THIAZINE DYE MEDIATED PHOTOKILLING OF HIV 1 VIRUSES

描述(改编自申请人的摘要) 调查人员发现，光活性染料亚甲基蓝(MB) 加光(MB+L)非常有效地灭活HIV-1病毒在 体外培养的人外周血单个核细胞。 MB非常低(约24 NM)，所需的光线也很低 短时间的强度。此外，甲基溴有一个有利的 用于血液成分的吸收光谱。因此，一个 采用这种方法可能有助于灭活HIV病毒。 在血液成分中。MB+L抑制HIV-1的可能性很大 传播是由单线态氧机制介导的，而 致命的损伤发生在一个非常特定的目标上。与其他 可能的光活性方法，其中的作用机制似乎 在病毒膜上，基于对MB+L的广泛机制研究 在申请人的实验室里杀死RNA病毒，调查人员 认为HIV-1病毒的MB+L致死靶点是一个核糖核酸 二级结构区域，可能位于特定的必需RNA/蛋白质 交互界面。申请者想要进行基础研究 这将有助于我们理解MB+L介导的机制基础 使艾滋病毒失活，并利用所获得的知识来设计和测试 将在没有病毒灭活的情况下实现改进的策略 对血液成分造成严重损害，如血小板 浓缩液。研究人员提出了一系列实验 旨在： 了解MB+L传播艾滋病病毒的站点和性质。 申请者计划确定艾滋病毒的传染性是否会因 用MB+L处理分离的HIV病毒粒子。如果病毒粒子被破坏， 将确定病毒RNA是否受损，并确定 发生致命性损害的特定区域(残留物)并确定 如果它的形成是由单线态氧介导的。它也将是 确定MB+L是否损害病毒蛋白或病毒包膜脂类 确定所涉及的机制以及 损坏。还将确定细胞中是否存在艾滋病毒RNA (装配前)被MB+L损坏。 调查人员还将设计和测试一系列亚甲基蓝 以(噻嗪)为基础的衍生物，努力提高特异性和 在消除艾滋病毒感染风险方面的有效性。 最后，调查人员计划开发一种程序来应用 在上述基础研究中获得的知识将进一步 在血液成分存在的情况下消除感染艾滋病毒的风险 而不会对血液成分造成严重损害。

DESCRIPTION (Adapted from the applicant's abstract) The investigators have found that the photoactive dye methylene blue (MB) plus light (MB+L) very effectively inactivates the HIV-1 virus in cultured human peripheral blood mononuclear cells.The effective dose of MB is very low (about 24 nM) and the light required is also of low intensity for a short duration. In addition, MB has a favorable absorption spectra for use with blood components. Therefore an adaptation of this approach may be useful to inactivate the HIV virus in blood components. It is highly likely that MB+L inhibition of HIV-1 propagation is mediated by a singlet oxygen mechanism and that the lethal lesion occurs at a very specific target. In contrast to other possible photoactive approaches where the mechanism of action appears to be at the viral membrane, based on extensive mechanistic studies on MB+L killing of RNA viruses in the applicant's laboratory, the investigators think that the MB+L lethal target of the HIV-1 virus is at a RNA secondary structure area, probably at a specific essential RNA/protein interaction interface. The applicants want to conduct basic studies which will help us understand the mechanistic basis to MB+L mediated inactivation of HIV and utilize the knowledge gained to design and test strategies that will achieve improvement in viral inactivation without causing significant damage to blood components, such as platelet concentrates. The investigators propose a series of experiments designed to: Understand the site and nature of MB+L damage to HIV propagation. Applicants plan to determine whether HIV infectivity is diminished by treating isolated HIV virions with MB+L. If the virions are damaged, it will be determined if the viral RNA is damaged and ascertain the specific region (residue) where the lethal lesion occurs and determine if its formation is mediated by singlet oxygen. It will also be determined if MB+L damages viral proteins or viral envelope lipids and ascertain the mechanisms involved as well as the specificity of the damage. It will also be determined if the HIV RNA present in cells (prior to assembly) is damaged by MB+L. The investigators will also design and test a series of methylene blue based (thiazine) derivatives in an effort to improve the specificity and efficacy in eliminating the risk of HIV infections. Finally the investigators plan to develop a procedure to apply the knowledge obtained in the above basic studies that will further eliminate the risk of HIV infection in the presence of blood components without significantly damaging the blood components.