lu, Y., Liu, W., Verhaak, R. R., Mills, G. B., Akbani, R., Zhang, N., Broom, B. M., Casasent, T. D., Wakefield, C., Unruh, A. K., Baggerly, K., Coombes, K., Weinstein, J. N., Haussler, D., Benz, C. C., Stuart, J. M., Benz, S. C., Zhu, J., Szeto, C. C., Scott, G. K., Yau, C., Paul, E. O., Carlin, D., Wong, C., Sokolov, A., Thusberg, J., Mooney, S., Sam Ng, S., Goldstein, T. C., Ellrott, K., Grifford, M., Wilks, C., Ma, S., Craft, B., Yan, C., Hu, Y., Meerzaman, D., Gastier-Foster, J. M., Bowen, J., Ramirez, N. C., Black, A. D., Pyatt, R. E., White, P., Zmuda, E. J., Frick, J., Lichtenberg, T., Brookens, R., George, M. M., Gerken, M. A., Harper, H. A., Leraas, K. M., Wise, L. J., Tabler, T. R., McAllister, C., Barr, T., Hart-Kothari, M., Tarvin, K., Saller, C., Sandusky, G., Mitchell, C., Iacocca, M. V., Brown, J., Rabeno, B., Czerwinski, C., Petrelli, N., Dolzhansky, O., Abramov, M., Voronina, O., Potapova, O., Marks, J. R., Suchorska, W. M., Murawa, D., Kycler, W., Ibbs, M., Korski, K., Spychala, A., Murawa, P., Brzezinski, J. J., Perz, H., Lazniak, R., Teresiak, M., Tatka, H., Leporowska, E., Bogusz-Czerniewicz, M., Malicki, J., Mackiewicz, A., Wiznerowicz, M., Xuan Van Le, X. V., Kohl, B., Nguyen Viet Tien, N. V., Thorp, R., Nguyen Van Bang, N. V., Sussman, H., Bui Duc Phu, B. D., Hajek, R., Nguyen Phi Hung, N. P., Tran Viet The Phuong, V. T., Huynh Quyet Thang, H. Q., Khan, K. Z., Penny, R., Mallery, D., Curley, E., Shelton, C., Yena, P., Ingle, J. N., Couch, F. J., Lingle, W. L., King, T. A., Gonzalez-Angulo, A. M., Mills, G. B., Dyer, M. D., Liu, S., Meng, X., Patangan, M., Waldman, F., Stoeppler, H., Rathmell, W. K., Thorne, L., Huang, M., Boice, L., Hill, A., Morrison, C., Gaudioso, C., Bshara, W., Daily, K., Egea, S. C., Pegram, M. D., Gomez-Fernandez, C., Dhir, R., Bhargava, R., Brufsky, A., Shriver, C. D., Hooke, J. A., Campbell, J. L., Mural, R. J., Hu, H., Somiari, S., Larson, C., Deyarmin, B., Kvecher, L., Kovatich, A. J., Ellis, M. J., King, T. A., Hu, H., Couch, F. J., Mural, R. J., Stricker, T., White, K., Olopade, O., Ingle, J. N., Luo, C., Chen, Y., Marks, J. R., Waldman, F., Wiznerowicz, M., Bose, R., Chang, L., Beck, A. H., Gonzalez-Angulo, A. M., Pihl, T., Jensen, M., Sfeir, R., Kahn, A., Chu, A., Kothiyal, P., Wang, Z., Snyder, E., Pontius, J., Ayala, B., Backus, M., Walton, J., Baboud, J., Berton, D., Nicholls, M., Srinivasan, D., Raman, R., Girshik, S., Kigonya, P., Alonso, S., Sanbhadti, R., Barletta, S., Pot, D., Sheth, M., Demchok, J. A., Shaw, K. R., Yang, L., Eley, G., Ferguson, M. L., Tarnuzzer, R. W., Zhang, J., Dillon, L. A., Buetow, K., Fielding, P., Ozenberger, B. A., Guyer, M. S., Sofia, H. J., Palchik, J. D. 2012; 490 (7418): 61-70

  Abstract

  We analysed primary breast cancers by genomic DNA copy number arrays, DNA methylation, exome sequencing, messenger RNA arrays, microRNA sequencing and reverse-phase protein arrays. Our ability to integrate information across platforms provided key insights into previously defined gene expression subtypes and demonstrated the existence of four main breast cancer classes when combining data from five platforms, each of which shows significant molecular heterogeneity. Somatic mutations in only three genes (TP53, PIK3CA and GATA3) occurred at >10% incidence across all breast cancers; however, there were numerous subtype-associated and novel gene mutations including the enrichment of specific mutations in GATA3, PIK3CA and MAP3K1 with the luminal A subtype. We identified two novel protein-expression-defined subgroups, possibly produced by stromal/microenvironmental elements, and integrated analyses identified specific signalling pathways dominant in each molecular subtype including a HER2/phosphorylated HER2/EGFR/phosphorylated EGFR signature within the HER2-enriched expression subtype. Comparison of basal-like breast tumours with high-grade serous ovarian tumours showed many molecular commonalities, indicating a related aetiology and similar therapeutic opportunities. The biological finding of the four main breast cancer subtypes caused by different subsets of genetic and epigenetic abnormalities raises the hypothesis that much of the clinically observable plasticity and heterogeneity occurs within, and not across, these major biological subtypes of breast cancer.

  View details for DOI 10.1038/nature11412

  View details for Web of Science ID 000309446800032

  View details for PubMedID 23000897

  View details for PubMedCentralID PMC3465532

• Whole genome in vivo RNAi screening identifies the leukemia inhibitory factor receptor as a novel breast tumor suppressor BREAST CANCER RESEARCH AND TREATMENT Iorns, E., Ward, T. M., Dean, S., Jegg, A., Thomas, D., Murugaesu, N., Sims, D., Mitsopoulos, C., Fenwick, K., Kozarewa, I., Naceur-Lombarelli, C., Zvelebil, M., Isacke, C. M., Lord, C. J., Ashworth, A., Hnatyszyn, H. J., Pegram, M., Lippman, M. 2012; 135 (1): 79-91

  Abstract

  Cancer is caused by mutations in oncogenes and tumor suppressor genes, resulting in the deregulation of processes fundamental to the normal behavior of cells. The identification and characterization of oncogenes and tumor suppressors has led to new treatment strategies that have significantly improved cancer outcome. The advent of next generation sequencing has allowed the elucidation of the fine structure of cancer genomes, however, the identification of pathogenic changes is complicated by the inherent genomic instability of cancer cells. Therefore, functional approaches for the identification of novel genes involved in the initiation and development of tumors are critical. Here we report the first whole human genome in vivo RNA interference screen to identify functionally important tumor suppressor genes. Using our novel approach, we identify previously validated tumor suppressor genes including TP53 and MNT, as well as several novel candidate tumor suppressor genes including leukemia inhibitory factor receptor (LIFR). We show that LIFR is a key novel tumor suppressor, whose deregulation may drive the transformation of a significant proportion of human breast cancers. These results demonstrate the power of genome wide in vivo RNAi screens as a method for identifying novel genes regulating tumorigenesis.

  View details for DOI 10.1007/s10549-012-2068-7

  View details for Web of Science ID 000307333700008

  View details for PubMedID 22535017

• Tumor Biology Trumps Anatomy in Breast Cancer Brain Metastases ONCOLOGY-NEW YORK Pegram, M. D. 2012; 26 (7): 666-?

  View details for Web of Science ID 000306619500012

  View details for PubMedID 22888569

• Possible available treatment option for early stage, small, node-negative, and HER2-overexpressing breast cancer BREAST CANCER Araki, K., Saji, S., Gallas, M., Pegram, M., Sasaki, Y. 2012; 19 (2): 95-103

  Abstract

  Trastuzumab is known for its clinical activity in women with HER2-overexpressing breast cancer. Randomized clinical trials have shown significant improvement in disease-free and overall survival with trastuzumab administered in conjunction with adjuvant chemotherapy for early-stage HER2-positive breast cancer. However, there is no direct evidence of clinical benefit from adjuvant trastuzumab in patients with node-negative, HER2-overexpressing, small (T1a-b) breast cancers. Previous literature shows that most breast cancers with node-negative small tumors have a good prognosis, but HER2-overexpressing disease might still be worse in this population. Some recent retrospective studies showed that an adjuvant trastuzumab-based regimen has a better prognostic effect, even in patients with node-negative, HER2-overexpressing, small breast cancers, although absolute survival differences were small. On the basis of the available literature, we believe that trastuzumab should be considered for patients with minimal HER2-overexpressing disease, although tools for accurate selection of patients at risk of relapse still need to be developed.

  View details for DOI 10.1007/s12282-011-0296-z

  View details for Web of Science ID 000303380800002

  View details for PubMedID 21863310

• Phase 2 study of neoadjuvant treatment with NOV-002 in combination with doxorubicin and cyclophosphamide followed by docetaxel in patients with HER-2 negative clinical stage II-IIIc breast cancer BREAST CANCER RESEARCH AND TREATMENT Montero, A. J., Diaz-Montero, C. M., Deutsch, Y. E., Hurley, J., Koniaris, L. G., Rumboldt, T., Yasir, S., Jorda, M., Garret-Mayer, E., Avisar, E., Slingerland, J., Silva, O., Welsh, C., Schuhwerk, K., Seo, P., Pegram, M. D., Glueck, S. 2012; 132 (1): 215-223

  Abstract

  NOV-002 (a formulation of disodium glutathione disulfide) modulates signaling pathways involved in tumor cell proliferation and metastasis and enhances anti-tumor immune responsiveness in tumor models. The addition of NOV-002 to chemotherapy has been shown to increase anti-tumor efficacy in animal models and some early phase oncology trials. We evaluated the clinical effects of NOV-002 in primary breast cancer, whether adding NOV-002 to standard preoperative chemotherapy increased pathologic complete response rates (pCR) at surgery, and determined whether NOV-002 mitigated hematologic toxicities of chemotherapy and whether levels of myeloid derived suppressor cells (MDSC) were predictive of response. Forty-one women with newly diagnosed stages II-IIIc HER-2 negative breast cancer received doxorubicin-cyclophosphamide followed by docetaxel (AC ? T) every 3 weeks and concurrent daily NOV-002 injections. The trial was powered to detect a doubling of pCR rate from 16 to 32% with NOV-002 plus AC ? T (? = 0.05, ? = 80%). Weekly complete blood counts were obtained as well as circulating MDSC levels on day 1 of each cycle were quantified. Of 39 patients with 40 evaluable tumors, 15 achieved a pCR (38%), meeting the primary endpoint of the trial. Concurrent NOV-002 resulted in pCR rates for AC ? T chemotherapy higher than previously reported. Patients with lower levels of circulating MDSCs at baseline and on the last cycle of chemotherapy had significantly higher probability of a pCR (P = 0.02). Further evaluation of NOV-002 in a randomized study is warranted.

  View details for DOI 10.1007/s10549-011-1889-0

  View details for Web of Science ID 000300278400021

  View details for PubMedID 22138748

• Trastuzumab Treatment in Multiple Lines: Current Data and Future Directions CLINICAL BREAST CANCER Pegram, M., Liao, J. 2012; 12 (1): 10-18

  Abstract

  Trastuzumab improves response rate, time to progression, and overall survival when combined with first-line chemotherapy in patients with human epidermal growth factor receptor 2-positive (HER2-positive) metastatic breast cancer (MBC). However, the benefits of continuing trastuzumab beyond disease progression have not been clearly established. The literature was reviewed to obtain data on trastuzumab use beyond disease progression. In general, data from retrospective and observational studies suggest that there may be clinical benefit when trastuzumab is used beyond disease progression. These results are supported by prospective non-randomized studies. Response rates and survival outcomes have generally been superior in patients who have continued trastuzumab after disease progression compared with those who have not. Moreover, recent data from two prospective randomized phase III trials have shown that adding trastuzumab to the treatment regimen in patients with MBC who have progressed on trastuzumab-based therapy significantly prolongs progression-free survival. Emerging evidence from randomized controlled trials supports the potential clinical utility of continuing trastuzumab-based therapy beyond progression and supports the National Comprehensive Cancer Network recommendation to consider this treatment approach. Future treatment of HER2-positive MBC may involve trastuzumab being used in successive regimens in combination with other targeted therapies.

  View details for DOI 10.1016/j.clbc.2011.07.003

  View details for Web of Science ID 000299862300003

  View details for PubMedID 22154153

• Randomized Phase II Trial of Letrozole plus Anti-MUC1 Antibody AS1402 in Hormone Receptor-Positive Locally Advanced or Metastatic Breast Cancer CLINICAL CANCER RESEARCH Ibrahim, N. K., Yariz, K. O., Bondarenko, I., Manikhas, A., Semiglazov, V., Alyasova, A., Komisarenko, V., Shparyk, Y., Murray, J. L., Jones, D., Senderovich, S., Chau, A., Erlandsson, F., Acton, G., Pegram, M. 2011; 17 (21): 6822-6830

  Abstract

  AS1402 is a humanized immunoglobulin G1 antibody that targets the aberrantly glycosylated antigen MUC1, which is overexpressed in 90% of breast tumors and contributes to estrogen-mediated growth and survival of breast cancer cells in vitro by modulating estrogen receptor (ER) activity. Aromatase inhibitors have been reported to enhance antibody-dependent cell-mediated cytotoxicity elicited by antibodies in vitro. We compared the outcomes of patients with breast cancer treated with letrozole with or without AS1402.The study population included 110 patients with locally advanced or metastatic hormone receptor-positive breast cancer randomized to receive 2.5 mg letrozole only once daily or with a weekly 9 mg/kg AS1402 infusion. The primary endpoint was overall response rate. Secondary endpoints included progression-free survival, time to progression, and safety. AS1402 exposure and influence of allotypes of Fc?RIIIa, Fc?RIIa, and MUC1 were evaluated.The study was stopped early because of a trend toward worse response rates and a higher rate of early disease progression in the AS1402 + letrozole arm. Final analysis revealed no significant difference in efficacy between the study arms. Evaluated gene polymorphisms did not define patient subgroups with improved outcomes. Addition of AS1402 to letrozole was associated with manageable toxicity.Because adding AS1402 to letrozole did not improve outcomes compared with letrozole only, blocking ER may be a better strategy for harnessing MUC1 modulation of the ER to a clinical advantage. Fc?RIIIa, Fc?RIIa, and MUC1 allotype did not predict outcome for patients treated with letrozole with or without AS1402.

  View details for DOI 10.1158/1078-0432.CCR-11-1151

  View details for Web of Science ID 000296624000025

  View details for PubMedID 21878535

• HER2 discordance between primary breast cancer and its paired metastasis: tumor biology or test artefact? Insights through meta-analysis BREAST CANCER RESEARCH AND TREATMENT Houssami, N., Macaskill, P., Balleine, R. L., Bilous, M., Pegram, M. D. 2011; 129 (3): 659-674

  Abstract

  The proto-oncogene, HER2, has prognostic and predictive relevance in invasive breast cancer (IBC). HER2 testing of primary IBC guides treatment selection and is assumed to reflect HER2 status of associated metastases, although HER2 discordance between IBC and metastasis has been reported. Systematic review and meta-analysis of HER2 status in IBC and its paired loco-regional or distant metastasis were done. Quality appraisal considered whether (within-subject) testing conditions were maintained for paired primary and metastasis. Random effects logistic regression models were used to estimate pooled within-subject HER2 discordant proportions and to examine study-level covariates, including tumor-related and testing-related variables, potentially associated with HER2 discordance differences across (between) studies. Modelled paired HER2 data for primary and metastatic cancer (2520 subjects, 26 studies) showed a pooled HER2 discordance of 5.5% (3.6-8.5%). Sensitivity analysis, excluding the only study not maintaining same conditions for paired testing, gave a pooled estimate of 5.2% (3.5-7.8%). Pooled discordant proportion was not associated with differences between studies in test type, test scoring or interpretation criteria, subjects' median age, study time-frame, or HER2 positivity in primary cancer (all P > 0.05). However, type of metastasis was significantly associated with estimated HER2 discordance (P = 0.0017): studies of primary tumor paired with distant metastases had higher discordance [11.5% (6.9-18.6%)] than studies of primary paired with lymph node metastases only [4.1% (2.4-7.2%)], or those paired with nodal or various metastases [3.3% (2.0-5.6%)]; P < 0.01. HER2 discordant proportion was higher where paired metastases were metachronous relative to synchronous to primary IBC (P = 0.0024). Sensitivity analysis provided weak evidence (P = 0.074) that discordance in the direction of change from HER2-negative primary cancer to HER2-positive paired metastasis was more likely than the reverse. Study-level meta-analysis suggests factors associated with the type of metastasis as underlying mechanisms for observed HER2 discordance between primary IBC and paired metastasis. Test-related factors did not account for differences across studies in the HER2 discordant proportion.

  View details for DOI 10.1007/s10549-011-1632-x

  View details for Web of Science ID 000294680600001

  View details for PubMedID 21698410

• Genetic polymorphisms of multiple DNA repair pathways impact age at diagnosis and TP53 mutations in breast cancer CARCINOGENESIS Smith, T. R., Liu-Mares, W., Van Emburgh, B. O., Levine, E. A., Allen, G. O., Hill, J. W., Reis, I. M., Kresty, L. A., Pegram, M. D., Miller, M. S., Hu, J. J. 2011; 32 (9): 1354-1360

  Abstract

  Defective DNA repair may contribute to early age and late stage at time of diagnosis and mutations in critical tumor suppressor genes, such as TP53 in breast cancer. Using DNA samples from 436 breast cancer cases (374 Caucasians and 62 African-Americans), we tested these associations with 18 non-synonymous single-nucleotide polymorphisms (nsSNPs) in four DNA repair pathways: (i) base excision repair: ADPRT V762A, APE1 D148E, XRCC1 R194W/R280H/R399Q and POLD1 R119H; (ii) double-strand break repair: NBS1 E185Q and XRCC3 T241M; (iii) mismatch repair: MLH1 I219V, MSH3 R940Q/T1036A and MSH6 G39E and (iv) nucleotide excision repair: ERCC2 D312N/K751Q, ERCC4 R415Q, ERCC5 D1104H and XPC A499V/K939Q. Younger age at diagnosis (<50) was associated with ERCC2 312 DN/NN genotypes [odds ratio (OR) = 1.76; 95% confidence interval (CI) = 1.10, 2.81] and NBS1 185 QQ genotype (OR = 3.09; 95% CI = 1.47, 6.49). The XPC 939 QQ genotype was associated with TP53 mutations (OR = 5.80; 95% CI = 2.23, 15.09). There was a significant trend associating younger age at diagnosis (<50) with increasing numbers of risk genotypes for ERCC2 312 DN/NN, MSH6 39 EE and NBS1 185 QQ (P(trend) < 0.001). A similar significant trend was also observed associating TP53 mutations with increasing numbers of risk genotypes for XRCC1 399 QQ, XPC 939 QQ, ERCC4 415 QQ and XPC 499 AA (P(trend) < 0.001). Our pilot data suggest that nsSNPs of multiple DNA repair pathways are associated with younger age at diagnosis and TP53 mutations in breast cancer and larger studies are warranted to further evaluate these associations.

