CORE--Antiretroviral Drug Susceptibility and Drug Interactions

核心B的总体目标是为IPCP项目提供药物相互作用的药敏试验和评价。BBI Biotech已经对HIV-1进行了许多药物敏感性试验，这些试验有助于开发一种新型抗逆转录病毒药物PA 457，该药物刚刚完成了成功的I期临床试验。最初，核心将筛选5种NK-1 R拮抗剂（阿瑞匹坦、CJ-12255、CJ-96345、RP-67,580和L733060），用于抗逆转录病毒治疗。 活性和细胞毒性。有效化合物，即，具有可接受的抗逆转录病毒活性与细胞毒性比率的那些将在项目1和2以及核心B中进一步表征。为了表征抗病毒应答的广度，除阿瑞匹坦（计划用于初始动物研究和临床试验）外，还将针对一组25种HIV分离株（包括M型、O型、HIV-2原代分离株和耐药分离株）检测先导化合物。将确定每种分离株的50%有效剂量（EC 50）。由于大多数抗逆转录病毒疗法包含多种药物，阿瑞匹坦和其他先导化合物将分别针对不同类别的抗逆转录病毒药物进行药物相互作用测试。待检测相互作用的药物将与PBMC中的HIV-1（R5和X4病毒）一起或单独培养。将在中效方程中使用HIV p24产生的减少来计算EC 50和联合指数（CI），以表明协同作用或拮抗作用。为了评价 体外抗性诱导，THP-1细胞将与HIV-1 Ba-L和低剂量阿瑞匹坦一起培养。药物敏感性试验将每月进行一次，以寻找正在形成的耐药性。如果EC 50增加，将对病毒包膜进行测序，以确定共受体结合区域的变化，并将病毒和细胞分别送往项目1和2进行进一步研究。为了确定阿瑞匹坦耐药性是否可以在体内发展，分离自 将每隔一个月对项目3中用阿瑞匹坦处理的猴进行测定，并测定EC 50。为了将治疗效果与患者病毒群体的可能变化相关联，将分析项目4入组前、治疗结束和阿瑞匹坦治疗后1个月的分离株对阿瑞匹坦和共受体使用的敏感性。如果共受体使用或药物敏感性发生变化，将对治疗前和治疗后分离株的包膜进行测序，以鉴定耐药标志物。核心B提供的数据将确定阿瑞匹坦体内和体外抗病毒作用的广度和稳定性，并表征具有抗逆转录病毒活性的其他NK-1 R拮抗剂，用于未来研究。

The overall goal of Core B is to provide drug susceptibility testing and evaluation for drug interactions for the IPCP program. BBI Biotech has performed numerous drug susceptibility assays for HIV-1, which have contributed to the development of a novel anti-retroviral drug, PA457, that just completed a successful Phase I clinical trial. Initially, the Core will screen five NK-1R antagonists (aprepitant, CJ-12255, CJ-96345, RP-67,580, and L733060), for anti-retroviral activity and cytotoxicity in PBMC using HIV-1 Ba-L. Effective compounds, i.e., those with acceptable ratio of antiretroviral activity to cytotoxicity, will be further characterized in Projects 1 and 2, and Core B. To characterize the breadth of the antiviral response, the lead compounds, in addition to aprepitant (planned for initial animal study and clinical trial), will be tested against a panel of 25 HTV isolates consisting of type M, type O, HIV-2 primary isolates and drug resistant isolates. The 50% Effective Dose (EC50) will be determined against each isolate. Because most antiretroviral therapies contain multiple drugs, aprepitant and the other lead compounds will each be tested for drug interactions against different classes of anti-retrovirals. Drugs to be tested for interactions will be cultured with HIV-1 (R5 and X4 viruses) in PBMC together or individually. Reduction in HIV p24 production will be used in the median-effect equation to calculate the EC50s and combination indices (CI) to indicate synergism or antagonism. To evaluate in vitro resistance induction, THP-1 cells will be cultured with HIV-1 Ba-L and low doses of aprepitant. Drug susceptibility assays will be performed once a month to look for developing resistance. If the EC50 increases, the viral envelope will be sequenced to identify changes in co-receptor binding area and virus and cells sent to Projects 1 and 2, respectively, for further study. To determine whether aprepitant resistance can develop in vivo, SFV isolated from monkeys treated with aprepitant in Project 3 will be assayed every other month and EC50 determined. To associate treatment effects with possible changes in patient viral population, isolates from pre-entry, end of treatment, and one month after aprepitant treatment from Project 4 will be assayed for susceptibility to aprepitant and co-receptor usage. If there are changes in co-receptor usage or drug susceptibility the envelope will be sequenced in pretreatment and post treatment isolates to identify resistance markers. The data provided by Core B will determine the breadth and stability of the anti-viral effects of aprepitant both in vivo and in vitro and characterize additional NK-1R antagonists with anti-retroviral activities for future studies.