Supplemental Fig. 1. Combination-resistant and combination tolerant tumors re-activated ERK1/2 before progression. Supplemental Fig. 2. Combination-resistant tumor-derived cell lines progressed through cell cycle during combo treatment. Supplemental Fig 3. CRTs harbored tandem duplication of kinase domain but remained sensitive to ERK1/2 inhibition. Supplemental Fig. 4. Both intermittently and continuously treated tumors reactivated of ERK1/2 in vitro. Supplemental Fig. 5. Pathway analysis identified Plk11 as selective driver of resistance in CRTs.
ARTICLE ABSTRACT
Combined BRAF and MEK inhibition is a standard of care in patients with advanced BRAF-mutant melanoma, but acquired resistance remains a challenge that limits response durability. Here, we quantitated in vivo ERK1/2 activity and tumor response associated with resistance to combined BRAF and MEK inhibition in mutant BRAF xenografts. We found that ERK1/2 pathway reactivation preceded the growth of resistant tumors. Moreover, we detected a subset of cells that not only persisted throughout long-term treatment but restored ERK1/2 signaling and grew upon drug removal. Cell lines derived from combination-resistant tumors (CRT) exhibited elevated ERK1/2 phosphorylation, which were sensitive to ERK1/2 inhibition. In some CRTs, we detected a tandem duplication of the BRAF kinase domain. Monitoring ERK1/2 activity in vivo was efficacious in predicting tumor response during intermittent treatment. We observed maintained expression of the mitotic regulator, polo-like kinase 1 (Plk1), in melanoma resistant to BRAF and MEK inhibitors. Plk1 inhibition induced apoptosis in CRTs, leading to slowed growth of BRAF and MEK inhibitor–resistant tumors in vivo. These data demonstrate the utility of in vivo ERK1/2 pathway reporting as a tool to optimize clinical dosing schemes and establish suppression of Plk1 as potential salvage therapy for BRAF inhibitor and MEK inhibitor–resistant melanoma.