  View details for DOI 10.1093/carcin/bgr117

  View details for Web of Science ID 000294495500009

  View details for PubMedID 21700777

• Multicenter Phase III Randomized Trial Comparing Docetaxel and Trastuzumab With Docetaxel, Carboplatin, and Trastuzumab As First-Line Chemotherapy for Patients With HER2-Gene-Amplified Metastatic Breast Cancer (BCIRG 007 Study): Two Highly Active Therapeutic Regimens JOURNAL OF CLINICAL ONCOLOGY Valero, V., Forbes, J., Pegram, M. D., Pienkowski, T., Eiermann, W., von Minckwitz, G., Roche, H., Martin, M., Crown, J., Mackey, J. R., Fumoleau, P., Rolski, J., Mrsic-Krmpotic, Z., Jagiello-Gruszfeld, A., Riva, A., Buyse, M., Taupin, H., Sauter, G., Press, M. F., Slamon, D. J. 2011; 29 (2): 149-156

  Abstract

  Docetaxel-trastuzumab (TH) is effective therapy for HER2-amplified metastatic breast cancer (MBC). Preclinical findings of synergy between docetaxel, carboplatin, and trastuzumab (TCH) prompted a phase III randomized trial comparing TCH with TH in patients with HER2-amplified MBC.Two hundred sixty-three patients were randomly assigned to receive eight 3-week cycles of TH (trastuzumab plus docetaxel 100 mg/m(2)) or TCH (trastuzumab plus carboplatin at area under the serum concentration-time curve 6 and docetaxel 75 mg/m(2)). Trastuzumab was given at 4 mg/kg loading dose followed by a 2 mg/kg dose once per week during chemotherapy, and then 6 mg/kg once every 3 weeks until progression.Patient characteristics were balanced between groups. There was no significant difference between TH and TCH in terms of the primary end point, time to progression (medians of 11.1 and 10.4 months, respectively; hazard ratio, 0.914; 95% CI, 0.694 to 1.203; P = .57), response rate (72% for both groups), or overall survival (medians of 37.1 and 37.4 months, respectively; P = .99). Rates of grades 3 or 4 adverse effects for TH and TCH, respectively, were neutropenic-related complications, 29% and 23%; thrombocytopenia, 2% and 15%; anemia, 5% and 11%; sensory neuropathy, 3% and 0.8%; fatigue, 5% and 12%; peripheral edema, 3.8% and 1.5%; and diarrhea, 2% and 10%. Two patients given TCH died of sepsis, and one patient given TH experienced sudden cardiac death. Absolute left ventricular ejection fraction decline > 15% was seen in 5.5% of patients on the TH arm and 6.7% of patients on the TCH arm.Adding carboplatin did not enhance TH antitumor activity.TH (docetaxel, 100 mg/m(2)) and TCH (docetaxel, 75 mg/m(2)) demonstrated efficacy with acceptable toxicity in women with HER2-amplified MBC.

  View details for DOI 10.1200/JCO.2010.28.6450

  View details for Web of Science ID 000285965400018

  View details for PubMedID 21115860

• Proceedings of the First Global Workshop on Breast Cancer: Pathways to the Evaluation and Clinical Development of Novel Agents for Breast Cancer CLINICAL BREAST CANCER Albain, K. S., Carey, L., Gradishar, W. J., Gralow, J. R., Lipton, A., Rugo, H., Tripathy, D., Peck, S., Abair, T., Pegram, M. 2010; 10 (6): 421-439

  Abstract

  The number of treatment options available to patients with breast cancer is larger and more complex than ever before. This is due in part to increased understanding of breast cancer tumor biology and the signaling pathways involved in tumor development and progression, which drives new areas of breast cancer research and the development of novel agents. Therapies targeting HER2 signaling, angiogenesis, DNA repair, and many other essential cellular processes that are dysregulated in cancer have produced significant improvements in disease outcome, although careful patient selection and toxicity management are required to maximize their therapeutic potential. Multigene assays have added to the ability to predict disease outcome and degree of response to adjuvant chemotherapy, but the application of these assays in the right clinical context is necessary. Unfortunately, despite the use of appropriate and effective local and adjuvant therapies, some patients with early-stage breast cancer will eventually develop metastatic disease. Most of these patients will have received standard therapies in the adjuvant setting and/or will develop resistance to these therapies at some point during treatment. Thus, implementation of novel strategies is necessary to overcome resistance and improve disease outcome. This in turn will require creative clinical trial designs, more efficient accrual, and rapid translation of results into the clinical setting. This summary highlights selected challenges in the current management of breast cancer and discusses expert perspectives, key questions, areas of debate, and future directions.

  View details for DOI 10.3816/CBC.2010.n.056

  View details for Web of Science ID 000284915800002

  View details for PubMedID 21147685

• Population pharmacokinetics of the humanised monoclonal antibody, HuHMFG1 (AS1402), derived from a phase I study on breast cancer BRITISH JOURNAL OF CANCER Royer, B., Yin, W., Pegram, M., Ibrahim, N., Villanueva, C., Mir, D., Erlandsson, F., Pivot, X. 2010; 102 (5): 827-832

  Abstract

  HuHMFG1 (AS1402) is a humanised monoclonal antibody that has undergone a phase I trial in metastatic breast cancer. The aim of this study was to characterise the pharmacokinetics (PKs) of HuHMFG1 using a population PK model.Data were derived from a phase I study of 26 patients receiving HuHMFG1 at doses ranging from 1 to 16 mg kg(-1). Data were analysed using NONMEM software and covariates were included. A limited sampling strategy (LSS) was developed using training and a validation data set.A linear two-compartment model was shown to be adequate to describe data. Covariate analysis indicated that weight was not related to clearance. An LSS was successfully developed on the basis of the model, in which one sample is collected immediately before the start of an infusion and the second is taken at the end of infusion.A two-compartment population PK model successfully describes HuHMFG1 behaviour. The model suggests using a fixed dose of HuHMFG1, which would simplify dosing. The model could be used to optimise dose level and dosing schedule if more data on the correlation between exposure and efficacy become available from future studies. The derived LSS could optimise further PK assessment of this antibody.

  View details for DOI 10.1038/sj.bjc.6605560

  View details for Web of Science ID 000275170300007

  View details for PubMedID 20160731

  View details for PubMedCentralID PMC2833251

• Targeted Therapy in Metastatic Breast Cancer: The HER2/neu Oncogene. Breast care (Basel, Switzerland) Harbeck, N., Pegram, M. D., Rüschoff, J., Möbus, V. 2010; 5 (s1): 3?7

  Abstract

  SUMMARY: Besides surgery, radiation, chemotherapy, and endocrine treatment, immunotherapy has become an established part of systemic therapy in treating metastatic breast cancer. One of the most interesting targets for the design of anticancer therapeutics is the HER2/ErbB2 receptor which is overexpressed in about 20-25% of breast cancers. Given the poor prognosis of women whose tumors express ErbB2 (HER2) at high levels, accurate determination of the ErbB2 status should be routinely performed in women with newly diagnosed invasive breast cancer. Efficacy and safety data of numerous trials led to the approval of the monoclonal antibody trastuzumab as the first ErbB2-targeting therapy in ErbB2-positive breast cancer. However, the majority of patients who achieve an initial response to trastuzumab-based regimens for metastatic disease develop resistance within 1 year. This underlines the need for alternative or additional anti-ErbB2-targeting strategies.

  View details for DOI 10.1159/000285714

  View details for PubMedID 20847829

  View details for PubMedCentralID PMC2931093

• Lapatinib: new opportunities for management of breast cancer. Breast cancer (Dove Medical Press) Liao, J., Gallas, M., Pegram, M., Slingerland, J. 2010; 2: 79-91

  Abstract

  Approximately 20% of new diagnosed breast cancers overexpress the human epidermal growth factor receptor 2 (EGFR2), also known as erythroblastic leukemia viral oncogene homolog 2 (ERBB2) protein, as a consequence of ERBB2 gene amplification, resulting in a poor prognosis. Clinical outcome can be substantially improved by ERBB2-targeted therapy. Lapatinib is a potent, orally bioavailable small molecule that reversibly and selectively inhibits epidermal growth factor receptor (EGFR1 or ERBB1) and ERBB2 tyrosine kinases. Lapatinib binds the adenosine triphosphate-binding site of the receptor's intracellular domain to inhibit tumor cell growth. This review summarizes the pharmacology, pharmacokinetics, efficacy, and tolerability of lapatinib, and reviews both Food and Drug Administration-approved and investigational uses of lapatinib in breast cancer therapy. The drug is generally well tolerated in patients, with diarrhea and rashes being the most common (usually mild or moderate) adverse effects. Unlike trastuzumab, lapatinib has infrequent adverse effects on cardiac function. Lapatinib has substantial activity for advanced ERBB2-positive breast cancer, particularly in combination with capecitabine, following progression after anthracyclines, taxanes, and trastuzumab. Lapatinib combined with capecitabine yielded significant improvements in time to progression and response rate compared with capecitabine alone. This drug can also be combined with letrozole for the treatment of postmenopausal women with ERBB2-positive breast cancer, for whom hormonal therapy is indicated. Lapatinib has shown early promise in treatment of central nervous system metastasis and is being further evaluated in various clinical settings.

  View details for DOI 10.2147/BCTT.S5929

  View details for PubMedID 24367169

  View details for PubMedCentralID PMC3846530

• Lapatinib Combined With Letrozole Versus Letrozole and Placebo As First-Line Therapy for Postmenopausal Hormone Receptor-Positive Metastatic Breast Cancer JOURNAL OF CLINICAL ONCOLOGY Johnston, S., Pippen, J., Pivot, X., Lichinitser, M., Sadeghi, S., Dieras, V., Gomez, H. L., Romieu, G., Manikhas, A., Kennedy, M. J., Press, M. F., Maltzman, J., Florance, A., O'Rourke, L., Oliva, C., Stein, S., Pegram, M. 2009; 27 (33): 5538-5546

  Abstract

  Cross-talk between human epidermal growth factor receptors and hormone receptor pathways may cause endocrine resistance in breast cancer. This trial evaluated the effect of adding lapatinib, a dual tyrosine kinase inhibitor blocking epidermal growth factor receptor and human epidermal growth factor receptor 2 (HER2), to the aromatase inhibitor letrozole as first-line treatment of hormone receptor (HR) -positive metastatic breast cancer (MBC).Postmenopausal women with HR-positive MBC were randomly assigned to daily letrozole (2.5 mg orally) plus lapatinib (1,500 mg orally) or letrozole and placebo. The primary end point was progression-free survival (PFS) in the HER2-positive population. Results In HR-positive, HER2-positive patients (n = 219), addition of lapatinib to letrozole significantly reduced the risk of disease progression versus letrozole-placebo (hazard ratio [HR] = 0.71; 95% CI, 0.53 to 0.96; P = .019); median PFS was 8.2 v 3.0 months, respectively. Clinical benefit (responsive or stable disease >or= 6 months) was significantly greater for lapatinib-letrozole versus letrozole-placebo (48% v 29%, respectively; odds ratio [OR] = 0.4; 95% CI, 0.2 to 0.8; P = .003). Patients with centrally confirmed HR-positive, HER2-negative tumors (n = 952) had no improvement in PFS. A preplanned Cox regression analysis identified prior antiestrogen therapy as a significant factor in the HER2-negative population; a nonsignificant trend toward prolonged PFS for lapatinib-letrozole was seen in patients who experienced relapse less than 6 months since prior tamoxifen discontinuation (HR = 0.78; 95% CI, 0.57 to 1.07; P = .117). Grade 3 or 4 adverse events were more common in the lapatinib-letrozole arm versus letrozole-placebo arm (diarrhea, 10% v 1%; rash, 1% v 0%, respectively), but they were manageable.This trial demonstrated that a combined targeted strategy with letrozole and lapatinib significantly enhances PFS and clinical benefit rates in patients with MBC that coexpresses HR and HER2.

  View details for DOI 10.1200/JCO.2009.23.3734

  View details for Web of Science ID 000271954200010

  View details for PubMedID 19786658

• Single-agent lapatinib for HER2-overexpressing advanced or metastatic breast cancer that progressed on first- or second-line trastuzumab-containing regimens ANNALS OF ONCOLOGY Blackwell, K. L., Pegram, M. D., Tan-Chiu, E., Schwartzberg, L. S., Arbushites, M. C., Maltzman, J. D., Forster, J. K., Rubin, S. D., Stein, S. H., Burstein, H. J. 2009; 20 (6): 1026-1031

  Abstract

  This phase II study evaluated the efficacy and safety of lapatinib in patients with human epidermal growth factor receptor 2 (HER2)-positive advanced or metastatic breast cancer that progressed during prior trastuzumab therapy.Women with stage IIIB/IV HER2-overexpressing breast cancer were treated with single-agent lapatinib 1250 or 1500 mg once daily after protocol amendment. Tumor response according to RECIST was assessed every 8 weeks. HER2 expression was assessed in tumor tissue by immunohistochemistry and FISH.Seventy-eight patients were enrolled in the study. Investigator and independent review response rates [complete response (CR) or partial response (PR)] were 7.7% and 5.1%, and clinical benefit rates (CR, PR, or stable disease for >or=24 weeks) were 14.1% and 9.0%, respectively. Median time to progression was 15.3 weeks by independent review, and median overall survival was 79 weeks. The most common treatment-related adverse events were rash (47%), diarrhea (46%), nausea (31%), and fatigue (18%).Single-agent lapatinib has clinical activity with manageable toxic effects in HER2-overexpressing breast cancer that progressed on trastuzumab-containing therapy. Studies of lapatinib-based combination regimens with chemotherapy and other targeted therapies in metastatic and earlier stages of breast cancer are warranted.

  View details for DOI 10.1093/annonc/mdn759

  View details for Web of Science ID 000266343900009

  View details for PubMedID 19179558

• Phase I dose escalation pharmacokinetic assessment of intravenous humanized anti-MUC1 antibody AS1402 in patients with advanced breast cancer BREAST CANCER RESEARCH Pegram, M. D., Borges, V. F., Ibrahim, N., Fuloria, J., Shapiro, C., Perez, S., Wang, K., Stark, F. S., Luck, N. C. 2009; 11 (5)

  Abstract

  MUC1 is a cell-surface glycoprotein that establishes a molecular barrier at the epithelial surface and engages in morphogenetic signal transduction. Alterations in MUC1 glycosylation accompany the development of cancer and influence cellular growth, differentiation, transformation, adhesion, invasion, and immune surveillance. A 20-amino-acid tandem repeat that forms the core protein of MUC1 is overexpressed and aberrantly glycosylated in the majority of epithelial tumors. AS1402 (formerly R1550) is a humanized IgG1k monoclonal antibody that binds to PDTR sequences within this tandem repeat that are not exposed in normal cells. AS1402 is a potent inducer of antibody-dependent cellular cytotoxicity (ADCC), specifically against MUC1-expressing tumor cells. The objective of this study was to determine the safety, tolerability, and pharmacokinetic (PK) characteristics of AS1402 monotherapy in patients with locally advanced or metastatic MUC1-positive breast cancer that had progressed after anthracyclines- and taxane-based therapy.Patients received AS1402 over a 1- to 3-hour intravenous (i.v.) infusion at doses between 1 and 16 mg/kg, with repeated dosing every 1 to 3 weeks (based on patient-individualized PK assessment) until disease progression. Serum AS1402 levels were measured at multiple times after i.v. administration. Human anti-human antibody (HAHA) responses were measured to determine the immunogenicity of AS1402. Noncompartmental pharmacokinetic parameters were determined and were used to assess dose dependency across the dose range studied.Twenty-six patients were treated. AS1402 was generally well tolerated. Two grade 3/4 drug-related adverse events were reported, both at the 3-mg/kg dose. Neither was observed in expanded or subsequent dosing cohorts. No anti-human antibodies were detected. Plasma concentrations of AS1402 appeared to be proportional to dose within the 1- to 16-mg/kg dose range assessed, with a mean terminal half-life of 115.4 +/- 37.1 hours.Repeated iv administration of AS1402 was well tolerated, with a maximum tolerated dose (MTD) exceeding 16 mg/kg, the highest dose administered in this study. The half-life and exposure of AS1402 were such that weekly dosing could achieve plasma concentrations corresponding to the maximal ADCC activity observed in vitro. A phase II study is ongoing to evaluate the clinical activity of AS1402 in patients with advanced breast cancer.ClinicalTrials.gov Identifier: NCT00096057.

  View details for DOI 10.1186/bcr2409

  View details for Web of Science ID 000273342300018

  View details for PubMedID 19811637

• Phase I dose escalation and pharmacokinetic study of lapatinib in combination with trastuzumab in patients with advanced ErbB2-positive breast cancer 28th Annual San Antonio Breast Cancer Symposium Storniolo, A. M., Pegram, M. D., Overmoyer, B., Silverman, P., Peacock, N. W., Jones, S. F., Loftiss, J., Arya, N., Koch, K. M., Paul, E., Pandite, L., Fleming, R. A., Lebowitz, P. F., Ho, P. T., Burris, H. A. AMER SOC CLINICAL ONCOLOGY. 2008: 3317?23

  Abstract

  The combination of lapatinib and trastuzumab has been observed to have a synergistic, antiproliferative effect against ErbB2-positive breast cancer cells in vitro. This phase I study assessed the safety, clinical feasibility, optimally tolerated regimen (OTR), pharmacokinetics (PK), and preliminary clinical activity of this combination in patients with ErbB2-positive advanced breast cancer.Cohorts of three patients with ErbB2-positive advanced breast cancer were treated with escalating doses of lapatinib (750 to 1,500 mg) administered once daily (continuous) in combination with trastuzumab (4 mg/kg loading dose then 2 mg/kg weekly) to determine the OTR. Once the OTR was determined, additional patients were enrolled to provide the PK profile of both agents alone and in combination.A total of 54 patients were treated: 27 in the dose-escalation group and 27 in the PK group. Overall, adverse events were mild to moderate in severity, with no drug-related grade 4 events. The most frequent drug-related grade 3 events included diarrhea (17%), fatigue (11%), and rash (6%). The OTR was 1,000 mg lapatinib with standard weekly trastuzumab. One patient had a complete response and seven patients had partial responses. The PK parameters (maximum concentration in plasma and area under the curve) of lapatinib and trastuzumab in combination were not significantly different than when either was administered alone.The OTR of the lapatinib/trastuzumab combination was lapatinib 1,000 mg per day with standard weekly trastuzumab. At these doses, the regimen was well tolerated and clinically active in this heavily pretreated ErbB2-positive breast cancer population.

  View details for DOI 10.1200/JCO.2007.13.5202

  View details for Web of Science ID 000258046700008

  View details for PubMedID 18490651

• Can we circumvent resistance to ErbB2-targeted agents by targeting novel pathways? CLINICAL BREAST CANCER Pegram, M. 2008; 8: S121-S130

  Abstract

  The recent development of targeted therapies using monoclonal antibodies has added new dimensions to the rapidly evolving field of breast cancer treatment. In particular, the incorporation of trastuzumab into regimens containing existing chemotherapeutic agents has significantly improved clinical outcomes for patients with breast cancer in the adjuvant and metastatic settings. De novo and acquired resistance to this treatment, however, is widespread. A substantial amount of research has therefore been dedicated to the elucidation of molecular mechanisms that could explain resistance to this otherwise effective therapy. Potential mechanisms for resistance to trastuzumab include steric inhibition imposed by other extracellular factors, molecular changes in the target receptor itself (ErbB2), alterations in the regulation of downstream signaling components, and crosstalk with other pathways that could compensate for attenuated ErbB2 signaling. In addition, preclinical and clinical studies have been performed to identify potential methods for overcoming trastuzumab resistance, including targeting alternate ErbB2 epitopes and the combined inhibition of multiple signaling components and/or pathways (vertical or horizontal inhibition). Studies continue to evaluate the most promising approaches for overcoming mechanisms of resistance to trastuzumab and other ErbB2-targeted therapies. This review will summarize the most recent research designed to address this substantial clinical problem and provide clinicians with relevant background for understanding some of the potential molecular mechanisms for resistance to targeted therapies in the treatment of patients with breast cancer.

