Targeting Points of Vulnerability in individual Melanoma Tumors for Reducing Acquired Drug Resistance

Targeted drug therapy for melanoma patients carrying the BRAFV600E driver mutation with vemurafenib provides a temporary remission, followed by severe relapses due to the dominance of drug-resistant tumors. Acquired drug resistance can develop from persister tumor cells that maintain their cell viability during prolonged drug treatment. Here we propose a functional approach to circumvent this problem, by identifying the soft-spots in the cell death machinery of patients’ metastases, the targeting of which strongly reduces the number of persister cells. This challenge required a smart solution, as there exists huge heterogeneity in the composition of the cell death map among patients’ tumors. To this end, we developed a personalized prescreening platform that maps the soft-spots in each tumor carrying the BRAF mutation, before patients are treated with the drug. The platform is based on applying siRNA libraries targeting 81 genes of apoptosis, autophagy and necroptosis pathways, one at a time, to tumors derived from Melanoma and identifying the hits that reduce the number of persister cells upon BRAF inhibitor treatment. We have demonstrated the feasibility of this approach in an analysis of a cohort of 12 metastatic melanoma early passage cell cultures carrying BRAF mutation supplied by the Sheba Medical Center. By applying this platform we identified a large heterogeneity in the number and position of the soft spots among patients. In some tumors, the soft spots were spread over the three PCD modules, including clusters of hits targeting autophagic genes that participate in the ubiquitin like conjugation pathways. In other tumors the number of soft spot was low, mapped to one or two of the PCD modules at different positions. After validating the soft spots we were able to replace the siRNAs with small molecule inhibitors where available. Most importantly, by reducing the number of persister cell over long term of vemurafenib treatment, by simultaneously targeting one of the identified soft spots, we could reduce the number of drug resistant clones emerging after 4-5 weeks. We also measured the outcome of double hits either by testing all possible combinations, or in a rationally designed choice of targets by superimposing the soft-spots on the integrated map of programmed cell death that we have delineated in our lab. This approach has the potential to dramatically change the state-of-the-art of combinatorial drug therapy in precision cancer treatment, by reducing the number of the persister cells surviving the initial treatment by several orders of magnitude, thereby lessening the odds of developing drug resistance at later stages, and preventing tumor relapses.