Supplementation with a-ketoglutarate improved the efficacy of anti-PD1 melanoma treatment through epigenetic modulation of PD-L1

Patients with advanced melanoma have shown an improved outlook after receiving anti-PD1 therapy, but the low response rate restricts clinical benefit; therefore, enhancing anti-PD1 therapy efficacy remains a major challenge. Here, our findings show significantly higher abundance of a-KG in healthy controls, anti-PD1-sensitive melanoma-bearing mice, and anti-PD1-sensitive melanoma patients; moreover, supplementation with a-KG enhanced the efficacy of anti-PD1 immunotherapy and increased PD-L1 expression in melanomas via STAT1/3. We also found that supplementation with a-KG significantly increased methylcytosine dioxygenase TET2/3, which led to an increased 5-hydroxymethylcytosine (5-hmC) level in the PD-L1 promoter. As a consequence, STAT1/3 had been recognized and stabilized in PD-L1 promoter to upregulate PD-L1 expression. Importantly, single-cell sequencing of preclinical samples and analysis of clinical data revealed that the TET2/3-STAT1/3-CD274 signaling was associated with sensitivity to anti-PD1 treatment in melanoma. Taken together, we provide a novel insight into a-KG's function in melanoma anti-PD1 treatment and supplement of a-KG is a novel promising strategy to improve the efficacy of anti-PD1 therapy. Overall design: For the tumor immunotherapy model, B16F10 (100 µl RPMI 1640 containing 5 × 10^5 cells) were collected and injected into the right abdomen of female C57BL/6 mice (7-8-week-old) (SLAC Laboratory Animal Co., Ltd., Shanghai, China). When the tumor volume reached approximately 50 cubic millimeters, the tumors were randomly grouped (six in each group) and injected intraperitoneally with corn oil (vehicle) (Aladdin, China), 100 mg/kg a-KG (Sigma-Aldrich, USA), IgG2a (BioXCell, BE0089, USA) or PD1 mAb (BioXCell, BE0146, USA) for 9-11 days.Tumor tissues were collected from the vehicle, a-KG, anti-PD1 mAb and a-KG+anti-PD1 groups of mice, with four independent tumor tissue samples collected from each group.