  View details for Web of Science ID 000254738400004

  View details for PubMedID 18777951

• Biodistribution and predictive value of (IF)-I-18-fluorocyclophosphamide in mice bearing human breast cancer xenografts JOURNAL OF NUCLEAR MEDICINE Kesner, A. L., Hsueh, W., Htet, N. L., Pio, B. S., Czernin, J., Pegram, M. D., Phelps, M. E., Silverman, D. H. 2007; 48 (12): 2021-2027

  Abstract

  In mice bearing human breast cancer xenografts, we examined the biodistribution of (18)F-fluorocyclophosphamide ((18)F-F-CP) to evaluate its potential as a noninvasive prognostic tool for predicting the resistance of tumors to cyclophosphamide therapy.(18)F-F-CP was synthesized as we recently described, and PET data were acquired after administration of (18)F-F-CP in mice bearing human breast cancer xenografts (MCF-7 cells). Tracer biodistribution in reconstructed images was quantified by region-of-interest analysis. Distribution was also assessed by harvesting dissected organs, tumors, and blood, determining (18)F content in each tissue with a gamma-well counter. The mice were subsequently treated with cyclophosphamide, and tumor size was monitored for at least 3 wk after chemotherapy administration.The distribution of harvested activity correlated strongly with distribution observed in PET images. Target organs were related to routes of metabolism and excretion. (18)F-F-CP uptake was highest in kidneys, lowest in brain, and intermediate in tumors, as determined by both image-based and tissue-based measurements. (18)F-F-CP uptake was not inhibited by coadministration of an approximately x700 concentration of unlabeled cyclophosphamide. PET measures of (18)F-F-CP uptake in tumor predicted the magnitude of the response to subsequent administration of cyclophosphamide.Noninvasive assessment of (18)F-F-CP uptake using PET may potentially be helpful for predicting the response of breast tumors to cyclophosphamide before therapy begins.

  View details for DOI 10.2967/jnumed.107.045716

  View details for Web of Science ID 000252895100020

  View details for PubMedID 18006620

• American Society of Clinical Oncology/College of American Pathologists guideline recommendations for human epidermal growth factor receptor 2 testing in breast cancer JOURNAL OF CLINICAL ONCOLOGY Wolff, A. C., Hammond, M. E., Schwartz, J. N., Hagerty, K. L., Allred, D. C., Cote, R. J., Dowsett, M., Fitzgibbons, P. L., Hanna, W. M., Langer, A., McShane, L. M., Paik, S., Pegram, M. D., Perez, E. A., Press, M. F., Rhodes, A., Sturgeon, C., Taube, S. E., Tubbs, R., Vance, G. H., de Vijver, M. v., Wheeler, T. M., Hayes, D. F. 2007; 25 (1): 118-145

  Abstract

  To develop a guideline to improve the accuracy of human epidermal growth factor receptor 2 (HER2) testing in invasive breast cancer and its utility as a predictive marker.The American Society of Clinical Oncology and the College of American Pathologists convened an expert panel, which conducted a systematic review of the literature and developed recommendations for optimal HER2 testing performance. The guideline was reviewed by selected experts and approved by the board of directors for both organizations.Approximately 20% of current HER2 testing may be inaccurate. When carefully validated testing is performed, available data do not clearly demonstrate the superiority of either immunohistochemistry (IHC) or in situ hybridization (ISH) as a predictor of benefit from anti-HER2 therapy.The panel recommends that HER2 status should be determined for all invasive breast cancer. A testing algorithm that relies on accurate, reproducible assay performance, including newly available types of brightfield ISH, is proposed. Elements to reliably reduce assay variation (for example, specimen handling, assay exclusion, and reporting criteria) are specified. An algorithm defining positive, equivocal, and negative values for both HER2 protein expression and gene amplification is recommended: a positive HER2 result is IHC staining of 3+ (uniform, intense membrane staining of > 30% of invasive tumor cells), a fluorescent in situ hybridization (FISH) result of more than six HER2 gene copies per nucleus or a FISH ratio (HER2 gene signals to chromosome 17 signals) of more than 2.2; a negative result is an IHC staining of 0 or 1+, a FISH result of less than 4.0 HER2 gene copies per nucleus, or FISH ratio of less than 1.8. Equivocal results require additional action for final determination. It is recommended that to perform HER2 testing, laboratories show 95% concordance with another validated test for positive and negative assay values. The panel strongly recommends validation of laboratory assay or modifications, use of standardized operating procedures, and compliance with new testing criteria to be monitored with the use of stringent laboratory accreditation standards, proficiency testing, and competency assessment. The panel recommends that HER2 testing be done in a CAP-accredited laboratory or in a laboratory that meets the accreditation and proficiency testing requirements set out by this document.

  View details for DOI 10.1200/JCO.2006.09.2775

  View details for Web of Science ID 000243725900020

  View details for PubMedID 17159189

• American Society of Clinical Oncology/College of American Pathologists guideline recommendations for human epidermal growth factor receptor 2 testing in breast cancer ARCHIVES OF PATHOLOGY & LABORATORY MEDICINE Wolff, A. C., Hammond, M. E., Schwartz, J. N., Hagerty, K. L., Allred, D. C., Cote, R. J., Dowsett, M., Fitzgibbons, P. L., Hanna, W. M., Langer, A., McShane, L. M., Paik, S., Pegram, M. D., Perez, E. A., Press, M. F., Rhodes, A., Sturgeon, C., Taube, S. E., Tubbs, R., Vance, G. H., de Vijver, M. v., Wheeler, T. M., Hayes, D. F. 2007; 131 (1): 18-43

  Abstract

  To develop a guideline to improve the accuracy of human epidermal growth factor receptor 2(HER2) testing in invasive breast cancer and its utility as a predictive marker.The American Society of Clinical Oncology and the College of American Pathologists convened an expert panel, which conducted a systematic review of the literature and developed recommendations for optimal HER2 testing performance. The guideline was reviewed by selected experts and approved by the board of directors for both organizations.Approximately 20% of current HER2 testing may be inaccurate. When carefully validated testing is performed, available data do not clearly demonstrate the superiority of either immunohistochemistry(IHC) or in situ hybridization (ISH) as a predictor of benefit from anti-HER2 therapy.The panel recommends that HER2 status should be determined for all invasive breast cancer. A testing algorithm that relies on accurate, reproducible assay performance, including newly available types of brightfield ISH, is proposed. Elements to reliably reduce assay variation (for example, specimen handling, assay exclusion, and reporting criteria) are specified. An algorithm defining positive, equivocal, and negative values for both HER2 protein expression and gene amplification is recommended: a positive HER2 result is IHC staining of 3 + (uniform, intense membrane staining of 30% of invasive tumor cells), a fluorescent in situ hybridization (FISH) result of more than six HER2 gene copies per nucleus or a FISH ratio (HER2 gene signals to chromosome 17 signals) of more than 2.2; a negative result is an IHC staining of 0 or 1 +, a FISH result of less than 4.0 HER2 gene copies per nucleus, or FISH ratio of less than 1.8. Equivocal results require additional action for final determination. It is recommended that to perform HER2 testing, laboratories show 95% concordance with another validated test for positive and negative assay values. The panel strongly recommends validation of laboratory assay or modifications, use of standardized operating procedures, and compliance with new testing criteria to be monitored with the use of stringent laboratory accreditation standards, proficiency testing, and competency assessment. The panel recommends that HER2 testing be done in a CAP-accredited laboratory or in a laboratory that meets the accreditation and proficiency testing requirements set out by this document.

  View details for Web of Science ID 000243464500004

  View details for PubMedID 19548375

• Predicting chemotherapy response to paclitaxel with F-18-fluoropaclitaxel and PET JOURNAL OF NUCLEAR MEDICINE Hsueh, W., Kesner, A. L., Gangloff, A., Pegram, M. D., Beryt, M., Czernin, J., Phelps, M. E., Silverman, D. H. 2006; 47 (12): 1995-1999

  Abstract

  Paclitaxel is used as a chemotherapy drug for the treatment of various malignancies, including breast, ovarian, and lung cancers. To evaluate the potential of a noninvasive prognostic tool for specifically predicting the resistance of tumors to paclitaxel therapy, we examined the tumoral uptake of (18)F-fluoropaclitaxel ((18)F-FPAC) in mice bearing human breast cancer xenografts by using small-animal-dedicated PET and compared (18)F-FPAC uptake with the tumor response to paclitaxel treatment.PET data were acquired after tail vein injection of approximately 9 MBq of (18)F-FPAC in anesthetized nude mice bearing breast cancer xenografts. Tracer uptake in reconstructed images was quantified by region-of-interest analyses and compared with the tumor response, as measured by changes in tumor volume, after treatment with paclitaxel.Mice with tumors that progressed demonstrated lower tumoral uptake of (18)F-FPAC than mice with tumors that did not progress or that regressed (r = 0.55, P < 0.02; n = 19), indicating that low (18)F-FPAC uptake was a significant predictor of chemoresistance. Conversely, high (18)F-FPAC uptake predicted tumor regression. This relationship was found for mice bearing xenografts from cell lines selected to be either sensitive or intrinsically resistant to paclitaxel in vitro.PET data acquired with (18)F-FPAC suggest that this tracer holds promise for the noninvasive quantification of its distribution in vivo in a straightforward manner. In combination with approaches for examining other aspects of resistance, such quantification could prove useful in helping to predict subsequent resistance to paclitaxel chemotherapy of breast cancer.

  View details for Web of Science ID 000242563900040

  View details for PubMedID 17138742

• Clinical activity of Pertuzumab (rhuMAb 2C4), a HER dimerization inhibitor, in advanced ovarian cancer: Potential predictive relationship with tumor HER2 activation status 41st Annual Meeting of the American-Society-of-Clinical-Oncology Gordon, M. S., Matei, D., Aghajanian, C., Matulonis, U. A., Brewer, M., Fleming, G. F., Hainsworth, J. D., Garcia, A. A., Pegram, M. D., Schilder, R. J., Cohn, D. E., Roman, L., Derynck, M. K., Ng, K., Lyons, B., Allison, D. E., Eberhard, D. A., Pham, T. Q., Dere, R. C., Karlan, B. Y. AMER SOC CLINICAL ONCOLOGY. 2006: 4324?32

  Abstract

  Ovarian cancers (OCs) frequently have HER2 activation in the absence of HER2 overexpression. Pertuzumab, a humanized antibody that prevents HER2 dimerization and inhibits multiple HER-mediated pathways, was studied in a phase II, multicenter trial in advanced, refractory OC.Sixty-one patients (cohort 1) with relapsed OC received a loading dose of 840 mg pertuzumab intravenously followed by 420 mg every 3 weeks; 62 patients (cohort 2) received 1,050 mg every 3 weeks. Response rate was the primary end point. Fresh tumor biopsies were obtained in cohort 1 to assay for phosphorylated HER2 (pHER2).Median age was 57 years and median number of prior chemotherapy regimens was five. Fifty-five patients in cohort 1 and 62 patients in cohort 2 were assessable for efficacy. There were five partial responses (response rate [RR] = 4.3%; 95% CI, 1.7% to 9.4%), eight patients (6.8%) with stable disease (SD) lasting at least 6 months, and 10 patients with CA-125 reduction of at least 50% (includes two partial responses and four patients with SD > or = 6 months; total clinical activity, 14.5%). Median progression-free survival (PFS) was 6.6 weeks. Eight of 28 tumor biopsies (28.6%) were pHER2+ by enzyme-linked immunosorbent assay (ELISA; without gene amplification). Median PFS for pHER2+ patients was 20.9 weeks (n = 8) versus 5.8 weeks for pHER2- (n = 20; P = .14) and 9.1 weeks for unknown pHER2 status (n = 27). Pertuzumab was well tolerated with diarrhea in 69.1% (11.4% grade 3, no grade 4). Five patients had asymptomatic left ventricular ejection fraction decreases to less than 50% (one confirmed by central facility).Pertuzumab is well tolerated with a RR of 4.3% in heavily-pretreated OC patients. Further studies on pHER2 as a diagnostic are warranted.

  View details for DOI 10.1200/JCO.2005.05.4221

  View details for Web of Science ID 000240645300014

  View details for PubMedID 16896006

• Application and potential limitations of animal models utilized in the development of trastuzumab (Herceptin((R))): A case study ADVANCED DRUG DELIVERY REVIEWS Pegram, M., Ngo, D. 2006; 58 (5-6): 723-734

  Abstract

  The preclinical and clinical development of trastuzumab, a humanized monoclonal antibody directed against a juxtamembrane epitope in the HER2 receptor ectodomain, relied heavily on the use of animal models to validate HER2 as a potential MAb target. The identification of HER2 (neu) as a proto-oncogene was first established in a carcinogen-induced brain tumor in the rat. Transgenic mouse technology led to an understanding of the role of HER2 in pathogenesis of breast cancer. Transfection studies of human HER2 cDNA into murine xenograft models further explored the role HER2 plays in tumor progression and metastasis. A murine subrenal capsule fresh human tumor explant assay was utilized to test efficacy of various murine monoclonal anti-HER2 antibodies, and the data were helpful in choosing the most efficacious for subsequent human engineering for clinical use. HER2-overexpressing xenograft models in athymic mice were used to test the efficacy of anti-HER2 antibodies, develop dose-response relationships, measure drug interactions between trastuzumab and chemotherapy, and optimize dosing schedules of chemotherapeutics combined with trastuzumab. In this work, we will highlight the utility of animal models exploited in the development of trastuzumab - noting not only their contribution to drug development but also their limitations in translation of preclinical data into the clinic. It is likely that the experience we gained in the case of preclinical animal models to study in vivo effects of trastuzumab have parallels in the development of other monoclonal antibodies since overcoming the species boundaries (i.e. cross-reactivity with antigenic determinant, development of cross-species neutralizing antibodies, and cross-species interaction with activating Fc receptors on immune effector cells) are major limitations in the design and interpretation of preclinical/translational experiments designed to fulfill various regulatory requirements prior to initiation of phase I human clinical trials.

  View details for DOI 10.1016/j.addr.2006.05.003

  View details for Web of Science ID 000240573100008

  View details for PubMedID 16876287

• Inhibitors of growth factor receptors, signaling pathways and angiogenesis as therapeutic molecular agents 1st International Symposium on Cancer Metastasis and the Lymphovascular System Holash, J., Thurston, G., Rudge, J. S., Yancopoulos, G. D., Adjei, A. A., Bergers, G., Pytowski, B., Pegram, M., Gordon, M. S. SPRINGER. 2006: 243?52

  View details for DOI 10.1007/s10555-006-8504-6

  View details for Web of Science ID 000238268800007

  View details for PubMedID 16770536

• Docetaxel, cisplatin, and trastuzumab as primary systemic therapy for human epidermal growth factor receptor 2-positive locally advanced breast cancer 26th Annual San Antonio Breast Cancer Symposium Hurley, J., Doliny, P., Reis, I., Silva, O., Gomez-Fernandez, C., Velez, P., Pauletti, G., Pegram, M. D., Slamon, D. J. AMER SOC CLINICAL ONCOLOGY. 2006: 1831?38

  Abstract

  To evaluate the efficacy and safety of docetaxel, cisplatin, and trastuzumab as primary systemic therapy for human epidermal growth factor receptor 2 (HER2) -positive, locally advanced breast cancer (LABC).Forty-eight patients with immunohistochemistry-confirmed HER2-positive LABC or inflammatory breast cancer received 12 weeks of docetaxel, cisplatin, and trastuzumab with filgrastim, followed by surgery, adjuvant doxorubicin and cyclophosphamide, and locoregional radiotherapy with or without tamoxifen. The primary end point was pathologic complete response (pCR) in breast.Baseline mean tumor size was 9.2 cm (range, 4 to 32 cm). pCR occurred in breast in 11 patients (23%; 95% CI, 12% to 37%) and breast and axilla in eight patients (17%; 95% CI, 8% to 30%). pCR rates in breast (HER2 positive, seven of 30 patients, 23% v HER2 negative, four of 18 patients, 22%; P > .05) and breast and axilla (four of 30 patients, 13% v four of 18 patients, 22%, respectively; P > .05) were similar regardless of HER2 status by fluorescence in situ hybridization (FISH). At a median follow-up time of 43 months, 4-year progression-free survival (PFS) rate was 81% (95% CI, 64% to 90%); overall survival (OS) rate was 86% (95% CI, 71% to 94%). In patients with pCR in breast and axilla, PFS and OS rates were 100% (95% CI, inestimable). In patients without pCR, PFS rate was 76% (95% CI, 57% to 88%; P = .15, log-rank test), and OS rate was 83% (95% CI, 66% to 92%; P = .21). Survival rates were similar regardless of FISH status. There were only two grade 4 adverse events.Twelve weeks of docetaxel, cisplatin, and trastuzumab is clinically active and leads to excellent survival in patients with large, HER2-positive tumors.

  View details for DOI 10.1200/JCO.2005.02.8886

  View details for Web of Science ID 000237124300010

  View details for PubMedID 16549824

• Survivin expression in breast cancer predicts clinical outcome and is associated with HER2, VEGF, urokinase plasminogen activator and PAI-1 41st Annual Meeting of the American-Society-of-Clinical-Oncology Ryan, B. M., Konecny, G. E., Kahlert, S., Wang, H. J., Untch, M., Meng, G., Pegram, M. D., Podratz, K. C., Crown, J., Slamon, D. J., Duffy, M. J. OXFORD UNIV PRESS. 2006: 597?604

  Abstract

  Survivin, a novel inhibitor of apoptosis, is one of the most cancer-specific proteins identified to date. In this study we (a) evaluated the association between survivin and HER2, vascular endothelial growth factor (VEGF) and uPA/PAI-1 expression and (b) defined its effect on clinical outcome in a large breast cancer patient cohort.Survivin expression was measured by ELISA in primary breast cancer tissue extracts from 420 patients with long-term clinical follow-up.Survivin was detected in 378 (90%) of the 420 primary breast cancer cases. Increased survivin levels were significantly associated with high nuclear grade (P < 0.0001), negative hormone receptor status (P = 0.0028), HER2 overexpression (P = 0.0094), VEGF expression (P < 0.0001), high uPA (P = 0.0002) and PAI-1 levels (P = 0.0002). Using the 25th percentile (1.4 ng/mg) as a cut-off point, patients expressing elevated survivin had a significantly worse disease-free survival (DFS: P = 0.0007, RR 1.97) and overall survival (OS: P = 0.0009, RR 2.11) compared with patients expressing lower levels of survivin. In multivariate analysis, this prognostic value of survivin was independent of both traditional and novel clinicopathologic factors for both DFS (P = 0.0076, RR 1.72) and OS (P = 0.0155, RR 1.76).The independent prognostic relevance of survivin, when combined with previous data from model systems implicating survivin in the inhibition of apoptosis, suggests that survivin may be a suitable target for future therapeutic strategies.

