A Novel Oncogenic Role for Cdc20 Implications in Adult T Cell Leukemia Treatment

描述（由申请人提供）：在多种人类癌症中观察到Cdc20的过表达或超激活。在成人t细胞白血病（ATL）患者中，病毒癌蛋白Tax部分通过激活关键的M期驱动因子APCCdc20来促进细胞转化，从而增强增殖，并如本研究所提出的那样，避免细胞凋亡。本研究的目的是揭示APCCdc20泛素E3连接酶复合物在允许癌细胞逃避化疗药物诱导的细胞凋亡，从而促进白血病发生和赋予多药耐药中的新致癌作用。我的初步数据显示，在Cdc20最活跃的M期，促凋亡蛋白Bim的蛋白水平较低。此外，内源性Cdc20缺失后，多个细胞系中Bim蛋白丰度显著升高，但mRNA水平未显著升高。进一步研究发现，Cdc20直接与Bim结合，并促进其泛素化，随后被26S蛋白酶体降解。引人注目的是，在Cdc20活性升高的人类T细胞嗜淋巴病毒I型（HTLV-I）感染的T细胞系中，Bim的表达非常低，并且这些细胞始终对各种抗有丝分裂剂和DNA损伤剂具有抗性。在本提案中，我假设Cdc20依赖性的Bim抑制对于ATL细胞逃避凋亡至关重要，特别是在治疗药物的挑战下。因此，抑制Cdc20活性或引入bh3模拟试剂可使ATL细胞在体内外对常规化疗药物增敏。在本提案中，我计划：1)将APCCdc20描述为负调节Bim稳定性的上游E3连接酶；2)确定APCCdc20活化在抑制细胞凋亡中的生理作用；3)确定靶向抑制Cdc20或引入bh3模拟物是否能抑制ATL或其他过表达Cdc20的癌症（包括头颈癌）在体内的发展。我职业生涯的长期目标是应用分子和细胞生物学的见解来理解在人类恶性肿瘤发展中重要的解除管制的蛋白质水解途径的生理意义，并寻找适当的药物靶点。这个K99/R00奖将为我提供保护的时间来追求这个提案的新颖假设，获得新的技能来执行实验和解决问题。此外，该奖项将使我能够集中精力独立进行基础和转化研究，并培养未来的年轻科学家。

DESCRIPTION (provided by applicant): Overexpression or hyper-activation of Cdc20 has been observed in a variety of human cancers. In adult T-cell leukemia (ATL) patients, the viral oncoprotein Tax promotes cellular transformation in part through activating APCCdc20, a critical M phase driver, to enhance proliferation, and as proposed in this study, to evade apoptosis. The goal of this proposal is to uncover a novel oncogenic role of the APCCdc20 ubiquitin E3 ligase complex in allowing cancer cells to evade chemotherapeutic agents-induced cellular apoptosis, thereby promoting leukemia-genesis and conferring multidrug resistance. My preliminary data showed that the protein levels of the pro-apoptotic protein, Bim, were low in M phase when Cdc20 is most active. Moreover, Bim protein abundance but not mRNA level was significantly elevated in multiple cell lines after depletion of endogenous Cdc20. Further studies revealed that Cdc20 directly binds to Bim and promotes its ubiquitination and subsequent degradation by the 26S proteasome. Strikingly, Bim expression is remarkably low in Human T-cell Lymphotropic Virus type I (HTLV-I)-infected T cell lines with elevated Cdc20 activity, and consistently, these cells are resistant to various anti-mitotic and DNA damage agents. In this proposal, I hypothesize that Cdc20- dependent repression of Bim is critical for ATL cells to evade apoptosis, especially under the challenge of therapeutic drugs. Therefore, inhibiting Cdc20 activity or introducing BH3-mimetic reagents could sensitize ATL cells to conventional chemotherapeutic drugs both in vitro and in vivo. In this proposal, I plan to: 1) Characterize APCCdc20 as an upstream E3 ligase that negatively regulates Bim stability; 2) Determine the physiological role of APCCdc20 activation in suppressing apoptosis; 3) Determine whether targeted inhibition of Cdc20 or introducing BH3-mimetic suppresses the development of ATL or other Cdc20-overexpressing cancer including head and neck cancer in vivo. The long-term goals of my career are to apply the insights of molecular and cellular biology to understand the physiological significance of deregulated proteolytic pathways that are important in the development of human malignancies, and to search for proper druggable targets. This K99/R00 award will provide protected time for me to pursue the novel hypotheses of this proposal, obtain new skill sets to execute experiments and solve problems. In addition, the award will allow me to focus my efforts on independently conducting basic and translational research, and to train future young scientists.