Gabi Tarcic

Phase IB study of vemurafenib in combination with irinotecan and cetuximab in patients with metastatic colorectal cancer with BRAFV600E mutation

In vitro, EGFR inhibition, combined with the BRAF inhibitor vemurafenib, causes synergistic cytotoxicity for BRAFV600E metastatic colorectal cancer, further augmented by irinotecan. The safety and efficacy of vemurafenib, irinotecan, and cetuximab in BRAF-mutated malignancies are not defined. In this 3+3 phase I study, patients with BRAFV600E-advanced solid cancers received cetuximab and irinotecan with escalating doses of vemurafenib. Nineteen patients (18 with metastatic colorectal cancer and 1 with appendiceal cancer) were enrolled. Three patients experienced dose-limiting toxicities. The MTD of vemurafenib was 960 mg twice daily. Six of 17 evaluable patients (35%) achieved a radiographic response by Response Evaluation Criteria in Solid Tumors 1.1 criteria, consistent with in vivo models demonstrating tumor regressions with the triplet regimen. Median progression-free survival was 7.7 months. BRAFV600E circulating cell-free DNA (cfDNA) trends correlated with radiographic changes, and acquired mutations from cfDNA in genes reactivating MAPK signaling were observed at progression. SIGNIFICANCE Vemurafenib, in combination with irinotecan and cetuximab, was well tolerated in patients with refractory, BRAF-mutated metastatic colorectal cancer, and both survival outcomes and response rates exceeded prior reports for vemurafenib and for irinotecan plus cetuximab in BRAFV600E metastatic colorectal cancer. In vivo models demonstrated regressions with the triplet, in contrast with vemurafenib and cetuximab alone. cfDNA predicted radiographic response and identified mutations reactivating the MAPK pathway upon progression. Cancer Discov; 6(12); 1352-65. ©2016 AACR.This article is highlighted in the In This Issue feature, p. 1293.

A Novel High-Throughput Multispectral Cell Segmentation Algorithm

An increasingly common component of molecular diagnostics is the analysis of protein localization at the single-cell level using fluorescent microscopy. Manually extracting quantitative data from a large population of cells is unreasonably time-consuming and existing automatic systems are rather limited.

Revisited analysis of a SHIVA01 trial cohort using functional mutational analyses successfully predicted treatment outcome

It still remains to be demonstrated that using molecular profiling to guide therapy improves patient outcome in oncology. Classification of somatic variants is not straightforward, rendering treatment decisions based on variants with unknown significance (VUS) hard to implement. The oncogenic activity of VUS and mutations identified in 12 patients treated with molecularly targeted agents (MTAs) in the frame of SHIVA01 trial was assessed using Functional Annotation for Cancer Treatment (FACT). MTA response prediction was measured in vitro, blinded to the actual clinical trial results, and survival predictions according to FACT were correlated with the actual PFS of SHIVA01 patients. Patients with positive prediction had a median PFS of 5.8 months versus 1.7 months in patients with negative prediction (P < 0.05). Our results highlight the role of the functional interpretation of molecular profiles to predict MTA response.

Abstract LB-041: Novel platform to profile variants of uncertain significant (VUS) with preliminary retrospective clinical validation in non-small cell lung cancer (NSCLC)

Background: With the expanding usage of next generation sequencing (NGS) during treatment of NSCLC, an increasing number of variants of uncertain significance are being detected. Therefore, exists an urgent unmet need for a functional assay to reveal the significance of these findings both for naive patients and those with acquired resistance to targeted therapy. We present feasibility data and retrospective clinical validation for a novel high-throughput cell-based assay (FACT) which profiles alterations by measuring downstream activation of signaling pathways to determine oncogenic grading and prioritize treatment with targeted therapies. Methods: Patients with NSCLC adenocarcinoma that underwent NGS test (tissue based/cell-free tumor DNA) were consented and enrolled. Alterations were profiled using the FACT platform to quantify oncogenic signaling activity and inhibition by targeted therapies. Patient outcomes were compared to in vitro drug sensitivities retrospectively. Results: 9 patients with NSCLC were analyzed using the FACT platform. 17 alterations were functionally profiled (EGFR - 12, ERBB2 - 2, BRAF - 1, PDGFRA - 1 and JAK2 - 1). 12 (70%) mutations were found to be oncogenic, 11 (65%) oncogenic alterations were in actionable genes (EGFR/ERBB2), 7 patients (78%) were treated with targeted therapies. 3 patients had uncommon double point mutations in EGFR profiled on the same DNA construct (cis) and separately (trans) showing distinct oncogenic profiles. Drug inhibition was measured with used targeted therapies and showed concordance with retrospective clinical responses. Conclusions: Functional profiling of alterations found in NSCLC is feasible through the FACT platform and can provide useful insights on the oncogenicity of uncommon variants. Profiling of drug sensitivity was successfully performed in the majority of patients and in vitro sensitivity correlated with observed clinical outcomes. Citation Format: Benjamin Miron, Gabi Tarcic, Oded Edelheit, Nitza Burck, Michael Vidne, Laila Roisman, Nir Peled. Novel platform to profile variants of uncertain significant (VUS) with preliminary retrospective clinical validation in non-small cell lung cancer (NSCLC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr LB-041. doi:10.1158/1538-7445.AM2017-LB-041

