Ruoshi Shi1, Sebastiao N Martins Filho2, Ming Li3, Aline Fares4, Jessica Weiss3, Nhu-An Pham3, Olga Ludkovski3, Vibha Raghavan3, Quan Li3, Deepti Ravi2, Michael Cabanero2, Nadeem Moghal3, Natasha B Leighl4, Penelope Bradbury4, Adrian Sacher4, Frances A Shepherd4, Kazuhiro Yasufuku5, Ming-Sound Tsao6, Geoffrey Liu7. 1. University Health Network, Princess Margaret Cancer Centre, Toronto, Ontario, Canada; Department of Medical Biophysics, University of Toronto, Toronto, Ontario, Canada. 2. University Health Network, Princess Margaret Cancer Centre, Toronto, Ontario, Canada; Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada. 3. University Health Network, Princess Margaret Cancer Centre, Toronto, Ontario, Canada. 4. University Health Network, Princess Margaret Cancer Centre, Toronto, Ontario, Canada; Department of Medicine, University of Toronto, Toronto, Ontario, Canada. 5. University Health Network, Princess Margaret Cancer Centre, Toronto, Ontario, Canada; Department of Surgery, University of Toronto, Toronto, Ontario, Canada. 6. University Health Network, Princess Margaret Cancer Centre, Toronto, Ontario, Canada; Department of Medical Biophysics, University of Toronto, Toronto, Ontario, Canada; Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada. Electronic address: ming.tsao@uhn.ca. 7. University Health Network, Princess Margaret Cancer Centre, Toronto, Ontario, Canada; Department of Medical Biophysics, University of Toronto, Toronto, Ontario, Canada; Department of Medicine, University of Toronto, Toronto, Ontario, Canada; Dalla Lana School of Public Health and Institute of Medical Science, University of Toronto, Toronto, Ontario, Canada.
Abstract
BACKGROUND: Anaplastic lymphoma kinase (ALK) targeted therapies have demonstrated remarkable efficacy in ALK-positive lung adenocarcinomas. However, patients inevitably develop resistance to such therapies. To investigate novel mechanisms of resistance to second generation ALK inhibitors, we characterized and modeled ALK inhibitor resistance of ALK-positive patient-derived xenograft (PDX) models established from advanced-stage lung adenocarcinoma patients who have progressed on one or more ALK inhibitors. METHODS: Whole exome sequencing was performed to identify resistance mechanisms to ALK inhibitors in PDXs generated from biopsies at the time of relapse. ALK fusion status was confirmed using fluorescent in situ hybridization, immunohistochemistry, RNA-sequencing, RT-qPCR and western blot. Targeted therapies to overcome acquired resistance were then tested on the PDX models. RESULTS: Three PDX models were successfully established from biopsies of two patients who had progressed on crizotinib and/or alectinib. The PDX models recapitulated the histology and ALK status of their patient tumors, as well as their matched patients' clinical treatment outcome to ALK inhibitors. Whole exome sequencing identified MET amplification and previously unreported BRAF V600E mutation as independent mechanisms of resistance to alectinib. Importantly, PDX treatment of inhibitors specific for these targets combined with ALK inhibitor overcame resistance. CONCLUSIONS: Bypass signaling pathway through c-MET and BRAF are independent mechanisms of resistance to alectinib. Individualized intervention against these resistance pathways could be viable therapeutic options in alectinib-refractory lung adenocarcinoma.
BACKGROUND:Anaplastic lymphoma kinase (ALK) targeted therapies have demonstrated remarkable efficacy in ALK-positive lung adenocarcinomas. However, patients inevitably develop resistance to such therapies. To investigate novel mechanisms of resistance to second generation ALK inhibitors, we characterized and modeled ALK inhibitor resistance of ALK-positive patient-derived xenograft (PDX) models established from advanced-stage lung adenocarcinomapatients who have progressed on one or more ALK inhibitors. METHODS: Whole exome sequencing was performed to identify resistance mechanisms to ALK inhibitors in PDXs generated from biopsies at the time of relapse. ALK fusion status was confirmed using fluorescent in situ hybridization, immunohistochemistry, RNA-sequencing, RT-qPCR and western blot. Targeted therapies to overcome acquired resistance were then tested on the PDX models. RESULTS: Three PDX models were successfully established from biopsies of two patients who had progressed on crizotinib and/or alectinib. The PDX models recapitulated the histology and ALK status of their patienttumors, as well as their matched patients' clinical treatment outcome to ALK inhibitors. Whole exome sequencing identified MET amplification and previously unreported BRAFV600E mutation as independent mechanisms of resistance to alectinib. Importantly, PDX treatment of inhibitors specific for these targets combined with ALK inhibitor overcame resistance. CONCLUSIONS: Bypass signaling pathway through c-MET and BRAF are independent mechanisms of resistance to alectinib. Individualized intervention against these resistance pathways could be viable therapeutic options in alectinib-refractory lung adenocarcinoma.