M6A Demethylase ALKBH5 Regulates PD-L1 Expression and Tumor Immunoenvironment in Intrahepatic Cholangiocarcinoma

Abstract

N6-methyladenosine (m6A) has been reported as an important mechanism of posttranscriptional regulation. Programmed death-ligand 1 (PD-L1) is a primary immune inhibitory molecule expressed on tumor cells that promotes immune evasion. Here we report ALKBH5 as an important m6A demethylase that orchestrates PD-L1 expression in intrahepatic cholangiocarcinoma (ICC). Regulation of PD-L1 expression by ALKBH5 was confirmed in human ICC cell lines. Sequencing of the m6A methylome identified PD-L1 mRNA as a direct target of m6A modification whose levels were regulated by ALKBH5. Furthermore, ALKBH5 and PD-L1 mRNA were shown to interact. ALKBH5 deficiency enriched m6A modification in the 3′UTR region of PD-L1 mRNA, thereby promoting its degradation in a YTHDF2-dependent manner. In vitro and in vivo, tumor-intrinsic ALKBH5 inhibited the expansion and cytotoxicity of T cells by sustaining tumor cell PD-L1 expression. The ALKBH5-PD-L1–regulating axis was further confirmed in human ICC specimens. Single-cell mass cytometry analysis unveiled a complex role of ALKBH5 in the tumor immune microenvironment by promoting the expression of PD-L1 on monocytes/macrophages and decreasing the infiltration of myeloid-derived suppressor-like cells. Analysis of specimens from patients receiving anti-PD1 immunotherapy suggested that tumors with strong nuclear expression patterns of ALKBH5 are more sensitive to anti-PD1 immunotherapy. Collectively, these results describe a new regulatory mechanism of PD-L1 by mRNA epigenetic modification by ALKBH5 and the potential role of ALKBH5 in immunotherapy response, which might provide insights for cancer immunotherapies. Significance: This study identifies PD-L1 mRNA as a target of ALKBH5 and reveals a role for ALKBH5 in regulating the tumor immune microenvironment and immunotherapy efficacy.

N6-Methyladenosine in Cancer Immunotherapy: An Undervalued Therapeutic Target

“…Knockdown of FTO sensitizes melanoma cells to interferon gamma (IFNg) and sensitizes melanoma to anti-PD-1 treatment in mice (125). Tumor-intrinsic ALKBH5 inhibited the expansion and cytotoxicity of T cells by sustaining tumor cell PD-L1 expression, which was further confirmed in human intrahepatic cholangiocarcinoma (ICC) specimens (83). Importantly, a small-molecule ALKBH5 inhibitor enhanced the efficacy of cancer immunotherapy in melanoma (82), suggesting that the combination of FTO/ALKBH5 inhibition with anti-PD-1 blockade may reduce the resistance to immunotherapy in melanoma.…”

“…ALKBH5, unlike FTO, is not changing the TME by affecting the IFN-g pathway since the population of CD8+ T lymphocytes remains unchanged when the demethylase is depleted (82). In addition, single-cell mass cytometry analysis unveiled a role of ALKBH5 in TME by promoting the expression of PD-L1 on monocytes/macrophages and decreasing the infiltration of MDSCs (83).…”

m6A Modification in Non-Coding RNA: The Role in Cancer Drug Resistance

“…found that tumor-intrinsic ALKBH5 stopped T-cell expansion and cytotoxicity by maintaining PD-L1 expression in tumor cells. Moreover, the results of clinical sample analysis from patients receiving anti-PD1 immunotherapy showed that strong nuclear expression patterns of ALKBH5 are more sensitive to anti-PD1 immunotherapy ( 155 ). Knockdown YTHDF1/2 in NSCLC cells could upregulate PD-L1 expression and multiple immune-related genes.…”

N6-Methyladenosine and Rheumatoid Arthritis: A Comprehensive Review

“…In addition, ALKBH5 regulated PD-L1 mRNA in intrahepatic cholangiocarcinoma (ICC). The lack of ALKBH5 decreased the expression of PD-L1 on monocytes-macrophages by a YTHDF2-dependent manner (81). Also, the deletion of the m6A demethylase Alkbh5 sensitized tumors to cancer immuno therapy (82).…”