  View details for DOI 10.1093/annonc/mdj121

  View details for Web of Science ID 000236251200009

  View details for PubMedID 16403812

• Activity of the dual kinase inhibitor lapatinib (GW572016) against HER-2-overexpressing and trastuzumab-treated breast cancer cells CANCER RESEARCH Konecny, G. E., Pegram, M. D., Venkatesan, N., Finn, R., Yang, G. R., Rahmeh, M., Untch, M., Rusnak, D. W., Spehar, G., Mullin, R. J., Keith, B. R., Gilmer, T. M., Berger, M., Podratz, K. C., Slamon, D. J. 2006; 66 (3): 1630-1639

  Abstract

  Lapatinib (GW572016) is a selective inhibitor of both epidermal growth factor receptor (EGFR) and HER-2 tyrosine kinases. Here, we explore the therapeutic potential of lapatinib by testing its effect on tumor cell growth in a panel of 31 characterized human breast cancer cell lines, including trastuzumab-conditioned HER-2-positive cell lines. We further characterize its activity in combination with trastuzumab and analyze whether EGFR and HER-2 expression or changes induced in the activation of EGFR, HER-2, Raf, AKT, or extracellular signal-regulated kinase (ERK) are markers of drug activity. We report that concentration-dependent antiproliferative effects of lapatinib were seen in all breast cancer cell lines tested but varied significantly between individual cell lines with up to 1,000-fold difference in the IC(50)s (range, 0.010-18.6 micromol/L). Response to lapatinib was significantly correlated with HER-2 expression and its ability to inhibit HER-2, Raf, AKT, and ERK phosphorylation. Long-term in vivo lapatinib studies were conducted with human breast cancer xenografts in athymic mice. Treatment over 77 days resulted in a sustained and significant reduction in xenograft volume compared with untreated controls. For the combination of lapatinib plus trastuzumab, synergistic drug interactions were observed in four different HER-2-overexpressing cell lines. Moreover, lapatinib retained significant in vitro activity against cell lines selected for long-term outgrowth (>9 months) in trastuzumab-containing (100 microg/mL) culture medium. These observations provide a clear biological rationale to test lapatinib as a single agent or in combination with trastuzumab in HER-2-overexpressing breast cancer and in patients with clinical resistance to trastuzumab.

  View details for DOI 10.1158/0008-5472.CAN-05-1182

  View details for Web of Science ID 000235095900049

  View details for PubMedID 16452222

• Usefulness of 3 '-[F-18]fluoro-3 '-deoxythymidine with positron emission tomography in predicting breast cancer response to therapy MOLECULAR IMAGING AND BIOLOGY Pio, B. S., Park, C. K., Pietras, R., Hsueh, W. A., Satyamurthy, N., Pegram, M. D., Czernin, J., Phelps, M. E., Silverman, D. H. 2006; 8 (1): 36-42

  Abstract

  The usefulness of 2-deoxy-2-[F-18]fluoro-D-glucose (FDG)-positron emission tomography (PET) in monitoring breast cancer response to chemotherapy has previously been reported. Elevated uptake of FDG by treated tumors can persist however, particularly in the early period after treatment is initiated. 3'-[F-18]Fluoro-3'-deoxythymidine (FLT) has been developed as a marker for cellular proliferation and, in principle, could be a more accurate predictor of the long-term effect of chemotherapy on tumor viability. We examined side-by-side FDG and FLT imaging for monitoring and predicting tumor response to chemotherapy.Fourteen patients with newly diagnosed primary or metastatic breast cancer, who were about to commence a new pharmacologic treatment regimen, were prospectively studied. Dynamic 3-D PET imaging of uptake into a field of view centered over tumor began immediately after administration of FDG or FLT (150 MBq). After 45 minutes of dynamic acquisition, a clinically standard whole-body PET scan was acquired. Patients were scanned with both tracers on two separate days within one week of each other (1) before beginning treatment, (2) two weeks following the end of the first cycle of the new regimen, and (3) following the final cycle of that regimen, or one year after the initial PET scans, whichever came first. (Median and mean times of early scans were 5.0 and 6.6 weeks after treatment initiation; median and mean times for late scans were 26.0 and 30.6 weeks after treatment initiation.) Scan data were analyzed on both tumor-by-tumor and patient-by-patient bases, and compared to each patient's clinical course.Mean change in FLT uptake in primary and metastatic tumors after the first course of chemotherapy showed a significant correlation with late (av. interval 5.8 months) changes in CA27.29 tumor marker levels (r = 0.79, P = 0.001). When comparing changes in tracer uptake after one chemotherapy course versus late changes in tumor size as measured by CT scans, FLT was again a good predictor of eventual tumor response (r = 0.74, P = 0.01). Tumor uptake of FLT was near-maximal by 10 minutes after injection. The time frame five to 10 minutes postinjection of FLT produced standardized uptake value (SUV) values highly correlated with SUV values obtained after 45-minute uptake (r = 0.83, P < 0.0001), and changes in these early SUVs after the first course of chemotherapy correlated with late changes in CA27.29 (r = 0.93, P = 0.003).A 10-minute FLT-PET scan acquired two weeks after the end of the first course of chemotherapy is useful for predicting longer-term efficacy of chemotherapy regimens for women with breast cancer.

  View details for DOI 10.1007/s11307-005-0029-9

  View details for Web of Science ID 000235474100006

  View details for PubMedID 16362149

• Reducing the global breast cancer burden: the importance of patterns of care research. Clinical breast cancer Albain, K. S., De la Garza Salazar, J., Pienkowski, T., Aapro, M., Bergh, J., Caleffi, M., Coleman, R., Eiermann, W., Icli, F., Pegram, M., Piccart, M., Snyder, R., Toi, M., Hortobagyi, G. N. 2005; 6 (5): 412-420

  Abstract

  Breast cancer treatment guidelines are not uniformly followed in clinical practice, with evidence for substantial variations in treatment patterns, quality of care, and patient outcomes among and within countries. The factors that drive treatment decisions are unclear. Furthermore, the impact of different treatment strategies on survival is poorly understood outside the clinical trial setting. Sources of patterns of care information often have limitations in completeness, quality, timeliness of reporting, and relevance to the larger population. Patterns of care studies frequently lack details on cancer stage at diagnosis, tumor biology, and treatment received. It is difficult to compare data between studies and/or track changes over time because of variations in data sources and collection techniques. Thus, the design and implementation of a global registry is sorely needed in order to prospectively evaluate worldwide patterns of care and outcomes in patients with breast cancer. Components of this registry should include random selection of centers of variable practice settings in multiple countries and accurate and rapid data reporting at prestudy and follow-up timepoints. Data collected would include tumor and demographic factors, staging information, treatment rendered, and survival. Variables that influenced the treatment selected would be assessed. This unique international effort would allow the development of strategies to improve diagnostic and treatment-related standards of care and survival outcomes, thus reducing the breast cancer burden worldwide.

  View details for PubMedID 16381624

• Estimation of paclitaxel biodistribution and uptake in human-derived xenografts in vivo with F-18-fluoropaclitaxel JOURNAL OF NUCLEAR MEDICINE Gangloff, A., Hsueh, W. A., Kesner, A. L., Kiesewetter, D. O., Pio, B. S., Pegram, M. D., Beryt, M., Townsend, A., Czernin, J., Phelps, M. E., Silverman, D. H. 2005; 46 (11): 1866-1871

  Abstract

  Paclitaxel (PAC) is widely used as a chemotherapy drug in the treatment of various malignancies, including breast, ovarian, and lung cancers. We examined the biodistribution of (18)F-fluoropaclitaxel ((18)F-FPAC) in mice with and without human breast cancer tumor xenografts by use of small-animal-dedicated PET (microPET) and clinically practical semiquantitative methods. We compared the PET data to data derived from direct harvesting and analysis of blood, organs, and breast carcinoma xenografts.PET data were acquired after tail vein injection of (18)F-FPAC in nude mice. Tracer biodistribution in reconstructed images was quantified by region-of-interest analysis. Biodistribution also was assessed by harvesting and analysis of dissected organs, tumors, and blood after coadministration of (18)F-FPAC and (3)H-PAC. (18)F content in each tissue was assessed with a gamma-well counter, and (3)H content was quantified by scintillation counting of solubilized tissue after (18)F radioactive decay.The distributions of (18)F-FPAC and (3)H-PAC were very similar, with the highest concentrations in the small intestine, the lowest concentrations in the brain, and intermediate concentrations in tumor. Uptake in these and other tissues was not inhibited by the presence of more pharmacologic doses of unlabeled PAC. Administration of the P-glycoprotein modulator cyclosporine doubled the uptake of both (18)F-FPAC and (3)H-PAC into tumor.PET studies with (18)F-FPAC can be used in conjunction with clinically practical quantification methods to yield estimates of PAC uptake in breast cancer tumors and normal organs noninvasively.

  View details for Web of Science ID 000233095800019

  View details for PubMedID 16269601

• Anti-erbB-2 antibody trastuzumab in the treatment of HER2-amplified breast cancer INVESTIGATIONAL NEW DRUGS Yeon, C. H., Pegram, M. D. 2005; 23 (5): 391-409

  Abstract

  Human epidermal growth factor receptor-2 (HER2/erbB-2) is a member of a family of four transmembrane receptor tyrosine kinases that regulate cell growth, survival and differentiation via multiple signal transduction pathways. Amplification of the HER2 gene occurs in 20-25% of human breast cancers. This amplification event is an independent adverse prognostic factor as well as a predictive factor for increased response to doxorubicin-based combination chemotherapy, response to trastuzumab and decreased response to hormonal therapy. Methods for detecting protein overexpression or gene amplification in clinical tumor specimens include immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) techniques, with the latter considered by some to be more accurate. Trastuzumab (Herceptin) is a recombinant humanized monoclonal antibody which targets an epitope in the extracellular domain of the HER2 protein. Preclinical models demonstrated that this antibody has significant anti-tumor activity as a single agent and has synergy with certain chemotherapeutic drugs. Phase II and III clinical trials performed in women with metastatic breast cancer that overexpress HER2 have shown that trastuzumab has clinical activity when used as first-, second- or third-line monotherapy, and improves survival when used as first-line therapy in combination with chemotherapy. Newer combinations with numerous chemotherapeutic drugs have also shown significant clinical activity in phase II studies. In all of these trials, trastuzumab was generally well-tolerated, but cardiac toxicity (particularly when the antibody was combined with anthracyclines) was an unexpected adverse effect. Although trastuzumab is currently usually administered on a weekly intravenous schedule, evidence suggests that a triple dose of the drug given once every three weeks has a pharmacokinetic profile expected to be equally efficacious. Neither the optimal schedule nor the optimal duration of trastuzumab therapy has yet been clearly defined in controlled clinical trials. Current clinical investigations of trastuzumab include its use in both the adjuvant and neoadjuvant settings as well as in combination with other chemotherapy drugs or new biologic targeted agents.

  View details for DOI 10.1007/s10637-005-2899-8

  View details for Web of Science ID 000231245000001

  View details for PubMedID 16133791

• Image-detected breast cancer: State of the art diagnosis and treatment JOURNAL OF THE AMERICAN COLLEGE OF SURGEONS Silverstein, M. J., Lagios, M. D., Recht, A., Allred, D. C., Harms, S. E., Holland, R., Holmes, D. R., Hughes, L. L., Jackman, R. J., Julian, T. B., Kuerer, H. M., Mabry, H. C., McCready, D. R., McMasters, K. M., Page, D. L., Parker, S. H., Pass, H. A., Pegram, M., Rubin, E., Stavros, A. T., Tripathy, D., Vicini, F., Whitworth, P. W. 2005; 201 (4): 586-597

  View details for DOI 10.1016/j.jamcolisurg.2005.05.032

  View details for Web of Science ID 000232433500016

  View details for PubMedID 16183499

• Epidermal growth factor receptor and signal transduction: potential targets for anti-cancer therapy ANTI-CANCER DRUGS Pal, S. K., Pegram, M. 2005; 16 (5): 483-494

  Abstract

  Agents targeting the epidermal growth factor receptor (EGFR) pathway hold particular promise for the treatment of patients with advanced disease, for whom standard chemotherapy is generally palliative. Expression of EGFR on numerous types of solid tumors, and the association of EGFR activation with tumorigenic processes including proliferation, anti-apoptosis and metastatic spread, make this pathway a particularly compelling target for rational drug design. The two classes of anti-EGFR agents in late-stage clinical testing include antibodies directed toward the extracellular EGFR domain (cetuximab, panitumumab) and small molecule tyrosine kinase inhibitors (gefitinib, erlotinib), which inactivate the receptor enzyme activity. However, important issues remain to be addressed. These include the development of appropriate predictive markers for response, such as improved tests for EGFR activity, correlation of rash with response and potential pharmacogenomic approaches; the sequencing and combination of these agents with chemotherapy and irradiation; and the possible role of these agents in the treatment of patients with earlier stage disease.

  View details for Web of Science ID 000229224300003

  View details for PubMedID 15846113

• Targeted therapy: Wave of the future JOURNAL OF CLINICAL ONCOLOGY Pegram, M. D., Pietras, R., Bajamonde, A., Klein, P., Fyfe, G. 2005; 23 (8): 1776-1781

  View details for DOI 10.1200/JCO.2005.11.029

  View details for Web of Science ID 000227587200028

  View details for PubMedID 15755985

• Gemcitabine in combination with trastuzumab and/or platinum salts in breast cancer cells with HER2 overexpression. Oncology (Williston Park, N.Y.) Konecny, G. E., Pegram, M. D. 2004; 18 (14): 32-36

  Abstract

  Trastuzumab (Herceptin) is an effective treatment in patients with HER2-overexpressing metastatic breast cancer. Risk of trastuzumab-induced cardiotoxicity raises concerns regarding combined use with anthracyclines or other potentially cardiotoxic agents following anthracycline treatment. We characterized interactions between trastuzumab and gemcitabine (Gemzar) and the combination of gemcitabine and cisplatin or carboplatin (Paraplatin) as such combinations might help reduce the risk of cardiotoxicity. Multiple drug effect/combination index isobologram analysis was used to study the efficacy of chemotherapeutic drug plus trastuzumab combinations in HER2-overexpressing breast cancer cell lines. Combination index values were derived from parameters of the median effect plots, and statistical tests were used to determine whether the mean combination index at multiple effect levels significantly differed from a combination index value of 1.0 (values < 1.0 indicate synergy; values > 1.0, antagonism; values equal to 1.0, additivity). At a wide range of clinically achievable drug concentrations, interactions between trastuzumab and gemcitabine were synergistic at low concentrations of gemcitabine and antagonistic at high concentrations. A consistent synergistic interaction was observed with the three-drug combination of trastuzumab plus gemcitabine plus carboplatin or cisplatin. Available clinical data on the use of trastuzumab plus gemcitabine, and trastuzumab plus gemcitabine/paclitaxel, as well as clinical data on the use of gemcitabine/cisplatin in breast cancer, are discussed. These findings indicate that trastuzumab plus gemcitabine and trastuzumab plus gemcitabine plus cisplatin or carboplatin are rational combinations to evaluate in clinical trials.

  View details for PubMedID 15685824

• Anti-VEGF antibody bevacizumab (Avastin) with 5FU/LV as third line treatment for colorectal cancer. Techniques in coloproctology Emmanouilides, C., Pegram, M., Robinson, R., Hecht, R., Kabbinavar, F., Isacoff, W. 2004; 8: s50-2

  Abstract

  To evaluate the activity and safety of bevacizumab when given with standard 5FU/leukovorin (LV) regimens in patients with metastatic colorectal cancer who have failed irinotecan and oxaliplatin-based treatments.Bevacizumab was given at 5 mg/kg as an IV infusion every 2 weeks. Patients received 5FU according to Roswell Park or the de Gramont regimen.Nineteen patients enrolled, median age 60, median PS: 1. Most common toxicity attributable to bevacizumab was mild hypertension, epistaxis and mild proteinuria; 1 patient had a CNS haemorrhage. The median number of cycles was 1 (8 weeks). Clinical benefit as disease stabilisation lasting 2-6 months was noted in 9 patients, whereas 10 progressed (median f/u: 5 months). TTP was 16 weeks, and the overall survival has not been reached (24+ weeks).Bevacizumab may result in growth arrest and clinical benefit in a substantial proportion of patients with colorectal cancer and no alternative treatment.

  View details for PubMedID 15655642

• HER-2/neu gene amplification and response to paclitaxel in patients with metastatic breast cancer 37th Annual Meeting of the American-Society-of-Clinical-Oncology Konecny, G. E., Thomssen, C., Luck, H. J., Untch, M., Wang, H. J., Kuhn, W., Eidtmann, H., du Bois, A., Olbricht, S., Steinfeld, D., MOBUS, V., von Minckwitz, G., Dandekar, S., Ramos, L., Pauletti, G., Pegram, M. D., JANICKE, F., Slamon, D. J. OXFORD UNIV PRESS INC. 2004: 1141?51

  Abstract

  HER-2/neu overexpression appears to be associated with improved response to anthracycline-based chemotherapy, but its association with response to taxane-based chemotherapy is unclear. In this retrospective subset analysis of patients with metastatic breast cancer enrolled in a randomized treatment trial, we investigated the response of patients with known HER-2/neu status to treatment with taxane-based epirubicin-paclitaxel (ET) chemotherapy compared with treatment with epirubicin-cyclophosphamide (EC) chemotherapy.HER-2/neu status (positive [i.e., HER-2/neu amplification] or negative [i.e., no HER-2/neu amplification]) of archival specimens of primary tumors from 297 patients with metastatic breast cancer was determined by use of fluorescence in situ hybridization. Associations between HER-2/neu status and the efficacy of randomly assigned chemotherapy (ET versus EC) were investigated. All statistical tests were two-sided. Results: Patients with HER-2/neu-positive tumors had a statistically significantly greater objective response rate than patients with HER-2/neu-negative tumors to treatment with ET (76% versus 50%, respectively; P =.005) but not to treatment with EC (46% versus 33%; P =.130). The objective response rate associated with ET was greater than that associated with EC for both HER-2/neu-positive tumors (76% versus 46%; P =.004) and HER-2/neu-negative tumors (50% versus 33%; P =.002). However, the improvement in the objective response rate associated with ET, compared with that associated with EC, was greater for patients with HER-2/neu-positive tumors (adjusted odds ratio [OR] = 3.64, 95% confidence interval [CI] = 1.48 to 8.92; P=.005) than for patients with HER-2/neu-negative tumors (adjusted OR = 1.92, 95% CI = 1.01 to 3.64; P=.046). Among patients with HER-2/neu-positive tumors, those who received ET had better progression-free survival and overall survival than those who received EC (for progression-free survival, adjusted relative risk [RR] = 0.65, 95% CI = 0.42 to 1.02; P=.062; for overall survival, adjusted RR = 0.60, 95% CI = 0.36 to 1.02; P=.059). However, among patients with HER-2/neu-negative tumors, those who received ET and those who received EC had similar progression-free survival and overall survival.HER-2/neu amplification does not adversely influence response to first-line chemotherapy with either ET or EC. Furthermore, a taxane-containing regimen such as ET may provide a preferential benefit to patients with HER-2/neu-positive tumors.

