Adaptation of three different NRAS mutant melanoma cell lines (IPC298, MELJUSO and SKMEL30) to a combination of CDK4/6 and MEK inhibitors over 33 days.

To study the effects of MEKi and CDK4/6i, we selected three NRAS mutant melanoma cell-lines: IPC298, MELJUSO and SKMEL30. We determined the GI50 (drug concentration at which cell growth is reduced by half) of Binimetinib (MEKi) and opted for a concentration of 1 ??M for Palbociclib (CDK4/6i) as this inhibitor led to incomplete dose-response curves. Upon treatment over 33 days, we observed distinct cellular responses. IPC298 suffered very little from the treatment showing constant proliferation and no morphological change. SKMEL30 stopped proliferating and accumulated pigmentation before restoring proliferation after around 14 days. Finally, MELJUSO stopped dividing and started to stretch to acquire a filiform phenotype after 4 days. As distinct morphological changes were observed at these time points, we profiled the transcriptome of the cell lines at day 0, 4, 14 and 33 using total RNA-seq. We investigated the cellular signaling supporting the adaptation to the treatment by profiling the kinome of the senescent MELJUSO and adaptative SKMEL30 cell lines using PamGene technology at day 0, 1, 4 and 33. Finally, we characterized the resistant miRNA interactome of the SKMEL30 and IPC298 at 33 days by combining small RNA-seq and the qCLASH method.