FTO activates PD-L1 promotes immunosuppression in breast cancer via the m6A/YTHDF3/PDK1 axis under hypoxic conditions

Abstract

Introduction

Altered epigenetic reprogramming enables breast cancer cells to adapt to hypoxic stress. Hypoxic microenvironment can alter immune cell infiltration and function, limiting the effectiveness of immunotherapy.

Objectives

The study aimed to identify how fat mass and obesity-associated protein (FTO) helps breast cancer cells cope with the hypoxic microenvironment and the mechanisms behind breast cancer cell resistance to tumor immunity.

Methods

Clinical samples were utilized to analyze the impact of FTO on breast cancer progression and the effect of programmed cell death protein 1/ programmed cell death 1 ligand 1(PD-1/PD-L1) immune checkpoint inhibitor treatment. Utilized MeRIP-seq and mRNA-seq to analyze the downstream genes regulated by FTO under hypoxia. Methylation modification regulation of PDK1 by FTO was clarified using RIP. Then mouse models were utilized to analyze the efficacy of inhibiting FTO and 3-Phosphoinositide-dependent protein kinase 1(PDK1) in combination with PD-1/PD-L1 immune checkpoint inhibitor treatment.

Result

N6-Methyladenosine(m⁶A) demethylase FTO was transcriptionally activated by hypoxia inducible factor 1α(HIF-1α). PDK1 was identified as a potential target of FTO under hypoxic conditions through high-throughput sequencing. Mechanistically, overexpression of FTO decreases m⁶A modification sites on PDK1 mRNA, preventing YTH domain family 3(YTHDF3) from recognizing and binding to these sites, thereby inhibiting the degradation of PDK1 mRNA. Overexpression of PDK1 activates the AKT/STAT3 pathway, leading to enhanced PD-L1 expression. Targeting the FTO and PDK1-AKT pathways with FB23 and BX-912 inhibit breast cancer growth, enhance cytotoxic T lymphocyte (CTL) activity, and enhance the effectiveness of the PD-1/PD-L1 checkpoint inhibitor Atezolizumab.

Conclusion

This study reveals that HIF-1α promotes FTO transcription under hypoxic conditions, thereby increasing PD-L1 expression through the PDK1/AKT/STAT3 axis. Inhibition of FTO and PDK1 under hypoxic conditions could serve as a promising immunotherapeutic strategy for breast cancer.