  View details for DOI 10.1093/jnci/djh198

  View details for Web of Science ID 000223172200009

  View details for PubMedID 15292386

• Results of two open-label, multicenter phase II studies of docetaxel, platinum salts, and trastuzumab in HER2-positive advanced breast cancer JOURNAL OF THE NATIONAL CANCER INSTITUTE Pegram, M. D., Pienkowski, T., Northfelt, D. W., Eiermann, W., Patel, R., Fumoleau, P., Quan, E., Crown, J., Toppmeyer, D., Smylie, M., Riva, A., Blitz, S., Press, M. F., Reese, D., Lindsay, M. A., Slamon, D. J. 2004; 96 (10): 759-769

  Abstract

  Preclinical data indicate that docetaxel, platinum salts, and the combination of both drugs are highly synergistic with the anti-HER2 antibody trastuzumab. The University of California at Los Angeles-Oncology Research Network (UCLA-ORN) and the Breast Cancer International Research Group (BCIRG) have conducted two phase II studies to evaluate docetaxel and trastuzumab in combination with either cisplatin or carboplatin for the treatment of women with advanced breast cancer that overexpresses HER2.Each study enrolled 62 patients with HER2-overexpressing tumors. Patients received a median of six cycles of docetaxel at 75 mg/m2 of body surface area and cisplatin (BCIRG 101 study) at 75 mg/m2 or carboplatin (UCLA-ORN study) at AUC = 6 mg/mL. min given on day 1 and then every 21 days. Trastuzumab was given on day 1, cycle 1 (4 mg/kg) and then continued weekly at 2 mg/kg for 1 year or until disease progression. Tumor measurements were obtained at baseline, after three cycles of chemotherapy, and then every 3 months. HER2 gene amplification was determined by fluorescence in situ hybridization.Patient characteristics were comparable between trials with the exception that 15% of the patients in the UCLA-ORN study had received previous adjuvant taxane therapy. Both regimens were well tolerated, with manageable toxicities. Hematologic toxicities were more frequent in patients in the UCLA-ORN study than in patients in the BCIRG 101 study, whereas the reverse pattern was observed for non-hematologic toxicities. One patient in each study developed reversible congestive heart failure. Responses were observed in 49 of 62 patients in the BCIRG 101 study (overall response rate = 79%, 95% confidence interval [CI] = 66% to 89%) and in 34 of 59 evaluable patients in the UCLA-ORN study (overall response rate = 58%, 95% CI = 44% to 70%). Median times to progression were 9.9 months (95% CI = 8.3 to 13.1 months) and 12.7 months (95% CI = 8.6 to 15.5 months) for patients in the BCIRG 101 and UCLA-ORN studies, respectively. Overall response rates were higher and median time to progression was longer in the subset of patients whose tumors harbored HER2 gene amplification.Combinations of docetaxel, a platinum salt, and trastuzumab are feasible and active in patients with advanced breast cancers that overexpress HER2. The BCIRG is conducting ongoing randomized studies of the three-drug combination in both the metastatic and adjuvant settings.

  View details for DOI 10.1093/jnci/djh133

  View details for Web of Science ID 000221621400009

  View details for PubMedID 15150304

• Rational combinations of trastuzumab with chemotherapeutic drugs used in the treatment of breast cancer JOURNAL OF THE NATIONAL CANCER INSTITUTE Pegram, M. D., Konecny, G. E., O'Callaghan, C., Beryt, M., Pietras, R., Slamon, D. J. 2004; 96 (10): 739-749

  Abstract

  Trastuzumab, a humanized anti-HER2 antibody, increases the clinical benefit of first-line chemotherapy in patients with metastatic breast cancers that overexpress HER2. We characterized interactions between trastuzumab and chemotherapeutic agents commonly used in the treatment of breast cancer.Multiple drug effect/combination index isobologram analysis was used to study the efficacy of chemotherapeutic drug plus trastuzumab combinations tested against four HER2-overexpressing breast cancer cell lines (SK-BR-3, BT-474, MDA-MB-361, and MDA-MB-453). Combination index values were derived from parameters of the median effect plots, and statistical tests were used to determine whether the mean combination index values at multiple effect levels were statistically significantly different from a combination index value of 1.0. Values less than 1.0 indicate synergistic interactions, values greater than 1.0 indicate antagonistic interactions, and values equal to 1.0 indicate additive interactions.At a wide range of clinically achievable drug concentrations, synergistic interactions were observed in all four breast cancer cell lines for trastuzumab plus carboplatin (mean combination index values ranged from 0.32 [P<.001] to 0.53 [P<.001]), 4-hydroxycyclophosphamide (mean combination index values ranged from 0.38 [P<.001] to 0.73 [P =.010]), docetaxel (mean combination index values ranged from 0.30 [P<.001] to 0.62 [P<.001]), and vinorelbine (mean combination index values ranged from 0.24 [P<.001] to 0.78 [P<.034]). Additive interactions were observed in all four cell lines with trastuzumab plus doxorubicin, epirubicin, and paclitaxel. Interactions between trastuzumab and gemcitabine were synergistic at low concentrations of gemcitabine and antagonistic at high concentrations. A synergistic interaction was observed with a three-drug combination of docetaxel plus carboplatin plus trastuzumab in SK-BR-3 cells (mean combination index value = 0.09; P<.001).Consistent synergistic interactions of trastuzumab plus carboplatin, 4-hydroxycyclophosphamide, docetaxel, or vinorelbine across a wide range of clinically relevant concentrations in HER2-overexpressing breast cancer cells indicate that these are rational combinations to test in human clinical trials.

  View details for DOI 10.1093/jnci/djh131

  View details for Web of Science ID 000221621400007

  View details for PubMedID 15150302

• Association between HER-2/neu and vascular endothelial growth factor expression predicts clinical outcome in primary breast cancer patients CLINICAL CANCER RESEARCH Konecny, G. E., Meng, Y. G., Untch, M., Wang, H. J., Bauerfeind, I., Epstein, M., Stieber, P., Vernes, J. M., Gutierrez, J., Hong, K., Beryt, M., Hepp, H., Slamon, D. J., Pegram, M. D. 2004; 10 (5): 1706-1716

  Abstract

  Activation or overexpression of HER-2/neu is associated with up-regulation of vascular endothelial growth factor (VEGF) in human breast cancer cells in vitro. Preclinical experiments indicate that increased expression of VEGF may in part mediate the biologically aggressive phenotype of HER-2/neu-overexpressing human breast cancer. It was the purpose of this study to: (a). evaluate the association between HER-2/neu and VEGF expression in a large clinical cohort of primary breast cancer patients; (b). compare the prognostic significance of VEGF isoforms; and (c). analyze the combined effects of HER-2/neu and VEGF on clinical outcome.HER-2/neu and VEGF were measured by ELISA in primary breast tumor tissue lysates from 611 unselected patients with a median clinical follow-up of 50 months. At least six VEGF isoforms consisting of 121, 145, 165, 183, 189, or 206 amino acids are generated as a result of alternative splicing. The VEGF(121-206) ELISA uses antibodies that bind to VEGF(121) and, therefore, detects all of the VEGF isoforms with 121 and more amino acids. The VEGF(165-206) ELISA uses antibodies that bind to VEGF(165) and, therefore, detects all of the VEGF isoforms with 165 and more amino acids. VEGF(121-206) and VEGF(165-206) were analyzed both as continuous and categorical variables, using detectable expression as a cutoff for positivity. Cell lines with defined HER-2/neu expression levels were used to establish a cutoff point for HER-2/neu overexpression in breast tumor samples.Our findings indicate a significant positive association between HER-2/neu and VEGF expression. VEGF(121-206) and VEGF(165-206) expression was detectable in 88 (77.2%) and 100 (87.7%), respectively, of the 114 patients with HER-2/neu-overexpressing tumors, in contrast to 271 (54.5%) and 353 (71.0%), respectively, of the 497 patients with nonoverexpressing tumors (chi(2) test: P < 0.001 for both VEGF(121-206) and VEGF(165-206)). VEGF(121-206) and VEGF(165-206) demonstrate a comparable prognostic significance for survival in unselected primary breast cancer patients (univariate analysis: VEGF(121-206), P = 0.0068; VEGF(165-206), P = 0.0046; multivariate analysis: VEGF(121-206), P = 0.1475; VEGF(165-206), P = 0.1483). When the analyses were performed separately for node-negative and node-positive patients, VEGF(121-206) and VEGF(165-206) were of prognostic significance for survival only in node-positive patients (univariate analysis: VEGF(121-206), P = 0.0003; VEGF(165-206), P = 0.0038; multivariate analysis: VEGF(121-206), P = 0.0103; VEGF(165-206), P = 0.0150). A biological concentration-effect relationship between VEGF expression and survival (VEGF(121-206), P = 0.0280; VEGF(165-206,) P = 0.0097) suggests that VEGF levels, as determined by ELISA, could be of importance as a predictive marker for therapeutic strategies that target VEGF. Combining HER-2/neu and VEGF(121-206)/VEGF(165-206) results in additional prognostic information for survival (VEGF(121-206), P = 0.0133; VEGF(165-206), P = 0.0092).The positive association between HER-2/neu and VEGF expression implicates VEGF in the aggressive phenotype exhibited by HER-2/neu overexpression, and supports the use of combination therapies directed against both HER-2/neu and VEGF for treatment of breast cancers that overexpress HER-2/neu.

  View details for Web of Science ID 000220089100022

  View details for PubMedID 15014023

• Quantitative association between, HER-2/neu and steroid hormone receptors in hormone receptor-positive primary breast cancer JOURNAL OF THE NATIONAL CANCER INSTITUTE Konecny, G., Pauletti, G., Pegram, M., Untch, M., Dandekar, S., Aguilar, Z., Wilson, C., Rong, H. M., Bauerfeind, I., Felber, M., Wang, H. J., Beryt, M., Seshadri, R., Hepp, H., Slamon, D. J. 2003; 95 (2): 142-153

  Abstract

  HER-2/neu, which encodes a receptor tyrosine kinase, is amplified and overexpressed in 20%-25% of human breast cancers. Such tumors are often resistant to hormone therapy. Despite a general inverse association between HER-2/neu amplification/overexpression and estrogen receptor (ER) and/or progesterone receptor (PR) expression, a fraction of patients are both HER-2/neu- and hormone receptor (HR)-positive. The efficacy of hormone therapy in this group is currently a matter of debate. To better understand the relationship between HER-2/neu positivity and HR expression, we analyzed HER-2/neu, ER, and PR as continuous variables in breast cancer cell lines and two cohorts of primary breast cancer patients.HER-2/neu and ER/PR expression was analyzed by enzyme-linked immunosorbent assay (ELISA) and enzyme immunoassay (EIA), respectively, in 14 human breast cancer cell lines, some of which had been transfected with the HER-2/neu gene. For the clinical study population, HER-2/neu protein levels were assessed by ELISA (cohort A, n = 665), and HER-2/neu gene copy number was determined using fluorescence in situ hybridization (cohort B, n = 894). ER/PR expression was analyzed by EIA (cohort A) or radioligand binding (cohort B). Associations between HER-2/neu and ER/PR expression were analyzed using Spearman's rho correlation and the chi-square test, and absolute levels were compared using the Mann-Whitney U test. All statistical tests were two-sided.HR-positive human breast cancer cell lines transfected with the HER-2/neu gene expressed statistically significantly lower levels of ER and PR than parental lines. In the clinical cohorts, levels of HER-2/neu overexpression and gene amplification were inversely correlated with ER/PR levels (Cohort A [n = 112]: for ER, r = -0.34, P<.001; for PR, r = -0.24, P =.010. Cohort B [n = 188]: for ER, r = -0.39, P<.001; for PR, r = -0.26, P<.001). Among patients with HR-positive tumors, HER-2/neu-positive tumors had statistically significantly lower ER/PR levels than HER-2/neu-negative ones (Cohort A: for ER, median = 25 fmol/mg [interquartile range [IQR] = 13-78] versus median = 38.5 fmol/mg [IQR = 17-99] and P =.031; for PR, median = 35 fmol/mg [IQR = 12-119] versus median = 88.5 fmol/mg [IQR = 22-236] and P<.001. Cohort B: for ER, median = 44 fmol/mg [IQR = 13-156] versus median = 92 fmol/mg [IQR = 35-235] and P<.001; for PR, median = 36 fmol/mg [IQR = 13-108] versus median = 84 fmol/mg [IQR = 24-250] and P<.001). Patients with higher levels of HER-2/neu overexpression or amplification had statistically significantly lower levels of ER/PR than patients with lower levels of HER-2/neu overexpression or amplification.Because absolute HR levels are strongly related to response to hormone therapy in primary and advanced breast cancer, reduced ER/PR expression may be one mechanism to explain the relative resistance of HER-2/neu-positive:HR-positive tumors to hormone therapy.

  View details for Web of Science ID 000180350800011

  View details for PubMedID 12529347

• Targeted prodrug treatment of HER-2-positive breast tumor cells using trastuzumab and paclitaxel linked by A-Z-CINN Linker. Journal of experimental therapeutics & oncology Gilbert, C. W., McGowan, E. B., Seery, G. B., Black, K. S., Pegram, M. D. 2003; 3 (1): 27-35

  Abstract

  Targeting drugs for delivery and release has the potential to increase the efficacy of treatment. A bifunctional linker, A-Z-CINN Linker was used to create a targeted prodrug, A-Z-CINN 310. A-Z-CINN Linker links to a potent chemotherapeutic agent, paclitaxel, via an energy-reversible ester bond and also binds a targeting agent, the monoclonal antibody trastuzumab (Herceptin). This study demonstrates the effectiveness of a single-treatment use of A-Z-CINN 310 in decreasing tumor volume and tumor cell density of human HER-2-positive BT-474 mammary tumor cells implanted in scid mice, compared to treatment with simultaneously administered trastuzumab and paclitaxel and with saline control. After treatment with A-Z-CINN 310, some mice received light exposure at 6 h for 5 min adjacent to the tumor to cause light-accelerated release of paclitaxel. Changes in tumor volume were measured for 28 days following treatment; changes in histology were measured at 31 days. Animals treated with A-Z-CINN 310, then light, showed dose-dependent decreases in tumor volume and tumor cell density which were more rapid and extensive than those seen with A-Z-CINN 310 without light or a 10-fold higher concentration of co-administered trastuzumab plus paclitaxel. This suggests that targeted delivery of paclitaxel using A-Z-CINN 310 kills tumor cells by localized release of paclitaxel at the tumor site, which can be accelerated by light treatment. These results indicate that a targeted prodrug therapy containing trastuzumab as the targeting agent and A-Z-CINN-paclitaxel as the prodrug results in a conjugate that is more effective in killing tumor cells than equivalent concentrations of co-administered trastuzumab and paclitaxel. Targeting of a drug can reduce the dose needed for effective therapy and can increase local bioavailability. This makes targeted therapy using an A-Z-CINN prodrug delivery system feasible for treating both primary and metastatic tumors.

  View details for PubMedID 12724856

• Platinum-taxane combinations in metastatic breast cancer: an evolving role in the era of molecularly targeted therapy BREAST CANCER RESEARCH AND TREATMENT Crown, J., Pegram, M. 2003; 79: S11-S18

  Abstract

  Metastatic breast cancer is a partially chemotherapy-sensitive neoplasm. Docetaxel appears to be the most active single agent for this condition. The platinum coordination complexes are also active, but are seldom used. An emerging literature suggests that taxane-platinum combinations have substantial activity. Trastuzumab, a molecularly targeted therapy for metastatic breast cancer is active as a single agent in HER2-overexpressing disease, and has been demonstrated to augment the activity of other agents in random assignment trials. Preclinical data suggest a powerful synergistic interaction between trastuzumab and both platinum and docetaxel. In early trials, platinum-taxane-trastuzumab combinations have exhibited promising clinical activity. The potential for cardiac toxicity when trastuzumab is combined with the anthracyclines suggests a further rationale for the development of non-anthracycline regimens, especially in the adjuvant setting. Randomized, multicenter, phase III trials in the metastatic and adjuvant setting are currently underway to test the hypothesis that synergistic combinations of docetaxel, platinums, and trastuzumab will result in superior safety as well as efficacy.

  View details for Web of Science ID 000184150400003

  View details for PubMedID 12868801

• A human ovarian carcinoma murine xenograft model useful for preclinical trials GYNECOLOGIC ONCOLOGY Elkas, J. C., Baldwin, R. L., Pegram, M., Tseng, Y., Slamon, D., Karlan, B. Y. 2002; 87 (2): 200-206

  Abstract

  To establish a murine xenograft model of human ovarian carcinoma.A slurry of fresh human tumor from patients with intraperitoneal malignancies was heterotransplanted intraperitoneally into nude (nu/nu) and severely combined immunodeficient mice (CB-17, SCID). Xenograft growth was assessed by serial examination and necropsy. The xenografts were passaged to new animals when tumors were palpably greater than 1 cm(3). Histopathologic analysis of the xenografts was performed at each passage as well as immunohistochemical staining for p53 mutations. Persistent expression of human genes by the xenografts at higher passages was assessed by RT-PCR amplification of the human beta-globin gene. This xenograft model was used in the preclinical evaluation of an adenoviral vector containing a beta-galactosidase reporter gene and a wild-type p53 gene.Tumor growth was not established in any of the nude mice heterotransplanted with tissue from six different ovarian cancer patients. Eleven of 13 specimens established xenograft growth when injected in SCID mice. Nine xenografts have been subsequently passaged between 6 and 24 animal generations to date. All xenografts retained histopathologic similarities to their original human tumors and the p53 expression patterns remained stable through higher passages. Within 24 h after intraperitoneal administration of an adenoviral vector, transduction of the reporter gene was evident in the xenografts. In addition, administration of an adenoviral vector containing a wild-type p53 gene significantly decreased the tumor burden compared to controls (P < 0.04).This murine xenograft model of human ovarian carcinoma appears to be reliable and reproducible and has utility for the study of novel therapeutics.

  View details for DOI 10.1006/gyno.2002.6819

  View details for Web of Science ID 000179129200008

  View details for PubMedID 12477452

• Long term follow-up of patients with recurrent ovarian cancer after Ad p53 gene replacement with SCH 58500 9th International Conference on Gene Therapy of Cancer Buller, R. E., Shahin, M. S., Horowitz, J. A., Runnebaum, I. B., Mahavni, V., Petrauskas, S., KREIENBERG, R., Karlan, B., Slamon, D., Pegram, M. NATURE PUBLISHING GROUP. 2002: 567?72

  Abstract

  We have previously reported the safety, efficient gene transfer, and favorable CA125 responses of individuals with recurrent ovarian cancer treated by p53 gene replacement with the adenoviral vector SCH 58500. The purpose of the present investigation was to evaluate the long-term follow-up of these heavily pretreated patients subsequent to SCH 58500 dosing.Patients (n=36) were treated with either single-dose SCH 58500 in the phase I study or with multiple doses (MD) of SCH 58500 over multiple cycles in combination of platinum-based chemotherapy in the phase I/II portion of the study. Five patients were initially treated in the single-dose group and re-enrolled in the MD group. The MD group was evaluated both without the re-enrolled patients as MD1 (n=19), and as MD2 (n=24), which included them. Patients who were only treated on the single-dose arm were designated as SD (n=12). Most patients received additional chemotherapy at the discretion of their physicians on completion of the trial. The current analysis is a retrospective sequential cohort survival analysis.The first patient was treated in March 1997 and the last patient completed SCH 58500 in September 1998. There was no difference in age at diagnosis, Karnofsky performance status, interval between diagnosis to SCH 58500, prior cycles or regimen of chemotherapy, platinum-free interval, percent platinum refractory patients, pretreatment CA125, or largest tumor volume between groups. Both MD groups had a slightly longer chemotherapy-free interval before SCH 58500 than the SD group. Median survival of individuals who received MD SCH 58500 with chemotherapy was 12-13.0 months, compared to only 5 months for those treated with SD SCH 58500. There are 10 long-term survivors more than 20 months after MD treatment for recurrent disease compared to only 2 long-term survivors after SD SCH 58500.The 12- to 13.0-month median survival in a heavily pretreated population with recurrent ovarian cancer compares favorably to the 16-month median survival for individuals treated with paclitaxel at the time of initial recurrence of this disease and is more than double the 5-month survival seen with palliative radiotherapy or paclitaxel failure. These data suggest that further study of SCH58500 is clearly indicated.

