USP47在慢性粒细胞性白血病发病和治疗中的作用和机制

Ubiquitin specific proteases (USP) play an important role in regulating many cellular processes including cell cycle, signal transduction, immune response and DNA repair. Recently, it has been shown that many USP are closely associated with the development of cancer. And USP is becoming an attactive cancer targets. In this project, we found that USP47 expressed at higher level in CML bone marrow mononuclear cells than their normal counterpart. The expression of USP47 is changed with the treatment of tyrosine kinase inhibitor imatinib. Imatinib treatment resulted in the decrease of USP47. However, when the relapse occurred, the expression of USP47 were upregulated again. BCR/ABL, the driver force of CML, could significantly upregulate the expression of USP47. When the expression or the activity of USP47 was inhibited by RNA interference or by USP47 inhibitor, the proliferation and tumor formation of K562 cells were greatly reduced in vitro and in vivo. USP47 inhibitor could also inhibit the proliferation of primary CML cells and the colony formation of CML CD34+ cells. Based on these findings, we will next investigate the role of USP47 in the development of CML using the USP47 knockout mice as a mouse model, identify the interaction protein of USP47 from K562 cells with immunoprecipitation and mass spectrometry analysis, elucidate how BCR/ABL upregulates the expression of USP47 and identify novel USP47 inhibitors. These studies may shed new light on the pathogenesis of CML, provide novel drug target and novel USP47 targeting lead compounds for the treatment of CML.

近年来的研究发现，多种泛素化特异蛋白酶（USPs）分子与肿瘤的发生和发展密切相关，可望成为肿瘤治疗的新的靶点。最近，我们研究发现USP家族成员—USP47在慢性粒细胞白血病（CML）细胞中高表达，并且其表达水平和CML患者对酪氨酸激酶抑制剂的疗效密切相关；CML相关融合蛋白BCR/ABL显著上调USP47的表达，而利用小分子干扰RNA或小分子抑制化合物抑制USP47可在体内、外显著抑制CML细胞增殖，抑制原代CML CD34+细胞的克隆形成能力。本项目拟在这些重要发现的基础上，进一步应用USP47基因敲除小鼠模型，在分子、细胞和整体动物水平上研究USP47在CML发病和治疗中的作用和机制，尤其是对CML白血病干细胞的作用，可望为CML的发病机制研究提供新的见解，并为其治疗提供新的靶标和先导化合物，为研发治疗CML的新策略提供重要基础。

为克服对酪氨酸激酶抑制剂（TKIs）的耐药性（包括BCR-ABL依赖性和非依赖性耐药），寻找新的药物靶点是治疗慢性粒细胞白血病（CML）所必需的。我们发现去泛素化酶USP47是一种在原代CML细胞中高表达的蛋白，是克服TKI耐药的潜在靶点。USP47在深度分子生物学缓解患者中的高表达预示着其具有较大复发的可能性。USP47在CML细胞中可通过RAS/ERK和STAT5信号通路激活上调。功能分析表明，USP47基因敲除抑制了体内外对伊马替尼敏感或耐药的CML细胞的增殖。Usp47基因敲除显著抑制BCR-ABL和BCR-ABL（T315I突变）诱导的小鼠CML的发生发展，并伴有Lin-Sca1+c-Kit+CML干/祖细胞减少。机制研究表明，USP47通过稳定Y-box结合蛋白1促进CML细胞DNA损伤修复。P22077对TKI敏感或耐药的CML细胞及原代CML细胞均具有较好的杀伤作用，并降低原代CML CD34+细胞的集落形成。P22077的使用也显著提高了CML小鼠的存活时间。此外，P22077能够减少BCR-ABL诱导的CML小鼠的白血病干/祖细胞和微小残留病变。综上所述，靶向USP47是克服TKI抵抗和根除CML白血病干/祖细胞的新策略。