XML file - 99K, Kinase activity and selectivity for NVP-BGJ398; CCLE cell lines sensitive to NVP-BGJ398; GeneSet expression signatures; FGFR genetic alterations and concomitant mutations

PDF file - 1.4MB, Response to NVP-BGJ398; FGFR pathway modulation by NVP-BGJ398; NVP-BGJ398 - predictive features; FGFR2-c3 mRNA expression in cell lines and FGFR2 DNA copy number in tumors; FGF19 DNA copy number in CCLE cell lines and FGFR3 DNA copy number versus transcript expression in bladder cancer cell lines

XML file - 56K, High throughput pharmacological screen of BGJ398 in 517 cancer cell lines

Large panels of comprehensively characterized human cancer models, including the Cancer Cell Line Encyclopedia (CCLE), have provided a rigorous framework with which to study genetic variants, candidate targets, and small-molecule and biological therapeutics and to identify new marker-driven cancer dependencies. To improve our understanding of the molecular features that contribute to cancer phenotypes, including drug responses, here we have expanded the characterizations of cancer cell lines to include genetic, RNA splicing, DNA methylation, histone H3 modification, microRNA expression and reverse-phase protein array data for 1,072 cell lines from individuals of various lineages and ethnicities. Integration of these data with functional characterizations such as drug-sensitivity, short hairpin RNA knockdown and CRISPR-Cas9 knockout data reveals potential targets for cancer drugs and associated biomarkers. Together, this dataset and an accompanying public data portal provide a resource for the acceleration of cancer research using model cancer cell lines.

The Cancer Cell Line Encyclopedia presents the first results from a large-scale screen of some 947 cancer cell lines with 24 anticancer drugs, with the aim of identifying specific genomic alterations and gene expression profiles associated with selective sensitivity or resistance to potential therapeutic agents. Cancer cell lines are widely used as preclinical models to gain mechanistic and therapeutic insight. Two manuscripts in this issue describe the large-scale genetic and pharmacological characterization of human cancer cell lines. Each group characterized collections of several-hundred cell lines using different platforms and analytical methods. Their results are complementary, and confirm that many human cell lines capture the genomic diversity of their respective cancers. Initial findings include the identification of a number of potential markers of drug sensitivity and resistance. For example, Garnett et al. report an association between EWS-FLI1 gene translocations, frequently found in Ewing's sarcoma, and sensitivity to PARP inhibitors, a class of drug currently in clinical trials for other cancer types. Barretina et al. report a possible association between SLFN11 expression and sensitivity to topoisomerase inhibitors. The systematic translation of cancer genomic data into knowledge of tumour biology and therapeutic possibilities remains challenging. Such efforts should be greatly aided by robust preclinical model systems that reflect the genomic diversity of human cancers and for which detailed genetic and pharmacological annotation is available1. Here we describe the Cancer Cell Line Encyclopedia (CCLE): a compilation of gene expression, chromosomal copy number and massively parallel sequencing data from 947 human cancer cell lines. When coupled with pharmacological profiles for 24 anticancer drugs across 479 of the cell lines, this collection allowed identification of genetic, lineage, and gene-expression-based predictors of drug sensitivity. In addition to known predictors, we found that plasma cell lineage correlated with sensitivity to IGF1 receptor inhibitors; AHR expression was associated with MEK inhibitor efficacy in NRAS-mutant lines; and SLFN11 expression predicted sensitivity to topoisomerase inhibitors. Together, our results indicate that large, annotated cell-line collections may help to enable preclinical stratification schemata for anticancer agents. The generation of genetic predictions of drug response in the preclinical setting and their incorporation into cancer clinical trial design could speed the emergence of ‘personalized’ therapeutic regimens2.

