Search

Supplementary Data: List of TRIO investigators, Supplementary Materials and Methods, Supplementary Results, Supplementary Discussion, Supplementary References, Supplementary Tables, and Supplementary Figures.

Association of NR1I3 haplotype with TTP (Supplementary Online only). CAT/CAT carriers, median TTP 1.92 months vs 6.21 months for CAT/â^' or â^'/â^' carriers.

Purpose: This phase I/II single-arm study evaluated the safety, pharmacokinetics, pharmacodynamics, and activity of foretinib, an oral multikinase inhibitor of MET, ROS, RON, AXL, TIE-2, and VEGFR2, in the first-line setting in advanced hepatocellular carcinoma patients.Experimental Design: In the phase I part, advanced hepatocellular carcinoma patients were dose escalated on foretinib (30–60 mg) every day using the standard 3+3 design. Once the maximum tolerated dose (MTD) was determined, an additional 32 patients were dosed at the MTD in the phase II expansion cohort for assessment of efficacy and safety. Exploratory analyses were conducted to assess potential biomarkers that might correlate with clinical efficacy and survival.Results: The MTD of foretinib was established as 30 mg every day. The most frequent adverse events were hypertension, decreased appetite, ascites, and pyrexia. When dosed at 30 mg every day in the first-line setting, foretinib demonstrated promising antitumor activity. According to the modified mRECIST, the objective response rate was 22.9%, the disease stabilization rate 82.9%, and the median duration of response 7.6 months. The median time to progression was 4.2 months and the median overall survival (OS) was 15.7 months. Fifteen candidate biomarkers whose levels in the circulation were significantly altered in response to foretinib treatment were elucidated. Multivariate analyses identified IL6 and IL8 as independent predictors of OS.Conclusions: Foretinib demonstrated promising antitumor activity and good tolerability in the first-line setting in Asian advanced hepatocellular carcinoma patients. Baseline plasma levels of IL6 or IL8 might predict the response to foretinib. Clin Cancer Res; 23(10); 2405–13. ©2016 AACR.

Table S1: Summary of pharmacokinetic parameters (days 1 and 15); Table S2: Significant changes in circulating biomarkers from baseline; Table S3: Multivariate models for selected covariates, Overall Survival.

Background GSK2849330, an anti‐HER3 monoclonal antibody that blocks HER3/Neuregulin 1 (NRG1) signaling in cancer cells, is engineered for enhanced antibody‐dependent cellular cytotoxicity and complement‐dependent cytotoxicity. This phase I, first‐in‐human, open‐label study assessed the safety, pharmacokinetics (PK), pharmacodynamics, and preliminary activity of GSK2849330 in patients with HER3‐expressing advanced solid tumors. Patients and Methods Patients with various tumor types were prospectively selected for HER3 expression by immunohistochemistry; a subset was also screened for NRG1 mRNA expression. In the dose‐escalation phase, patients received GSK2849330 1.4–30 mg/kg every 2 weeks, or 3 mg/kg or 30 mg/kg weekly, intravenously (IV). In the dose‐expansion phase, patients received 30 mg/kg GSK2849330 IV weekly. Results Twenty‐nine patients with HER3‐expressing cancers, of whom two expressed NRG1, received GSK2849330 (dose escalation: n = 18, dose expansion: n = 11). GSK2849330 was well tolerated. No dose‐limiting toxicities were observed. The highest dose, of 30 mg/kg weekly, expected to provide full target engagement, was selected for dose expansion. Treatment‐emergent adverse events (AEs) were mostly grade 1 or 2. The most common AEs were diarrhea (66%), fatigue (62%), and decreased appetite (31%). Dose‐proportional plasma exposures were achieved, with evidence of HER3 inhibition in paired tissue biopsies. Of 29 patients, only 1 confirmed partial response, lasting 19 months, was noted in a patient with CD74‐NRG1‐rearranged non‐small cell lung cancer (NSCLC). Conclusion GSK2849330 demonstrated a favorable safety profile, dose‐proportional PK, and evidence of target engagement, but limited antitumor activity in HER3‐expressing cancers. The exceptional response seen in a patient with CD74‐NRG1‐rearranged NSCLC suggests further exploration in NRG1‐fusion–positive cancers. Implications for Practice This first‐in‐human study confirms that GSK2849330 is well tolerated. Importantly, across a variety of HER3‐expressing advanced tumors, prospective selection by HER3/NRG1 expression alone was insufficient to identify patients who could benefit from treatment with this antibody‐dependent cell‐mediated cytotoxicity– and complement‐dependent cytotoxicity–enhanced anti‐HER3 antibody. The only confirmed durable response achieved was in a patient with CD74‐NRG1‐rearranged lung cancer. This highlights the potential utility of screening for NRG1 fusions prospectively across tumor types to enrich potential responders to anti‐HER3 agents in ongoing trials., This phase I, first‐in‐human study assessed the safety, pharmacokinetics, pharmacodynamics, and preliminary activity of GSK2849330 in patients with HER3‐expressing advanced solid tumors.

Background Idiopathic pulmonary fibrosis (IPF) causes irreversible loss of lung function. The lysophosphatidic acid receptor 1 (LPA1) pathway is implicated in IPF etiology. Safety and efficacy of BMS-986020, a high-affinity LPA1 antagonist, was assessed vs placebo in a phase 2 study in patients with IPF. Methods IM136003 was a phase 2, parallel-arm, multicenter, randomized, double-blind, placebo-controlled trial. Adults with IPF (FVC, 45%-90%; diffusing capacity for carbon monoxide, 30%-80%) were randomized to receive placebo or 600 mg BMS-986020 (once daily [qd] or bid) for 26 weeks. The primary end point was rate of change in FVC from baseline to week 26. Results Of 143 randomized patients, 108 completed the 26-week dosing phase. Thirty-five patients discontinued prematurely. Patient baseline characteristics were similar between treatment groups (placebo: n = 47; 600 mg qd: n = 48; 600 mg bid: n = 48). Patients treated with BMS-986020 bid experienced a significantly slower rate of decline in FVC vs placebo (−0.042 L; 95% CI, −0.106 to −0.022 vs −0.134 L; 95% CI, −0.201 to −0.068, respectively; P = .049). Dose-related elevations in hepatic enzymes were observed in both BMS-986020 treatment groups. The study was terminated early because of three cases of cholecystitis that were determined to be related to BMS-986020 after unblinding. Conclusions BMS-986020 600 mg bid treatment for 26 weeks vs placebo significantly slowed the rate of FVC decline. Both regimens of BMS-986020 were associated with elevations in hepatic enzymes. Trial Registry ClinicalTrials.gov; No.: NCT01766817; URL: www.clinicaltrials.gov

Fibrosis, resulted from the imbalance of fibrogenesis and fibrolysis, is a key readout of disease progression in nonalcoholic steatohepatitis (NASH) and reflects mortality risk. Non-invasive biomarkers capable of diagnosing fibrosis stages and monitoring fibrosis changes in NASH patients are urgently needed. This study is to evaluate collagen formation and degradation biomarkers, reflective of fibrogenesis or fibrolysis, in patients with biopsy proven NASH. Collagen formation biomarker PRO-C3 and PRO-C6 levels were significantly higher in patients with advanced fibrosis stage 3–4 than those with fibrosis stage 0–2. Elevated PRO-C3 levels were also associated with severe lobular inflammation and ballooning, but not with steatosis. Multivariate logistic regression analysis identified PRO-C3 and PRO-C6 to be independently related to fibrosis stage. PRO-C3 showed similar performance to identify patients with advanced fibrosis in discovery and validation cohorts. Furthermore, in a longitudinal study cohort with paired biopsies, mean PRO-C3 increased with worsening of fibrosis and decreased with fibrosis improvement. The results suggest that PRO-C3 may be a potentially useful biomarker in identifying patients with advanced fibrosis and active fibrogenesis, as well as in assessing changes in fibrosis over time. It is worthy of further evaluation to confirm its diagnostic value and clinical utility.

Background: In two clinical trials of the vascular endothelial growth factor (VEGF) receptor inhibitor pazopanib in advanced renal cell carcinoma (mRCC), we found interleukin-6 as predictive of pazopanib benefit. We evaluated the prognostic significance of candidate cytokines and angiogenic factors (CAFs) identified in that work relative to accepted clinical parameters. Methods: Seven preselected plasma CAFs (interleukin-6, interleukin-8, osteopontin, VEGF, hepatocyte growth factor, tissue inhibitor of metalloproteinases (TIMP-1), and E-selectin) were measured using multiplex ELISA in plasma collected pretreatment from 343 mRCC patients participating in the phase 3 registration trial of pazopanib vs placebo (NCT00334282). Tumour burden (per sum of longest diameters (SLD)) and 10 other clinical factors were also analysed for association with overall survival (OS; based on initial treatment assignment). Results: Osteopontin, interleukin-6, and TIMP-1 were independently associated with OS in multivariable analysis. A model combining the three CAFs and five clinical variables (including SLD) had higher prognostic accuracy than the International Metastatic Renal Cell Carcinoma Database Consortium criteria (concordance-index 0.75 vs 0.67, respectively), and distinguished two groups of patients within the original intermediate risk category. Conclusions: A prognostic model incorporating osteopontin, interleukin-6, TIMP-1, tumour burden, and selected clinical criteria increased prognostic accuracy for OS determination in mRCC patients.

