METTL3-VISTA axis-based combination immunotherapy for APC truncation colorectal cancer.

Abstract

Objective: Although immune checkpoint blockade (ICB) therapy represents a bright spot in antitumor immunotherapy, its clinical benefits in colorectal cancer (CRC) are limited. Therefore, a new target for mediating CRC immunosuppression is urgently needed. Adenomatous polyposis coli (APC) mutations have been reported as early-stage characteristic events in CRC, but the role of truncated APC in the CRC immune microenvironment remains unclear and its clinical significance has yet to be explored.

Design: Adenocarcinoma formation in the colon of the APC^Min/+ mouse model, which displays features associated with the translation of truncated APC proteins, was induced by azoxymethane/dextran sodium sulfate. Multiplexed immunohistochemical consecutive staining on single slides and flow cytometry were used to explore the activation of immune cells and the expression of the immune checkpoint V-domain immunoglobulin suppressor of T-cell activation (VISTA) in the CRC tissues of APC^WT and APC^Min/+ mice. The construction of truncated APC vectors and an initial subserosal graft tumor mouse model was employed to mimic the tumor microenvironment (TME) during APC mutation. Methylated RNA immunoprecipitation-quantitative PCR assays were performed to investigate the N6-methyladenosine (m6A)-dependent transcriptional regulation of hypoxia-inducible factor-1 alpha (HIF1α) by methyltransferase-like protein 3 (METTL3). Mettl3^fl/fl vil1-cre^+/- mice were used to demonstrate that targeting METTL3 is a mediator that mitigates the deleterious effects of the APC978∆-HIF1α axis on antitumor immunity. A chimeric VISTA humanized mouse model was used to evaluate the drug efficacy of the VISTA-targeted compound onvatilimab.

Results: We showed that APC978∆, a truncated APC protein, mediated overexpression of METTL3, resulting in m6A methylation of HIF1α messenger RNA and high expression of HIF1α. Furthermore, HIF1α promotes the migration of myeloid-derived suppressor cells to the TME by binding to the promoters of MCP-1 and MIF. In addition, HIF1α enhances the expression of the immune checkpoint VISTA on CRC cells, weakening tumor immune monitoring.

Conclusions: We elucidate that an underappreciated function of truncated APC in CRC is its ability to drive an immunosuppressive program that boosts tumor progression. Our work could provide a new perspective for the clinical application of immunotherapy in patients with CRC resistant to ICB therapy.