SUMO化修饰在急性髓系白血病发生发展中的作用及靶向研究

Post-translational modification (PTM) is an essential mechanism for regulating protein function. Targeting PTM, which is expected to reduce off-target effect caused by tumor cells heterogeneity, is one of the main strategies to solve the bottleneck problem of molecular therapy. SUMOylation, as a core mechanism of PTM, is closely correlated to tumorigenesis. Targeting SUMOyaltion has shown a favorable application potential in hematological malignancies. However, due to lacking of ideal animal models and large-scale analysis methods among recent studies, the regulation of SUMOylation in normal and malignant hematopoiesis is still poorly understood. In the present study, we aim to further evaluate the SUMOylation/DeSUMOylation effects on adult hematopoiesis and leukemogenesis by conducting two conditional knockout mice strains Ubc9fl/fl;Mx1Cre and Senp1fl/fl;Mx1Cre. In addition, we plan to screen out the targeted protein panels modified by SUMO paralogs in acute myeloid leukemia cells, through High Performance Liquid Chromatography Tandam Mass Spectrometry. Furthermore, we intend to utilize the biological targeting capabilities of microvesicles from CD123 CART cells which are active-loaded with the CRISPR/Cas9 system against leukemia cells to identify the possibility of precisely and specifically targeting SUMOylation process in leukemia. We believe that our study will have a unique scientific significance in the field, and improve the broad prospects in clinical application.

蛋白质翻译后修饰 (PTM) 是蛋白质功能调节的重要机制。靶向PTM有望突破肿瘤异质性所致的不可靶向性，是解决分子治疗瓶颈的主要策略之一。SUMO化作为一种PTM的核心机制，其异常与肿瘤发生发展密切相关。靶向SUMO化修饰，已在血液肿瘤中初显良好应用潜力。然而，由于缺乏理想的动物模型和高通量技术分析手段，国内外对SUMO化机制的研究尚在起步阶段，对正常和恶性造血调控更是知之甚少。本项目运用条件性敲除小鼠模型Ubc9fl/fl;Mx1Cre和Senp1fl/fl;Mx1Cre 系统观察评估SUMO/DeSUMO化对正常成体造血和白血病发生发展的影响；通过SUMO化液相质谱筛查白血病特异性下游靶标；利用CD123CAR-T细胞中微泡的天然生物学靶向特性，负载白血病特异性SUMO化CRISPR/Cas9基因编辑载体，探索实现靶向白血病细胞SUMO化修饰的可能性，具有重要科学意义和广阔的应用前景。

靶向蛋白质翻译后修饰这一肿瘤细胞共享生存机制，有望解决肿瘤异质性造成的治疗瓶颈问题，成为一种新的治疗策略。近些年来，虽然关于SUMO化修饰在血液肿瘤中的研究越来越多，但是目前研究对整体SUMO化修饰水平及SUMO化修饰途径相关酶在肿瘤中的作用尚不清楚。UBC9作为SUMO化修饰途径中唯一的E2结合酶，可以调控机体整体SUMO化修饰水平。通过对数据库中病⼈资料的分析，我们发现UBC9在⾼危组AML病⼈中相对于低危组病⼈低表达，且⾼表达Ubc9病⼈⽣存期低于低表达组，另外我们检测了本中心86例AML患者UBC9的表达量，发现UBC9低表达与AML患者高危险分层及初诊时肝 /脾肿大相关。通过 Ubc9条件性敲除小鼠模型 ，发现Ubc9单倍剂量不足可以抑制 MLL-AF9小鼠白血病细胞的凋亡，促进其侵袭并增加恶性程度。同时在体外也发现UBC9基因敲降后可以导致人AML细胞系活性和侵袭能力的增强。UBC9基因敲降在体内和体外均可以促进 AML的进展，可能与UBC9基因敲降使 PARP1的 SUMO化修饰水平发生下降，从而激活 NF-κB信号通路有关。我们的研究表明UBC9有望为AML的治疗提供新策略。为了明确靶向白血病特异性SUMO化靶点的可能性，我们构建了CAR-囊泡介导的CRISPR/Cas9靶向基因编辑平台并进行了有效性验证。