Understanding of cancer-immune system interaction led to development of immunotherapy; anti-programmed cell death protein-1 (PD-1)/programmed cell death ligand-1 (PD-L1) antibodies are now used in non small cell lung cancer (NSCLC) treatment. MAPK cascade is a key intracellular network for tumor proliferation and recent data suggest that it is implicated in interplay of tumor and T-CD8+ cytotoxic lymphocytes (CTL).
We evaluated PD-L1 mRNA level by Real Time qPCR (RT-qPCR) and its protein production, togheter with MAPK proteins, by western blot (WB), in NSCLC cell lines. Then, we studied the changes in PD-L1 and major histocompatibility complex class-I (MHC-I) expression and cytokines’ production, after MAPK-inhibition or -stimulation, by MEK-inhibitor, cobimetinib, or phorbol 12-myristate 13-acetate (PMA), respectively. In addition, we explored the effect of cobimetinib on cytokines’ genes by RT-qPCR on cDNA, obtained from retro-transcription of RNA extracted from T-lymphocytes, derived from Peripheral blood mononuclear cells (PBMC) of healthy volunteers, by density gradient separation, and activated with anti-CD3/anti-CD28 coated beads.
WB and RT-qPCR for PD-L1 in NSCLC cells revealed a consistent correlation between mRNA and protein levels, togheter with activated MAPK and MEK1/2 signals, and suggested that ectopic PD-L1 mainly depends on trascriptional regulation. PDL-1 levels were significantly decreased by cobimetinib and increased by PMA, suggesting that MAPK can regulate PD-L1. Moreover, MEK-inhibition resulted on cancer cells in increased synthesis of MHC-I, IFN-gamma, IL-6, IL-1B, and TNFalpha, involved in CTL activation, and on activated human pheripheral T-lymphocytes in increment of mRNA levels of IL-12, TNFalpha and IFNgamma, that are pro-inflammatory cytokines typical of CTL subset, that seems more involved in immune response against cancer.
These results demonstrate that MEK-inhibion induces the establishment of a pro-inflammatory microenvironment and may represent a potential mechanism to convert otherwise resistant cancers through treatment combination strategies of MEK-inhibitors and anti-PD-L1/PD-1 antibodies in NSCLC.
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AOU Università della Campania “Luigi Vanvitelli”
All authors have declared no conflicts of interest.