Abstract The non-receptor protein tyrosine phosphatase (PTP) SHP2 is an important component of RTK signaling in response to growth factor stimulus and sits just upstream of the RAS-MAPK signaling cascade. The first oncogenic phosphatase to be identified, SHP2 is dysregulated in multiple human diseases including the developmental disorders Noonan and Leopard syndromes, as well as leukemia, lung cancer and neuroblastoma where aberrant activity of SHP2 leads to uncontrolled MAPK signaling. Cancer-associated activating mutations in SHP2 impart an “auto-on” state of the enzyme, boosting basal activity by shifting the equilibrium away from the auto-inhibited state. Reduction of SHP2 activity through genetic knockdown suppresses tumor growth, validating SHP2 as a target for cancer therapy. SHP099, a recently reported potent and selective allosteric inhibitor of SHP2, stabilizes the auto-inhibited form of SHP2 through interactions with the N-terminal SH2 and C-terminal PTP domains of the protein. SHP099 suppresses MAPK signaling in RTK amplified cancers resulting in suppressed proliferation in vitro and inhibition of tumor growth in mouse tumor xenograft models. Together, these data demonstrate the therapeutic potential of SHP2 inhibition in the treatment of cancer and other RAS/MAPK-linked diseases. Citation Format: Michael G. Acker, Ying-Nan P. Chen, Matthew J. LaMarche, Ho Man Chan, Peter Fekkes, Jorge Garcia-Fortanet, Jonathan R. LaRochelle, Brandon Antonakos, Christine Hiu-Tung Chen, Zhuoliang Chen, Vesselina G. Cooke, Jason R. Dobson, Zhan Deng, Fei Feng, Brant Firestone, Michelle Fodor, Cary Fridrich, Hui Gao, Huai-Xiang Hao, Jaison Jacob, Samuel Ho, Kathy Hsiao, Zhao B. Kang, Rajesh Karki, Mitsunori Kato, Jay Larrow, Laura R. La Bonte, Gang Liu, Shumei Liu, Dyuti Majumdar, Matthew J. Meyer, Mark Palermo, Minying Pu, Edmund Price, Subarna Shakya, Michael D. Shultz, Kavitha Venkatesan, Ping Wang, Markus Warmuth, Sarah Williams, Guizhi Yang, Jing Yuan, Ji-Hu Zhang, Ping Zhu, Stephen C. Blacklow, Timothy Ramsey, Nicholas J. Keen, William R. Sellers, Travis Stams, Pascal D. Fortin. Conformational activation and allosteric inhibition of SHP2 in RTK-driven cancers [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 2084. doi:10.1158/1538-7445.AM2017-2084

Elucidation of the mutational landscape of human cancer has progressed rapidly and been accompanied by the development of therapeutics targeting mutant oncogenes. However, a comprehensive mapping of cancer dependencies has lagged behind and the discovery of therapeutic targets for counteracting tumor suppressor gene loss is needed. To identify vulnerabilities relevant to specific cancer subtypes, we conducted a large-scale RNAi screen in which viability effects of mRNA knockdown were assessed for 7,837 genes using an average of 20 shRNAs per gene in 398 cancer cell lines. We describe findings of this screen, outlining the classes of cancer dependency genes and their relationships to genetic, expression, and lineage features. In addition, we describe robust gene-interaction networks recapitulating both protein complexes and functional cooperation among complexes and pathways. This dataset along with a web portal is provided to the community to assist in the discovery and translation of new therapeutic approaches for cancer.

SHP099, a selective inhibitor of signalling meditator SHP2 with drug-like properties, has an allosteric mechanism of action whereby it stabilizes SHP2 in an auto-inhibited conformation, and suppresses RAS–ERK signalling and proliferation in receptor-tyrosine-kinase-driven cancer cell lines and mouse tumour xenograft models.