Background Combined targeted strategy with letrozole (Le) and lapatinib (La) improves progression-free survival (PFS) in patients with metastatic breast cancer (MBC) co-expressing hormone receptor-positive (HR+) and HER2+ but not in HR+/HER2-negative (HER-) disease (Johnston J Clin Oncol 2009). However, among HER2+ tumors, quantitative levels of HER2 are heterogeneous with a broad dynamic range corresponding to approximately 163.7 to 17446.7 amol/µg as previously reported (Nuciforo SABCS 2014). In addition, within HER2- tumors, quantitative measurement of HER family proteins may identify those patients most likely to benefit from the addition of La to Le. In this retrospective study, we tested the prognostic and predictive ability of HER proteins quantification in clinically HER2+ tumor samples from the EGF30008 study. Methods Formalin-fixed paraffin-embedded primary tumor tissues sections from HER2+ MBC population were used. After laser microdissection, tissue lysates were prepared for selected reaction monitoring mass spectrometry (SRM-MS) analysis. Absolute quantitation was accomplished through simultaneous detection of endogenous target and synthetic labeled heavy peptide identical to analytical targets (EGFR, HER2, HER3). HER2 protein levels were correlated with PAM50 molecular subtypes, ERBB2 and ESR1 genes by nCounter. PFS and overall survival (OS) were analyzed by Kaplan–Meier and log-rank test. To determine whether HER2 protein levels were predictive of La benefit, we tested the interaction term of HER2 protein as a continuous variable by treatment arm in a Cox model. Results Within the HER2+ study cohort (n=219), 107 had an available tumor block; 84 cases had sufficient material for HER expression measurement by SRM-MS. Average HER2 levels were 2321.1 amol/ug (median, 817.6). HER2 levels were lower in Le+La (n=43; mean, 1761 amol/ug) compared to Le (n=41; mean, 2908 amol/ug) arms, although the difference was non-significant (p=0.108). No expression of EGFR and HER3 was observed. HER2 protein levels were significantly different among PAM50 subtypes with HER2-enriched (HER2E) tumors showing the highest expression followed by Basal-like, Luminal A, Luminal B, and Normal-like (p Conclusions Levels of HER2 protein in HER2+ MBC are extremely heterogeneous. An association between HER2 protein and gene expression by nCounter was observed. HER2E tumors by PAM50 showed the highest levels of HER2 protein. Within the group of HER2+ MBC by standard IHC/FISH, tumors with high HER2 protein had a statistically non-significant worse outcome and do not seem to benefit from La. Further validation of these findings is warranted. Citation Format: Nuciforo P, Thyparambil S, Galván P, Vilaro M, Jimenez J, Liao W-L, Cecchi F, Blackler A, Press MF, Gagnon R, Ellis C, Hembrough T, Johnston S, Prat A. Quantitative HER family proteins assessment as prognostic and predictive biomarkers in the EGF30008 clinical trial. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P3-07-08.

Background Combination of letrozole and lapatinib improved progression-free survival (PFS) compared with letrozole and placebo in patients with hormone receptor-positive (HR+)/HER2+ metastatic breast cancer (MBC), but not HR+/HER2-negative (HER2-) disease (JCO 2009). However, HR+ disease is clinically and biologically heterogeneous with all intrinsic molecular subtypes (Luminal A, Luminal B, HER2-enriched [HER2E] and Basal-like) identified. Here, we tested retrospectively the prognostic and predictive ability of intrinsic subtype in tumor samples of the EGF30008 trial. Methods Expression profiling from FFPE tumor tissues was performed on the nCounter platform. Tumors were classified into each intrinsic subtype using the research-based PAM50 classifier (JCO 2009). Cox proportional hazard models for PFS and overall survival (OS) were used to generate point estimates of hazard ratios (HR) and corresponding 95% confidence intervals (CIs). Changes in likelihood ratio χ2 values were used to measure and compare the relative amount of information of each variable. Variables evaluated were: age, prior endocrine therapy, presence of visceral disease, number of metastatic sites, performance status, clinical HER2 status, and treatment. To determine whether the intrinsic subtypes were predictive of lapatinib benefit, we tested the interaction term of subtype by treatment arm in a Cox model that also included the main effects. Kaplan-Meier plots were used to depict the proportion of patients free from progression as a function of time. Results Tumor samples from 821 patients (63.8%) were profiled (85.7% primary and 14.3% metastatic tumor samples). Clinical-pathological features of this patient subset were well balanced compared with the original set. Within the entire cohort, all subtypes were identified: Luminal A (46.5%); Luminal B (29.7%); HER2E (7.4%); Basal-like (3.4%) and normal-like (12.9%). Within HER2+ disease, 28.6% of samples were HER2E. Intrinsic subtype was found the strongest prognostic factor independently associated with PFS and OS in all patients, and in patientswith HER2-negative or HER2+ disease (P Conclusions HR-positive disease is biologically heterogeneous and intrinsic subtypes are strongly prognostic in a first-line MBC setting. HR+/HER2- disease with a HER2E profile may benefit from lapatinib. The clinical value of intrinsic subtyping in HR+ MBC warrants further investigation, but patients with Luminal A/HER2-negative MBC disease might be good candidates for letrozole monotherapy in the first-line setting regardless of visceral disease and number of metastases. Citation Format: Prat A, Cheang MCU, Galván P, Nuciforo P, Paré L, Adamo B, Viladot M, Press MF, Gagnon R, Ellis C, Johnston S. Prognostic and predictive abilities of intrinsic subtype in hormone receptor-positive metastatic breast cancer from the EGF30008 phase III clinical trial. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P2-08-16.

Purpose: Tumor-derived circulating cell–free DNA (cfDNA) is a potential alternative source from which to derive tumor mutation status. cfDNA data from four clinical studies of the BRAF inhibitor (BRAFi) dabrafenib or the MEK inhibitor (MEKi) trametinib were analyzed to determine the association between BRAF mutation status in cfDNA and tumor tissue, and the association of BRAF cfDNA mutation status with baseline factors and clinical outcome. Experimental Design: Patients with BRAF V600 mutation–positive melanoma were enrolled in each study after central confirmation of BRAF status in tumor using a PCR-based assay. BRAF mutation status in cfDNA from patient plasma collected at baseline, 732 of 836 (88%) enrolled patients in total, was determined. Results: BRAF mutations were detectable in cfDNA in 76% and 81% of patients with BRAF V600E/V600K–positive tumors, respectively. Patients negative for BRAF mutations in cfDNA had longer progression-free survival (PFS) and overall survival in each of the four studies, compared with patients with detectable cfDNA BRAF mutations. The presence of BRAF-mutant cfDNA was an independent prognostic factor for PFS after multivariate adjustment for baseline factors in three of four studies. Patients negative for BRAF mutation–positive cfDNA in plasma had higher response rates to dabrafenib and trametinib. Conclusions: BRAF mutations in cfDNA are detectable in >75% of late-stage melanoma patients with BRAF mutation–positive tumors. The lack of circulating, BRAF mutation–positive cfDNA is clinically significant for metastatic melanoma patients, and may be a prognostic marker for better disease outcome. Clin Cancer Res; 22(3); 567–74. ©2015 AACR.

Purpose: The interaction of programmed death-1 ligand (PD-L1) with its receptor (PD-1) on T cells inactivates antitumor immune responses. PD-L1 expression has been associated with poor outcomes in renal cell carcinoma (RCC) but has not been investigated in advanced RCC patients receiving VEGF-targeted therapy. Experimental Design: Formalin-fixed paraffin-embedded specimens were collected at baseline from patients in the COMPARZ trial. Tumor cell PD-L1 expression by IHC was evaluated using H-score (HS). Dual PD-L1/CD68 staining was used to differentiate PD-L1 tumor expression from tumor-associated macrophages. Intratumor CD8-positive T cells were quantified morphometrically. Associations between biomarkers and survival were investigated using the log-rank test. Results: HS data were available from 453 of 1,110 patients. Sixty-four percent of patients had negative PD-L1 expression (HS = 0). Patients with HS > 55 (n = 59, 13%) had significantly shorter overall survival (OS) than those with HS ≤ 55 in both pazopanib and sunitinib arms (median 15.1 vs. 35.6 and 15.3 vs. 27.8 months, respectively, P = 0.03). In both arms, median OS was shortest in patients with HS > 55 and intratumor CD8-positive T-cell counts > 300 (9.6 and 11.9 months with pazopanib and sunitinib, respectively). Median OS in patients with HS ≤ 55 and CD8-positive T-cell counts ≤ 300 was 36.8 and 28.0 months with pazopanib and sunitinib, respectively. Progression-free survival results were similar to OS results. Conclusions: Increased tumor cell PD-L1, or PD-L1 plus tumor CD8-positive T-cell counts, were associated with shorter survival in patients with metastatic RCC receiving VEGF-targeted agents. These findings may have implications for future design of randomized clinical trials in advanced RCC. Clin Cancer Res; 21(5); 1071–7. ©2014 AACR.

