B-cell acute lymphoblastic leukemia (B-ALL) is an aggressive malignancy characterized by the aberrant accumulation of immature and dysfunctional B cells in bone marrow (BM). Although chemotherapy and other therapies have been widely applied, some patients such as relapsed or drug-refractory (R/R) B-ALL patients exhibit limited response. YT521-B homologous domain-containing protein 1 (YTHDC1) is a nuclear reader of N⁶-methyladenosine (m⁶A) RNA modification, which has been implicated in different malignancies including leukemia. In the current study, we show that YTHDC1 is highly expressed in B-ALL cells. YTHDC1 knockdown attenuated B-ALL cell proliferation and cell cycle progression in vitro, and prolonged survival of mice in the human B-ALL xenograft model in vivo attributable to compromised leukemogenesis. Mechanistically, YTHDC1 knockdown significantly increased the accumulation of endogenous and chemotherapeutic agents-induced DNA damage in B-ALL cells. Furthermore, we identified that YTHDC1 binds to and stabilizes m⁶A-modified KMT2C mRNA. KMT2C is a key enzyme catalyzing histone H3K4 methylation required for the expression of DNA damage response (DDR)-related genes, implying that YTHDC1 inhibitors might improve chemotherapy by attenuating DDR via reducing KMT2C. Indeed, with molecular docking and biochemical experiments, we identified EPZ-5676 as a YTHDC1 inhibitor, and combination of EPZ-5676 with Cytarabine (Ara-c) significantly improved the efficacy of chemotherapy in B-ALL mouse models using YTHDC1^high primary and lined B-ALL cells. Collectively, YTHDC1 is required for DDR in B-ALL cells by upregulating DDR-related gene expression via stabilizing m⁶A-modified KMT2C mRNA, thereby leading to increased histone H3K4 methylation, and targeted inhibition of YTHDC1 is a potentially new therapeutic strategy against B-ALL, especially YTHDC1^high B-ALL.

B 细胞急性淋巴细胞白血病 （B-ALL） 是一种侵袭性恶性肿瘤，其特征是未成熟和功能失调的 B 细胞在骨髓 （BM） 中异常积累。尽管化疗和其他疗法已得到广泛应用，但一些患者，如复发或药物难治性 （R/R） B-ALL 患者表现出有限的反应。YT521-B 同源结构域包含蛋白 1 （YTHDC1） 是 N⁶-甲基腺苷 （m⁶A） RNA 修饰的核读取器，与包括白血病在内的不同恶性肿瘤有关。在目前的研究中，我们表明 YTHDC1 在 B-ALL 细胞中高度表达。YTHDC1 敲除在体外减弱了 B-ALL 细胞增殖和细胞周期进程，并延长了人 B-ALL 异种移植模型中小鼠的体内存活率，这归因于白血病发生受损。从机制上讲，YTHDC1 敲除显著增加了 B-ALL 细胞中内源性和化疗药物诱导的 DNA 损伤的积累。此外，我们发现 YTHDC1 结合并稳定 m⁶A 修饰的 KMT2C mRNA。KMT2C 是催化组蛋白 H3K4 甲基化的关键酶，是 DNA 损伤反应 （DDR） 相关基因表达所必需的，这意味着 YTHDC1 抑制剂可能通过减少 KMT2C 来减弱 DDR，从而改善化疗。事实上，通过分子对接和生化实验，我们确定 EPZ-5676 是一种 YTHDC1 抑制剂，EPZ-5676 与阿糖胞苷 （Ara-c） 的组合显着改善了使用 YTHDC1^高原代和衬里 B-ALL 细胞的 B-ALL 小鼠模型中的化疗疗效。 总的来说，YTHDC1 是通过稳定 m⁶A 修饰的 KMT2C mRNA 上调 DDR 相关基因表达，从而导致组蛋白 H3K4 甲基化增加，并且靶向抑制 YTHDC1 是一种针对 B-ALL，尤其是 YTHDC1^高 B-ALL 的潜在新治疗策略。