Background: The existence of immunologically 'cold tumors' frequently found across a wide spectrum of tumor types represents a significant challenge for cancer immunotherapy. Cold tumors have poor baseline pan-leukocyte infiltration, including a low prevalence of cytotoxic lymphocytes, and not surprisingly respond unfavorably to immune checkpoint (IC) inhibitors. We hypothesized that cold tumors harbor a mechanism of immune escape upstream and independent of ICs that may be driven by tumor biology rather than differences in mutational neoantigen burden.
Methods: Using a bioinformatic approach to analyze TCGA (The Cancer Genome Atlas) RNA sequencing data we identified genes upregulated in cold versus hot tumors across four different smoking-related cancers, including squamous carcinomas from the oral cavity (OCSCC) and lung (LUSC), and adenocarcinomas of the bladder (BLCA) and lung (LUAD). Biological significance of the gene most robustly associated with a cold tumor phenotype across all four tumor types, glutathione peroxidase 2 (GPX2), was further evaluated using a combination of in silico analyses and functional genomic experiments performed both in vitro and in in vivo with preclinical models of oral cancer.
Results: Elevated RNA expression of five metabolic enzymes including GPX2, aldo-keto reductase family 1 members AKR1C1, AKR1C3, and cytochrome monoxygenases (CP4F11 and CYP4F3) co-occurred in cold tumors across all four smoking-related cancers. These genes have all been linked to negative regulation of arachidonic acid metabolism-a well-established inflammatory pathway-and are also known downstream targets of the redox sensitive Nrf2 transcription factor pathway. In OCSCC, LUSC, and LUAD, GPX2 expression was highly correlated with Nrf2 activation signatures, also elevated in cold tumors. In BLCA, however, GPX2 correlated more strongly than Nrf2 signatures with decreased infiltration of multiple leukocyte subtypes. GPX2 inversely correlated with expression of multiple pro- inflammatory cytokines/chemokines and NF-kB activation in cell lines and knockdown of GPX2 led to increased secretion of prostaglandin E2 (PGE2) and interleukin-6. Conversely, GPX2 overexpression led to reduced PGE2 production in a murine OCSCC model (MOC1). GPX2 overexpressing MOC1 tumors had a more suppressive tumor immune microenvironment and responded less favorably to anti-cytotoxic T-lymphocytes-associated protein 4 IC therapy in mice.
Conclusion: GPX2 overexpression represents a novel potentially targetable effector of immune escape in cold tumors.
Keywords: head and neck neoplasms; immune evation; immune tolerance; immunotherapy; tumor escape.
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