IN VITRO INACTIVATION OF VIRUSES IN BLOOD COMPONENTS

血液成分中病毒的体外灭活

• 批准号: 2771384
• 负责人: DANIEL MERUELO
• 金额: $ 36.77万
• 依托单位: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-09-01 至 2000-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2771384
• 关键词: Retroviridae; SDS polyacrylamide gel electrophoresis; antiviral agents; blood transfusion; chemical structure function; covalent bond; crosslink; cyclic compound; drug screening /evaluation; erythrocytes; polymerase chain reaction; virus RNA; virus envelope; virus protein; western blottings

DESCRIPTION (Adapted from the applicant's abstract) This application proposes studies to evaluate the potential for development of hypericin (HY) and of numerous structurally related aromatic polycyclic diones, as virus inactivators in red cell concentrates for transfusion. The studies are based on encouraging preliminary data which show lack of adverse effects to red blood cells (RBC) at effective virucidal doses. They aim to define the optimal conditions for obtaining complete inactivation of very high titers of cell free virus in a blood environment. Hypericin is an effective virucidal agent which directly inactivates a broad range of enveloped viruses. Hypericin is a photodynamic molecule, which is also active in vivo, presumably due to a low red/ox potential. Thus, HY is currently being evaluated in clinical trials in AIDS patients, and phase I of the trials confirmed the transfusibility of HY. The problem of dual transmission of HIV infection via cell-free virus and through latently infected PBMC will be addressed by evaluating combinations of HY and leukocyte filtration in achieving complete sterility, and the possibility of inflicting photodynamic lesions to infected PBMC. The application will address aspects of the mechanism of the virucidal action of HY. The modifications in viral proteins HIV p55gag, p24 and RT, which are targeted for photodynamic cross-linking by HY will be characterized, and other affected virus proteins identified. The potential involvement of viral RNA in RNA- protein covalent complexes will be evaluated. The investigators propose clarify whether the action of HY covalently cross links viral genomic RNA, with RT and other core proteins. Amplification of RNA from HY treated virions using RNA PCR will be utilized to identify genomic regions cross linked by HY. The question whether HY itself or only photodynamically generated excited oxygen species affect the capsids will be examined by isolation of intact HIV cores and examination of their susceptibility to HY and light. These studies have implications for potential inactivation of non enveloped viruses (such as B19 parvovirus) in blood by HY. RT activity and capsid protein migration patterns on SDS-PAGE and Western blots will be used as assays. Structural HY analogs with methyl group substitutions and alterations in the polycyclic aromatic skeleton will be evaluated for potentially improved efficacies and for reduced binding to RBC. The virucidal activities of HY ion pairs with cationic metals, basic amines and amino acids, and their interactions with physiological transport proteins, will be studied in comparison with the standard HY-Na+. Potential for adverse effects to blood components including clinically significant parameters of red cell surface membrane integrity, (2,3-DPG, hemolysis, normal expression of ABO, Rh, minor blood group antigens and possible deposition of IgG or complement on RBC), by treatment with HY will be examined. Potential methods for removal of HY from blood after virus inactivation, incorporating hydrophobic resins and other methods will be evaluated. The applicant organization will collaborate with the following sites: 1) New York Blood Center, 2) Oklahoma Blood Institution, and 3) Weizmann Institute of Science.

描述（改编自申请人的摘要）本申请 提出研究评估金丝桃素的开发潜力 (HY) 和许多结构相关的芳香族多环二酮， 作为浓缩红细胞中的病毒灭活剂用于输血。这 研究基于令人鼓舞的初步数据，这些数据表明缺乏 在有效杀病毒剂量下对红细胞（RBC）产生不良影响。 他们的目标是确定获得完整的最佳条件 血液中极高滴度的无细胞病毒失活 环境。金丝桃素是一种有效的杀病毒剂，可直接 灭活多种包膜病毒。金丝桃素是一种 光动力分子，在体内也很活跃，大概是由于 低红/氧化电位。因此，HY 目前正在评估 在艾滋病患者中进行的临床试验，第一阶段试验证实了 HY 的可输血性。 HIV感染双重传播问题 通过无细胞病毒和潜伏感染的 PBMC 将得到解决 通过评估 HY 和白细胞过滤的组合来实现 完全不育，以及造成光动力的可能性 感染 PBMC 的损伤。该应用程序将解决以下方面 HY的杀病毒作用机制。病毒的修饰 HIV p55gag、p24 和 RT 蛋白，这些蛋白是光动力靶向的 HY 交联将被表征，而其他受影响的病毒 鉴定出的蛋白质。病毒RNA在RNA-中的潜在参与 将评估蛋白质共价复合物。调查人员建议 阐明 HY 的作用是否共价交联病毒基因组 RNA，以及 RT 和其他核心蛋白。 HY RNA 的扩增 使用 RNA PCR 处理的病毒粒子将用于鉴定基因组 区域通过 HY 交叉连接。问题是 HY 本身还是仅仅 光动力产生的激发氧物种影响衣壳 将通过分离完整的 HIV 核心并检查 他们对 HY 和光的敏感性。这些研究具有重要意义 用于潜在灭活无包膜病毒（例如 B19 HY 引起的血液中的细小病毒）。 RT 活性和衣壳蛋白迁移 SDS-PAGE 和蛋白质印迹上的图案​​将用作测定。 具有甲基取代和改变的结构 HY 类似物 将评估多环芳族骨架的潜在潜力 提高功效并减少与红细胞的结合。杀病毒的 HY 离子对与阳离子金属、碱性胺和氨基的活性 酸及其与生理转运蛋白的相互作用， 将与标准 HY-Na+ 进行比较研究。潜力 对血液成分的不良影响，包括临床上显着的影响 红细胞表面膜完整性参数（2,3-DPG、溶血、 ABO、Rh、次要血型抗原的正常表达和可能 IgG 或补体沉积在红细胞上），通过 HY 治疗将 检查了。病毒感染后从血液中去除 HY 的潜在方法 失活、掺入疏水性树脂等方法将 进行评估。 申请组织将与以下站点合作： 1) 纽约血液中心，2) 俄克拉荷马州血液研究所，3) 魏茨曼 科学研究所。