Targeting Mitotic Kinases Inhibitors for Cancer Therapy

描述(申请人提供)：伊马替尼最近成功地用于治疗费城染色体阳性的慢性髓细胞白血病，使酪氨酸和丝氨酸/苏氨酸激酶成为癌症治疗的主要靶点。由于目前只有一小部分人类基因组可以被合理选择和有效的抑制剂靶向，因此迫切需要开发有效的策略来发现和优化新的抑制剂。为了实现这一目标，我们最近开发了一个三磷酸腺苷竞争性激酶抑制剂的化合物文库。使用我们收集的新化合物(约2,000种)，我们测试了16个培养的肿瘤细胞系的这些化合物诱导凋亡死亡的能力。这项研究揭示了一种化合物ON1231320，它对所有16种肿瘤细胞株都有显著的细胞毒性，对正常细胞几乎没有或几乎没有细胞毒性。对一组285个激酶的抑制分析表明，该化合物对PLK2/SNK具有显著的特异性，PLK2/SNK是一种参与中心体复制和有丝分裂进程的激酶。最重要的是，PLK2最近被认为是连接细胞代谢和细胞周期的一种蛋白。在癌细胞中经常观察到线粒体功能障碍，并增加对糖酵解的依赖(称为Warburg效应)，识别在线粒体功能障碍条件下促进细胞存活的途径具有治疗意义。最近的一项研究表明，靶向去除人结肠癌细胞系HCT116中的SCO2基因会导致线粒体呼吸的消融，而PLK2是SCO2-/-细胞中表达最高的基因。即使线粒体呼吸缺陷的人类癌细胞中PLK2水平略有下降，也会导致它们在小鼠体内形成异种移植的能力丧失。我们的结果表明，ON1231320是一种有效的肿瘤细胞死亡诱导剂，在体内具有良好的安全性。我们建议研究该化合物在体外和体内对肿瘤生长的影响，以确定ON1231320将如何作为一种新的癌症化疗药物。该方案的目的是：1.确定ON01231320抑制PLK2的动力学；2.确定ON1231320对PLK2介导的下游信号转导事件的影响，并评价其在缺氧、遗传毒性和呼吸缺陷的情况下对肿瘤细胞生长的影响；3.鉴定ON1231320在异种移植模型中的PK特性，以确定抑制肿瘤生长所需的PLK2的程度，并评估ON1231320的浓度与化合物的抗肿瘤活性的关系，以及评价以PBMN为替代物的有效性；4.进一步探讨ON1231320联合奥沙利铂、5-氟尿嘧啶、紫杉醇、阿霉素等细胞毒性抗癌药物诱导乳腺癌和结直肠癌模型细胞凋亡和肿瘤消退的作用。

DESCRIPTION (provided by applicant): The recent success of imatinib for the treatment of Philadelphia chromosome positive chronic myelogenous leukemia has made tyrosine and serine/threonine kinases major targets for cancer therapy. As only a small fraction of the human kinome can currently be targeted by a reasonably selective and potent inhibitor, there is an urgent need to develop strategies for efficient discovery and optimization of new inhibitors. Towards this goal, we have recently developed a compound library of ATP-competitive kinase inhibitors. Using our collection of novel compounds (approximately 2,000), we tested a panel of 16 cultured tumor cell lines for the ability of these compounds to induce apoptotic death. This search revealed a compound, ON1231320, that had remarkable cytotoxicity against the entire panel of 16 tumor cell lines, with little or no cytotoxicity towards normal cells. Kinase inhibition assays against a panel of 285 kinases revealed that this compound had a remarkable specificity towards Plk2/Snk, a kinase involved in centrosome duplication and mitotic progression. Most importantly, Plk2 has recently been implicated as one of the kinases that links cellular metabolism to cell cycle. Mitochondrial dysfunction with increased dependence on glycolysis is frequently observed in cancer cells (known as the Warburg effect) and identification of pathways that promote cell survival under conditions of mitochondrial dysfunction have therapeutic implications. In a recent study, it has been shown that targeted ablation of SCO2 gene in HCT116 human colon cancer cell line results in the ablation of mitochondrial respiration and that PLK2 is the most highly expressed gene in SCO2-/-cells. Even a modest reduction in PLK2 levels in human cancer cells with defects in mitochondrial respiration results in the elimination of their ability to form xenografts in mice. Our results show that ON1231320 is a potent inducer of tumor cell death and has an excellent safety profile in vivo. We propose to study the effects of this compound on tumor growth in vitro and in vivo to determine how ON1231320 will serve as a novel cancer chemotherapeutic. The aims of the proposal are: 1. To determine the kinetics of inhibition of Plk2 by ON01231320; 2. To determine the effects of ON1231320 on downstream signaling events mediated by Plk2 and evaluate its effect on tumor cell growth under hypoxic, genotoxic and defective respiratory contexts; 3. Characterize the PK properties of ON1231320 in xenograft models to determine the degree of inhibition of Plk2 that is required for inhibition of tumor growth and assess how ON1231320 concentrations are related to antitumor activity of the compound and to evaluate the validity of using PBMN as surrogates; and 4. To further explore the effects of combination therapy using ON1231320 and cytotoxic anti-cancer agents such as oxaliplatin, 5-fluorouracil, paclitaxel and doxorubicin to induce apoptosis and tumor regression in breast and colorectal cancer model systems.