生长抑制特异蛋白GAS2促急性T淋巴细胞白血病细胞生长的研究

T-cell acute lymphoblastic leukemia (T-ALL) is a heterogeneous disease as multiple genetic abnormalities may occur in one patient. Although prognosis has been improved by current chemotherapy, some patients still suffer from drug resistance and relapse. The constitutive activation of NOTCH is considered as the most prominent factor to initiate T-ALL, recently the important role of CXCL12/CXCR4 axis in the disease maintenance has been reported. Growth arrest-specific 2 (GAS2) is a component of microfilament as well as the endogenous inhibitor of calpain2, which plays important role in regulations of cell-morphology, cell-cycle and apoptosis. Our data have shown that GAS2 is aberrantly expressed in various leukemic cells, which is a novel biomarker of leukemia. GAS2 silencing impaired the in vitro growth and migration of Jurkat cells and induced apoptosis of Jurkat cells as well. Overexpression of GAS2 promoted the growth of HPB-ALL cells and enhanced their leukemogenesis ability in immunodeficient mice (NSG mice). Mechanistically, GAS2 regulated the expression of CXCR4 post-transcriptionally. In the proposal, the effect of GAS2 on the growth, migration, apoptosis and leukemogenesis ability of T-ALL cells will be investigated, the role of CXCR4 in GAS2 promoted T-ALL growth will be delineated, and how GAS2 regulates the expression of CXCR4 will be studied. The performance of these studies will deepen the understanding of the molecular pathogenesis of T-ALL and enrich the knowledge of the molecular function of GAS2 protein, which possibly leads to better management of the disease.

药物耐受和复发仍困扰部分急性T淋巴细胞白血病（T-ALL）患者，对T-ALL致病机制的深入理解将有助改善治疗。文献表明CXCR4对维持T-ALL至关重要。生长抑制特异蛋白GAS2是微丝组分和钙蛋白酶（Calpain2）内源抑制剂；参与细胞形态、周期和凋亡的调控。我们发现GAS2是多种白血病的新生物标志物，在T-ALL中异常高表达。但GAS2在T-ALL中的功能和作用机制尚不明确。预研显示GAS2沉默抑制Jurkat细胞生长、迁移并诱发凋亡；过表达GAS2促进HPB-ALL细胞体外生长和在免疫缺陷小鼠（NSG）体内生成白血病的能力；GAS2在转录后水平调控CXCR4表达。本项目将在体内外研究GAS2对T-ALL细胞多种生物学性状的影响；明晰CXCR4在GAS2行使其功能中的作用；解析GAS2调控CXCR4表达的分子机制。将有助于深化对T-ALL致病机制的认知，并为改善该疾病治疗提供新思路。

急性T淋巴细胞白血病（T-cell acute lymphoblastic leukemia, T-ALL）是一种异质性恶性血液病，化疗或骨髓移植是该疾病的主要治疗方法，尚缺乏有效靶向治疗药物。因此，研究T-ALL细胞生长/生存的分子机制，可能为T-ALL的治疗提供新线索。生长抑制特异蛋白GAS2（Growth arrest-specific 2）是微丝成分，也是钙蛋白酶（Calpain2）的内源性抑制剂。我们的前期研究显示GAS2在T-ALL细胞中高表达，且促进这些细胞的体外增殖，但GAS2异常高表达的原因、GAS2对T-ALL细胞体内生长的影响尚不清晰且GAS2在正常造血中的作用也不明确；此外，对GAS2在白血病细胞中作用机制的理解也非常有限。针对这些问题，本项目发现：（1）GAS2在T-ALL患者骨髓细胞中异常高表达，部分与GAS2启动子区域的低甲基化（hypomethylation）修饰相关；（2）GAS2沉默显著抑制Jurkat细胞在免疫缺陷小鼠中的成白血病能力；（3）GAS2沉默特异性地抑制T-ALL患者骨髓细胞的增殖而对正常骨髓细胞增殖影响较小；（4）免疫沉淀和荧光显微术显示GAS2与CXCR4相互作用，且GAS2通过Calpain2调控CXCR4蛋白表达，研究证实GAS2/CXCR4轴调控T-ALL细胞的生长、迁移和成白血病能力；（5）依据对照和GAS2沉默Jurkat细胞RNA-Seq数据的提示，证实沉默GAS2或CXCR4都抑制NOTCH1/c-MYC的表达，且c-MYC过表达可以部分“挽救”GAS2沉默或CXCR4沉默导致的生长抑制；（6）造血细胞中特异缺陷Gas2并未对小鼠造血发生和T-细胞生成造成明显影响，提示GAS2可能是T-ALL治疗的新靶标。综上，项目的实施加深对T-ALL细胞生长/生存分子机制的认知，可能为T-ALL治疗提供新线索。