Background GSK609 an agonist non-T cell depleting IgG4 monoclonal antibody (mAb) against inducible co-stimulatory receptor (ICOS) exhibits T cell mediated immune stimulating and anti-tumor activity. INDUCE-1 is the first in human study investigating GSK609 alone and in combinations which include pembrolizumab in select tumor types including recurrent/metastatic (R/M) HNSCC. Methods Safety, PK, PD, and preliminary antitumor activity of GSK609 are being evaluated at doses from 0.001 to 10 mg/kg every 3 weeks (Q3W). Blood samples collected prior to dosing and on-study are evaluated for PK and PD effects on lymphocytes and ICOS receptor occupancy (RO). Tumor biopsies at Screening and Week 6 are evaluated for changes in tumor immune infiltrates (TIL) by a multiplexed immuno-fluorescence and gene expression platforms. Results The GSK609 PK disposition shows low clearance, limited central volume of distribution, and mean systemic half-life of 19 days which is consistent with other humanized mAbs. Evidence of target engagement and tumor size reduction are observed in a R/M HNSCC expansion cohort (EC) at 0.3 mg/kg with 200 mg pembrolizumab. Dose and concentration-RO analyses suggest ≥0.1 mg/kg GSK609 maintains ≥ 70% RO on peripheral CD4+ and CD8+ T cells. Quantitative TIL evaluation of paired tumor biopsies demonstrates favorable immune microenvironment in the tumor at exposures observed in patients treated with 0.3mg/kg. TIL and tumor-based gene expression data demonstrate non-linear, dose-dependent changes in select immune activation markers. Exposure-response assessments reveal no difference in baseline-to-Week 9 target lesion change across exposures in the EC. Furthermore, cross-cohort pooled exposure-response analysis of ≥Grade 2 AEs demonstrates similar safety outcomes across the exposures/doses. Additionally, population PK modeling suggests comparable exposures can be maintained by fixed dosing as well. Conclusions The current data provide preliminary evidence of GSK609 target engagement and biological activity at clinically tolerable doses and support further exploration of the 0.3mg/kg or 24mg fixed dose. Clinical trial identification NCT02723955 (Rel. 31March2016). Legal entity responsible for the study GlaxoSmithKline. Funding GlaxoSmithKline. Disclosure M. Maio: Honoraria (self): BMS, MSD, Roche, Merck, Eli Lilly; Honoraria (institution), Patients’ fee to the University Hospital of Siena: BMS, MSD, AZ, Roche, Merck; Advisory / Consultancy: BMS, MSD, Roche, Merck, Eli Lilly; Travel / Accommodation / Expenses: BMS, MSD, Roche, Merck, Eli Lilly; Non-remunerated activity/ies, Press conferences: Merck, BMS. T. Bauer: Advisory / Consultancy, Self: Guardant Health; Loxo; Pfizer; Advisory / Consultancy, Institution: Ignyta; Moderna Therapeutics; Pfizer; Speaker Bureau / Expert testimony, Self: Bayer; Research grant / Funding (institution): Daiichi Sankyo; Medpacto, Inc.; Incyte; Mirati Therapeutics; MedImmune; Abbvie; AstraZeneca; Leap Therapeutics; MabVax; Stemline Therapeutics; Merck; Lilly; GlaxoSmithKline; Novartis, Pfizer; Genentech/Roche; Deciphera; Merrimack; Immunogen; Millennium; I; Travel / Accommodation / Expenses: Astellas Pharma; AstraZeneca; Celgene; Clovis Oncology; EMD Serono; Genentech; Lilly; Merck; Novartis; Pharmacyclics; Sysmex. D. Rischin: Advisory / Consultancy, All uncompensated : MSD, Regeneron, GSK, BMS; Research grant / Funding (institution): Regeneron, Roche, MSD, GSK, BMS; Travel / Accommodation / Expenses: MSD. V. Moreno: Advisory / Consultancy: Merck, BMS; Travel / Accommodation / Expenses: Regeneron/Sanofi, BMS. J.M. Trigo Perez: Advisory / Consultancy: Regeneron/Sanofi, BMS; Speaker Bureau / Expert testimony: Regeneron/Sanofi, BMS; Travel / Accommodation / Expenses: Regeneron/Sanofi, BMS. M. Chisamore: Full / Part-time employment: Merck & Co. Inc; Shareholder / Stockholder / Stock options: Merck & Co. Inc. J. Sadik Shaik: Full / Part-time employment: GlaxoSmithKline; Shareholder / Stockholder / Stock options: GlaxoSmithKline. F. Rigat: Full / Part-time employment: GlaxoSmithKline; Shareholder / Stockholder / Stock options: GlaxoSmithKline. C. Ellis: Full / Part-time employment: GlaxoSmithKline; Shareholder / Stockholder / Stock options: GlaxoSmithKline. H. Chen: Full / Part-time employment: GlaxoSmithKline; Shareholder / Stockholder / Stock options: GlaxoSmithKline. R. Gagnon: Full / Part-time employment: GlaxoSmithKline; Shareholder / Stockholder / Stock options: GlaxoSmithKline. S. Scherer: Full / Part-time employment: GlaxoSmithKline; Shareholder / Stockholder / Stock options: GlaxoSmithKline. D. Turner: Full / Part-time employment: GlaxoSmithKline; Shareholder / Stockholder / Stock options: GlaxoSmithKline. S. Yadavilli: Full / Part-time employment: GlaxoSmithKline; Shareholder / Stockholder / Stock options: GlaxoSmithKline. M. Ballas: Full / Part-time employment: GlaxoSmithKline; Shareholder / Stockholder / Stock options: GlaxoSmithKline. A. Hoos: Full / Part-time employment: GlaxoSmithKline; Shareholder / Stockholder / Stock options: GlaxoSmithKline. E. Angevin: Advisory / Consultancy: Merck Sharp & Dohme, GlaxoSmithKline, Celgene Research, MedImmune; Travel / Accommodation / Expenses: AbbVie, Roche, Sanofi, Pfizer, MedImmune, Innate Pharma, Celgene, BMS; Research grant / Funding (institution): Abbvie, Aduro, Agios, Amgen, Argen-x, Astex, AstraZeneca, Aveo pharmaceuticals, Bayer, Beigene, Blueprint, BMS, Boeringer Ingelheim, Celgene, Chugai, Clovis, Daiichi Sankyo, Debiopharm, Eisai, Eos, Exelixis, Forma, Gamamabs, Genentech, Gortec, GSK, H3 bio. All other authors have declared no conflicts of interest.

Purpose: Inactivation of von Hippel-Lindau (VHL) gene in clear-cell renal cell carcinoma (RCC) leads to increased levels of hypoxia-inducible factors (HIF) and overexpression of HIF target genes, such as VEGF and others. VEGF-targeted agents are standard in advanced clear-cell RCC but biomarkers of activity are lacking. Experimental Design: We analyzed tumor tissue samples from metastatic clear-cell RCC patients who received pazopanib as part of clinical trial VEG102616. We evaluated several components of the VHL/HIF pathway: VHL gene inactivation (mutation and/or methylation), HIF-1α and HIF-2α immunohistochemistry staining, and HIF-1α transcriptional signature. We evaluated the association of these biomarkers with best overall response rate (ORR) and progression-free survival (PFS) to pazopanib, a standard first-line VEGF-targeted agent. Results: The VEG102616 trial enrolled 225 patients, from whom 78 samples were available for tumor DNA extraction. Of these, 70 patients had VHL mutation or methylation. VHL gene status did not correlate with ORR or PFS. Similarly, HIF-1α (65 samples) and HIF-2α (66 samples) protein levels (high vs. low) did not correlate with ORR or PFS to pazopanib. The HIF-1α transcriptional signature (46 samples) was enriched in tumors expressing high HIF-1α levels. However, the HIF-1α gene expression signature was not associated with clinical outcome to pazopanib. Conclusions: In patients with advanced clear-cell RCC, several potential biomarkers along the VHL/HIF-1α/HIF-2α axis were not found to be predictive for pazopanib activity. Additional efforts must continue to identify biomarkers associated with clinical outcome to VEGF-targeted agents in metastatic RCC. Clin Cancer Res; 19(18); 5218–26. ©2013 AACR.

Background The results from a phase III trial conducted outside of Japan demonstrated a significant improvement in time to progression (TTP) when lapatinib was combined with capecitabine compared with capecitabine alone in patients with HER2-positive advanced or metastatic breast cancer. In this clinical study of lapatinib in combination with capecitabine, efficacy, safety, pharmacokinetics (PK) and biomarkers were investigated in Japanese patients with HER2-positive advanced or metastatic breast cancer treated with prior trastuzumab. Methods Eligible women received lapatinib 1250 mg once daily and capecitabine 1000 mg/m2 twice daily on days 1 through 14 of a 21-day cycle. The primary endpoint was the clinical benefit rate (CBR: complete response, partial response or stable disease for at least 24 weeks). Results Lapatinib in combination with capecitabine was well tolerated in the 51 patients enrolled in this study. CBR was 59 % (95 % CI 44.2, 72.4), and the median TTP in the Kaplan-Meier estimate was 36 weeks (95 % CI 27.1, 48.0). The majority of drug-related adverse events were mild to moderate (grade 1 or 2); the most common adverse events reported were palmar-plantar erythrodysesthesia syndrome (76 %), diarrhea (67 %) and stomatitis (41 %). Conclusions Lapatinib in combination with capecitabine in Japanese HER2-positive breast cancer patients was well tolerated. Overall, our findings on the efficacy, safety and PK were similar to those reported from the overseas studies.

