RNA helicase DHX15 exemplifies a unique dependency in acute leukemia. RNA helicase DHX15 exemplifies a unique dependency in acute leukemia

RNA-binding proteins (RBPs) have emerged as essential regulators to control gene expression and modulate multiple cancer traits. T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematological malignancy derived from transformation of T-cell progenitors that normally undergo discrete steps of differentiation in the thymus. Yet implications of essential RBPs during T cell neoplastic transformation remain largely unexplored. Systematic evaluation of RBPs identifies the RNA helicase DHX15, a key subunit in the intron lariat spliceosome (ILS) complex, as a potential T-ALL dependency factor. Using cell-based functional assays and the NOTCH1-induced murine T-ALL model, we demonstrate the essential importance of DHX15 in tumor cell survival and leukemogenesis. Moreover, single-cell transcriptomics reveals that DHX15 depletion in T-cell progenitors impairs normal T-cell development and, more specifically, hinders burst proliferation during CD4-CD8-(DN)-to-CD4+CD8+(DP) transition. Mechanistically, abrogation of DHX15 induces altered RNA splicing and downregulation of SLC7A6 and SLC38A5, thereby suppressing glutamine import and mTORC1 activity. Sustained mTORC1 hyperactivation significantly rescues the DHX15-deficient phenotypes, supporting mTORC1 as a prominent downstream effector mediating the pro-proliferative role of DHX15. Moreover, we propose a DHX15 signature modulator drug ciclopirox and demonstrate the prominent anti-T-ALL efficacy. We therefore highlight DHX15 as a crucial regulator during T-cell leukemogenesis, and provide mechanistic insights that DHX15, although functioning at the end of splicing catalysis, modulates a subset of pathogenic mis-splicing events. These findings exemplify a promising therapeutic approach that splicing perturbation by targeting DHX15 may achieve robust anti-tumor efficacy. Overall design: We used human T-ALL as a model system, and performed multi-omics analysis including RNA-seq and RIP-seq to study the regulatory mechanism of RNA-helicase DHX15. Meanwhile, to further clarify the heterogeneities of thymocyte subsets and the abnormal gene expression profiles in the absence of Dhx15, we performed single-cell RNA sequencing (scRNA-seq) using 10X Genomics platform on isolated single cells from thymus with or without Dhx15 expression

RNA结合蛋白（RNA-binding proteins, RBPs）已被证实为一类关键的基因表达调控因子，可调控多种癌症相关表型。T细胞急性淋巴细胞白血病（T-cell acute lymphoblastic leukemia, T-ALL）是一种侵袭性血液系统恶性肿瘤，起源于正常情况下在胸腺中经历阶段性分化的T细胞祖细胞的恶性转化。然而，关键RBPs在T细胞肿瘤性转化过程中的作用仍未得到充分研究。
通过系统筛选RBPs，本研究发现RNA解旋酶DHX15——内含子套索剪接体（intron lariat spliceosome, ILS）复合物的关键亚基——是潜在的T-ALL依赖型靶点。借助基于细胞的功能实验与NOTCH1诱导的小鼠T-ALL模型，我们证实了DHX15在肿瘤细胞存活及白血病发生过程中的核心重要性。
此外，单细胞转录组学分析显示，在T细胞祖细胞中敲除DHX15会损害正常T细胞发育，更具体地说，会阻滞CD4-CD8-（双阴性，DN）向CD4+CD8+（双阳性，DP）转化阶段的爆发性增殖。机制层面，DHX15的功能缺失会引发RNA剪接异常，并下调SLC7A6与SLC38A5的表达，进而抑制谷氨酰胺摄取与mTOR复合物1（mTORC1）的活性。持续激活mTORC1可显著挽救DHX15缺失导致的细胞表型，证实mTORC1是介导DHX15促增殖功能的关键下游效应分子。
此外，我们鉴定出一种靶向DHX15调控特征的调节剂环吡酮（ciclopirox），并证实其具有显著的抗T-ALL疗效。综上，本研究明确DHX15是T细胞白血病发生过程中的关键调控因子，并揭示了其作用机制：尽管DHX15在剪接催化的终末阶段发挥功能，但它可调控一组致病性异常剪接事件。上述研究结果表明，靶向DHX15的剪接干扰策略有望实现高效的抗肿瘤疗效。
总体实验设计：本研究以人源T-ALL为模型系统，开展包括RNA测序（RNA-seq）与RNA免疫沉淀测序（RIP-seq）在内的多组学分析，以解析RNA解旋酶DHX15的调控机制。同时，为进一步阐明胸腺细胞亚群的异质性以及Dhx15缺失时的异常基因表达谱，我们借助10X基因组学（10X Genomics）平台，对来自表达或缺失Dhx15的胸腺组织的分离单细胞进行了单细胞RNA测序（single-cell RNA sequencing, scRNA-seq）。