Novel Substrate Competitive Bcr-Abl Inhibitor Active Against Gleevec-Resistant CM

描述（申请人提供）：伊马替尼是一种BCR-ABL酪氨酸激酶抑制剂，用于治疗人CML，取得了巨大成功。然而，长期使用伊马替尼治疗的患者中有很大一部分由于BCR-ABL激酶结构域的突变而产生耐药。我们最近开发了一种化合物（ON 012380），该化合物在不同于伊马替尼的位点与BCR-ABL结合，并在5-10 nM的浓度下诱导Ph+ CML细胞凋亡（比伊马替尼有效10-50倍）。更有趣的是，发现这种化合物在诱导迄今为止鉴定的所有伊马替尼耐药CML突变体的死亡方面非常有效。在本申请中，我们建议对该化合物的作用机制和受该化合物影响的信号传导途径的性质进行详细的生物化学表征。目标是：1.确定ON 012380抑制BCR-ABL的动力学，并对诱变BCR-ABL克隆进行体外筛选，以了解可能损害ON 012380结合的氨基酸取代。2.确定ON 012380对（a）BCR-ABL野生型和伊马替尼耐药突变体对激酶活性的影响;（B）下游信号传导，如MAPK、AKT和STAT 5激活;（c）细胞周期进展;和（d）表达野生型或突变型BCR-ABL蛋白的肿瘤细胞中激活的凋亡途径的性质。3.确定ON 012380是否诱导过表达林恩的伊马替尼耐药细胞的细胞死亡，如果是，确定作用机制。4.按照有效性研究所用的途径和时间表进行药代动力学研究，以及5.在已建立的CML异种移植模型中进行疗效试验。

DESCRIPTION (provided by applicant): Imatinib, which is an inhibitor of BCR-ABL tyrosine kinase and used for the treatment of human CML, has been a spectacular success. However, a significant proportion of patients chronically treated with imatinib develop resistance due to acquisition of mutations in the kinase domain of BCR-ABL. We have recently developed a compound (ON012380) that binds to BCR-ABL at a site different from imatinib and induces apoptosis of Ph+ CML cells at a concentration of 5-10 nM (which is 10-50 fold more potent than imatinib). More interestingly, this compound was found to be very effective in inducing the death of all of the imatinib-resistant mutants of CML identified so far. In this application, we propose to carry out a detailed biochemical characterization of the mechanism of action of this compound and the nature of signaling pathways that are affected by this compound. The aims are: 1. To determine the kinetics of inhibition of BCR-ABL by ON012380 and carry out in vitro screen of mutagenized BCR-ABL clones to gain an understanding of the amino acid substitutions that are likely to impair the binding of ON012380. 2. To determine the effects of ON012380 on (a) wild-type and imatinib-resistant mutants of BCR-ABL on the kinase activity; (b) downstream signaling such as MAPK, AKT and STAT5 activation (c) cell cycle progression; and (d) the nature of apoptotic pathways activated in tumor cells that express wild-type or mutant BCR-ABL protein. 3. Determine whether ON012380 induces cell death of Lyn overexpressing, imatinib resistant cells and if so, determine the mechanism of action. 4. Conduct pharmacokinetic studies by the route and schedule used for efficacy studies, and 5. Conduct efficacy trials in established xenograft models of CML.