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.

中文翻译：

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在缺氧条件下，FTO 激活 PD-L1 通过 m6A/YTHDF3/PDK1 轴促进乳腺癌的免疫抑制

 介绍

改变的表观遗传重编程使乳腺癌细胞能够适应低氧应激。缺氧微环境会改变免疫细胞的浸润和功能，从而限制免疫治疗的有效性。

 目标

该研究旨在确定脂肪量和肥胖相关蛋白 （FTO） 如何帮助乳腺癌细胞应对缺氧微环境以及乳腺癌细胞对肿瘤免疫产生抵抗力的机制。

 方法

利用临床样本分析 FTO 对乳腺癌进展的影响以及程序性细胞死亡蛋白 1/程序性细胞死亡 1 配体 1 （PD-1/PD-L1） 免疫检查点抑制剂治疗的影响。利用 MeRIP-seq 和 mRNA-seq 分析缺氧下 FTO 调控的下游基因。使用 RIP 澄清 FTO 对 PDK1 的甲基化修饰调控。然后利用小鼠模型分析抑制 FTO 和 3-磷酸肌醇依赖性蛋白激酶 1 （PDK1） 联合 PD-1/PD-L1 免疫检查点抑制剂治疗的疗效。

 结果

N6-甲基腺苷 （m⁶A） 脱甲基化酶 FTO 被缺氧诱导因子 1α （HIF-1α） 转录激活。通过高通量测序，PDK1 被确定为低氧条件下 FTO 的潜在靶点。从机制上讲，FTO 过表达减少了 PDK1 mRNA 上的 m⁶A 修饰位点，阻止 YTH 结构域家族 3 （YTHDF3） 识别和结合这些位点，从而抑制 PDK1 mRNA 的降解。PDK1 过表达激活 AKT/STAT3 通路，导致 PD-L1 表达增强。用 FB23 和 BX-912 靶向 FTO 和 PDK1-AKT 通路可抑制乳腺癌生长，增强细胞毒性 T 淋巴细胞 （CTL） 活性，并增强 PD-1/PD-L1 检查点抑制剂 Atezolizumab 的有效性。

 结论

本研究揭示了 HIF-1α 在缺氧条件下促进 FTO 转录，从而通过 PDK1/AKT/STAT3 轴增加 PD-L1 的表达。在低氧条件下抑制 FTO 和 PDK1 可能是一种很有前途的乳腺癌免疫治疗策略。