64P Identification of the functional significance of mutations using the novel Precision Cancer Analysis System

C. Lazzari1, N. Karachaliou2, A. Verlicchi3, I. Chaib4, S. Marin4, A. Gkountakos5, S. Pilotto6, J.L. Ramı́rez Serrano4, F. de Marinis1, R. Rosell7. 1 Divisione di Oncologica Toracica, Istituto Europeo di Oncologia – IEO, Milan, Italy , 2 Oncology, IOR, Quirón-Dexeus University Institute, Barcelona, Spain, 3 Oncology, Ospedale Sta Maria delle Croci, Ravenna, Italy, 4 Laboratory of Cellular and Molecular Biology, Catalan Institute of Oncology (ICO Badalona), Hospital Germans Trias i Pujol, Badalona, Spain, 5 Laboratory of Cellular and Molecular Biology, University of Crete, School of Medicine, Heraklion, Greece, 6 Department of Medical Oncology, University of Verona, Azienda Ospedaliera Universitaria Integrata, Verona, Italy, 7Medical Oncology Service, Catalan Institute of Oncology (ICO Badalona), Hospital Germans Trias i Pujol, Badalona, Spain

Abstract 1379: Identification of the functional significance of mutations using the novel precision cancer analysis system

Mounting evidence indicates that growth of pathologically identical cancers in each individual patient is fueled by different sets of driving mutations. The need to identify these drivers stems from the recognized necessity for tailoring therapy and scheduling future surveillance. This personalized medical approach has been shown to result in better treatment outcomes. We present a novel Precision Cancer Analysis system (PCAS) capable of identifying activated signaling pathways by means of a transfected cell-based fluorescent reporter assay yielding a quantitative output of particular pathway activation levels. Being a functional platform PCAS reveals activated pathways regardless of the type of mutation behind it, i.e. whether it is already a known mutation or a variant of unknown significance (VUS) mutation. 20 cancer patients were sequenced for a panel of 37 genes and analyzed by the PCAS. This system quantifies oncogenic activity in the majority of the oncogenic signaling pathways altered by the patients’ mutations through a functional assay and does not rely on prior knowledge of the mutations. The system produces a quantitative output enabling grading the different mutants of the same patient, providing prioritization for better drug selection. In 3 tested genes, 16 different mutations were identified- 4 in EGFR, 4 in PIK3CA and 8 in KRAS. Of these 10 were classified as known mutations for which functional annotation exists, and 6 were VUS. In addition to correctly annotating all known mutations, the PCAS further quantified oncogenic activity in all the VUS tested. Measuring the functional mechanism behind known mutations and VUS provides another layer of critical information to the physician. These results clearly demonstrate the value of a functional assay in accurately identifying the optimal course of treatment, particularly by its ability to add actionable information to VUS. The study produced a comprehensive delineation of the oncogenic activity of each patient9s individual mutations demonstrating the ability of the PCAS to 1) accurately deliver comparable actionable information as found by NGS, 2) functionally characterize mutations annotated as VUS, and 3) monitor oncogenic activity of signaling pathways induced by different mutations and mutation-combinations enabling informed treatment decisions. Citation Format: Gabi Tarcic, Nir Peled, Zohar Barbash, Naama Barabash-Katzir, Shlomo Yaakobi, Naama Barabash-Katzir, Hani Nevo, Michael Vidne, Mariusz Adamek, Mordechai R. Kramer, Nikolai Goncharenko, Yakov Fellig, Karen Meir, Keith Mostov, Yoram Altschuler. Identification of the functional significance of mutations using the novel precision cancer analysis system. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1379.