Author Manuscript Published OnlineFirst on December 28, 2017; DOI: 10.1158/1078-0432.CCR-17-3001 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. Structural alterations of MET trigger response to MET kinase inhibition in lung adenocarcinoma patients Dennis Plenker1,2*, Miriam Bertrand3*, Adrianus J. de Langen4*, Richard Riedel5*, Carina Lorenz1,2*, Andreas H. Scheel6, Judith Müller3, Johannes Brägelmann1,2, Juliane Daßler- Plenker7, Carsten Kobe8, Thorsten Persigehl9, Alexander Kluge10, Thomas Wurdinger11,12,13, Pepijn Schellen14, Gunther Hartmann7, Tobias Zacherle3, Roopika Menon3, Erik Thunnissen15, Reinhard Büttner6, Frank Griesinger16, Jürgen Wolf5, Lukas Heukamp17, Martin L. Sos1,2,18*, Johannes M. Heuckmann3* Correspondence Martin L. Sos ([email protected]), Molecular Pathology, Institute of Pathology and Department of Translational Genomics, University of Cologne, Weyertal 115b, 50931 Cologne, Germany. Tel.: +49 221 478 96175 Fax: +49 221 478 97902 Johannes M. Heuckmann ([email protected]), NEO New Oncology GmbH, Gottfried-Hagen-Str. 20, 51105 Cologne, Germany. Tel.: +49 221 8882380 Fax: +49 221 88823822 Affiliations 1 Molecular Pathology, Institute of Pathology, University of Cologne, Kerpener Str. 62, 50937 Cologne, Germany. 2 Department of Translational Genomics, Medical Faculty, University of Cologne, Weyertal 115b, 50931 Cologne, Germany 3 NEO New Oncology GmbH, Gottfried-Hagen-Str. 20, 51105 Köln, Germany. 4 Department of Pulmonary Diseases, VU University Medical Center, Amsterdam, The Netherlands. 5 Department of Internal Medicine, Center for Integrated Oncology Köln Bonn, University Hospital Cologne, Cologne, 50931 Cologne, Germany. 6 Institute of Pathology, Center of Integrated Oncology, University Hospital Cologne, 50937 Cologne, Germany 7 Department of Clinical Chemistry and Clinical Pharmacology, University Hospital Bonn, Sigmund-Freud-Str. 25, 53127 Bonn, Germany 8 Department of Nuclear Medicine, University Hospital of Cologne, Cologne, Germany. 9 Department of Radiology, University Hospital of Cologne, Germany. 10 Institute for diagnostic and interventional Radiology, Pius-Hospital, Medical Campus University of Oldenburg, Germany 11 Department of Neurosurgery, VU University Medical Center, Cancer Center Amsterdam, De Boelelaan 1117, 1081 HV Amsterdam, the Netherlands. 12 Brain Tumor Center Amsterdam, VU University Medical Center, Cancer Center Amsterdam, De Boelelaan 1117, 1081 HV Amsterdam, the Netherlands. 13 Department of Neurology Massachusetts General Hospital and Neuroscience Program, Harvard Medical School, 149 13(th) Street, Charlestown, MA 02129, USA. 1 Downloaded from clincancerres.aacrjournals.org on September 26, 2021. © 2017 American Association for Cancer Research. Author Manuscript Published OnlineFirst on December 28, 2017; DOI: 10.1158/1078-0432.CCR-17-3001 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. 14 Department of Neurosurgery, VU University Medical Center, Cancer Center Amsterdam, De Boelelaan 1117, 1081 HV Amsterdam, the Netherlands; Brain Tumor Center Amsterdam, VU University Medical Center, Cancer Center Amsterdam, De Boelelaan 1117, 1081 HV Amsterdam, the Netherlands. 15 Dept of Pathology, VU University Medical Center, Amsterdam, The Netherlands. 16 Lung Cancer Network NOWEL, Georgstrasse 12, 26121, Oldenburg, Germany; Department of Hematology and Oncology, Pius-Hospital, University Department Internal Medicine-Oncology, Medical Campus University of Oldenburg, Germany. 17 NEO New Oncology GmbH, Gottfried-Hagen-Str. 20, 51105 Köln, Germany; Institute for Hematopathology, Fangdieckstr. 75a, Hamburg, Germany; Lung Cancer Network NOWEL, Georgstrasse 12, 26121, Oldenburg, Germany. 18 Center for Molecular Medicine Cologne, University of Cologne, 50931 Cologne, Germany. Current affiliation of D. Plenker: Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11724, USA Running title: MET alterations trigger response to MET inhibition Disclosures of Potential Conflict of Interest A.H.S. attended advisory boards of MSD, BMS, Roche and received sponsored travels from QuIP GmbH, NordiQC within the last 12 months. R.B. is an employee of Targos Molecular Pathology. J.W. is a consultant/advisory board member for AstraZeneca, Bristol-Myers Squibb, Boehringer-Ingelheim, Clovis, MSD, Novartis, Pfizer, and Roche. M.L.S received a commercial research grant from Novartis. R.R. attended advisory boards of Boehringer Ingelheim and received sponsored travels from Lilly Oncology and Boehringer Ingelheim within the last 12 months. M.B., J.M., R.M., T.Z., L.H. and J.M.H. are employees of NEO New Oncology GmbH. M.L.S received a commercial research grant from Novartis. 