PTPases as Therapeutic Targets

PTPases 作为治疗靶点

• 批准号: 6948210
• 负责人: Taolin Yi
• 金额: $ 27.23万
• 依托单位: CLEVELAND CLINIC LERNER COM-CWRU
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-09-30 至 2007-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6948210
• 关键词: antineoplastics; athymic mouse; carcinogenesis inhibitor; cell line; chemical group; drug discovery /isolation; enzyme activity; enzyme substrate complex; enzyme therapy; gene expression; gene mutation; in situ hybridization; mass spectrometry; metastasis; neoplasm /cancer therapy; nuclear magnetic resonance spectroscopy; pathologic process; pentamidine; phosphatase inhibitor; polymerase chain reaction; protein structure; protein tyrosine phosphatase

DESCRIPTION (provided by applicant): PRL family phosphatases (PRL-1, PRL-2 and PRL-3) are highly attractive targets for developing inhibitors as novel anti-cancer therapeutics because their over-expression has oncogenic effects and plays a potentially pathogenic role in human malignancies. Our recent studies provided the first evidence that pentamidine (PE), an anti-protozoan drug with an unknown mechanism of action, is an effective inhibitor of PRLs and has anti-cancer effects. The goal of this study is to test our hypothesis that PE has growth inhibitory effects against tumor cells via inactivating PRLs through direct binding to the PTPases and has therapeutic potential against malignancies associated with PRL over-expression. Aim 1. To evaluate the roles of PRL PTPases in mediating PE anti-cancer effects, we will 1) determine the capacities of Peinsensitive mutants of PRL-1, PRL-2 and PRL-3 individually and in combinations to confer resistance to Peinduced growth inhibition in WM9 human melanoma cell line to define the contributions of each PRL in PE effects against the cancer cells and in cancer cell growth; 2) determine PE activity to reverse transformed phenotypes induced by over-expression of PRLs or PTPalpha oncogene in NIH3T3 cells to further elucidate PE anti-cancer mechanism; 3) determine PE effects on lung metastasis of A549 transfectants of PRLs in nude mice to provide prove of concept evidence regarding PE anti-metastasis activity and the roles of individual PRLs in tumor metastasis; 4) identify human malignancies over-expressing PRLs by RNA in situ hybridization to assess the significance of individual PRLs as anti-cancer targets. Aim 2. To elucidate inhibition mechanism of PE against PRLs, we will 1) identify key chemical groups in PE essential for PRL inhibition and PTPase specificity by evaluating 10 PE-derivatives in which PE chemical groups have been selectively modified; 2) characterize PE / PRL complex formation and PE effects on PRL-1/substrate interaction by mass spectrometry and biochemical analysis; 3) analyze solution structures of PRLs and their complexes with PE by NMR. Results of the study will provide proof of principle evidence to establish PE as a PRL-targeted agent with anti-cancer potential, reveal the inhibition mechanism of PE against PRLs at molecular levels, provide insights for rational design and development of more specific and less toxic novel PRL inhibitors and identify human malignancies potentially responsive to PRL-targeted therapy.

描述（由申请人提供）： PRL家族磷酸酶（PRL-1、PRL-2和PRL-3）是用于开发作为新型抗癌疗法的抑制剂的高度有吸引力的靶标，因为它们的过表达具有致癌效应并且在人类恶性肿瘤中起潜在的致病作用。我们最近的研究提供了第一个证据，证明喷他脒（PE），一种作用机制未知的抗原生动物药物，是一种有效的PRL抑制剂，具有抗癌作用。本研究的目的是验证我们的假设，即PE通过直接结合PTPases灭活PRL对肿瘤细胞具有生长抑制作用，并具有治疗与PRL过表达相关的恶性肿瘤的潜力。目标1.为了评估PRL PTPases在介导PE抗癌作用中的作用，我们将1）确定PRL-1、PRL-2和PRL-3的Pes不敏感突变体单独和组合赋予WM 9人黑色素瘤细胞系对Pes诱导的生长抑制的抗性的能力，以确定每种PRL在PE对抗癌细胞的作用和在癌细胞生长中的贡献; 2）测定PE对NIH 3 T3细胞中PRL或PTPalpha癌基因过表达诱导的转化表型的逆转活性，以进一步阐明PE抗癌机制; 3）确定PE对PRL的A549转染子在裸鼠中的肺转移的影响，以提供关于PE抗肿瘤的概念证据的证明。转移活性和单个PRL在肿瘤转移中的作用; 4）通过RNA原位杂交鉴定过表达PRL的人类恶性肿瘤，以评估单个PRL作为抗癌靶点的意义。目标二。为了阐明PE对PRL的抑制机制，我们将1）通过评价10种PE化学基团被选择性修饰的PE衍生物来鉴定PE中对PRL抑制和PTL特异性至关重要的关键化学基团; 2）通过质谱和生化分析来表征PE / PRL复合物的形成和PE对PRL-1/底物相互作用的影响; 3）用NMR分析PRL及其与PE复合物的溶液结构。研究结果将为确立PE作为具有抗癌潜力的PRL靶向药物提供原则性证据，在分子水平上揭示PE对PRL的抑制机制，为合理设计和开发更具特异性和毒性更低的新型PRL抑制剂提供见解，并确定对PRL靶向治疗可能有反应的人类恶性肿瘤。