BRAF and MEK inhibitors increase PD1-positive melanoma cells leading to a potential lymphocyte-independent synergism with anti-PD1 antibody

Purpose: BRAF and MEK inhibitors (BRAF/MEKi) favor melanoma-infiltrating lymphocytes, providing the rationale for current combinatorial trials with anti-PD1 antibody. A portion of melanoma cells may express PD1, and anti-PD1 antibody could have a direct anti-tumor effect. Here, we explore if BRAF/MEKi modulate rates of PD1⁺ melanoma cells, supporting an additional - lymphocyte-independent - basis for their therapeutic combination with anti-PD1 antibody. Experimental design: With data mining and flow cytometry, we assessed PD1, PDL1/2 expression on melanoma cell lines (CCLE, N=61; validation cell lines, N=7) and melanoma tumors (TCGA, N=214). We explored in-vitro how BRAF/MEKi affect rates of PD1⁺, PDL1/2⁺ melanoma cells, and characterized the proliferative and putative stemness features of PD1⁺ melanoma cells. We tested the functional lymphocyte-independent effect of anti-PD1 antibody alone and in combination with BRAF/MEKi in-vitro and in an in-vivo immunodeficient murine model. Results: PD1 is consistently expressed on a small subset of melanoma cells, but PD1⁺ cells increase to relevant rates during BRAF/MEKi treatment (7.3% [5.6-14.2] vs 1.5% [0.7-3.2], p=0.0156; N=7), together with PDL2⁺ melanoma cells (8.5% [0.0-63.0] vs 1.5% [0.2-43.3], p=0.0312; N=7). PD1⁺ cells proliferate less than PD1^- cells (avg. 65% less; t=7 days), and are preferentially endowed with stemness features. In-vivo, the direct anti-melanoma activity of PD1-blockage as monotherapy was negligible, but its association with BRAF/MEKi significantly delayed the development of drug resistance and tumor relapse. Conclusions: BRAF/MEKi increase the rates of PD1⁺ melanoma cells that may sustain tumor relapse, providing a lymphocyte-independent rational to explore combinatory strategies with anti-PD1 antibody.

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