Enzyme-linked and electrochemiluminescence immunoassays were developed for quantification of amino (N-) terminal fragments of the skeletal muscle protein titin (N-ter titin) and qualified for use in detection of urinary N-ter titin excretion. Urine from normal subjects contained a small but measurable level of N-ter titin (1.0 ± 0.4 ng/ml). A 365-fold increase (365.4 ± 65.0, P = 0.0001) in urinary N-ter titin excretion was seen in Duchene muscular dystrophy (DMD) patients. Urinary N-ter titin was also evaluated in dystrophin deficient rodent models. Mdx mice exhibited low urinary N-ter titin levels at 2 weeks of age followed by a robust and sustained elevation starting at 3 weeks of age, coincident with the development of systemic skeletal muscle damage in this model; fold elevation could not be determined because urinary N-ter titin was not detected in age-matched wild type mice. Levels of serum creatine kinase and serum skeletal muscle troponin I (TnI) were also low at 2 weeks, elevated at later time points and were significantly correlated with urinary N-ter titin excretion in mdx mice. Corticosteroid treatment of mdx mice resulted in improved exercise performance and lowering of both urinary N-ter titin and serum skeletal muscle TnI concentrations. Low urinary N-ter titin levels were detected in wild type rats (3.0 ± 0.6 ng/ml), while Dmdmdx rats exhibited a 556-fold increase (1652.5 ± 405.7 ng/ml, P = 0.002) (both at 5 months of age). These results suggest that urinary N-ter titin is present at low basal concentrations in normal urine and increases dramatically coincident with muscle damage produced by dystrophin deficiency. Urinary N-ter titin has potential as a facile, non-invasive and translational biomarker for DMD.

Purpose: This phase I/II single-arm study evaluated the safety, pharmacokinetics, pharmacodynamics, and activity of foretinib, an oral multikinase inhibitor of MET, ROS, RON, AXL, TIE-2, and VEGFR2, in the first-line setting in advanced hepatocellular carcinoma patients. Experimental Design: In the phase I part, advanced hepatocellular carcinoma patients were dose escalated on foretinib (30–60 mg) every day using the standard 3+3 design. Once the maximum tolerated dose (MTD) was determined, an additional 32 patients were dosed at the MTD in the phase II expansion cohort for assessment of efficacy and safety. Exploratory analyses were conducted to assess potential biomarkers that might correlate with clinical efficacy and survival. Results: The MTD of foretinib was established as 30 mg every day. The most frequent adverse events were hypertension, decreased appetite, ascites, and pyrexia. When dosed at 30 mg every day in the first-line setting, foretinib demonstrated promising antitumor activity. According to the modified mRECIST, the objective response rate was 22.9%, the disease stabilization rate 82.9%, and the median duration of response 7.6 months. The median time to progression was 4.2 months and the median overall survival (OS) was 15.7 months. Fifteen candidate biomarkers whose levels in the circulation were significantly altered in response to foretinib treatment were elucidated. Multivariate analyses identified IL6 and IL8 as independent predictors of OS. Conclusions: Foretinib demonstrated promising antitumor activity and good tolerability in the first-line setting in Asian advanced hepatocellular carcinoma patients. Baseline plasma levels of IL6 or IL8 might predict the response to foretinib. Clin Cancer Res; 23(10); 2405–13. ©2016 AACR.

Background: In EGF30008 the addition of lapatinib (La) to first-line treatment with letrozole (Le) significantly improved progression-free survival (PFS) in patients (pts) with hormone receptor positive (HR+), HER2 positive (HER2+) metastatic breast cancer (hazard ratio (HR) = 0.71; p=0.019), but not in HR+, HER2 negative (HER2−) pts. This study assessed the ability of VeriStrat (VS), a mass-spectrometry blood-based pretreatment assay correlating with clinical outcome from EGFR-TKI therapy, to stratify pt outcome in EGF30008. The VS assay assigns VS Good (Good), VS Poor (Poor), or Indeterminate labels to a serum sample based on a specific 8-peak signature in the mass-spectra (Taguchi F et al., JNCI 2007). Methods: Blinded to clinical data, pretreatment serum was analyzed using standard VS procedurei. Statistical analyses were performed using Mantel-Haenszel and Cox proportional hazards methods; correlations were evaluated using Fisher's exact and χ2 tests. Results: Of the 1286 pts randomized (HER2+=219; HER2−=952; HER2 unknown=115), 1163 pts had serum available; a VS label was assigned to 1046 pts (961 Good; 80 Poor; 5 Indeterminate); 117 were not evaluable (hemolyzed). In the overall population there was no significant difference in PFS between Good and Poor groups within the Le+La arm (p=0.53). In contrast, PFS of the Good group was longer than that of the Poor group within the Le only arm (HR=0.36, p In the HER2+ population both Good (n= 169) and Poor (n= 13) groups received significant PFS benefit from addition of La to Le: in Good HR=0.71, p=0.046, medians 3.0 mo (Le) vs 8.0 mo (Le+La); in Poor HR=0.17, p=0.02, medians 2.3 mo (Le) vs 8.6 mo (Le+La). In the HER2− population median PFS for the Poor group (n=58) increased from 3.1 to 11.0 mo (HR=0.57, p=0.068) with addition of La to Le, whereas the Good group (n=702) received no significant benefit, median 13.6 mo on Le and 13.8 mo on Le+La (PFS HR= 0.85, p=0.09). In multivariate analysis VS remained independently significant and predictive of differential treatment effect in the overall population. No significant correlation of VS classification with HER2 status (p=0.35) or prior adjuvant hormonal therapy (p=0.33) was found. VeriStrat was also not significantly correlated with baseline HER2 ECD levels (low Conclusions: In the overall population VeriStrat predicted PFS in Le alone but not in the Le+La combination, with a significant differential treatment effect. VeriStrat identified a subset of pts with inferior PFS on Le therapy. Adding La to Le improved outcome in HER2+ pts, and interestingly suggested an improvement in the VS Poor subset of HER2− pts. HER2− VS Good pts do not gain benefit with the addition of La. No correlation between VS and investigated predictors of benefit from the addition of La to Le in HER2− pts was observed. If these results can be validated in a prospective study, the addition of La to Le may be a potential treatment option for HER2− HR+ VS poor pts. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr S1-4.

BACKGROUND: Changes in serum human epidermal growth factor receptor 2 (HER2) levels associated with clinical outcomes, including objective response rate, progression-free survival (PFS), and overall survival have been reported in patients with metastatic breast cancer (MBC) receiving trastuzumab and chemotherapy. This study investigated whether baseline or changes in serum HER2 correlated with overall response rate (ORR) and/or PFS in patients with MBC receiving first-line lapatinib monotherapy. METHODS: The EGF20009 study investigated lapatinib monotherapy in 138 HER2-positive patients with MBC previously untreated for their metastatic disease. Serum was collected and assessed at baseline and every 4 weeks for 16 weeks after treatment initiation. Disease assessment was performed at weeks 8 and 12 and every 12 weeks thereafter. A ≥20% decrease or increase in serum HER2 was defined as a significant change. RESULTS: Seventy-nine percent of patients had elevated baseline serum HER2. Baseline serum HER2 was associated with ORR (P = .043) but not PFS. Patients with a ≥20% decrease from baseline of serum HER2 at weeks 4, 8, 12, and 16 had a significantly increased ORR and prolonged PFS. Conversely, those with a ≥20% increase from baseline had a significantly lower ORR and shorter PFS. CONCLUSION: Significant decreases in serum HER2 levels during the first 16 weeks of lapatinib monotherapy were associated with better clinical outcome (longer PFS and increased ORR) in HER2-positive MBC patients. Cancer 2011;. © 2011 American Cancer Society.

Background: HER2-positive metastatic breast cancer (MBC) relapsing after trastuzumab-based therapy may require continued HER2 receptor inhibition to control the disease and preserve the patients' quality-of-life. Efficacy and safety of lapatinib monotherapy was evaluated in Japanese breast cancer patients after trastuzumab-based therapies. Methods: In studies, EGF100642 and EGF104911 evaluated the efficacy and safety of oral lapatinib given 1500 mg once daily in patients with advanced or MBC. All patients progressed on anthracyclines and taxanes; HER2-positive patients had also progressed on trastuzumab. Results: For HER2-positive tumours (n=100), objective response rate was 19.0% (95% confidence interval (CI): 11.8–28.1) and clinical benefit rate (CBR) was 25.0% (95% CI: 16.9–34.7). One out of 22 HER2-negative tumour was documented as complete response (n=22). The median time-to-progression (TTP) in the HER2-positive and HER2-negative groups was 13.0 and 8.0 weeks (P=0.007); median overall survival was 58.3 and 40.0 weeks, respectively. The most frequent adverse event was diarrhoea. TTP and CBR were significantly associated with HER2 expression. Patients with tumours harbouring an H1047R PIK3CA mutation or low expression of PTEN derived clinical benefit from lapatinib. Conclusion: Lapatinib monotherapy had shown anti-tumour activity in Japanese patients with HER2-positive MBC that relapsed after trastuzumab-based therapy, including those with brain metastases. Patients benefiting from lapatinib may have biomarker profiles differing from that reported for trastuzumab.

