Role of Cytochromes P450 in Acetaminophen Nephrotoxicity

Tarloff, Joan

Abstract

A variety of drugs and chemicals induce reversible and/or irreversible kidney damage. The applicant is interested in: (1) defining the mechanism(s) whereby cytotoxicity occurs, (2) defining factors that contribute to organ-specific injury, (3) identifying those factor(s) that differentiate reversible from irreversible injury, and (4) defining mechanism(s) involved in tissue repair and/or regeneration. Renal injury during or following drug therapy is not an uncommon problem, and presents a serious burden on health care costs in the United States. A more thorough understanding of the mechanisms involved in organ- and cell-specific injury might assist in developing drugs that do not produce renal injury. The present proposal is designed to test the hypothesis that acetaminophen (APAP) bioactivation in the kidney occurs, at least in part, via cytochrome P450-dependent pathways. Following acute overdosage. APAP produces both hepatotoxicity and nephrotoxicity. Hepatic damage occurs following bioactivation of APAP to a reactive intermediate in a reaction catalyzed by cytochrome P450-dependent mixed function oxidases. The role of cytochrome P450 in the development of renal damage has been difficult to assess, at least in part due to an absence of selective modulators of cytochrome P450. It is important to identify the pathways involved in APAP nephrotoxicity as a first step in identifying the mechanisms that contribute to cellular toxicity. The goal of this project is to quantify the contribution of oxidative vs. deacetylase-dependent bioactivation of APAP in the development of nephrotoxicity. Experiments have been designed to examine to establish an in vitro system of renal cortical tubular suspensions and correlate the expression of nephrotoxicity in vivo following APAP treatment with cytotoxicity observed in vitro in these renal tubules. The relative contribution of the major pathways of APAP bioactivation will be assessed using specifically labeled APAP and selective inhibitors of these bioactivation pathways. Animals will be treated with APAP labeled in the ring or acetate position and covalent binding to renal proteins determined at various times following treatment. In addition, selective inhibitors of deacetylase-dependent APAP bioactivation (e.g., bis-p-nitrophenyl-phosphate) and oxidative metabolism (e.g., 1-aminobenzotriazole), will be used in vitro to correlate covalent binding of APAP-derived material with cytotoxicity. These experiments will assist in establishing the pathways involved in APAP bioactivation in the kidney and will clarify the role of cytochrome P450 in the development of APAP nephrotoxicity.