  View details for DOI 10.1037/sj.cgt.7700473

  View details for Web of Science ID 000176492400002

  View details for PubMedID 12082456

• A phase I/II trial of rAd/p53 (SCH 58500) gene replacement in recurrent ovarian cancer 30th Annual Meeting of the Society-of-Gynecologic-Oncologists Buller, R. E., Runnebaum, I. B., Karlan, B. Y., Horowitz, J. A., Shahin, M., Buekers, T., Petrauskas, S., KREIENBERG, R., Slamon, D., Pegram, M. NATURE PUBLISHING GROUP. 2002: 553?66

  Abstract

  To determine the safety, gene transfer, host immune response, and pharmacokinetics of a replication-deficient adenovirus encoding human, recombinant, wild-type p53 (SCH 58500) delivered into the peritoneal cavity (i.p.) alone and sequentially in combination with platinum-based chemotherapy, of patients with recurrent ovarian, primary peritoneal, or fallopian tube cancer containing aberrant or mutant p53.SCH 58500 was administered i.p. to three groups of patients with heavily pretreated recurrent disease. Group 1 (n=17) received a single dose of SCH 58500 escalated from 7.5 x 10(10) to 7.5 x 10(12) particles. Group 2 (n=9) received two or three doses of SCH 58500 given alone for one cycle, and then with chemotherapy for two cycles. The SCH 58500 dose was further escalated to 2.5 x 10(13) particles/dose in group 2. A third group (n=15) received a 5-day regimen of SCH 58500 given at 7.5 x 10(13) particles/dose per day i.p. alone for cycle 1 and then with intravenous carboplatin/paclitaxel chemotherapy for cycles 2 and 3.No dose-limiting toxicity resulted from the delivery of 236/287 (82.2%) planned doses of SCH 58500. Fever, hypotension abdominal complaints, nausea, and vomiting were the most common adverse events. Vector-specific transgene expression in tumor was documented by RT-PCR in cells from both ascitic fluid and tissue biopsies. Despite marked increases in serum adenoviral antibody titers, transgene expression was measurable in 17 of 20 samples obtained after two or three cycles of SCH 58500. Vector was detectable in peritoneal fluid by 24 hours and persisted for as long as 7 days whereas none was detected in urine or stool. There was poor correlation between CT scans and CA125 responses. CA125 responses, defined as a greater than 50% decrement in serum CA125 from baseline, were documented in 8 of 16 women who completed three cycles of the multidose regimen.CT scans are not a valid measure of response to i.p. SCH 58500 due to extensive adenoviral-induced inflammatory changes. Intraperitoneal SCH 58500 is safe, well tolerated, and combined with platinum-based chemotherapy can be associated with a significant reduction of serum CA125 in heavily pretreated patients with recurrent ovarian, primary peritoneal, or fallopian tube cancer.

  View details for DOI 10.1038/sj.cgt.7700472

  View details for Web of Science ID 000176492400001

  View details for PubMedID 12082455

• Combined biological therapy of breast cancer using monoclonal antibodies directed against HER2/neu protein and vascular endothelial growth factor SEMINARS IN ONCOLOGY Pegram, M. D., Reese, D. M. 2002; 29 (3): 29-37

  Abstract

  Vascular endothelial growth factor (VEGF) is one of the most important endothelial mitogens involved in the development and differentiation of the vascular system. Vascular endothelial growth factor is a highly conserved, homodimeric glycoprotein with multiple isoforms. The most abundant isoform, VEGF165, binds to vascular endothelial growth factor receptors-1 (Flt-1) and 2 (KDR/Flk-1) with picomolar affinity. Recently, correlation between microvessel density and engineered expression of VEGF in human breast xenografts was observed. A role of VEGF in breast cancer progression is evident from clinical studies showing elevated serum VEGF in invasive breast cancers. Vascular endothelial growth factor in breast tumor cytosols is correlated with microvessel density, and VEGF165 content correlates with disease-free and overall survival in primary breast cancers. Preliminary data indicate a transcriptional upregulation of VEGF in HER2-overexpressing breast cancer cells. We hypothesize that the upregulation of VEGF in HER2-overexpressing breast cancers contributes to the aggressive phenotype observed in HER2-positive cases and that the "angiogenic switch" associated with HER2 can be attenuated by trastuzumab. Although tumor angiogenesis in breast cancer is complex, the VEGF/vascular endothelial growth factor receptor (VEGFR) system provides a useful model for testing new angiogenesis inhibitors that target this pathway. The VEGF/VEGFR system provides a number of opportunities for therapeutic intervention in breast cancer. Understanding the biology of this system is paramount to fully exploiting VEGF as a therapeutic target in breast cancer. We hypothesize that new therapeutic molecules targeting VEGF and/or its receptors, such as recombinant humanized monoclonal anti-VEGF antibody (rhuMAb VEGF), may have unique activity against HER2-overexpressing breast cancers.

  View details for DOI 10.1053/sonc.2002.34053

  View details for Web of Science ID 000177095800005

  View details for PubMedID 12138395

• Inhibition of cell growth by NB1011 requires high thymidylate synthase levels and correlates with p53, p21, bax, and GADD45 induction MOLECULAR CANCER THERAPEUTICS Neuteboom, S. T., Karjian, P. L., Boyer, C. R., Beryt, M., Pegram, M., Wahl, G. M., SHEPARD, H. M. 2002; 1 (6): 377-384

  Abstract

  NB1011, a phosphoramidate derivative of (E)-5-(2-bromovinyl)-2'-deoxyuridine, is a novel small molecule anticancer agent. NB1011 is selectively active against tumor cells expressing high levels of thymidylate synthase (TS), a critical enzyme in DNA biosynthesis. NB1011 is different from the current TS-targeted drugs, which require inhibition of TS to be effective, because NB1011 cytotoxicity depends upon activation by TS. Here we report a dose-dependent, antitumor activity of NB1011 against established Tomudex-resistant breast cancer (MCF7TDX) xenografts in athymic mice. Against 5-fluorouracil-resistant colon carcinoma (H630R10) xenografts, NB1011 was as efficacious as irinotecan, a drug recently approved for the treatment of 5-fluorouracil-resistant colon cancer. To gain insight into the mechanisms NB1011 uses to suppress cellular growth, we analyzed the downstream molecular events in the high TS-expressing MCF7TDX and RKOTDX cell lines upon NB1011 treatment. NB1011 treatment increased the mRNA levels of p21, Bax, and GADD45. Furthermore, NB1011 induced p53, p21, and Bax proteins specifically in high TS-expressing tumor cells, whereas no induction was observed in low TS-expressing tumor cells (MCF7) or normal cells (WI38). Cell cycle analysis demonstrated that NB1011 treatment of MCF7TDX and RKOTDX cells resulted in an accumulation of cells in the G2-M phase of the cell cycle. Altogether, our data indicate that the induction of the p53 target genes p21, bax, and GADD45, with a concomitant deregulation of the cell cycle, may represent one of the mechanisms by which NB1011 exerts its growth-suppressive effects.

  View details for Web of Science ID 000178770300003

  View details for PubMedID 12477050

• Combining the anti-HER2 antibody trastuzumab with taxanes in breast cancer: results and trial considerations. Clinical breast cancer Pegram, M. D., O'Callaghan, C. 2001; 2: S15-9

  Abstract

  Overexpression of the p185/HER2 protein is seen in 20%-25% of primary breast cancers and is associated with poor prognosis. Recent phase II and III clinical trials demonstrate that trastuzumab is active against breast tumors, both as a single agent and in combination with chemotherapy. In patients with HER2-overexpressing metastatic breast cancer, use of trastuzumab in combination with chemotherapy is associated with a 20% reduction in relative risk of death and an increase in median survival from 20.3 to 25.1 months compared to chemotherapy alone. Side effects include fever and chills and an unexpected increase in doxorubicin/trastuzumab-associated cardiomyopathy. Clinical development is now focused on trastuzumab in combination with chemotherapy regimens that do not contain an anthracycline. Trastuzumab in combination with docetaxel is synergistic in vitro. Data from ongoing clinical trials are consistent with this finding. Preliminary data from 3 phase II studies suggest a 44%-63% response rate when the combination is used first or second line in HER2-overexpressing metastatic breast cancer. The combination of docetaxel with trastuzumab is well tolerated and has not been associated with significant cardiotoxicity. Given in vitro evidence that platinum salts act synergistically with trastuzumab and docetaxel, and phase II data suggesting clinical efficacy and good tolerability, the combination of platinum salt plus trastuzumab and docetaxel is now being assessed in adjuvant trials

  View details for PubMedID 11970740

• HER-2/neu and urokinase-type plasminogen activator and its inhibitor in breast cancer CLINICAL CANCER RESEARCH Konecny, G., Untch, M., Arboleda, J., Wilson, C., Kahlert, S., Boettcher, B., Felber, M., Beryt, M., Lude, S., Hepp, H., Slamon, D., Pegram, M. 2001; 7 (8): 2448-2457

  Abstract

  Recent studies suggest that HER-2/neu specifically promotes the invasive capacity of tumor cells by up-regulating secretion of the proteolytic enzyme, urokinase-type plasminogen activator (uPA), or its inhibitor, plasminogen activator inhibitor-1 (PAI-1), in colon and gastric cancer. It was the purpose of this study to: (a) evaluate the association between HER-2/neu and uPA and PAI-1 expression in a large primary breast cancer cohort; (b) perform the first multivariate analysis, including HER-2/neu, uPA, and PAI-1 in breast cancer; and (c) define the effect of HER-2/neu overexpression on uPA and PAI-1 expression in breast cancer cells.HER-2/neu, uPA, and PAI-1 were measured as continuous variables by ELISA in primary breast cancer tissue extracts from 587 patients with clinical follow-up and analyzed for correlations with clinical outcome. Furthermore, a full-length human HER-2/neu cDNA was introduced into five human breast cancer cell lines to define the effects of HER-2/neu overexpression on uPA and PAI-1 expression. In addition, we tested whether HER-2/neu antibodies could reverse any given alteration of uPA and PAI-1 levels.Our findings indicate a weak positive association between HER-2/neu and uPA (r = 0.147; P < 0.001) and no association between HER-2/neu and PAI-1 (r = 0.07; P = 0.085). HER-2/neu overexpression (> or =400 fmol/mg) and high levels of uPA/PAI-1 (> or =5.5 ng/mg and/or > or =14 ng/mg, respectively) were significantly associated with shorter disease-free survival (DFS; P < 0.001 and P = 0.003) and metastasis-free survival (MFS; P = 0.015 and P < 0.001). Multivariate analysis revealed prognostic independence between HER-2/neu and the uPA/PAI-1 axis for DFS and MFS. Both uPA and PAI-1 had no significant discriminatory effect among HER-2/neu-positive patients for DFS. The prognostic value of HER-2/neu overexpression for MFS, however, was significantly enhanced by elevated uPA expression (P = 0.053). Stable transfection of the HER-2/neu gene into multiple human breast cancer cell lines resulted in consistent down-regulation of uPA or PAI-1 expression. In addition, anti-HER-2/neu antibodies did not significantly affect uPA or PAI-1 expression in human cancer cell lines naturally overexpressing HER-2/neu.The present findings suggest that the invasive phenotype elicited by HER-2/neu overexpression in breast cancer is not a direct effect of uPA or PAI-1 expression. HER-2/neu and the uPA/PAI-1 axis have been shown to affect the invasive capacity of breast cancer independently. Determination of uPA can provide significant additional prognostic information for MFS in HER-2/neu-positive and -negative patients.

  View details for Web of Science ID 000170328300038

  View details for PubMedID 11489825

• Drug interactions and cytotoxic effects of paclitaxel in combination with carboplatin, epirubicin, gemcitabine or vinorelbine in breast cancer cell lines and tumor samples BREAST CANCER RESEARCH AND TREATMENT Konecny, G., Untch, M., Slamon, D., Beryt, M., Kahlert, S., Felber, M., LANGER, E., Lude, S., Hepp, H., Pegram, M. 2001; 67 (3): 223-233

  Abstract

  The purpose of this study was to analyze the drug interactions of paclitaxel (PTX) with epirubicin (EPI), carboplatin (CBDCA), gemcitabine (GEM) and vinorelbine (VIN) in human breast cancer cells and compare the cytotoxic activity of each drug combination in primary breast cancer samples. These experiments were intended to identify the most active agents in combination with PTX, and to provide a preclinical rational for future clinical investigations in breast cancer. Multiple drug effect/combination index (CI) isobologram analysis was applied to combinations of PTX with either CBDCA, EPI, GEM or VIN in MCF-7, MDA-MB-231 and SK-BR-3 human breast cancer cell lines. Drug concentrations were limited to the ranges achievable in humans in vivo, and the drugs were applied simultaneously at fixed molar ratios for each drug combination. Interactions were assessed at multiple effect levels (IC10-IC90). Additionally, the cytotoxic activity of these combinations was assessed in tumor samples of 50 primary breast cancer patients, utilizing the ATP-tumorchemosensitivity assay (ATP-TCA). Drug interactions were shown to be strongly dose-related in the human breast cancer cell lines investigated. At clinically relevant concentrations, CBDCA/PTX demonstrated synergistic (MCF-7) or additive (MDA-MB-231, SK-BR-3) interactions, and EPI/PTX showed additive (SK-BR-3, MCF-7) and antagonistic (MDA-MB-231) interactions. GEM/PTX and VIN/PTX, however, demonstrated antagonism over multiple dose effect levels at clinically relevant drug concentrations in all three cell lines tested. At plasma peak concentrations, EPI/PTX, CBDCA/PTX, GEM/PTX and VIN/PTX achieved > or = 90% tumor growth inhibition in 93, 86, 63 and 50%, respectively, of primary breast cancer samples investigated with the ATP-TCA. Cumulative dose-response plots of primary breast cancer tumor cells responding in vitro with > or = 90% growth inhibition showed a strong dose dependence for both EPI/PTX and CBDCA/PTX. In conclusion, the current data indicate favorable drug interactions for CBDCA/PTX at clinically relevant drug concentrations in breast cancer cells, and demonstrate superior in vitro cytotoxicity of EPI/PTX and CBDCA/PTX compared to GEM/PTX and VIN/PTX in primary breast cancer cultures.

  View details for Web of Science ID 000170903800003

  View details for PubMedID 11561768

• Association of urokinase-type plasminogen activator and its inhibitor with disease progression and prognosis in ovarian cancer CLINICAL CANCER RESEARCH Konecny, G., Untch, M., Pihan, A., Kimmig, R., Gropp, M., Stieber, P., Hepp, H., Slamon, D., Pegram, M. 2001; 7 (6): 1743-1749

  Abstract

  Urokinase-type plasminogen activator (uPA) and its inhibitor, plasminogen activator inhibitor (PAI)-1, have been shown to be related to poor prognosis in a variety of malignant solid tumors. Studies on the prognostic relevance of uPA and PAI-1 in ovarian cancer, however, have been inconclusive. The current study tests the hypothesis that elevated expression of uPA and PAI-1 is associated with prognosis and disease progression.uPA and PAI-1 were prospectively measured by quantitative ELISA in tumor samples from 103 ovarian cancer patients (82 primary invasive epithelial carcinomas, 9 low malignant potential tumors, and 12 recurrent ovarian carcinomas).uPA but not PAI-1 levels were consistently associated with malignant progression, with levels increased from low malignant potential tumors to primary tumors (uPA, P = 0.04; PAI-1, P = 0.019), from early to advanced disease stages (uPA, P = 0.014; PAI-1, P = 0.23), and from primary to intra-abdominal metastatic tumors (uPA, P = 0.001; PAI-1, P = 0.16). High uPA and PAI-1 levels were associated with residual tumor volumes of >1 cm (P = 0.001 and P = 0.016, respectively). Among invasive International Federation of Gynecologists and Obstetrician stages I-IV tumors, elevated levels of uPA (>5.5 ng/mg) and PAI-I (>18.8 ng/ml) were associated with a shortened progression-free survival (uPA, P = 0.003; PAI-1, P = 0.039) and overall survival (uPA, P = 0.0002; PAI-1, P = 0.007). In multivariate analysis, uPA retained prognostic independence for progression-free survival (P = 0.037) and overall survival (P = 0.006).These data suggest that the uPA/PAI-1 axis may play an important role in the intra-abdominal spread and reimplantation of ovarian cancer cells. The prognostic relevance of uPA and PAI-1 supports their possible role in the malignant progression of ovarian cancer.

  View details for Web of Science ID 000169310600036

  View details for PubMedID 11410515

• Use of chemotherapy plus a monoclonal antibody against HER2 for metastatic breast cancer that overexpresses HER2. NEW ENGLAND JOURNAL OF MEDICINE Slamon, D. J., Leyland-Jones, B., Shak, S., Fuchs, H., Paton, V., Bajamonde, A., Fleming, T., Eiermann, W., Wolter, J., Pegram, M., Baselga, J., Norton, L. 2001; 344 (11): 783-792

  Abstract

  The HER2 gene, which encodes the growth factor receptor HER2, is amplified and HER2 is overexpressed in 25 to 30 percent of breast cancers, increasing the aggressiveness of the tumor.We evaluated the efficacy and safety of trastuzumab, a recombinant monoclonal antibody against HER2, in women with metastatic breast cancer that overexpressed HER2. We randomly assigned 234 patients to receive standard chemotherapy alone and 235 patients to receive standard chemotherapy plus trastuzumab. Patients who had not previously received adjuvant (postoperative) therapy with an anthracycline were treated with doxorubicin (or epirubicin in the case of 36 women) and cyclophosphamide alone (138 women) or with trastuzumab (143 women). Patients who had previously received adjuvant anthracycline were treated with paclitaxel alone (96 women) or paclitaxel with trastuzumab (92 women).The addition of trastuzumab to chemotherapy was associated with a longer time to disease progression (median, 7.4 vs. 4.6 months; P<0.001), a higher rate of objective response (50 percent vs. 32 percent, P<0.001), a longer duration of response (median, 9.1 vs. 6.1 months; P<0.001), a lower rate of death at 1 year (22 percent vs. 33 percent, P=0.008), longer survival (median survival, 25.1 vs. 20.3 months; P=0.01), and a 20 percent reduction in the risk of death. The most important adverse event was cardiac dysfunction of New York Heart Association class III or IV, which occurred in 27 percent of the group given an anthracycline, cyclophosphamide, and trastuzumab; 8 percent of the group given an anthracycline and cyclophosphamide alone; 13 percent of the group given paclitaxel and trastuzumab; and 1 percent of the group given paclitaxel alone. Although the cardiotoxicity was potentially severe and, in some cases, life-threatening, the symptoms generally improved with standard medical management.Trastuzumab increases the clinical benefit of first-line chemotherapy in metastatic breast cancer that overexpresses HER2.

  View details for Web of Science ID 000167440400001

  View details for PubMedID 11248153

• Rationale for trastuzumab (Herceptin) in adjuvant breast cancer trials SEMINARS IN ONCOLOGY Slamon, D., Pegram, M. 2001; 28 (1): 13-19

  Abstract

  The discovery of the HER2/neu proto-oncogene and its role in the pathogenesis of breast cancer tumors, and the development of the anti-HER2 monoclonal antibody, trastuzumab (Herceptin; Genentech, South San Francisco, CA), directed against the HER2 receptor represent major milestones in the research developments in breast cancer, making trastuzumab the first monoclonal antibody available for treatment of this disease. Clinical trials in HER2-positive patients have demonstrated that the combined use of targeted therapy with trastuzumab in conjunction with cytotoxic chemotherapy is associated with improved time to disease progression and overall survival. Unfortunately, findings also demonstrate an increased risk for cardiotoxicity when trastuzumab is combined with anthracyclines. For HER2/neu-overexpressing breast cancer patients, the adjuvant use of trastuzumab will become paramount; therefore, it must be evaluated in a randomized controlled trial. There is disagreement regarding the design of such a trial, largely because of the ubiquitous use of anthracyclines in the adjuvant setting and the opposing necessity of avoiding anthracycline plus trastuzumab combinations. Combination index values for various chemotherapeutic drugs in combination with trastuzumab demonstrate dramatic synergistic interactions with the platinum agents and with docetaxel (Taxotere; Aventis Pharmaceuticals, Inc, Parsippany, NJ). The greatest level of synergy has been demonstrated with the triple-drug combination of docetaxel, platinum, and trastuzumab in which synergy is demonstrated, even at low doses. The adjuvant trial design for the Breast Cancer International Research Group uses a control arm of doxorubicin/cyclophosphamide for four cycles followed by docetaxel for four cycles and the second arm contains the addition of trastuzumab to the taxane sequence. The third arm, a non-anthracycline-containing regimen, contains docetaxel, a platinum agent (either cisplatin or carboplatin), and trastuzumab. The rationale for the selection of this three-drug regimen is based on the biology of the system and preclinical and clinical findings that demonstrate a high potential for clinical synergy.