Metabolic genes are increasingly recognized as targets of somatic genetic alteration in human cancer often leading to profound changes in intracellular metabolite concentrations. 5-Methylthioadenosine Phosphorylase (MTAP) is a key enzyme in the methionine salvage pathway that metabolizes methylthioadenosine (MTA) to adenine and methionine. Its chromosomal position proximal to CDKN2A results in frequent collateral homozygous deletion in a wide range of human cancers. By interrogating data from a large scale deep-coverage pooled shRNA screen across 390 cancer cell line models we found that the viability of MTAP null cancer cells is strongly impaired upon shRNA-mediated depletion of the protein arginine methyltransferase PRMT5. In MTAP deleted cells there is marked accumulation of the substrate MTA and surprisingly, we find that MTA is a specific inhibitor of the catalytic activity of PRMT5. In keeping with these data, knockout of MTAP in an MTAP-proficient cell line led to increased MTA levels and rendered them sensitive to PRMT5 depletion. Moreover, reconstitution of MTAP in an MTAP-deficient cell line fully rescued PRMT5 dependence. Collectively, these findings indicate that the collateral loss of MTAP in CDNK2A deleted cancers leads to accumulation of MTA that thereby creates a hypomorphic PRMT5 state that is selectively sensitized towards further PRMT5 inhibition. Citation Format: Konstantinos Mavrakis, E Robert McDonald III, Michael R. Schlabach, Eric Billy, Gregory R. Hoffman, Antoine deWeck, David A. Ruddy, Kavitha Venkatesan, Greg McAllister, Rosalie deBeaumont, Samuel Ho, Yue Liu, Yan Yan-Neale, Guizhi Yang, Fallon Lin, Hong Yin, Hui Gao, David Randal Kipp, Songping Zhao, Joshua T. McNamara, Elizabeth R. Sprague, Young Shin Cho, Justin Gu, Ken Crawford, Vladimir Capka, Kristen Hurov, Jeffrey A. Porter, John Tallarico, Craig Mickanin, Emma Lees, Raymond Pagliarini, Nicholas Keen, Tobias Schmelzle, Francesco Hofmann, Frank Stegmeier, William R. Sellers. Disordered methionine metabolism in MTAP/CDKN2A-deleted cancers leads to marked dependence on PRMT5. [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 LB-017.

Abstract Metabolic genes are increasingly recognized as targets of somatic genetic alteration in human cancer often leading to profound changes in intracellular metabolite concentrations. 5-Methylthioadenosine Phosphorylase (MTAP) is a key enzyme in the methionine salvage pathway that metabolizes methylthioadenosine (MTA) to adenine and methionine. Its chromosomal position proximal to CDKN2A results in frequent collateral homozygous deletion in a wide range of human cancers. By interrogating data from a large scale deep-coverage pooled shRNA screen across 390 cancer cell line models we found that the viability of MTAP null cancer cells is strongly impaired upon shRNA-mediated depletion of the protein arginine methyltransferase PRMT5. In MTAP deleted cells there is marked accumulation of the substrate MTA and surprisingly, we find that MTA is a specific inhibitor of the catalytic activity of PRMT5. In keeping with these data, knockout of MTAP in an MTAP-proficient cell line led to increased MTA levels and rendered them sensitive to PRMT5 depletion. Moreover, reconstitution of MTAP in an MTAP-deficient cell line fully rescued PRMT5 dependence. Collectively, these findings indicate that the collateral loss of MTAP in CDNK2A deleted cancers leads to accumulation of MTA that thereby creates a hypomorphic PRMT5 state that is selectively sensitized towards further PRMT5 inhibition. Citation Format: Konstantinos Mavrakis, E Robert McDonald III, Michael R. Schlabach, Eric Billy, Gregory R. Hoffman, Antoine deWeck, David A. Ruddy, Kavitha Venkatesan, Greg McAllister, Rosalie deBeaumont, Samuel Ho, Yue Liu, Yan Yan-Neale, Guizhi Yang, Fallon Lin, Hong Yin, Hui Gao, David Randal Kipp, Songping Zhao, Joshua T. McNamara, Elizabeth R. Sprague, Young Shin Cho, Justin Gu, Ken Crawford, Vladimir Capka, Kristen Hurov, Jeffrey A. Porter, John Tallarico, Craig Mickanin, Emma Lees, Raymond Pagliarini, Nicholas Keen, Tobias Schmelzle, Francesco Hofmann, Frank Stegmeier, William R. Sellers. Disordered methionine metabolism in MTAP/CDKN2A-deleted cancers leads to marked dependence on PRMT5. [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 LB-017.

