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.
| Wen, Jin; Chowdhury, Pramit; Wills, Nick J et al. (2002) Toward the molecular flashlight: preparation, properties, and photophysics of a hypericin-luciferin tethered molecule. Photochem Photobiol 76:153-7 |
| Wills, N J; Park, J; Wen, J et al. (2001) Tumor cell toxicity of hypericin and related analogs. Photochem Photobiol 74:216-20 |
| Showalter, B M; Datta, A; Chowdhury, P K et al. (2001) Identification of a vibrational frequency corresponding to H-atom translocation in hypericin. Photochem Photobiol 74:157-63 |