  View details for DOI 10.1053/sonc.2001.22812

  View details for Web of Science ID 000167891900002

  View details for PubMedID 11301370

• Enzyme-catalyzed therapeutic agent (ECTA) design: activation of the antitumor ECTA compound NB1011 by thymidylate synthase NCI-EORTC Symposium on Chemotherapeutic Strategies for Treatment of Colorectal Cancer: Present and Future Developments Lackey, D. B., Groziak, M. P., Sergeeva, M., Beryt, M., Boyer, C., Stroud, R. M., Sayre, P., Park, J. W., Johnston, P., Slamon, D., SHEPARD, H. M., Pegram, M. PERGAMON-ELSEVIER SCIENCE LTD. 2001: 179?89

  Abstract

  The in vivo administration of enzyme-inhibiting drugs for cancer and infectious disease often results in overexpression of the targeted enzyme. We have developed an enzyme-catalyzed therapeutic agent (ECTA) approach in which an enzyme overexpressed within the resistant cells is recruited as an intracellular catalyst for converting a relatively non-toxic substrate to a toxic product. We have investigated the potential of the ECTA approach to circumvent the thymidylate synthase (TS) overexpression-based resistance of tumor cells to conventional fluoropyrimidine [i.e. 5-fluorouracil (5-FU)] cancer chemotherapy. (E)-5-(2-Bromovinyl)-2'-deoxy-5'-uridyl phenyl L-methoxyalaninylphosphoramidate (NB1011) is a pronucleotide analogue of (E)-5-(2-bromovinyl)-2'-deoxyuridine (BVdU), an antiviral agent known to be a substrate for TS when in the 5'-monophosphorylated form. NB1011 was synthesized and found to be at least 10-fold more cytotoxic to 5-FU-resistant, TS-overexpressing colorectal tumor cells than to normal cells. This finding demonstrates that the ECTA approach to the design of novel chemotherapeutics results in compounds that are selectively cytotoxic to tumor cell lines that overexpress the target enzyme, TS, and therefore may be useful in the treatment of fluoropyrimidine-resistant cancer.

  View details for Web of Science ID 000166498300006

  View details for PubMedID 11163332

• Docetaxel and Herceptin: Foundation for future strategies Symposium on Innovative Therapies in Breast Cancer Pegram, M. D. ALPHAMED PRESS. 2001: 22?25

  Abstract

  Randomized controlled studies have demonstrated that both docetaxel and Herceptin are capable of increasing survival in patients with metastatic breast cancer. The two agents show synergy in vitro, and their use in combination is not likely to be associated with the problem of enhanced cardiotoxicity. In two trials of Herceptin plus docetaxel in patients with advanced breast cancer, preliminary data are available for 35 patients. These early results show that the combination is well-tolerated. No symptomatic cardiotoxicity has occurred. The preliminary response rates (RR) in these first- and second-line patients are 44% in one study and 63% in the other. In the subgroups of patients who were HER-2 3+ overexpressers, the RRs are currently 55% and 73%. In an attempt to maximize the efficacy of Herceptin, its use has also been studied in combination with docetaxel and a platinum salt, producing a preliminary RR of 78% in patients positive for HER-2 on the fluorescence in situ hybridization assay. These data are sufficiently promising to justify a study of the role of Herceptin in combination with adjuvant chemotherapy regimens containing docetaxel or docetaxel plus a platinum. The combination of Herceptin with adjuvant therapy containing docetaxel and a platinum may provide a helpful alternative to the potentially cardiotoxic Herceptin/anthracycline-containing regimens currently under investigation.

  View details for Web of Science ID 000175117300005

  View details for PubMedID 11346681

• HER-2/neu overexpression and in vitro chemosensitivity to CMF and FEC in primary breast cancer BREAST CANCER RESEARCH AND TREATMENT Konecny, G., FRITZ, M., Untch, M., Lebeau, A., Felber, M., Lude, S., Beryt, M., Hepp, H., Slamon, D., Pegram, M. 2001; 69 (1): 53-63

  Abstract

  Available clinical and experimental data on the effect of HER-2/neu overexpression on chemosensitivity are controversial. It was the purpose of this in vitro study to define the association between HER-2/neu overexpression and the sensitivity to the chemotherapeutic drug combinations of cyclophosphamide, methotrexate and 5-fluorouracil (CMF) and 5-fluorouracil, epirubicin and cyclophosphamide (FEC) of breast cancer cells derived from 140 chemotherapy-naïve patients at the time of primary surgery. Both drug combinations were tested at six different concentrations ranging from 6.25-200% peak plasma concentration (PPC). Immunohistochemical detection of HER-2/neu overexpression was performed with the HER-2/neu antibodies, CB11, TAB250 and AO485, in the same tumor specimens. Immunoreactions were determined as negative (0/1+), weakly positive (2+) and strongly positive (3+). However, the antibodies varied in their degrees of sensitivity. Breast cancer samples with strong (3+) HER-2/neu overexpression demonstrated 90% growth inhibition (IC90) at significantly lower PPC values, using the CB11 (p = 0.048), TAB250 (p = 0.007) and AO485 (p < or =0.01) antibodies, and showed 50% growth inhibition (IC50) at significantly lower PPC values, using the CB11 antibody (p = 0.01) compared to their counterparts with lower levels of HER-2/neu expression. When analyzing the group of patients with intermediate and strong HER-2/neu overexpression (2+ and 3+), an association between HER-2/neu overexpression and increased chemosensitivity was seen with the TAB250 (p = 0.044) and AO485 (p = 0.032) antibodies, but not with the CB11 antibody (p =0.8) at the IC90 level. Differences in chemosensitivity between samples with strong HER-2/neu overexpression and those with lower levels were then analyzed separately for CMF and FEC. Both regimens achieved 90% tumor growth inhibition at lower PPC values in samples with strong HER-2/neu overexpression (3+) compared to their counterparts with lower expression levels (AO485 p = 0.011 for CMF, and p = 0.09 for FEC). Cumulative concentration-response plots of tumors responding in vitro with 90% tumor cell inhibition showed a stronger dose dependence for both CMF and FEC among tumor samples with strong HER-2/neu overexpression compared to those with lower levels of expression. In conclusion, the data show that HER-2/neu overexpression was not associated with in vitro drug resistance to CMF or FEC. In contrast, tumors with strong HER-2/neu overexpression demonstrated increased dose-dependent in vitro sensitivity to both the FEC and CMF regimens.

  View details for Web of Science ID 000171802100006

  View details for PubMedID 11759828

• Trastuzumab and chemotherapeutics: Drug interactions and synergies SEMINARS IN ONCOLOGY Pegram, M. D., Lopez, A., Konecny, G., Slamon, D. J. 2000; 27 (6): 21-25

  Abstract

  Previous studies have shown a synergistic interaction between trastuzumab (Herceptin; Genentech, Inc, South San Francisco, CA) and the cytotoxic drug cisplatin in human breast cancer cells. To define the nature of the interaction between trastuzumab and other classes of cytotoxic drugs, we applied multiple drug effect/combination index isobologram analysis to a variety of chemotherapeutic drug/trastuzumab combinations in vitro. Synergistic interactions at clinically relevant drug concentrations were observed for trastuzumab in combination with cisplatin, docetaxel, thiotepa, 4-OH cyclophosphamide, vinorelbine, and etoposide. Additive cytotoxic effects were observed with trastuzumab plus doxorubicin, paclitaxel, methotrexate, and vinblastine. One drug, 5-fluorouracil was found to be antagonistic with trastuzumab in vitro. In vivo drug/trastuzumab studies were conducted with HER-2/neu-transfected MCF7 human breast cancer xenografts in athymic mice. Combinations of trastuzumab plus cisplatin, docetaxel, cyclophosphamide, doxorubicin, paclitaxel, methotrexate, etoposide, and vinblastine in vivo resulted in a significant reduction in xenograft volume compared to chemotherapy-alone controls (P < .05). The synergistic interaction of trastuzumab with specific chemotherapeutic agents suggests rational combinations for testing in human clinical trials.

  View details for Web of Science ID 000166958600005

  View details for PubMedID 11236023

• HER-2/neu as a predictive marker in breast cancer GEBURTSHILFE UND FRAUENHEILKUNDE Konecny, G., Pegram, M., Untch, M., Thomssen, C., JANICKE, F., Hepp, H., Slamon, D. J. 2000; 60 (12): 609-619

  View details for Web of Science ID 000166161900004

• Biological rationale for HER2/neu (c-erbB2) as a target for monoclonal antibody therapy SEMINARS IN ONCOLOGY Pegram, M., Slamon, D. 2000; 27 (5): 13-19

  Abstract

  The physical characteristics of tumor antigens that would make the most ideal targets for antibody therapeutics include cell surface expression; high, stable expression levels in tumor cells; low or absent expression in normal tissues; lack of a soluble form of the antigenic target; and lack of internalization of the antigen/antibody complex. HER2/neu is a 185-kd surface membrane protein that is overexpressed in approximately 25% of human breast cancers due to amplification of the HER2 gene. Overexpression of the gene results in ligand-independent activation of HER2 kinase, causing mitogenic signal transduction and increased cell proliferation. Consequently, patients with this alteration have a worse clinical prognosis. Trastuzumab (Herceptin; Genentech, Inc, So. San Francisco, CA), a humanized anti-HER2 monoclonal antibody, has significant clinical activity against metastatic breast cancers with HER2/neu overexpression, despite the fact that the p185HER2 protein product lacks some of the ideal characteristics of tumor antigens listed above. We propose that the efficacy of trastuzumab may be explained on the basis of its effects on signal transduction, which is independent from its immune mechanism(s) of action. Furthermore, trastuzumab is synergistic with some chemotherapeutic drugs, resulting in improved therapeutic efficacy. Thus, in the case of trastuzumab, a clear distinction may be drawn between the use of monoclonal antibodies as immuneactive agents and their use to achieve a desired cellular/biochemical activity.

  View details for Web of Science ID 000089873600003

  View details for PubMedID 11049052

• Docetaxel and trastuzumab: A combination with clinical relevance Symposium on a New Era in Oncology Konecny, G., Pegram, M., Slamon, D. KARGER. 2000: 7?9

  View details for Web of Science ID 000089400600004

• Correlation of drug response with the ATP tumorchemosensitivity assay in primary FIGO stage III ovarian cancer GYNECOLOGIC ONCOLOGY Konecny, G., Crohns, C., Pegram, M., Felber, M., Lude, S., Kurbacher, C., Cree, I. A., Hepp, H., Untch, M. 2000; 77 (2): 258-263

  Abstract

  Our purpose was to: (a) study the in vitro chemosensitivity of primary epithelial ovarian cancer to drug combinations with cisplatin (CDDP), carboplatin (CBDCA), paclitaxel (PTX), epirubicin (EPI), or cyclophosphamide (CTX) utilizing the ATP tumorchemosensitivity assay (ATP-TCA); (b) correlate the test results with clinical response in patients with FIGO stage III ovarian cancer; and (c) analyze the most useful parameters for interpretation of test results.CBDCA/CTX, CBDCA/PTX, CDDP/PTX, and EPI/PTX were tested in 93 fresh human primary epithelial ovarian cancer specimens. Correlations of in vitro drug sensitivity/resistance and clinical response were performed in 38 patients with FIGO stage III disease utilizing Fisher's exact test and by comparison of progression-free (PFS) and overall survival (OS) between those testing as sensitive or resistant. A progression-free interval of more than 12 months following surgery was classified as clinical response. ATP-TCA results were analyzed using the median effective dose, area under the curve, or a defined sensitivity index.Evaluable test results were achieved in 83 of 93 patients (89%). EPI/PTX had the highest in vitro activity (P < 0.001). In the clinical correlation, 29 of 38 patients (76%) were classified as in vitro sensitive (sensitivity index [SI] <250) and 9 patients as in vitro resistant (SI >250). The SI was superior for interpretation of test results. Patients testing as chemosensitive had a significantly longer mean PFS (28.5 vs 12.6 months, P = 0.033) and OS (46.1 vs 17.6, P = 0.03) compared to those patients predicted to be resistant. The assay demonstrated a sensitivity, specificity, and positive and negative predictive value of 95, 44, 66, and 89%, respectively (Fisher's exact test, P = 0. 007).The observed in vitro efficacy of EPI/PTX in primary epithelial ovarian cancer specimens warrants further clinical evaluation. The high evaluability rate and the observed correlation with PFS and OS, within the limitations of a nonrandomized study, support the use of the ATP chemosensitivity assay in future prospective assay-directed trials.

  View details for Web of Science ID 000087028300008

  View details for PubMedID 10785475

• The molecular and cellular biology of HER2/neu gene amplification/overexpression and the clinical development of herceptin (trastuzumab) therapy for breast cancer. Cancer treatment and research Pegram, M. D., Konecny, G., Slamon, D. J. 2000; 103: 57-75

  View details for PubMedID 10948442

• A model-based approach for assessing in vivo combination therapy interactions PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Lopez, A. M., Pegram, M. D., Slamon, D. J., Landaw, E. M. 1999; 96 (23): 13023-13028

  Abstract

  We present an approach for evaluating the efficacy of combination antitumor agent schedules that accounts for order and timing of drug administration. Our model-based approach compares in vivo tumor volume data over a time course and offers a quantitative definition for additivity of drug effects, relative to which synergism and antagonism are interpreted. We begin by fitting data from individual mice receiving at most one drug to a differential equation tumor growth/drug effect model and combine individual parameter estimates to obtain population statistics. Using two null hypotheses: (i) combination therapy is consistent with additivity or (ii) combination therapy is equivalent to treating with the more effective single agent alone, we compute predicted tumor growth trajectories and their distribution for combination treated animals. We illustrate this approach by comparing entire observed and expected tumor volume trajectories for a data set in which HER-2/neu-overexpressing MCF-7 human breast cancer xenografts are treated with a humanized, anti-HER-2 monoclonal antibody (rhuMAb HER-2), doxorubicin, or one of five proposed combination therapy schedules.

  View details for Web of Science ID 000083649400011

  View details for PubMedID 10557266

• Biologic effects of heregulin/neu differentiation factor on normal and malignant human breast and ovarian epithelial cells ONCOGENE Aguilar, Z., Akita, R. W., Finn, R. S., Ramos, B. L., Pegram, M. D., Kabbinavar, F. F., Pietras, R. J., Pisacane, P., Sliwkowski, M. X., Slamon, D. J. 1999; 18 (44): 6050-6062

  Abstract

  The heregulins are a family of ligands with ability to induce phosphorylation of the p185HER-2/neu receptor. Various investigators have reported a variety of responses of mouse and human breast and ovarian cells to this family of ligands including growth stimulation, growth inhibition, apoptosis and induction of differentiation in cells expressing the HER-2/neu receptor. Some of the disparity in the literature has been attributed to variations in the cell lines studied, ligand dose applied, methodologies utilized or model system evaluated (i.e. in vitro or in vivo). To evaluate the effects of heregulin on normal and malignant human breast and ovarian epithelial cells expressing known levels of the HER-2/neu receptor, this report presents the use of several different assays, performed both in vitro and in vivo, in vitro proliferation assays, direct cell counts, clonogenicity under anchorage-dependent and anchorage-independent conditions, as well as the in vivo effects of heregulin on human cells growing in nude mice to address heregulin activity. Using a total of five different biologic assays in nine different cell lines, across two different epithelia and over a one log heregulin dose range, we obtained results that clearly indicate a growth-stimulatory role for this ligand in human breast and ovarian epithelial cells. We find no evidence that heregulin has any growth-inhibitory effects in human epithelial cells. We also quantitated the amount of each member of the type I receptor tyrosine kinase family (RTK I, i.e. HER-1, HER-2, HER-3 and HER-4) in the cell lines employed and correlated this to their respective heregulin responses. These data demonstrate that HER-2/neu overexpression itself affects the expression of other RTK I members and that cells expressing the highest levels of HER-2/neu have the greatest response to HRG.

  View details for Web of Science ID 000083359100011

  View details for PubMedID 10557094

• Combination therapy with trastuzumab (Herceptin) and cisplatin for chemoresistant metastatic breast cancer: Evidence for receptor-enhanced chemosensitivity Medical Oncologist Consensus Conference on the Future Directions with Herceptin (Trastuzumab) for Breast Cancer Pegram, M. D., Slamon, D. J. W B SAUNDERS CO-ELSEVIER INC. 1999: 89?95

  Abstract

  The anti-HER-2/neu antibody trastuzumab (Herceptin; Genentech, San Francisco, CA) interferes with DNA repair induced by cisplatin and, as a result, promotes cytotoxicity in HER-2/neu-overexpressing tumor target cells in a synergistic fashion. This effect of trastuzumab, termed receptor-enhanced chemosensitivity, is specific for HER-2/neu-overexpressing cells, having no effect on cells without overexpression. Based on these findings, we conducted phase I and II clinical trials of trastuzumab plus cisplatin to determine the toxicity, pharmacokinetics, response rate, and response duration of this combination in patients with HER-2/neu-overexpressing metastatic breast cancer who had demonstrated disease progression (chemoresistance) while on active chemotherapy just prior to study entry. In phase I, four of 15 patients had objective clinical responses, including one complete response of several years' duration. Of 37 assessable patients enrolled in phase II, nine (24.3%) had objective clinical responses and an additional nine had minor responses or stable disease. The median time to progression among the responders was 8.4 months. The toxicity profile reflected that expected from cisplatin alone, with no apparent increase in toxicity caused by the addition of trastuzumab. Moreover, the pharmacokinetics of trastuzumab were unaltered by coadministration of cisplatin. We conclude that the combination of trastuzumab and cisplatin results in response rates higher than that reported for either single agent alone. Such receptor-enhanced chemosensitivity offers a new approach to target overexpressed growth factor receptors in a variety of cancers, which will lead to new, biologically based therapeutic strategies for clinical intervention.