5-Methylthioadenosine phosphorylase (MTAP) is a key enzyme in the methionine salvage pathway. The MTAP gene is frequently deleted in human cancers because of its chromosomal proximity to the tumor suppressor gene CDKN2A. By interrogating data from a large-scale short hairpin RNA-mediated screen across 390 cancer cell line models, we found that the viability of MTAP-deficient cancer cells is impaired by depletion of the protein arginine methyltransferase PRMT5. MTAP-deleted cells accumulate the metabolite methylthioadenosine (MTA), which we found to inhibit PRMT5 methyltransferase activity. Deletion of MTAP in MTAP-proficient cells rendered them sensitive to PRMT5 depletion. Conversely, reconstitution of MTAP in an MTAP-deficient cell line rescued PRMT5 dependence. Thus, MTA accumulation in MTAP-deleted cancers creates a hypomorphic PRMT5 state that is selectively sensitized toward further PRMT5 inhibition. Inhibitors of PRMT5 that leverage this dysregulated metabolic state merit further investigation as a potential therapy for MTAP/CDKN2A-deleted tumors.

In an attempt to identify novel therapeutics and mechanisms to differentially kill tumor cells using phenotypic screening, we identified N-benzyl indole carbinols (N-BICs), synthetic analogs of the natural product indole-3-carbinol (I3C). To understand the mode of action for the molecules we employed Cancer Cell Line Encyclopedia viability profiling and correlative informatics analysis to identify and ultimately confirm the phase II metabolic enzyme sulfotransferase 1A1 (SULT1A1) as the essential factor for compound selectivity. Further studies demonstrate that SULT1A1 activates the N-BICs by rendering the compounds strong electrophiles which can alkylate cellular proteins and thereby induce cell death. This study demonstrates that the selectivity profile for N-BICs is through conversion by SULT1A1 from an inactive prodrug to an active species that induces cell death and tumor suppression.

The authors implement a collection of patient-derived xenograft tumors to test cancer drug responses.

Abstract Patient selection biomarkers are essential for the successful and rapid development of emerging targeted anti-cancer therapeutics. In this study we have identified a novel patient selection strategy for NVP-CGM097, a p53-Mdm2 inhibitor currently in a Phase I clinical trial (NCT01760525). We have analyzed the sensitivity of over 500 cell lines from the “Cancer Cell Line Encyclopedia” to the p53-Mdm2 inhibitor in cell viability assays, and intersected response data with information on gene expression and genomic alterations. This analysis has led to the identification of a gene signature consisting of 13 genes, as a predictor for sensitivity to NVP-CGM097. Interestingly, these 13 genes are known p53 downstream target genes, supporting the hypothesis that the identified gene signature is reflective of the p53 pathway functionality in sensitive cell lines. We show the performance of the signature as ROC and Precision-Recall curves in cell lines as well as in a number of human primary tumor xenografts, both unselected as well as pre-selected for p53 wild-type status. Work is now ongoing to validate this gene signature using baseline tumor biopsy samples and RECIST-based efficacy readouts in the current Phase I clinical trial. Citation Format: Sebastien Jeay, Swann Gaulis, Stéphane Ferretti, Geneviève Albrecht, Louise Barys, Daniel Guthy, Ensar Halilovic, Moriko Ito, Masato Murakami, Astrid Pornon, Stephan Ruetz, Kavitha Venkatesan, Jianjun Yu, Michael Jensen, Marion Wiesmann, Jens Wuerthner, Diana Graus-Porta. A gene signature composed of 13 p53 target genes predicts for response to NVP-CGM097, a novel p53-Mdm2 inhibitor, in cell lines and in human primary tumor xenograft models. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2909. doi:10.1158/1538-7445.AM2014-2909