Purpose: Biomarkers from two randomized phase III trials were analyzed to optimize selection of patients for lapatinib therapy. Experimental Design: In available breast cancer tissue from EGF30001 (paclitaxel ± lapatinib in HER-2-negative/unknown metastatic breast cancer, n = 579) and EGF100151 (capecitabine ± lapatinib in HER-2-positive metastatic breast cancer, n = 399), HER-2 gene amplification by fluorescence in situ hybridization (FISH), HER-2 mRNA by reverse transcription-PCR (RT-PCR), HER-2 protein expression by HercepTest immunohistochemistry (IHC), epidermal growth factor receptor (EGFR) mRNA level by RT-PCR, and EGFR protein by IHC were analyzed and compared with clinical outcome. HER-2 was determined by FISH in an academic reference/research laboratory and in a large, high-volume commercial reference laboratory. Results: The HER-2 gene was amplified in 47% (344 of 733) and IHC was 3+ in 35% (279 of 798), with significant correlation (P < 0.01) between FISH and IHC. Positive EGFR immunostaining (IHC 1+, 2+, or 3+) in 28% (213 of 761) correlated with EGFR mRNA levels by RT-PCR (r = 0.59; P < 0.01). HER-2 gene amplification/overexpression was associated with improved clinical outcomes (progression-free survival; P < 0.001) in both trials. A significant improvement in outcome was seen in FISH-positive and IHC 0, 1+, or 2+ patients. HER-2 mRNA expression correlated with HER-2 FISH (r = 0.83) and IHC status (r = 0.72; n = 138). No correlation was found between EGFR expression (IHC or mRNA) and responsiveness to lapatinib regardless of HER-2 status. Although a significant correlation with lapatinib responsiveness was observed among “HER-2-negative” breast cancer patients in the large, high-volume commercial reference laboratory, this was not confirmed in the academic reference/research laboratory. Conclusions: Women with HER-2-positive metastatic breast cancer benefit from lapatinib, whereas women with HER-2-negative metastatic breast cancer derive no incremental benefit from lapatinib.

Lapatinib (GW572016) is a small-molecule dual inhibitor of epidermal growth factor receptor (ErbB1) and ErbB2 receptor kinase activities currently in phase III clinical trials. We used phosphoprotein and microarray analyses to carry out targeted pathway studies of phosphorylation and gene expression changes in human breast cancer cell lines in the presence or absence of lapatinib. Studies were done in four breast cancer cell lines, two of which were responsive and two of which were nonresponsive to lapatinib. Responsive cell lines, BT474 and SKBr3, constitutively overexpress ErbB2 and show an IC50 of 25 or 32 nmol/L for lapatinib, respectively. In contrast, nonresponsive MDA-MB-468 and T47D cells expressed a low basal level of ErbB2 and showed IC50 values in the micromolar range. Cells responsive to lapatinib exhibited strong differential effects on multiple genes in the AKT pathway. After 12 h of exposure to 1.0 μmol/L of lapatinib, AKT1, MAPK9, HSPCA, IRAK1, and CCND1 transcripts were down-regulated 7- to 25-fold in responsive BT474 and SKBr3 cells. In contrast, lapatinib weakly down-regulated the AKT pathway in nonresponsive breast cancer cell lines (

HER2/ERBB2 status is used to select patients for HER2-targeted therapy. HER2/ERBB2 amplification/overexpression of upper gastrointestinal (UGI) adenocarcinomas was determined locally or in two central laboratories to select patients for the TRIO-013/LOGiC trial of chemotherapy with or without lapatinib. Patients selected locally had central laboratory confirmation of HER2 amplification for inclusion in the primary efficacy population. HER2 was assessed with PathVysion or IQ PharmDx FISH and HercepTest immunohistochemistry assays. Associations with outcomes were retrospectively evaluated. Overall, HER2 status was determined in UGI cancers from 4,674 patients in a central laboratory for eligibility (1,995 cases) and for confirmation of local HER2 results (333 cases). Of 1,995 adenocarcinomas screened centrally, 322 (16.1%) had HER2-amplified disease with 29 (1.5%) showing HER2 genomic heterogeneity. Men and older patients had higher rates of amplification. Of 545 patients accrued to the trial (gastric, 87.3%; GEJ, 8.3% and esophageal cancer, 4.4%), 487 patients (89%) were centrally confirmed as having HER2-amplified disease. Concordance between central and local HER2 testing was 83%. Concordance between PathVysion and IQ PharmDx FISH assays was 99% and FISH in the two central laboratories was 95%. Lapatinib-treated Asian participants and those less than 60 years had significant improvement in progression-free survival (PFS), particularly among those whose cancers had 5.01–10.0 and >10.0-fold amplification of HER2. In conclusion, HER2 is commonly amplified in UGI adenocarcinomas with amplification highly correlated to overexpression, and HER2 amplification levels correlated with PFS. While HER2 genomic heterogeneity occurs, its prevalence is low. Mol Cancer Ther; 16(1); 228–38. ©2016 AACR.

In various pharmaceutical applications, repeated measurements are taken from each subject, and model parameters are estimated from the collected data. Examples include dose response modeling and PK/PD studies with serial blood sampling, among others. The quality of the information in an experiment is reflected in the precision of estimates of model parameters, which is traditionally measured by their variance-covariance matrix. In this article, we concentrate on the example of a clinical PK study where multiple blood samples are taken for each enrolled patient, which leads to nonlinear mixed effects regression models with multiple responses. The sampling scheme for each patient is considered a multidimensional point in the space of admissible sampling sequences. We demonstrate how to optimize the precision of parameter estimates by finding the best number and allocation of sampling times. It is shown that a reduced number of samples may be taken without significant loss of precision of parameter estimates. Moreover, our approach allows for taking experimental costs into account, which leads to a more meaningful comparison of sampling schemes and to potential cost savings.

Recently, a benzo-1,2,4-thiadiazine was shown to be a potent, specific inhibitor of the hepatitis C virus (HCV) RNA polymerase [J. Biol. Chem. 277 (2002) 32327]. Herein, we present several lines of evidence to demonstrate that thiadiazine compound 4 (C(21)H(21)N(3)O(4)S) is highly synergistic with interferon-alpha (IFN-alpha) and disrupts HCV replicon RNA synthesis with a distinct kinetic profile. A time course analysis after a single treatment with 5 microM compound 4 showed a loss of viral RNA consistent with replicon RNA half-life, suggesting inhibition of 90% of ongoing or newly initiated replicative intermediates. This finding is consistent with the mechanism of action recently reported for compound 4, an RNA synthesis initiation inhibitor [J. Biol. Chem. 278 (2003) 16602]. Further, unlike IFN-alpha, an immediate reduction of HCV replicon RNA synthesis was apparent upon addition of compound 4. Treatment with IFN-alpha showed a delay of approximately 4h prior to inhibition of viral RNA replication, consistent with its signaling kinetics.

Model fitting when the variance function depends on unknown parameters is a popular problem in many areas of research. Iterated estimators which are asymptotically equivalent to maximum likelihood estimators are proposed and their convergence is discussed. From a computational point of view, these estimators are very close to the iteratively reweighted least-squares methods. The additive structure of the corresponding information matrices allows us to apply convex design theory which leads to optimal design algorithms. We conclude with examples which illustrate how to bridge our general results with specific applied needs. In particular, a model with experimental costs is introduced and is studied within the normalized design paradigm.

To determine the health and functional impact of undiagnosed airflow obstruction for subjects in the general population, we used data obtained as part of the Third National Health and Nutrition Examination Survey (NHANES III). Categories of diagnosed and undiagnosed airflow obstruction were defined using questionnaire responses and spirometric results. Health and functional impact of airflow obstruction was assessed from responses to questions about general health status, walking 1/4 mile, lifting or carrying something as heavy as 10 lb, or needing help with personal care. Undiagnosed airflow obstruction (12.0%) was more common than doctor-diagnosed chronic obstructive pulmonary disease (COPD) (3.1%) or asthma (2.7%). Although undiagnosed airflow obstruction was usually very mild, approximately 5% of the entire sample had an FEV(1) less than 75% predicted. After adjusting for smoking, obesity, and comorbid conditions, the risk of impaired health and functional status with undiagnosed airflow obstruction was independently associated with severity of FEV(1) impairment. For males and females, ever smoking was strongly associated with all types of airflow obstruction, diagnosed or not. However, among females with airflow obstruction, 12.2% to 35.2% never smoked. Undiagnosed airflow obstruction is common in the general population of the United States and is associated with impaired health and functional status.airflow obstruction; spirometry; health impact; screening

Keliximab, a Primatized IgG1 CD4 mAb, was reconfigured to an IgG4 antibody. The gamma4 constant region was further modified by substituting glutamic acid for serine at position 235 in the CH2 domain (IgG4-E), to remove residual binding to Fcgamma receptors, and substitution of serine with proline at position 228 in the hinge region (IgG4-PE) for greater stability. Pharmacokinetic analysis in rats gave a t(1/2) of approximately 4 days for IgG4-E and 9 days for IgG4-PE, consistent with a greater stability of the IgG4-PE molecule. The effects on T cell subsets were assessed in chimpanzees given escalating doses of IgG4-PE: 0.05 mg/kg on Day 16, 1.5 mg/kg dose on Day 43, and 15 mg/kg on Day 85. Receptor modulation was observed at the two highest doses, but no depletion of T cells at any dose. The in vitro and in vivo results demonstrate the potential of this IgG4-PE mAb for use in human trials.

