Treatment with antibodies that target programmed cell death 1 (PD-1) or its ligand programmed death ligand 1 (PD-L1) has demonstrated durable efficacy for various malignant tumors. Programmed death ligand 2 (PD-L2), which is another ligand of PD-1, has recently been shown to be implicated in tumor immune escape. The regulation of PD-L2 expression in tumor cells has remained unclear, however. We here examined intrinsic and extrinsic regulation of PDL2 expression in NSCLC.
PD-L2 expression was evaluated by reverse transcription and real-time polymerase chain reaction analysis and by flow cytometry.
BEAS-2B cells stably expressing an activated mutant form of the epidermal growth factor receptor (EGFR) or the EML4-ALK fusion oncoprotein manifested increased expression of PD-L2 at both mRNA and protein levels. Furthermore, treatment of NSCLC cell lines that harbor such driver oncogenes with corresponding EGFR or ALK tyrosine kinase inhibitors or depletion of EGFR or ALK by siRNA transfection suppressed expression of PD-L2, demonstrating that activating EGFR mutations or EML4-ALK fusion intrinsically induce PD-L2 expression. We also found that interferon-γ extrinsically induced expression of PD-L2 via STAT1 signalingin NSCLC cells. Oncogene-driven expression of PD-L2 in NSCLC cells was inhibited by knockdown of the transcription factors STAT3 or c-FOS. Interferon-γ also activated STAT3 and c-FOS. Knockdown of STAT3 or c-FOS decreased Interferon-γ-induced expression of PD-L2, suggesting that these proteins may also contribute to the extrinsic induction of PD-L2 expression.
Expression of PD-L2 is induced intrinsically by activating EGFR mutations or EML4-ALK fusion as well as extrinsically by interferon-γ, with STAT3 and c-FOS possibly contributing to both intrinsic and extrinsic pathways. Our results thus provide insight into the complexity of tumor immune escape in NSCLC.
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K. Azuma: Personal fees: AstraZeneca; Grants and personal fees: Boehringer Ingelheim, Ono Pharmaceutical, MSD Oncology, Bristol-Myers Squibb, and Chugai Pharma outside the submitted work. I. Okamoto: Grants and personal fees: AstraZeneca, Taiho Pharmaceutical, Boehringer Ingelheim, Ono Pharmaceutical, MSD Oncology, Lilly, Bristol-Myers Squibb, Chugai Pharma; Grants: Astellas Pharma Novartis; Personal fees: Pfizer outside the submitted work. All other authors have declared no conflicts of interest.