2 Downloaded from clincancerres.aacrjournals.org on September 26, 2021. © 2017 American Association for Cancer Research. Author Manuscript Published OnlineFirst on December 28, 2017; DOI: 10.1158/1078-0432.CCR-17-3001 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. Abstract Purpose: We sought to investigate the clinical response to MET inhibition in patients diagnosed with structural MET alterations and to characterize their functional relevance in cellular models. Experimental Design: Patients were selected for treatment with crizotinib upon results of hybrid capture-based next generation sequencing. To confirm the clinical observations we analyzed cellular models that express these MET kinase alterations. Results: Three individual patients were identified to harbor alterations within the MET receptor. Two patients showed genomic rearrangements leading to a gene fusion of KIF5B or STARD3NL and MET. One patient diagnosed with an EML4-ALK rearrangement developed a MET kinase domain duplication as a resistance mechanism to ceritinib. All three patients showed a partial response to crizotinib that effectively inhibits MET and ALK among other kinases. The results were further confirmed using orthogonal cellular models. Conclusions: Crizotinib leads to a clinical response in patients with MET rearrangements. Our functional analyses together with the clinical data suggest that these structural alterations may represent actionable targets in lung cancer patients. Statement of translational relevance Oncogenically activated MET kinases have been implicated in the tumorigenesis of several cancer subtypes. We identified three structurally unique MET kinase alterations in lung adenocarcinoma patients. More specifically we characterize two different MET kinase fusions as well as a MET kinase domain duplication that developed in an ALK- 3 Downloaded from clincancerres.aacrjournals.org on September 26, 2021. © 2017 American Association for Cancer Research. Author Manuscript Published OnlineFirst on December 28, 2017; DOI: 10.1158/1078-0432.CCR-17-3001 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. rearranged tumor along with acquired resistance to ceritinib. Off-label use of the kinase inhibitor crizotinib led to a marked response of all patients. Together with these observations our cellular analyses provide a functional basis for the oncogenic role of these structural MET alterations. Our findings will have an immediate impact for both the diagnostic and the therapeutic routine of lung cancer patients. 4 Downloaded from clincancerres.aacrjournals.org on September 26, 2021. © 2017 American Association for Cancer Research. Author Manuscript Published OnlineFirst on December 28, 2017; DOI: 10.1158/1078-0432.CCR-17-3001 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. Introduction Over the past years we have witnessed a dramatic shift in the clinical routine of lung adenocarcinoma patients, driven by the identification of oncogenically activated and therapeutically actionable targets. Next to oncogenic mutations, structural rearrangements of receptor kinases involving ALK, ROS1 or RET represent an ever- increasing pool of druggable targets in lung cancer patients (1-6). Effective inhibition of these oncogenic drivers frequently results in dramatic clinical responses (1,4-6). Most common oncogenic alterations of MET kinase involve MET exon 14 skipping mutations but the therapeutic relevance of more complex structural rearrangements remains largely unknown (2,7-9). Here, we report the discovery of structural alterations of MET that may predict response to targeted MET inhibition in lung cancer patients. 5 Downloaded from clincancerres.aacrjournals.org on September 26, 2021. © 2017 American Association for Cancer Research. Author Manuscript Published OnlineFirst on December 28, 2017; DOI: 10.1158/1078-0432.CCR-17-3001 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. Materials and Methods Patient material All three patients consented to testing, therapy and registration of data for future publication. Genetic testing was conducted in concordance with local ethical guidelines and reviewed by the institutional ethics committee. The patient material was sequenced using hybrid capture-based next-generation sequencing (NEO, New Oncology GmbH). Hybrid-capture based sequencing NEOplus Genomic DNA was extracted from FFPE material, sheared (Covaris) and subjected to hybrid capture-based next-generation sequencing to detect point mutations, small insertions and deletions, copy number alterations and rearrangement/gene fusions in a single