Research Project
9680555
Kaposi Sarcoma (KS) is the most common cancer in the 36 million worldwide AIDS population. KS tumors are caused by Kaposi Sarcoma-Associated Herpes Virus (KSHV). Even though HAART therapy has resulted in a dramatic decline in KS in the US, this malignancy remains the most predominant tumor of AIDS. Paradoxically, KS tumors arise in some AIDS patients following HAART treatment, due to Immune Reconstitution Inflammatory Syndrome. KS is a systemic cancer that can form on the skin, oral cavity and internal organs. These tumors grow on the hard and soft palate, gingiva, tongue, uvula, pharynx, trachea and tonsils. KS oral tumors are painful, extremely angiogenic, and can ulcerate and interfere with mastication of food, cause tooth loss, impede speech, and hinder breathing. Oral KS thus not only compromises the quality of life but threatens survival. Of significance, there is no drug for treating KS, making this viral cancer an unmet medical need. Our long-term goal is to develop a drug for treating acute Oral KS by injection into tumors and sustained systemic treatment by oral-capsule delivery. Notably, a small number of KSHV lytically infected cells within KS tumors are critical for maintaining malignancy. We have discovered a KSHV protein, Processivity Factor-8 (PF-8), which is essential for KSHV DNA synthesis and lytic infection, making PF-8 an ideal drug target to treat KS. We showed PF-8 associates with KSHV DNA polymerase (Pol-8), enabling it to incorporate nucleotides continuously without dissociating from the template and that PF-8 is specific for Pol-8. We also revealed by crystallography that PF-8 forms head-head homodimers and identified domains required for dimerization and stabilization of Pol-8 on the DNA. Our FIRST AIM is to identify Leads by two approaches: Approach-1 is to extend our successful pilot screen of a 50,000 compound Chemical Diversity Library using our Rapid Plate Assay designed to specifically identify functional inhibitors of PF-8. Our pilot screen of 3,000 compounds has already produced four HIT small molecules that specifically block processive DNA synthesis and KSHV infection in cells. Approach-2 is to test peptidomimetics and small molecules synthesized by mutation-structure guided design to prevent formation of PF-8 homodimers. This approach is based on our crystal structure of PF-8 and recent site-directed mutagenesis which revealed that each of five individual and consecutive amino acids (STVHK) that compose the Beta-Sheet needed for homodimerization, is necessary for processive DNA synthesis. Our SECOND AIM is to use a proven iterative medicinal chemistry SAR approach to test for potency in blocking PF-8 processive DNA synthesis, inhibition of KSHV infection in cells, toxicity and validation of binding by IsoThermal Calorimetry (ITC). Our THIRD AIM is to evaluate the top Leads from AIM2 for In Vitro ADME and Off-Target Activity by tests that include: Aqueous Solubility, Metabolic Stability, Plasma Protein Binding (PPB), CYP 450 Inhibition and hERG Patch Clamp.