Epigenetic dysregulation is an emerging hallmark of cancers. We developed a high-information-content mass spectrometry approach to profile global histone modifications in human cancers. When applied to 115 lines from the Cancer Cell Line Encyclopedia, this approach identified distinct molecular chromatin signatures. One signature was characterized by increased histone 3 lysine 36 (H3K36) dimethylation, exhibited by several lines harboring translocations in NSD2, which encodes a methyltransferase. A previously unknown NSD2 p.Glu1099Lys (p.E1099K) variant was identified in nontranslocated acute lymphoblastic leukemia (ALL) cell lines sharing this signature. Ectopic expression of the variant induced a chromatin signature characteristic of NSD2 hyperactivation and promoted transformation. NSD2 knockdown selectively inhibited the proliferation of NSD2-mutant lines and impaired the in vivo growth of an NSD2-mutant ALL xenograft. Sequencing analysis of >1,000 pediatric cancer genomes identified the NSD2 p.E1099K alteration in 14% of t(12;21) ETV6-RUNX1-containing ALLs. These findings identify NSD2 as a potential therapeutic target for pediatric ALL and provide a general framework for the functional annotation of cancer epigenomes.

Abstract The mammalian fibroblast growth factor/fibroblast growth factor receptor (FGF/FGFR) family comprises 22 FGF ligands, which exert their action through 4 highly conserved transmembrane tyrosine kinase receptors (FGFR1, FGFR2, FGFR3 and FGFR4). This highly complex signaling network regulates fundamental developmental pathways and physiological processes in the adult organism. A growing body of evidence linking deregulation of the FGF/FGFR system, as a consequence of genetic alterations, to the pathogenesis of cancer makes these receptors attractive targets for therapeutic intervention. To this purpose we have developed NVP-BGJ398, an orally bioavailable, selective pan-FGFR kinase inhibitor which is currently in clinical Phase I dose escalation trial. In order to preclinically identify and validate patient stratification biomarkers that accurately predict for response to NVP-BGJ398 in the clinic, we have utilized the Cancer Cell Line Encyclopedia (CCLE). The CCLE is a collection of almost 1000 cancer cell lines representing multiple tumor types that, in a collaborative effort between The Novartis Institutes for BioMedical Research and the Broad Institute, has been comprehensively annotated in terms of genome-scale mRNA expression, gene copy number alterations and gene mutations (http:/www.broadinstitute.org/ccle). In addition, over half of these cell lines were subjected to high-throughput cell viability assays in the presence NVP-BGJ398, with the aim of generating cell line sensitivity data. Here we show the validation of predictive biomarkers of response in primary human tumor xenografts, revealed through the analysis of the CCLE, as well as the identification of potentially new indications for NVP-BGJ398. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 854. doi:1538-7445.AM2012-854

The systematic translation of cancer genomic data into knowledge of tumor biology and therapeutic possibilities remains challenging.Such efforts should be greatly aided by robust preclinical model systems that reflect the genomic

Abstract Patient stratification biomarkers that enable the translation of cancer genetic knowledge into clinical use are essential for the successful and rapid development of emerging targeted anticancer therapeutics. Here, we describe the identification of patient stratification biomarkers for NVP-BGJ398, a novel and selective fibroblast growth factor receptor (FGFR) inhibitor. By intersecting genome-wide gene expression and genomic alteration data with cell line–sensitivity data across an annotated collection of cancer cell lines called the Cancer Cell Line Encyclopedia, we show that genetic alterations for FGFR family members predict for sensitivity to NVP-BGJ398. For the first time, we report oncogenic FGFR1 amplification in osteosarcoma as a potential patient selection biomarker. Furthermore, we show that cancer cell lines harboring FGF19 copy number gain at the 11q13 amplicon are sensitive to NVP-BGJ398 only when concomitant expression of β-klotho occurs. Thus, our findings provide the rationale for the clinical development of FGFR inhibitors in selected patients with cancer harboring tumors with the identified predictors of sensitivity. Significance: The success of a personalized medicine approach using targeted therapies ultimately depends on being able to identify the patients who will benefit the most from any given drug. To this end, we have integrated the molecular profiles for more than 500 cancer cell lines with sensitivity data for the novel anticancer drug NVP-BGJ398 and showed that FGFR genetic alterations are the most significant predictors for sensitivity. This work has ultimately endorsed the incorporation of specific patient selection biomakers in the clinical trials for NVP-BGJ398. Cancer Discov; 2(12); 1118–33. ©2012 AACR. Read the Commentary on this article by Loch and Pollock, p. 1081 This article is highlighted in the In This Issue feature, p. 1065