  View details for Web of Science ID 000082526300011

  View details for PubMedID 10482199

• Herceptin in the treatment of metastatic breast cancer GYNAKOLOGE Konecny, G., Untch, M., Pegram, M. 1999; 32 (8): 624-631

  View details for Web of Science ID 000082011000006

• Inhibitory effects of combinations of HER-2/neu antibody and chemotherapeutic agents used for treatment of human breast cancers ONCOGENE Pegram, M., HSU, S., Lewis, G., Pietras, R., Beryt, M., Sliwkowski, M., Coombs, D., Baly, D., Kabbinavar, F., Slamon, D. 1999; 18 (13): 2241-2251

  Abstract

  Previous studies have demonstrated a synergistic interaction between rhuMAb HER2 and the cytotoxic drug cisplatin in human breast and ovarian cancer cells. To define the nature of the interaction between rhuMAb HER2 and other classes of cytotoxic drugs, we applied multiple drug effect/combination index (CI) isobologram analysis to a variety of chemotherapeutic drug/rhuMAb HER2 combinations in vitro. Synergistic interactions at clinically relevant drug concentrations were observed for rhuMAb HER2 in combination with cisplatin (CI=0.48, P=0.003), thiotepa (CI=0.67, P=0.0008), and etoposide (CI=0.54, P=0.0003). Additive cytotoxic effects were observed with rhuMAb HER2 plus doxorubicin (CI=1.16, P=0.13), paclitaxel (CI=0.91, P=0.21), methotrexate (CI=1.15, P=0.28), and vinblastine (CI=1.09, P=0.26). One drug, 5-fluorouracil, was found to be antagonistic with rhuMAb HER2 in vitro (CI=2.87, P=0.0001). In vivo drug/rhuMAb HER2 studies were conducted with HER-2/neu-transfected, MCF7 human breast cancer xenografts in athymic mice. Combinations of rhuMAb HER2 plus cyclophosphamide, doxorubicin, paclitaxel, methotrexate, etoposide, and vinblastine in vivo resulted in a significant reduction in xenograft volume compared to chemotherapy alone (P<0.05). Xenografts treated with rhuMAb HER2 plus 5-fluorouracil were not significantly different from 5-fluorouracil alone controls consistent with the subadditive effects observed with this combination in vitro. The synergistic interaction of rhuMAb HER2 with alkylating agents, platinum analogs and topoisomerase II inhibitors, as well as the additive interaction with taxanes, anthracyclines and some antimetabolites in HER-2/neu-overexpressing breast cancer cells demonstrates that these are rational combinations to test in human clinical trials.

  View details for Web of Science ID 000079525100008

  View details for PubMedID 10327070

• Remission of human breast cancer xenografts on therapy with humanized monoclonal antibody to HER-2 receptor and DNA-reactive drugs ONCOGENE Pietras, R. J., Pegram, M. D., Finn, R. S., Maneval, D. A., Slamon, D. J. 1998; 17 (17): 2235-2249

  Abstract

  HER-2 oncogene encodes a transmembrane growth factor receptor that is overexpressed in 25-30% of patients with primary breast and ovarian cancer. A murine monoclonal antibody, 4D5, to the extracellular domain of HER-2 receptor elicits cytostatic growth inhibition of tumor cells overexpressing HER-2 protein, but clinical use of this antibody is limited by genesis of human anti-mouse antibodies. To avoid this problem, a recombinant humanized 4D5 monoclonal antibody (rhuMAb HER-2) was developed and tested using a human tumor xenograft model. Human breast and ovarian cancer cells which overexpress HER-2 were inhibited in vivo by the rhuMAb HER-2 antibody. Tumor growth relative to control was reduced at all doses of antibody tested, and the magnitude of growth inhibition was directly related to dose of rhuMAb HER-2. Tumor growth resumed on termination of antibody therapy, indicating a cytostatic effect. To elicit a cytotoxic response, human breast tumor xenografts were treated with a combination of antibody and antitumor drugs, cisplatin or doxorubicin. The combination of antibody with either cisplatin or doxorubicin resulted in significantly greater growth inhibition, with the cisplatin combination demonstrating a greater response. In addition, therapy with cisplatin and antireceptor antibody elicited complete tumor remissions after 2-3 cycles of therapy. The schedule of administration of anti-receptor antibody and cisplatin was critical for occurrence of antibody-induced potentiation in cisplatin cytotoxicity. Enhanced killing of tumor cells was found only if antibody and drug were given in close temporal proximity. Since interference with DNA repair pathways may contribute to this receptor-enhanced chemosensitivity, repair of cisplatin-damaged reporter DNA (pCMV-beta) was determined in human breast cells. As in studies of antibody-enhanced cisplatin cytotoxicity in vivo, treatment with rhuMAb HER-2 blocked the repair of cisplatin-damaged DNA only if the antibody was administered in close temporal proximity to transfection of the drug-exposed reporter DNA. An alternative measure of DNA repair, unscheduled DNA synthesis, was also assessed. Treatment with either cisplatin or doxorubicin led to an increase in unscheduled DNA synthesis that was reduced by combined therapy with antireceptor antibody specific to HER-2-overexpressing breast cancer cells. Using a direct measure of DNA repair, therapy of HER-2-overexpressing cells with rhuMAb HER-2 also blocked the removal of cisplatin-induced DNA adducts. Expression of p21/WAF1, an important mediator of DNA repair, was disrupted in breast cancer cells with HER-2 overexpression, but not in control cells, after treatment with HER-2 antibody, thus suggesting cross-communication between the HER-2 signaling and DNA repair pathways. These data demonstrate an in vivo antiproliferative effect of rhuMAb HER-2 on tumors that overexpress HER-2 receptor and a therapeutic advantage in the administration of the antireceptor antibody in combination with chemotherapeutic agents.

  View details for Web of Science ID 000076698200009

  View details for PubMedID 9811454

• Phase II study of receptor-enhanced chemosensitivity using recombinant humanized anti-p185(HER2/neu) monoclonal antibody plus cisplatin in patients with HER2/neu-overexpressing metastatic breast cancer refractory to chemotherapy treatment JOURNAL OF CLINICAL ONCOLOGY Pegram, M. D., Lipton, A., Hayes, D. F., Weber, B. L., Baselga, J. M., Tripathy, D., Baly, D., Baughman, S. A., Twaddell, T., Glaspy, J. A., Slamon, D. J. 1998; 16 (8): 2659-2671

  Abstract

  To determine the toxicity, pharmacokinetics, response rate, and response duration of intravenous (i.v.) administration of recombinant, humanized anti-p185HER2 monoclonal antibody (rhuMAb HER2) plus cisplatin (CDDP) in a phase II, open-label, multicenter clinical trial for patients with HER2/neu-overexpressing metastatic breast cancer.The study population consisted of extensively pretreated advanced breast cancer patients with HER2/neu overexpression and disease progression during standard chemotherapy. Patients received a loading dose of rhuMAb HER2 (250 mg i.v.) on day 0, followed by weekly doses of 100 mg i.v. for 9 weeks. Patients received CDDP (75 mg/m2) on days 1, 29, and 57.Of 37 patients assessable for response, nine (24.3%) achieved a PR, nine (24.3%) had a minor response or stable disease, and disease progression occurred in 19 (51.3%). The median response duration was 5.3 months (range, 1.6-18). Grade III or IV toxicity was observed in 22 of 39 patients (56%). The toxicity profile reflected that expected from CDDP alone with the most common toxicities being cytopenias (n = 10), nausea/vomiting (n = 9), and asthenia (n = 5). Mean pharmacokinetic parameters of rhuMAb HER2 were unaltered by coadministration of CDDP.The use of rhuMAb HER2 in combination with CDDP in patients with HER2/neu-overexpressing metastatic breast cancer results in objective clinical response rates higher than those reported previously for CDDP alone, or rhuMAb HER2 alone. In addition, the combination results in no apparent increase in toxicity. Finally, the pharmacology of rhuMAb HER2 was unaffected by coadministration with CDDP.

  View details for Web of Science ID 000075215900012

  View details for PubMedID 9704716

• Cytotoxicity and specificity of directed toxins composed of diphtheria toxin and the EGF-like domain of heregulin beta 1 BIOCHEMISTRY Landgraf, R., Pegram, M., Slamon, D. J., Eisenberg, D. 1998; 37 (9): 3220-3228

  Abstract

  As a step in the design of directed toxins, aimed at cells that overexpress HER receptors, particularly breast carcinoma cells, we studied the properties of a chimera of diphtheria toxin (DT) and heregulin beta1. The EGF-like growth hormone heregulin is a ligand for the HER3 and HER4 receptors and their heterodimers with HER2. The 60-residue EGF-like domain (hrg) of heregulin elicits a biological response and binds to these receptors primarily through its N terminus. We tested a fusion protein in which hrg replaces the C-terminal receptor-binding domain of DT (DT(389)hrg) and an alternative design in which this domain is fused to the N terminus of DT(389). Of those two constructs, the N-terminal fusion was not active as a directed toxin but elicited a growth response. The C-terminal fusion of hrg to DT(389) yielded a functional toxin and showed cell line specific cytotoxicity that is consistent with heregulin specificity. The binding of hrg to its cognate receptor is not impaired as shown by receptor activation, direct binding, and competition with free hrg. Cytotoxicity is dependent on high-affinity binding of DT(389)hrg to HER3 and HER4 receptors and is not mediated by HER2 overexpression alone. For those cell lines exhibiting high-affinity binding sites, the level of cytotoxicity correlates with the rate of internalization. Thus DT(389)hrg chimeras offer a possible avenue toward directed toxins against cells that overexpress HER receptors.

  View details for Web of Science ID 000072523600057

  View details for PubMedID 9485477

• HER-2/neu as a predictive marker of response to breast cancer therapy BREAST CANCER RESEARCH AND TREATMENT Pegram, M. D., Pauletti, G., Slamon, D. J. 1998; 52 (1-3): 65-77

  Abstract

  Amplification of the HER-2/neu (c-erbB-2) gene resulting in overexpression of the p185HER-2 growth factor receptor occurs in approximately 25% of early stage breast cancers. HER-2/neu has been established as an important independent prognostic factor in early stage breast cancer in large cohorts of patients and in cohorts with very long (30 year) follow-up duration. New data are emerging to suggest that HER-2/neu may be useful not only as a prognostic factor but also as a predictive marker for projecting response to chemotherapeutics, antiestrogens, and therapeutic anti-HER-2/neu monoclonal antibodies. In this review we highlight recent data on HER-2/neu as a predictive marker of response to breast cancer therapy and discuss the clinical implications of this information. The difficulty in comparing results from different data sets due to the wide variety of reagents and technologies used to detect HER-2/neu amplification/overexpression in clinical specimens is also discussed. Finally, we report results from experimental models of HER-2/neu overexpression which have been used in an effort to understand the relationship between HER-2/neu and response to chemotherapeutics and antiestrogens in breast cancer.

  View details for Web of Science ID 000078753100006

  View details for PubMedID 10066073

• Comparative mutational analysis of DPC4 (Smad4) in prostatic and colorectal carcinomas ONCOGENE MacGrogan, D., Pegram, M., Slamon, D., Bookstein, R. 1997; 15 (9): 1111-1114

  Abstract

  Allelic deletions of chromosome 18q are reported to be common in prostate and colorectal cancers, suggesting that one or more tumor suppressor genes on 18q are involved in the genesis of these neoplasms. The DPC4 gene, a recently identified candidate tumor suppressor in 18q21, was examined for evidence of inactivation in prostatic carcinomas, and results compared to those of a parallel analysis of colorectal carcinomas, for which DPC4 mutation has been reported in approximately 10% of cases. In this study, only three (10%) of 29 informative primary prostate cancers showed allelic loss of chromosome 18q21 markers, and no point mutations or deletions of DPC4 were detected in the complete set of 45 primary or metastatic cases. In contrast, five (56%) of nine primary colorectal tumors displayed allelic loss of 18q markers and in one of these a somatically acquired G-->T missense mutation was found in exon 1. Of twelve colorectal tumor cell lines, one showed a G-->C missense mutation in exon 8 and two had partial homozygous deletions that would likely abrogate gene function. These data suggest that DPC4 is rarely if ever mutated during prostatic oncogenesis, whereas inactivation of this gene may contribute to the genesis of a subset of colorectal carcinomas.

  View details for Web of Science ID A1997XT16100013

  View details for PubMedID 9285566

• The effect of HER-2/neu overexpression on chemotherapeutic drug sensitivity in human breast and ovarian cancer cells ONCOGENE Pegram, M. D., Finn, R. S., Arzoo, K., Beryt, M., Pietras, R. J., Slamon, D. J. 1997; 15 (5): 537-547

  Abstract

  Recent studies indicate that oncogenes may be involved in determining the sensitivity of human cancers to chemotherapeutic agents. To define the effect of HER-2/neu oncogene overexpression on sensitivity to chemotherapeutic drugs, a full-length, human HER-2/neu cDNA was introduced into human breast and ovarian cancer cells. In vitro dose-response curves following exposure to 7 different classes of chemotherapeutic agents were compared for HER-2- and control-transfected cells. Chemosensitivity was also tested in vivo for HER-2- and control-transfected human breast and ovarian cancer xenografts in athymic mice. These studies indicate that HER-2/neu overexpression was not sufficient to induce intrinsic, pleomorphic drug resistance. Furthermore, changes in chemosensitivity profiles resulting from HER-2/neu transfection observed in vitro were cell line specific. In vivo, HER-2/neu-overexpressing breast and ovarian cancer xenografts were responsive to different classes of chemotherapeutic drugs compared to control-treated xenografts with no statistically significant differences between HER-2/neu-overexpressing and nonoverexpressing xenografts. We found no instance in which HER-2/neu-overexpressing xenografts were rendered more sensitive to chemotherapeutic drugs in vivo. HER-2/neu-overexpressing xenografts consistently exhibited more rapid regrowth than control xenografts following initial response to chemotherapy suggesting that a high rate of tumor cell proliferation rather than intrinsic drug resistance may be responsible for the adverse prognosis associated with HER-2/neu overexpression in human cancers.

  View details for Web of Science ID A1997XN25500005

  View details for PubMedID 9247307

• Thalidomide and chemotherapy combination: Preliminary results of preclinical and clinical studies INTERNATIONAL JOURNAL OF ONCOLOGY Nguyen, M., Tran, C., Barsky, S., Sun, J. R., McBride, W., Pegram, M., Pietras, R., Love, S., Glaspy, J. 1997; 10 (5): 965-969

  Abstract

  Angiogenesis has been shown to be important in tumor growth and metastasis. Thalidomide, an oral sedative, has recently been found to inhibit angiogenesis. We therefore set out to ask whether thalidomide can be used as therapy for breast cancer. In a mouse model of breast cancer, we found that thalidomide alone did not suppress tumor growth. However, mice treated with thalidomide in combination with cytoxan and adriamycin had significantly smaller tumors than those given the two chemotherapeutic agents alone (3,432 +/- 303 mm(3) versus 4,643 +/- 203 mm(3), p = 0.0005). We proceeded to administer thalidomide together with chemotherapy to seven breast cancer patients in the context of a Phase I trial. Side effects attributed to thalidomide were minimal, and included constipation and a rash. We concluded that an approach at cancer therapeutics combining an antiangiogenic agent such as thalidomide with conventional chemotherapy may be feasible and deserves further studies.

  View details for Web of Science ID A1997WW11900015

  View details for PubMedID 21533471

• HER-3 TYROSINE KINASE PATHWAY TARGETS ESTROGEN-RECEPTOR AND PROMOTES HORMONE-INDEPENDENT GROWTH IN HUMAN BREAST-CANCER CELLS ONCOGENE Pietras, R. J., Arboleda, J., Reese, D. M., WONGVIPAT, N., Pegram, M. D., Ramos, L., Gorman, C. M., Parker, M. G., Sliwkowski, M. X., Slamon, D. J. 1995; 10 (12): 2435-2446

  Abstract

  Growth of human breast cells is closely regulated by steroid hormone as well as peptide hormone receptors. Members of both receptor classes are important prognostic factors in human breast cancer. Clinical data indicate that overexpression of the HER-2 gene is associated with an estrogen receptor-negative phenotype. In this study, we demonstrate that introduction of a HER-2 cDNA, converting non-overexpressing breast cancer cells to those which overexpress this receptor, results in development of estrogen-independent growth which is insensitive to both estrogen and the antiestrogen, tamoxifen. Moreover, activation of the HER-2 receptor in breast cancer cells by the peptide growth factor, heregulin, leads to direct and rapid phosphorylation of ER on tyrosine residues. This is followed by interaction between ER and the estrogen-response elements in the nucleus and production of an estrogen-induced protein, progesterone receptor. In addition, overexpression of HER-2 receptor in estrogen-dependent tumor cells promotes ligand-independent down-regulation of ER and a delayed autoregulatory suppression of ER transcripts. These data demonstrate a direct link between these two receptor pathways and suggest one mechanism for development of endocrine resistance in human breast cancers.

  View details for Web of Science ID A1995RE54300022

  View details for PubMedID 7784095

• ANTIBODY TO HER-2/NEU RECEPTOR BLOCKS DNA-REPAIR AFTER CISPLATIN IN HUMAN BREAST AND OVARIAN-CANCER CELLS ONCOGENE Pietras, R. J., Fendly, B. M., CHAZIN, V. R., Pegram, M. D., Howell, S. B., Slamon, D. J. 1994; 9 (7): 1829-1838

  Abstract

  Approximately 30% of human breast and ovarian cancers have amplification and/or overexpression of HER-2/neu gene which encodes a cell surface growth-factor receptor. Overexpression of this receptor, p185HER-2/neu, is associated with poor outcome and may predict clinical response to chemotherapy. Antibodies to HER-2/neu receptor have a cytostatic effect in suppressing growth of cells with overexpression of p185HER-2/neu. To elicit a cytocidal effect, therapy with antireceptor antibody was used in combination with the DNA-damaging drug, cisplatin, and this combined treatment produced a synergistic decrease in cell growth. In addition, antibody mediated an increased sensitivity to cisplatin in drug-resistant ovarian carcinoma cells containing multiple copies of HER-2/neu gene. To evaluate the mechanism for this synergy, unscheduled DNA synthesis was measured in cancer cells using incorporation of [3H]thymidine and autoradiography, and formation and repair of cisplatin-induced DNA adducts was also measured. Treatment with cisplatin led to a marked, dose-dependent increase in unscheduled DNA synthesis which was significantly reduced by combined treatment with antireceptor antibody in HER-2/neu-overexpressing cells. Therapy with antibody to HER-2/neu receptor also led to a 35-40% reduction in repair of cisplatin-DNA adducts after cisplatin exposure and, as a result, promoted drug-induced killing in target cells. This phenomenon which we term receptor-enhanced chemosensitivity may provide a rationale for more selective targeting and exploitation of overexpressed growth factor receptors in cancer cells, thus leading to new strategies for clinical intervention.

  View details for Web of Science ID A1994NR68500004

  View details for PubMedID 7911565

• DIAGNOSIS OF INFECTIVE AND INFLAMMATORY DISORDERS BY FLOW CYTOMETRIC ANALYSIS OF BLOOD NEUTROPHILS AMERICAN JOURNAL OF CLINICAL PATHOLOGY Bentley, S. A., Pegram, M. D., Ross, D. W. 1987; 88 (2): 177-181

  Abstract

  The utility of neutrophil parameters provided by two flow cytometric hematologic analyzers (the H-1 and H6000, Technicon Instruments Corporation, Tarrytown, NY) was investigated for the diagnosis of infective and/or inflammatory disorders. The test population of 156 hospital patients was selected on the basis of a blood culture request. Positivity or negativity for infective and/or inflammatory disease was inferred from chart review. The parameters evaluated included the absolute neutrophil count, the lobularity index, and the left shift flag from the H-1, the percentage of high peroxidase cells from the H6000, the routine laboratory band count, and a reference band count. Significant intercorrelations were observed between these parameters. The diagnostic performance of the routine laboratory band count was significantly inferior to that of all other parameters. At equivalent points on their receiver operating characteristic curves, the diagnostic efficiencies of the remaining tests ranged from 61.5% for the lobularity index to 67% for the left shift flag and the percentage of high peroxidase cells. These differences were not significant statistically.

  View details for Web of Science ID A1987J571700007

  View details for PubMedID 3618549