Hypoxia‐inducible factor‐1α and nuclear factor‐κB play important roles in regulating programmed cell death ligand 1 expression by epidermal growth factor receptor mutants in non‐small‐cell lung cancer cells

Abstract

Some driver gene mutations, including epidermal growth factor receptor ( EGFR ), have been reported to be involved in expression regulation of the immunosuppressive checkpoint protein programmed cell death ligand 1 ( PD ‐L1), but the underlying mechanism remains obscure. We investigated the potential role and precise mechanism of EGFR mutants in PD ‐L1 expression regulation in non‐small‐cell lung cancer ( NSCLC ) cells. Examination of pivotal EGFR signaling effectors in 8 NSCLC cell lines indicated apparent associations between PD ‐L1 overexpression and phosphorylation of AKT and ERK , especially with increased protein levels of phospho‐IκBα (p‐IκBα) and hypoxia‐inducible factor‐1α (HIF‐1α). Flow cytometry results showed stronger membrane co‐expression of EGFR and PD ‐L1 in NSCLC cells with EGFR mutants compared with cells carrying WT EGFR . Additionally, ectopic expression or depletion of EGFR mutants and treatment with EGFR pathway inhibitors targeting MEK / ERK , PI 3K/ AKT , mTOR /S6, IκBα, and HIF ‐1α indicated strong accordance among protein levels of PD ‐L1, p‐IκBα, and HIF ‐1α in NSCLC cells. Further treatment with pathway inhibitors significantly inhibited xenograft tumor growth and p‐IκBα, HIF ‐1α, and PD ‐L1 expression of NSCLC cells carrying EGFR mutant in nude mice. Moreover, immunohistochemical analysis revealed obviously increased protein levels of p‐IκBα, HIF ‐1α, and PD ‐L1 in NSCLC tissues with EGFR mutants compared with tissues carrying WT EGFR . Non‐small‐cell lung cancer tissues with either p‐IκBα or HIF ‐1α positive staining were more likely to possess elevated PD ‐L1 expression compared with tissues scored negative for both p‐IκBα and HIF ‐1α. Our findings showed important roles of phosphorylation activation of AKT and ERK and potential interplay and cooperation between NF ‐κB and HIF ‐1α in PD ‐L1 expression regulation by EGFR mutants in NSCLC .

NF-κB and Its Role in Checkpoint Control

“…Recently, Guo and colleagues described a stronger expression of PD-L1 expression in EGFR mutant NSCLC cells in comparison to non-mutant EGFR NSCLC, which was associated with increased expression levels of phospho-IκBα and hypoxia-induced factor 1α (HIF-1α) [43]. Therefore, the authors postulated a potential interplay between NF-κB and HIF-1α in the regulation of PD-L1 expression [43]. Indeed, a direct binding of NF-κB to the HIF-1α promoter, and vice versa, a HIF-1α-dependent NF-κB activity were reported [99][100][101][102].…”

“…As outlined in Section 2, EGFR mutations were reported as genetic drivers of PD-L1 expression thereby contributing to tumor immune escape. [43]. EGFR-TKIs have been identified to reduce PD-L1 expression by inhibiting NF-κB.…”

“…This leads to the activation of T cells in the TME and finally to the targeting of tumor cells by releasing effector cytokines and cytotoxic granules [30][31][32][33]. The NF-κB signaling pathway is also involved in regulation of immune checkpoint expression in tumor cells, as NF-κB can induce PD-L1 expression thereby promoting T cell suppression and consequently tumorigenesis [34][35][36][37][38][39][40][41][42][43]. Additionally, PD-L1 expression on tumor cells regulates several cell-intrinsic mechanisms promoting tumor cell growth, metastasis, and resistance to Fas-ligand as well as chemotherapy-induced apoptosis [44][45][46].…”

Role and mechanism of programmed death‑ligand�1 in hypoxia‑induced liver cancer immune escape (Review)

“…Hypoxia, a common phenomenon in the tumor microenvironment, induces the expression of PD-L1 to promote immune escape (15)(16)(17). A number of immune cells with immunosuppressive activities, including tumor-associated regulatory T cells (Tregs), myeloid-derived suppressor cells (MDSCs) and tumor-associated macrophages (TAMs), are recruited to the tumor tissue to form an immunosuppressive microenvironment (18)(19)(20).…”

Pemetrexed sensitizes human lung cancer cells to cytotoxic immune cells

“…Given that PC9 cells have an EGFR mutation and that RAS activation can be induced by signals via receptor tyrosine kinase, RAS activation could lead to ERK activation. Indeed, several studies have revealed that MEK/ERK signaling, as well as NFκB, can increase the expression of PD‐L1 on cancer cells 26,27 . However, PEM treatment also increased the phosphorylation levels of ERK in A549 cells, but no change of their PD‐L1 expression was observed.…”