Studies on Probenecid Prodrugs that Protect against Mitochondrial Toxicity of Tenofovir

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ABSTRACT: Almost 38 million individuals worldwide are infected with the human immunodeficiency virus (HIV) causative agent of acquired immunodeficiency disease (AIDS), 1.2 million of them being Americans. With no vaccine and no cure available, HIV/AIDS patients must take anti-HIV drugs throughout the rest of their lives. Thus, the long-term use of highly active antiretroviral therapy (HAART)drugs that revolutionized HIV/AIDS AIDS patients can also live as long as non-infected people, has been accompanied by serious mitochondrial toxicities, with fatal consequences that led to discontinuation of many of the early effective nucleoside reverse transcriptase (RT) inhibitor (NRTIs) backbone components, like ddC and d4T. The development of the nucleotide RT inhibitors (NtRTIs) particularly tenofovir (TFV), has been innovative in the HIV RT inhibitors field, to the extent that it has gained widespread use worldwide and is incorporated into almost all first-line HIV/AIDS therapeutic combination regimens, as well as used for pre-exposure prophylactics (PrEP). TFV is marketed as two prodrugs tenofovir disoproxil fumarate (TDF) and tenofovir alafenamide (TAF). However, long-term use of the TDF prodrug causes kidney toxicity stemming from the active parent TFV. TFV-associated kidney toxicity is said to have constituted the single referrals for specialist renal services when TFD was first introduced. Moreover, this renal decline was irreversible in about 30 % of patients even after cessation of TFV therapy. Although, TAF, which was introduced more recently is more potent and less toxic than TDF, case studies show it has kidney toxicity potential which may manifest upon long-term use. Moreover, it causes troubling weight gain that could exacerbate metabolic syndrome. These problems mean that TAF cannot always replace TDF. The mechanisms of TFV toxicity are not well understood, but damage to renal proximal tubular mitochondrial has been strongly implicated. The goal of this research is to gain better understanding and develop effective approaches to addressing TFV kidney toxicity. The specific aims are: 1) to confirm that human mitochondrial membranes express functional organic anion transporters 1 and 3 (hOAT1 and hOAT3), and show their ability to import TFV into mitochondria, and, 2) to develop and investigate the protective ability of mitochondrially targeted hOAT1 and 3 inhibitor prodrugs against TFV toxicity. We have demonstrated the expression of the TFV transporters hOAT1 and hOAT3 in the mitochondrial membranes of human renal proximal tubule epithelial cells, supporting our hypothesis that TFV enters mitochondria through hOAT1 and 3 to cause mitochondrial toxicity. For aim 2, we have synthesized an initial hOAT1, and 3 inhibitor prodrug and shown that it is protective against TFV mitochondrial toxicity. A comprehensive multidisciplinary approach involving medicinal chemistry, biochemistry, functional genomics, and pharmacology will be applied, using human kidney proximal tubule cells. Our initially synthesized targeted prodrug reverses TFV-induced mitochondrial toxicity. The success of this project will give new insights into TFV-induced toxicity and provide biological probes and/or potential drug leads.