A typical calibration problem in bioassay involves several known concentrations of an analyte, with replicate determinations of the analyte at each level of concentration. A straight-line regression between the replicate determinations of the analyte and the known concentrations is sought. Common practice in this situation is to obtain several determinations of one analyte at each consecutive concentration level, instead of running the experiment in a completely randomized fashion. A proper model for this type of calibration experiment is a simple mixed linear model with two sources of variation: between analyte concentrations, and within analyte concentrations (model A). However, a practitioner may use a model based on the average determination at each concentration of analyte (model B), or helshe may adopt a model that assumes a completely independent error structure (model C) to fit a straight line. The purpose of this article is to compare the analysis based on models B and C to the supposedly...

Purpose The receptors for hepatocyte and vascular endothelial cell growth factors (MET and VEGFR2, respectively) are critical oncogenic mediators in gastric adenocarcinoma. The purpose is to examine the safety and efficacy of foretinib, an oral multikinase inhibitor targeting MET, RON, AXL, TIE-2, and VEGFR2 receptors, for the treatment of metastatic gastric adenocarcinoma. Patients and Methods Foretinib safety and tolerability, and objective response rate (ORR) were evaluated in patients using intermittent (240 mg/day, for 5 days every 2 weeks) or daily (80 mg/day) dosing schedules. Thirty evaluable patients were required to achieve alpha = 0.10 and beta = 0.2 to test the alternative hypothesis that single-agent foretinib would result in an ORR of ≥25%. Up to 10 additional patients could be enrolled to ensure at least eight with MET amplification. Correlative studies included tumor MET amplification, MET signaling, pharmacokinetics and plasma biomarkers of foretinib activity. Results From March 2007 until October 2009, 74 patients were enrolled; 74% male; median age, 61 years (range, 25–88); 93% had received prior therapy. Best response was stable disease (SD) in 10 (23%) patients receiving intermittent dosing and five (20%) receiving daily dosing; SD duration was 1.9–7.2 months (median 3.2 months). Of 67 patients with tumor samples, 3 had MET amplification, one of whom had SD. Treatment-related adverse events occurred in 91% of patients. Rates of hypertension (35% vs. 15%) and elevated aspartate aminotransferase (23% vs. 8%) were higher with intermittent dosing. In both patients with high baseline tumor phospho-MET (pMET), the pMET:total MET protein ratio decreased with foretinib treatment. Conclusion These results indicate that few gastric carcinomas are driven solely by MET and VEGFR2, and underscore the diverse molecular oncogenesis of this disease. Despite evidence of MET inhibition by foretinib, single-agent foretinib lacked efficacy in unselected patients with metastatic gastric cancer. Trial Registration ClinicalTrials.gov NCT00725712

Purpose: Dabrafenib is a selective inhibitor of V600-mutant BRAF kinase, which recently showed improved progression-free survival (PFS) as compared with dacarbazine, in metastatic melanoma patients. This study examined potential genetic markers associated with response and PFS in the phase I study of dabrafenib. Experimental Design: Baseline (pretreatment or archival) melanoma samples were evaluated in 41 patients using a custom genotyping melanoma-specific assay, sequencing of PTEN, and copy number analysis using multiplex ligation amplification and array-based comparative genomic hybridization. Nine patients had on-treatment and/or progression samples available. Results: All baseline patient samples had BRAFV600E/K confirmed. Baseline PTEN loss/mutation was not associated with best overall response to dabrafenib, but it showed a trend for shorter median PFS [18.3 (95% confidence interval, CI, 9.1–24.3) vs. 32.1 weeks (95% CI, 24.1–33), P = 0.059]. Higher copy number of CCND1 (P = 0.009) and lower copy number of CDKN2A (P = 0.012) at baseline were significantly associated with decreased PFS. Although no melanomas had high-level amplification of BRAF, the two patients with progressive disease as their best response had BRAF copy gain in their tumors. Conclusions: Copy number changes in CDKN2A, CCND1, and mutation/copy number changes in PTEN correlated with the duration of PFS in patients treated with dabrafenib. The results suggest that these markers should be considered in the design and interpretation of future trials with selective BRAF inhibitors in advanced melanoma patients. Clin Cancer Res; 19(17); 4868–78. ©2013 AACR.

Importance The value of the intrinsic subtypes of breast cancer (luminal A, luminal B, human epidermal growth factor receptor 2 [currently known as ERBB2, but referred to as HER2 in this study]-enriched, and basal-like) in the metastatic setting is currently unknown. Objective To evaluate the association of the intrinsic subtypes of breast cancer with outcome and/or benefit in hormone receptor (HR)-positive metastatic breast cancer. Design, Setting, and Participants Unplanned retrospective analysis of 821 tumor samples (85.7% primary and 14.3% metastatic) from theEGF30008phase 3 clinical trial (NCT00073528), in which postmenopausal women with HR-positive invasive breast cancer and no prior therapy for advanced or metastatic disease were randomized to letrozole with or without lapatinib, an epidermal growth factor receptor (EGFR)/HER2 tyrosine kinase inhibitor. Tumor samples were classified into each subtype using the research-based PAM50 classifier. Prior neoadjuvant/adjuvant antiestrogen therapy was allowed. Patients with extensive symptomatic visceral disease were excluded. Treatment effects were evaluated using interaction tests. Main Outcomes and Measures Primary and secondary end points were progression-free survival and overall survival. Results The median (range) age was 62 (31-94) years. Intrinsic subtype was the strongest prognostic factor independently associated with progression-free survival and overall survival in all patients, and in patients with HER2-negative (n = 644) or HER2-positive (n = 157) diseases. Median progression-free survival differed across the intrinsic subtypes of clinically HER2-negative disease: luminal A (16.9 [95% CI, 14.1-19.9] months), luminal B (11.0 [95% CI, 9.6-13.6] months), HER2-enriched (4.7 [95% CI, 2.7-10.8] months), and basal-like (4.1 [95% CI, 2.5-13.8] months). Median OS also differed across the intrinsic subtypes: luminal A (45 [95% CI, 41-not applicable {NA}] months), luminal B (37 [95% CI, 31-42] months), HER2-enriched (16 [95% CI, 10-NA] months), and basal-like (23 [95% CI, 12-NA] months). Patients with HER2-negative/HER2-enriched disease benefited from lapatinib therapy (median PFS, 6.49 vs 2.60 months; progression-free survival hazard ratio, 0.238 [95% CI, 0.066-0.863]; interactionP = .02). Conclusions and Relevance This is the first study to reveal an association between intrinsic subtype and outcome in first-line HR-positive metastatic breast cancer. Patients with HR-positive/HER2-negative disease with a HER2-enriched profile may benefit from lapatinib in combination with endocrine therapy. The clinical value of intrinsic subtyping in hormone receptor–positive metastatic breast cancer warrants further investigation, but patients with luminal A/HER2-negative metastatic breast cancer might be good candidates for letrozole monotherapy in the first-line setting regardless of visceral disease and number of metastases.

Purpose: A subgroup of human epidermal growth factor receptor 2 (HER2)–overexpressing breast tumors coexpresses p95HER2, a truncated HER2 receptor that retains a highly functional HER2 kinase domain but lacks the extracellular domain and results in intrinsic trastuzumab resistance. We hypothesized that lapatinib, a HER2 tyrosine kinase inhibitor, would be active in these tumors. We have studied the correlation between p95HER2 expression and response to lapatinib, both in preclinical models and in the clinical setting. Experimental Design: Two different p95HER2 animal models were used for preclinical studies. Expression of p95HER2 was analyzed in HER2-overexpressing breast primary tumors from a first-line lapatinib monotherapy study (EGF20009) and a second-line lapatinib in combination with capecitabine study (EGF100151). p95HER2 expression was correlated with overall response rate (complete + partial response), clinical benefit rate (complete response + partial response + stable disease ≥24 wk), and progression-free survival using logistic regression and Cox proportional hazard models. Results: Lapatinib inhibited tumor growth and the HER2 downstream signaling of p95HER2-expressing tumors. A total of 68 and 156 tumors from studies EGF20009 and EGF100151 were evaluable, respectively, for p95HER2 detection. The percentage of p95HER2-positive patients was 20.5% in the EGF20009 study and 28.5% in the EGF100151 study. In both studies, there was no statistically significant difference in progression-free survival, clinical benefit rate, and overall response rate between p95HER2-positive and p95HER2-negative tumors. Conclusions: Lapatinib as a monotherapy or in combination with capecitabine seems to be equally effective in patients with p95HER2-positive and p95HER2-negative HER2-positive breast tumors. Clin Cancer Res; 16(9); 2688–95. ©2010 AACR.

