The hypomethylating agent 5-azacytidine (AZA) is the first-line treatment for AML patients unfit for intensive chemotherapy. The effect of AZA results in part from T-cell cytotoxic responses against MHC-I-associated peptides (MAPs) deriving from hypermethylated genomic regions such as cancer-testis antigens (CTAs), or endogenous retroelements (EREs). However, evidence supporting higher ERE MAPs presentation after AZA treatment is lacking. Therefore, using proteogenomics, we examined the impact of AZA on the repertoire of MAPs and their source transcripts. AZA-treated AML upregulated both CTA and ERE transcripts, but only CTA MAPs were presented at greater levels. Upregulated ERE transcripts triggered innate immune responses against double-stranded RNAs but were degraded by autophagy, and not processed into MAPs. Autophagy resulted from the formation of protein aggregates caused by AZA-dependent inhibition of DNMT2. Autophagy inhibition had an additive effect with AZA on AML cell proliferation and survival, increased ERE levels, increased pro-inflammatory responses, and generated immunogenic tumor-specific ERE-derived MAPs. Finally, autophagy was associated with a lower abundance of CD8⁺ T-cell markers in AML patients expressing high levels of EREs. This work demonstrates that AZA-induced EREs are degraded by autophagy and shows that inhibiting autophagy can improve the immune recognition of AML blasts in treated patients.

去甲基化剂 5-氮杂胞苷 (AZA) 是不适合强化化疗的 AML 患者的一线治疗药物。 AZA 的作用部分是由于 T 细胞对源自高甲基化基因组区域的 MHC-I 相关肽 (MAP) 的细胞毒性反应，例如癌睾丸抗原 (CTA) 或内源性逆转录元件 (ERE)。然而，缺乏支持 AZA 治疗后 ERE MAP 升高的证据。因此，利用蛋白质基因组学，我们检查了 AZA 对 MAP 及其源转录本的影响。 AZA 治疗的 AML 上调了 CTA 和 ERE 转录本，但只有 CTA MAP 的水平更高。上调的 ERE 转录物触发了针对双链 RNA 的先天免疫反应，但会被自噬降解，并且不会被加工成 MAP。自噬是由 DNMT2 的 AZA 依赖性抑制引起的蛋白质聚集体的形成所致。 AZA 的自噬抑制对 AML 细胞增殖和存活、增加 ERE 水平、增加促炎反应以及产生免疫原性肿瘤特异性 ERE 衍生 MAP 具有附加作用。最后，在表达高水平 ERE 的 AML 患者中，自噬与 CD8 ⁺ T 细胞标记丰度较低有关。这项工作表明，AZA 诱导的 ERE 会被自噬降解，并表明抑制自噬可以提高治疗患者对 AML 母细胞的免疫识别。