Background Heterogeneous nuclear ribonucleoprotein A2B1 (HNRNPA2B1) is a reader protein of RNA N6-methyladenosine (m6A) modification, which could recognize and bind m6A-modified RNA, thereby regulating tumor development. However, its role in the tumor immune microenvironment (TIME) of colorectal cancer (CRC) remains elusive. Here, we analyzed the function and the regulatory mechanism of HNRNPA2B1 in CRC TIME.

Methods The association of HNRNPA2B1 expression with clinical features was explored in CRC tissue samples from the SYSU (N=30) and TCGA (N=1061) cohorts. The role of HNRNPA2B1 in CRC development was determined in vitro and in vivo assays. Flow cytometry was used to determine the immunological profile of tissue samples in the assays. RNA-seq, MeRIP-seq, and RIP-seq were employed to identify the target of HNRNPA2B1.

Results HNRNPA2B1 expression was upregulated in tumor tissue compared with paired normal tissues, and the higher expression was correlated with worse overall survival in CRC patients. Functionally, knockdown of HNRNPA2B1 in mouse CRC cell lines (MC38, CT26) inhibited cell growth in vitro and subcutaneous tumor growth in immunodeficient NSG mice. Furthermore, we observed that the tumor suppression effect was more obvious in immune-competent mice. The flow cytometry analysis of subcutaneous tumors revealed that knockdown of HNRNPA2B1 could increase tumor infiltration of CD8⁺, CD4⁺, and NK cells to enhance antitumor immunity. Mechanistically, together with MeRIP-seq, RIP-seq and RNA-seq, CLEC2D was identified as the downstream target of HNRNPA2B1. HNRNPA2B1 improved the stability of CLEC2D mRNA and enhanced its expression depending on m6A modification on CLEC2D mRNA, thereby impairing CD161⁺CD8⁺ T cell activity via CLEC2D-CD161 inhibitory signal. Finally, the knockdown of HNRNPA2B1 boosted the response to PD1 blockade in CRC.

Conclusions HNRNPA2B1 facilitates immune evasion by upregulating CLEC2D-CD161 inhibitory signal and serves as a potential target for immunotherapy in CRC.