Characterizing the tumor and immune landscape of patients with melanoma treated with combined checkpoint blockade and MAPK-targeted therapy.
Authors: Zisman Liron Pola , Gushterova Irena , LaSalle Thomas , Blaum Emily , Sharova Tatyana , Liu Mofei , Giobbie-Hurder Anita , Sade-Feldman Moshe , Yizhak Keren , Hacohen Nir , Sullivan Ryan J. 
Published: July 2024

e21503

Background: Melanoma therapy has been revolutionized by two novel therapeutic approaches: mitogen activated protein kinase (MAPK) targeted therapy (MTT) and immune checkpoint blockade therapy (ICB). It has been shown that MTT enhances anti-tumor immunity within the tumor microenviroment (TME), thus providing a strong rationale for its combination with immunotherapy. Regimens combining MTT with ICB there have had mixed results, and which patients (pts) should be treated with these combinations is unknown. Methods: 16 pts harboring BRAFV600 mutation were enrolled and treated with 2 weeks (wks) of MTT (dabrafenib plus trametinib) then 6 wks of concomitant MTT and pembrolizumab, followed by single-agent pembrolizumab. The primary endpoint was clinical benefit (CB) defined as partial/complete response or stable disease (per RECIST1.1) persisting at 24 wks. Serial biopsies were performed prior to MTT, following the 2-week lead-in of MTT, and following 6 wks of combination immune therapy. Single-cell RNA-seq profiling of CD45+ and CD45- cells was performed using both the smart-seq2 plate-based protocol and 10x genomics platform. Results: Out of the 16 pts, 14 received both MTT and ICB, of them only 5 pts had CB. 2 pts did not receive ICB due to MTT toxicity. A clustering analysis of all immune cells showed a significant recruitment of Cytotoxic CD8+ T cells after MTT treatment in pts with CB. In addition T cell clusters had a significantly higher expression of IFNG pathway genes at both baseline and after MTT compared to NCB pts. Tumor associated macrophages (TAM) in NCB pts possessed mainly a “M2” phenotype and expressed a significantly higher level of immune suppressor genes, such as HLA-G, PLAC8 and CD52. In contrast, most of the TAMs occupying the tumor of CB pts had M1 phenotype, and significantly expressed APOE, IL10, CXC12. Calculation of M1/M2 score correlated with chemokines expressed by tumor cells such as CCL3 and TIGIT. In addition, MAPK-pathway genes expressed by the tumor cells were down-regulated after MTT in both CB & NCB pts, and up-regulated after ICB, but only in NCB pts. Our analysis also revealed tumor programs that were significantly higher in NCB patients: the first expressing high MITF-SOX levels pre-therapy that decreased after MTT, the second expressing AXL, low at baseline and up regulated after combination treatment. These AXL expressing cells also had a significantly higher TGF-β expression levels. Moreover, MITF/AXL ratio in NCB patients decreased significantly after MTT and after ICB. Conclusions: Overall, abbreviated MTT with ICB did not lead to increased CB. Our findings highlights factors that modify the TME towards immune suppression, tumor programs resistance, MAPK up-regulation and can thus enhance drug resistance. Clinical trial information: NCT03149029 .