T-细胞急性淋巴细胞白血病（T-ALL）是一种侵袭性的血液恶性肿瘤，其特点是具有不对称富集的白血病始终细胞（LIC）活性的受限细胞亚群。然而，目前尚不清楚哪些信号通路促进LIC的维持和进展。在本研究中，我们通过综合实验方法来评估生物钟在调控T-ALL中分子机制和信号通路对细胞动力学的影响。该方法包括对shRNA修饰的T-ALL细胞系的基因表达谱分析以及白血病细胞的染色质免疫沉淀测序（ChIP-Seq）。我们还对患者源性异种移植模型细胞亚群进行遗传操作，以评估人源T-ALL中生物钟缺失对LIC活性的调控作用。我们报告，在通过shRNA介导的CLOCK和BMAL1基因敲低破坏昼夜节律时钟电路的情况下，PDX移植入免疫缺陷宿主小鼠后，从PDX获得的LIC的体外生长和体内活性均受到负面影响。此外，基因表达数据与白血病细胞的ChIP-Seq数据显示，生物钟直接促进了关键参与JAK/STAT信号通路的基因（如IL20RB）的表达，使T-ALL细胞对白细胞介素20（IL20）的反应性增强。总之，我们的数据支持生物钟驱动IL20R表达的概念，从而促进JAK/STAT信号通路的激活，并促进T-ALL中的LIC活性。结果表明，有选择性地靶向生物钟组分可能对T-ALL患者的治疗具有重要意义。© 2023年。意大利国家癌症研究所“Regina Elena”。

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematological malignancy, characterized by restricted cellular subsets with asymmetrically enriched leukemia initiating cell (LIC) activity. Nonetheless, it is still unclear which signaling programs promote LIC maintenance and progression.Here, we evaluated the role of the biological clock in the regulation of the molecular mechanisms and signaling pathways impacting the cellular dynamics in T-ALL through an integrated experimental approach including gene expression profiling of shRNA-modified T-ALL cell lines and Chromatin Immunoprecipitation Sequencing (ChIP-Seq) of leukemic cells. Patient-derived xenograft (PDXs) cell subsets were also genetically manipulated in order to assess the LIC activity modulated by the loss of biological clock in human T-ALL.We report that the disruption of the circadian clock circuitry obtained through shRNA-mediated knockdown of CLOCK and BMAL1 genes negatively impacted the growth in vitro as well as the activity in vivo of LIC derived from PDXs after transplantation into immunodeficient recipient mice. Additionally, gene expression data integrated with ChIP-Seq profiles of leukemic cells revealed that the circadian clock directly promotes the expression of genes, such as IL20RB, crucially involved in JAK/STAT signaling, making the T-ALL cells more responsive to Interleukin 20 (IL20).Taken together, our data support the concept that the biological clock drives the expression of IL20R prompting JAK/STAT signaling and promoting LIC activity in T-ALL and suggest that the selective targeting of circadian components could be therapeutically relevant for the treatment of T-ALL patients.© 2023. Italian National Cancer Institute ‘Regina Elena’.