IN VIVO CHEMILUMINESCENT ACTIVATION OF PHOTOSENSITIZERS

描述：金丝桃素是一种天然存在的光敏剂， 当被光激活时显示出有效的抗病毒和抗肿瘤活性。 金丝桃素的抗病毒活性对多种病毒有效 包膜病毒，包括HIV-1，艾滋病的病原体。 然而，在这方面， 金丝桃素用于抗病毒或癌症治疗的体内应用受到阻碍 它的光依赖性。 这些研究的长期目标是产生 化学发光作为体内光源用于激活杀病毒剂 以及金丝桃素和其他光敏剂的杀肿瘤活性。 的 本提案的目的是确定以下方面的可行性： 金丝桃素在细胞内的荧光激活作用。 化学发光 萤火虫荧光素酶和荧光素酶的反应可以激活抗病毒药物 金丝桃素在无细胞系统中的活性;然而， 能量转移被发现依赖于局部浓度的 能量供体和受体。 在第一个具体目标中， 能量供体和受体，即胡萝卜素和金丝桃素，将增加 通过化学系链 第一种拴系分子是 “笼状”的胡萝卜素，其中存在于胡萝卜素中的羧酸基团 被封端为活化酯。 第二种被束缚的分子 是在酯酶存在下保持其完整性的那些。 这 设计将利用非常有效的分子内能量 金丝桃素和荧光中间体之间的转移 荧光素酶的反应。 所有拴系化合物将 初步测试了光化学性质，有前途的分子将 进一步进行生物学表征。 在第二个具体目标中，稳定 在慢病毒控制下表达荧光素酶基因的细胞系 启动子将被分离，用病毒感染，并用拴系的 分子。 共聚焦和荧光显微镜将用于定位 金丝桃素系链和荧光素酶。 反应 荧光素酶和拴系的荧光素酶之间的关系将作为以下函数进行监测： 金丝桃素激活的荧光光谱法。 最后，细胞将 检测感染性病毒的产生。 这些研究的结果 将决定金丝桃素或其他 光敏剂可以在体内应用。 拟议的研究代表了 多学科方法开发新的抗病毒药物， 抗肿瘤疗法

DESCRIPTION: Hypericin is a naturally occurring photosensitizer that exhibits potent anti-viral and anti-tumor activity when activated by light. The antiviral activity of hypericin is effective against a number of enveloped viruses, including HIV-1, the causative agent of AIDS. However, in vivo application of hypericin for antiviral or cancer therapy is hampered by its photodependency. The long range goal of these studies is to generate chemiluminescence as an in vivo light source for activation of the virucidal and tumoricidal activity of hypericin and other photosensitizers. The objectiv of this proposal is to determine the feasibility of chemiluminescent activatio of hypericin in cells. The chemiluminescent reaction of firefly luciferase and luciferin can activate the antiviral activity of hypericin in a cell free system; however, the efficiency of energy transfer was found to depend on the localized concentrations of the energy donor and acceptor. In the first specific aim, the proximity of the energy donor and acceptor, i.e. luciferin and hypericin, will be increased by means of a chemical tether. The first type of tethered molecular will be a "caged" luciferin, wherein the carboxylic acid group present in luciferin is capped as an activated ester. The second type of tethered molecules will be those which maintain their integrity in the presenc of esterases. This design will take advantage of a very efficient intramolecular energy transfer between hypericin and the chemiluminescent intermediate of the reaction of luciferin with luciferase. All tethered compounds will be tested initially for photochemical properties, and promising molecules will be further characterized biologically. In the second specific aim, stable cell lines expressing the luciferase gene under the control of a lentivirus promoter will be isolated, infected with virus, and treated with tethered molecules. Confocal and fluorescence microscopy will be used to localize hypericin tethers and luciferase within virus-infected cells. Reactions between luciferase and tethered luciferin will be monitored as a function of hypericin activation using fluorescence spectroscopy. Finally, cells will be assayed for production of infectious virus. The results of these studies will determine if chemiluminescent activation of hypericin or other photosensitizers can be applied in vivo. The proposed studies represent a multidisciplinary approach to the development of novel antiviral and anti-tumo therapies.