Purpose The HER2 extracellular domain (ECD) is enzymatically cleaved from the cell membrane. Shed ECD in serum has been studied as both prognostic and predictive markers. Lapatinib is a dual inhibitor of HER2 and epidermal growth factor receptor kinases. We examined the prognostic and predictive role of HER2 ECD in a randomized trial of paclitaxel with placebo or lapatinib in women with HER2-negative or -unknown breast cancer. Patients and Methods Patients (n = 579) with newly diagnosed metastatic breast cancer (MBC) were randomly assigned to paclitaxel with placebo or lapatinib. HER2 status was determined centrally. ECD was centrally measured by enzyme linked immunoassay in available samples at baseline (b; n = 472), week 9, and every 12 weeks thereafter. Results were correlated to overall response rate (ORR) and progression-free survival (PFS). Results Elevated baseline ECD (bECD) levels (≥ 16 ng/mL) did not predict HER2 tumor status (sensitivity, 62%; specificity, 75%). In HER2-negative tumors, elevated bECD was not correlated with improved efficacy for lapatinib plus paclitaxel versus placebo plus paclitaxel (ORR: odds ratio, 1.6; 95% CI, 0.1 to 3.8; P = .365; PFS: hazard ratio, 0.94; 95% CI, 0.60 to 1.47; P = .797). ECD levels tended to decrease over time when bECD was elevated. ECD conversion from low to high was associated with worse PFS. Converting from high to low was associated with a better PFS. A consistently low ECD level had better PFS than a consistently elevated ECD. All associations were found to be independent of lapatinib. Conclusion HER2 bECD does not predict lapatinib benefit in patients with HER2-negative MBC. Changes in ECD status correlates with patient outcome regardless of treatment given. Measuring HER2 ECD is not currently recommended for predicting benefit to lapatinib.

501 Background: Reports have demonstrated an inverse relationship between suppression of immune surveillance mechanisms and activation of the vascular endothelial growth factor receptor (VEGFR) pathway suggesting that T cell repertoires may impact response to VEGFR blockade. We evaluated the association between clinical outcomes and T cell repertoire in metastatic RCC patients receiving a front-line anti-VEGFR, pazopanib. Methods: Pre-treatment RCC tumors were analyzed from VEG105192, a phase III study of mRCC patients randomized (2:1) to pazopanib (paz) 800 mg daily vs placebo (pbo) for TCR gamma (TCRG) and TCR beta (TCRB) CDR3 regions. Using the Adaptive Biotechnologies immunoSEQ Assay, we assessed TCR clonality, a measure of total repertoire represented by expanded clones, and entropy, a measure of evenness and diversity. PD-L1 was evaluated by immunohistochemistry (IHC) H-Scores. The goal of the study was to determine if the repertoire of T cell clones was associated with progression-free survival as a clinical endpoint. Results: In the cohort with available tissue, the median PFS was 10.8 and 5.5 months (mos) for paz and pbo, respectively. TCRB (n = 114) and TCRG (n = 43 pbo, 109 paz) clonality ranged from 0-0.31 and 0-0.98, and entropy from 1-12.1 and 0-10.37, respectively. TCRB and TCRG entropy were highly correlated (Spearman’s R = 0.92, n = 114). Samples from the pbo-treated group with higher TCRG entropy, defined as the top 25th percentile, were associated with an improved median PFS (12.8 months) when compared to the lower 75th percentile (3.1 months, P = 0.023); similar trends were seen for TCRB entropy. Neither entropy nor clonality was associated with maximal reduction in tumor volume in the paz-treated group. PD-L1 H-scores were not associated with entropy or clonality (P > 0.05). Conclusions: Our data suggests that RCC samples with higher entropy are associated with a favorable prognosis. Identification of tumors with restricted TCRB/G chain usage and less diverse repertoire, as represented by lower entropy and higher clonality, may impact responses to VEGFR blockade and requires further study. Clinical trial information: NCT00334282.

Microbiologists are conditioned to approach a scientific subject with a hypothesis, a protocol outlining how to proceed, and a clear idea of what they are looking for. Data mining is a relatively new concept to microbiologists, and requires a change in mind set,as a key element of this approach is to apply methods developed for other fields like statistics and bioinformatics to a search for novel facts within the accumulated data.The papers cited in Section 2 of this chapter are included here to encourage workers interested in the subject of antimicrobial resistance to approach their subject within a new paradigm. The three general methods presented in some detail, antibiotypes, multivariate analysis, and evolutionary genetics,are techniques designed to stimulate the investigator rather than present any one set approach to the subject. Interested parties are encouraged to take the first step, namely, search for colleagues with expertise in other fields to become familiar with the rich data generated by antimicrobial surveillance and other research programs from the viewpoint of their individual specialities and search for novel aspects that will shed light on a problem that will only become more critical over the coming years. Data mining is one approach that may offer novel insights into our understanding of resistance, and ultimately may result in providing potential solutions to slow the rate of resistance against organisms of medical and environmental importance.

Traditionally, large surveillance studies have been analyzed by the use of the MICs at which 90% of isolates tested are inhibited (MIC 90 s), MIC 50 s, frequency distributions, and percent susceptibility. In the past, these approaches have proved satisfactory for the monitoring of resistance. From these traditional uses, one can readily detect an increase in MICs for organism and drug combinations. Now that large surveillance studies have been conducted for a number of years and databases have grown to include a large number of datum points, new approaches to the extraction of useful information from these studies are needed. The present study proposes approaches, including the use of antibiotypes, principal components analysis, phylogenetics, and population genetic analysis, to the evaluation of data from large multinational surveillance studies. Application of these types of analyses can be used to describe genetic diversity, analyze changes in susceptibility patterns over time, and possibly, shed light on the origins and evolution of antimicrobial resistance. As global surveillance studies become more common and new questions concerning the evolution of resistance are raised, innovative approaches to analysis of the data will increase in importance.

4582 Background: No biomarker has been incorporated into clinically based prog algorithms for mRCC. In previous work in pts with ECOG PS ≤1 treated in a randomized placebo-controlled phase III tria...

e20019 Background: To investigate the previously reported association between baseline HGF and clinical response in patients (pts) treated with RAF inhibitors, HGF levels were assessed in pt blood ...

4072^ Background: Patient inclusion to the primary efficacy population of TRIO-013/LOGiC trial of lapatinib in combination with chemotherapy in advanced HER2-positive UGI adenocarcinomas required e...

5501 Background: BRCA mutational status, in combination with the accumulation of somatic mutations, has been associated with ovarian cancer prog and response to chemotherapy. AGO-OVAR16/VEG110655 i...

416 Background: The interaction of PDL1 (B7H1) with its receptor PD-1 on activated T cells contributes to suppression of antitumor immune responses. Tumor PDL1 expression has been associated with poor outcomes in RCC but has not been investigated as a biomarker of response in RCC patients receiving standard vascular endothelial growth factor (VEGF)-targeted therapy. Methods: Formalin-fixed paraffin-embedded tumor samples were collected at baseline from consenting patients enrolled in COMPARZ—a phase lll clinical trial comparing pazopanib and sunitinib as first-line interventions for metastatic RCC (Motzer et al, NEJM 2013). PDL1 expression was evaluated using the anti-PDL1 mouseIgG1 (clone 5H1; Thompson) on the Leica automated immunohistochemistry platform. Additional dual PDL1/CD68 staining was performed on tumor associated macrophages (TAMs). Tumor PDL1 expression was quantified by an H-score (HS). PDL1+ TAMs were assessed semiquantitatively. In addition, intra-tumor CD8+ T cells were quantified morphometrically. The association between PDL1 HS, CD8+T cell counts, and survival was investigated using Kaplan-Meier analysis. Results: HS data were available from 453 of 1110 patients. 64% of patients had negative (HS = 0) PDL1 expression (HS range 0-290), but PDL1 expression was associated with tumours containing higher numbers of infiltrating macrophages. Peripheral CD8+ T cells in the invasive margin surrounding the tumor were also observed. Patients with HS >50 (n = 61, 13%) had significantly shorter overall survival (OS) in both pazopanib (19.7 vs 31.6 mo) and sunitinib (15.3 vs 27.7 mo) arms. In both arms, patients with HS >50 with intratumoral CD8+T cell counts >300 had the shortest OS. Results were similar for progression-free survival and persisted on multivariate analysis. Conclusions: This is the largest report to show that tumors’PDL1 expression and CD8+ T cell counts are associated with treatment outcome in metastatic RCC patients. Increased PDL1, or increased PDL1 plus tumor CD8+ T cell counts, were associated with shorter OS. These findings may have major implications for future trial designs that involve PD-1 inhibitors.

