PD-1 blockade in combination with epigenetic therapy to antagonize tumor mesenchymal program and drive therapeutic efficacy in an esophageal adenocarcinoma model.

Abstract

e16159 Background: Immune checkpoint blockade (ICB) alone or in combination with chemotherapy has demonstrated a modest survival benefit in gastroesophageal cancer patients. In other cancer, epigenetic therapies have been shown to improve effectiveness of ICB via additive antiproliferative effects, enhanced immune activation, and reversal of T-cell exhaustion. Methods: To induce esophageal adenocarcinoma (EAC), esophagojejunostomy was performed on Sprague-Dawley rats. 32 weeks postoperative, 74 animals were randomized into 8 unique groups. Animals underwent three 14-day cycles of epigenetic therapy or placebo, ± radiation. Cycles consisted of 1 week of DNA methyltransferase treatment or placebo at 0.5 mpk, then 1 week of HDAC treatment or placebo at 2 mpk. One dose of 3 mpk PD1 inhibitor (AUNP-12) or placebo was given each cycle. Safety and efficacy were evaluated via health assessments, serial MRI, immunofluorescent labeling (IF) and RNA sequencing. Results: The study showed no differences in mortality (p = 0.556) between study groups. Pre- to post-treatment mean MRI tumor volume increased by 263.1% and 90.9% in the placebo and the placebo + radiation groups, respectively. Significant tumor reduction was observed in the dual therapy groups, 84.5% and 61.8% ± radiation, respectively. Independent epigenetic and ICB treatments ± radiation exhibited a decrease in tumor volume, but to a lesser extent than the dual regimens (p = 0.0046). IF demonstrated a significant increase in CD3CD8 + T cells in all treatment groups compared to placebo (p = < 0.0001). The degree of CD3CD8 + T cell intratumor infiltration was positively associated with the depth of tumor response as measured by MRI. Bulk tumor RNAseq analysis by DESeq2 revealed a modest transcriptional response, with the greatest number of differentially expressed genes occurring in epigenetic therapy (n = 113 genes) and the dual therapy ± radiation (n = 96 genes) groups. Gene set enrichment analysis of RNAseq data defined a conserved induction of interferon-related gene sets across all treatment groups. Dual therapy ± radiation revealed a pronounced downregulation of epithelial-mesenchymal transition (EMT) and hypoxia-related gene sets. Critically, interferon, EMT, and hypoxia-related gene sets were all found to have a significant association with the depth of tumor regression in response to therapy. Conclusions: This study establishes antitumor efficacy and molecular correlates associated with response for epigenetic therapy with ICB and radiation, to treat EAC. All treatment groups enhanced CD3CD8 + T-cell tumor infiltration and interferon signaling. Dual therapy ± radiation, downregulated EMT and hypoxia-related gene sets, both having a role in modulation of cancer hallmarks, including the acquisition of an immune suppressive tumor microenvironment, tumor invasion, and metastasis.