Mitochondrial Nucleotide Carriers of NRTI Metabolites

NRTI 代谢物的线粒体核苷酸载体

• 批准号: 7115805
• 负责人: TOD GULICK
• 金额: $ 37.48万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 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 2008-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7115805
• 关键词: antiviral agents; drug adverse effect; drug screening /evaluation; high performance liquid chromatography; immunologic assay /test; intracellular transport; laboratory mouse; liposomes; membrane transport proteins; mitochondria; mitochondrial DNA; mitochondrial disease /disorder; nucleoside analog; nucleotides; radiotracer; reverse transcriptase inhibitors; tissue /cell culture; transfection; yeasts; zidovudine

DESCRIPTION (provided by applicant): Nucleoside reverse transcriptase inhibitors (NRTI) are critical components of highly active anti-viral therapies. Phosphorylated derivatives (NRTI-P) mimic the cellular dNTP substrates for HIV RT, but produce chain termination, thereby limiting retroviral amplification. NRTIs produce reversible side effects that resemble those of patients with mitochondrial (mito) genetic disorders, including cardiomyopathy, myopathy, neuropathy and lactic acidosis, and these correlate with NRTI-induced loss of mito DNA in various tissues. Toxicity has been attributed to inhibition of DNA polymerase g, interfering with mito DNA maintenance. Alternative or additional mechanisms of mito damage may also be at play. Regardless of mechanism, NRTI toxicity is dependent on import into mito. We have isolated novel human genes and cDNAs that encode orphan members of the mito metabolite carrier family (OMC). Five carry a signature specific to nucleotide (nt) carriers, including a putative adenine nt transporter (ANT), two putative CoA transporters, and a non-specific exchanger of nucleotides (OMC27). We have shown that purified OMC27 and its yeast ortholog transport NRTI-P's AZT-TP and -DP, ddl-TP, and ddC-TP in proteoliposome radiochemical flux assays. Disruption of the orthologous yeast gene protects against AZT-induced mito dysfunction. This project examines the role of OMC27 in NRTI-mediated toxicity, with goals of understanding mechanisms by which NRTIs gain entry to the matrix, and identifying nucleoside analogs with known anti-viral activity that offer potential for improved therapeutic index by virture of mito exclusion and reduced mito toxicity. We will test the hypotheses that OMC27 (and yeast ortholog) are highly affinity transporters of NRTI-P, that this represents the primary pathway for mito uptake of NRTI-P; that OMC27 expression is critical to NRTI-induced mito failure; and that a useful therapeutic index for NRTIs may be evident from the ratio of NRTI anti-viral to MC-mediated uptake activities; using complementary in vitro, yeast and mammalian cell, and in vivo mouse studies.

描述（由申请人提供）： 核苷类逆转录酶抑制剂（NRTI）是高效抗病毒治疗的关键成分。 磷酸化衍生物（NRTI-P）模拟HIV RT的细胞dNTP底物，但产生链终止，从而限制逆转录病毒扩增。 NRTI产生可逆的副作用，类似于线粒体（mito）遗传疾病患者的副作用，包括心肌病、肌病、神经病和乳酸酸中毒，这些副作用与NRTI诱导的各种组织中mito DNA丢失相关。 毒性归因于DNA聚合酶g的抑制，干扰线粒体DNA的维持。 其他或额外的线粒体损伤机制也可能起作用。 无论机制如何，NRTI的毒性取决于输入到mito。 我们已经分离出新的人类基因和编码线粒体代谢物载体家族（OMC）孤儿成员的cDNA。 五个携带特异于核苷酸（nt）载体的特征，包括一个推定的腺嘌呤nt转运蛋白（ANT），两个推定的CoA转运蛋白和一个非特异性核苷酸交换蛋白（OMC 27）。 我们已经在蛋白脂质体放射化学通量测定中表明，纯化的OMC 27及其酵母直系同源物转运NRTI-P的AZT-TP和-DP、ddl-TP和ddC-TP。 破坏酵母基因可防止AZT诱导的线粒体功能障碍。 该项目研究了OMC 27在NRTI介导的毒性中的作用，目的是了解NRTI进入基质的机制，并鉴定具有已知抗病毒活性的核苷类似物，这些核苷类似物通过排除线粒体和降低线粒体毒性来提高治疗指数。 我们将测试OMC 27的假设，（和酵母直系同源物）是NRTI-P的高亲和力转运蛋白，这代表了NRTI-P的线粒体摄取的主要途径; OMC 27表达对NRTI诱导的线粒体衰竭至关重要;并且NRTI的有用治疗指数可以从NRTI抗病毒与MC介导的摄取活性的比率中得到证实;使用互补的体外、酵母和哺乳动物细胞以及体内小鼠研究。