3021 Background: The interaction between PDL1 (B7H1) and its receptor PD-1 on activated T cells plays an important role in the inhibition of T-cell responses and contributes to suppression of antitumor immune responses. Tumor PDL1 expression has been associated with poor outcomes in RCC. This study investigates the correlation between PDL1 tumor expression and outcomes in RCC pts treated with paz. Methods: Using IHC, we retrospectively analyzed baseline FFPE tumor samples for PDL1 from 2 paz RCC studies: a single arm phase II trial and randomized placebo (pbo)-controlled phase III study. PDL1 expression was analyzed by MedTox Laboratories using the anti-PDL1 MouseIgG1 clone 5H1 (Thompson) on the Leica automated IHC platform. Additional dual PDL1/CD68 staining was carried out to delineate tumor and macrophage PDL1 expression. Tumor PDL1 expression was quantified by H-Score (HS) and PDL1+ macrophages were assessed semi-quantitatively. Association between PDL1H scores and PFS was investigated by Kaplan-Meier analysis using optimal cutoff of PDL1tumor HS (minimum p value, log rank test). Results: The optimal cut-point of PD-L1 tumor HS, relative to PFS, was identified as HS > 3. In the phase II study (46 available samples out of 225), HS range was 0-150 and most samples had negative (HS = 0, n = 34, 74%) or low (HS 1-3, n=4, 9%) PDL1 expression. Pts with HS > 3 (n = 8, 17%) had significantly shorter PFS (2.6 mo) than those with HS ≤ 3 (12 mo; p = .0005). In the phase III study (N = 160 available samples: paz, 113 of 290; pbo, 47 of 145), HS range was 0-280. Most patients had negative (n = 122/160, 76%) or low (n = 9/160, 6%) PD-L1 expression, with 18% (29/160) having HS > 3. Pbo-arm pts with HS > 3 (n = 6/47, 13%) had shorter PFS (2.3 vs 5.5 mo p = .0207). Paz-arm pts with HS > 3 (n = 23/113, 20%) trended toward shorter PFS (7.3 vs 11 mo, p = .1405). Conclusions: PDL1 appears to be a prognostic marker with PDL1 HS > 3 associated with shorter PFS. Limitations of the study include the retrospective nature of the analysis with limited pt samples available, low or negative PDL1 expression in the vast majority of pts, and use of archival samples that may not accurately reflect PDL1 status at study entry. Additional results (tumor volume, OS) will be presented.

8518 Background: Tumor specific cfDNA levels in blood increase with tumor burden and decrease following treatment. cfDNA can harbor muts consistent with the tumor. Thus cfDNA could be a useful biomarker of therapeutic response. BREAK-2, an open label, single arm, Phase II study evaluated efficacy, safety and tolerability of dabrafenib in BRAF V600E/K mut+ metastatic melanoma pts. Exploratory objectives of BREAK-2 were to evaluate whether (i) tumor and cfDNA BRAF muts are correlated; (ii) cfDNA levels correlate with baseline tumor burden and (iii) cfDNA muts predict clinical outcome with dabrafenib. Methods: BRAF mut status was established for 92pts using an allele-specific PCR assay in tumor samples. Baseline plasma samples were available for 91/92 pts. cfDNA BRAF mut status was evaluated by Inostics GmBH using the BEAMing technology. Spearman correlation coefficients (R) were used to determine the association between cfDNA fraction (mut DNA molecules > 0.01%) and estimated baseline tumor burden, calculated by the sum of RECIST measurements of target lesions. Logistic regression and Cox proportional hazards models were used to assess the association between cfDNA mut status and objective response rate (ORR) and progression free survival (PFS), respectively. Results: The overall agreement between tumor and cfDNA BRAF V600E and V600K mut status was 83%, and 96% respectively. Higher cfDNA V600E mut fraction was associated with higher baseline tumor burden (R=0.73; p-value < 0.0001; n=60); lower ORR (O.R. = 0.83; 95% CI = 0.72, 0.96; p-value=0.0134; n=46) and shorter PFS (H.R.=1.09; p-value=0.0006; n=46). Median PFS was 27.4 weeks in the overall V600E pt population (n=76) and 20.0 weeks in the cfDNA V600E pt population (n=46). Otherwise, the response endpoints were comparable between the two populations. There was no correlation between V600K mut fraction (n=14) and any efficacy endpoints. Conclusions: cfDNA was useful for detecting BRAF muts in pts treated with dabrafenib and increasing V600E mut fraction was associated with reduced ORR and shorter PFS, suggesting higher amounts of mut cfDNA in V600E mut+ pts predicts poorer clinical outcome.

408 Background: Microarray and clinical data from a randomized phase II RCC clinical trial were evaluated to examine the HIF1α kidney cancer gene signature developed from cell lines by Shen et al (Cancer Discovery 2011). Methods: Archival FFPE tumor samples were collected from patients (pts) enrolled in a Phase II RCC study of pazopanib (Hutson, JCO 2009). RNA was isolated and microarray analysis was performed at Response Genetics, Inc. (Los Angeles, CA) using the Affymetrix (Affy) U133 plus Array. Affy data were normalized using the MAS5 algorithm. Genes (n=69) identified from experiments in 10 cancer cell lines were mapped to genes on the U133 array. Patterns of expression (exp) among the mapped genes were assessed using multivariate analyses. Putative patterns were assessed for association with HIF1α gene exp levels, and both patterns and HIF1α levels were assessed for association with response rate (RR) (RECIST), and PFS, using nonparametric and Kaplan-Meier analyses. Gene set enrichment analysis (GSEA) was applied to determine if mapped genes were enriched for HIF1α gene exp. Results: RNA was available for 46 of 225 patients; baseline characteristics were similar to the overall study. 41/46 patients had VHL heterozygous mutations; 5/46 patients were VHL wild type. HIF1a gene expression was detected in all 46 patients. 64/69 genes were mapped to the Affy platform. For genes which mapped to multiple probes, the probe closest to the 3` end was used. A potential 4 cluster pattern was identified. These clusters were associated with HIF1α gene exp, but neither clusters nor HIF1α gene exp were associated with clinical response to pazopanib. Conclusions: The observed pattern of gene exp among 46 pts with available data from a Phase II RCC study of pazopanib is consistent with the cell line derived gene set proposed by Shen et al. Patterns are associated with HIF1α exp; neither patterns nor HIF1α exp are associated with clinical response in this small subset of pts. These data support the on-going efforts in bench-to-bedside research.

404 Background: Understanding the molecular basis of ccRCC heterogeneity could optimize the use of targeted therapies. Using consensus clustering and logical analysis of data (LAD) of gene expression data (Agilent), Brannon et al (Genes Cancer 2010) identified two distinct subtypes of ccRCC [ccA & ccB] defined by a predictive gene set. Patients (pts) with ccA tumors had better disease-specific survival vs. ccB (8.6 vs 2.0 years, P = 0.002). We evaluated these results in an independent dataset. Methods: Gene expression analysis of archival FFPE samples obtained from pts in a randomized phase II trial of pazopanib in ccRCC (Hutson, JCO 2009) were done using Affy U133 plus chips. Corresponding probe sets from the Agilent and Affy platform were identified using Unigene ID’s (DAVID alignment tool). LAD was applied; pts were stratified into ccA/ccB. Kaplan-Meier analysis was used to compare progression-free survival (PFS) for pts stratified by ccA/ccB. Results: RNA was available for 46 pts. Of 110 genes, 108 were mapped to the Affy platform (probe closest to 3` was used). The 108 Affy genes were assessed for signal (present/absent calls) across the 46 samples. 74/108 genes were present in at least 20% (9/46) of pts, 96/108 genes were present in at least 1/46, and 12/108 were not present (0/46). No obvious patterns of expression were evident from heat maps of the 74 or 96 genes. LAD scoring was problematic due to loss of information imposed by using genes present. Of the total LAD patterns in the Brannon paper, 59% or 78% were available using 74 or 96 genes present. PFS based on ccA/ccB were not statistically distinct using either set of present genes. Conclusions: FFPE samples are routinely available from pts with ccRCC and could be used for gene expression studies which could inform clinical practice. Using the Affy platform and FFPE-RNA samples, stratification of ccRCC pts into ccA/ccB subtypes did not reveal PFS differences. While the analysis was confounded by differences in cross-study platform performance, results suggest that further investigation will be necessary to identify a robust set of genes that separates ccRCC into prognostic categories.

8501 Background: GSK436 is a highly potent, selective ATP-competitive BRAF inhibitor. BRF112680, a first in human study, assessed safety, pharmacokinetics, pharmacodynamics, and efficacy. Efficacy ...

This study was designed to evaluate the cardioprotective effects of a solubilized human complement receptor, sCR1, in the rat subjected to myocardial infarction. Following coronary artery occlusion for 0.5 h and reperfusion for 24 h (MI/R group), myocardial infarct size (determined by planimetric analysis) was 18.3 +/- 2.1% of the left ventricle (n = 16), while myeloperoxidase activity (a biochemical marker of neutrophil activation) was increased from 0.94 +/- 0.09 U/g tissue in the sham occluded + vehicle group to 2.96 +/- 0.17 U/g tissue in the MI/R + vehicle treated group (P0.01). Injection of sCR1 (5 mg/kg i.v., 5 min prior to coronary artery occlusion) produced plasma concentrations of 154 +/- 4 microgram/ml 1 min prior to coronary artery occlusion, and concentrations of 86 +/- 2 and 58 +/- 3 micrograms/ml at 40 min and 125 min after dosing (n = 6). sCR1 reduced myocardial infarct size to 11.3 +/- 2.2% of the left ventricle, and attenuated the increase in myeloperoxidase activity to 2.11 +/- 0.20 U/g tissue (n = 18; P0.01, compared to the MI/R + vehicle group). Administration of sCR1 5 min prior to reperfusion afforded a 25.3% non-significant reduction in myocardial injury. These results suggest a beneficial effect of sCR1 in myocardial ischemia/reperfusion injury by reducing the infiltration of neutrophils and attenuating the extent of myocardial injury.

10572 Background: L is a reversible, oral small molecule inhibitor of EGFR and HER2. L demonstrated a clinical overall response rate of 39%, median progression free survival (PFS) of 14.6 weeks and...

1036 Background: Improvement in progression-free (PFS: HR: 0.73; p=0.008) and overall survival (OS: HR: 0.74; p=0.026) was demonstrated with the combination of lapatinib (L) plus trastuzumab (T) in...