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 建议进行研究，以评估金丝桃素的开发潜力， (HY)和许多结构相关的芳族多环二酮， 作为输血用红细胞浓缩物中的病毒灭活剂。的 研究是基于令人鼓舞的初步数据，这些数据表明， 在有效的杀病毒剂量下对红细胞（RBC）的不利影响。 他们的目的是确定获得完整的最佳条件， 灭活血液中非常高滴度的无细胞病毒 环境金丝桃素是一种有效的杀病毒剂， 灭活多种包膜病毒。金丝桃素是一种 光动力分子，其在体内也是活性的，大概是由于 低的红/氧化电位。因此，HY目前正在进行评估， 在艾滋病患者中进行的临床试验，以及第一阶段的试验证实， HY的可输注性艾滋病毒感染的双重传播问题 通过无细胞病毒和通过潜伏感染的PBMC， 通过评估HY和白细胞过滤的组合， 完全无菌，以及施加光动力的可能性 感染的PBMC。该应用程序将解决 HY的杀病毒作用机制。病毒中的修饰 蛋白质HIV p55 gag，p24和RT，其被靶向用于光动力学 交联HY将被表征，而其他受影响的病毒 蛋白质鉴定病毒RNA参与RNA- 将评价蛋白质共价复合物。研究人员建议， 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。从HY扩增RNA 使用RNA PCR处理的病毒体将用于鉴定基因组 通过HY交联的区域。问题是HY本身或仅 光动力学产生的受激氧影响衣壳 将通过分离完整的HIV核心和检查 它们对HY和光的敏感性。这些研究对我们 用于潜在灭活无包膜病毒（如B19 细小病毒）。RT活性和衣壳蛋白迁移 SDS-PAGE和蛋白质印迹上的模式将用作测定。 具有甲基取代和改变的结构HY类似物， 多环芳族骨架将被评估为潜在的 提高功效和减少与RBC的结合。的杀病毒 HY离子对与阳离子金属、碱性胺和氨基的活性 酸，以及它们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，包括临床显著影响 红细胞表面膜完整性，（2，3-DPG，溶血， ABO、Rh、次要血型抗原表达正常， IgG或补体在RBC上的沉积）， 考察从病毒感染后的血液中去除HY的潜在方法 灭活、掺入疏水性树脂和其它方法将 被评价。 申请组织将与以下研究中心合作： 1)纽约血液中心，2）俄克拉荷马州血液机构，和3）魏茨曼 科学研究所。