Cutaneous drug reactions (CDR) are among the most frequent adverse medical events. Recent studies suggest that between 30 and 45 percent of all adverse drug reactions involve the skin. Of particular interest are the delayed-type hypersensitivity reactions that occur with sulfonamides and sulfones. While most investigators have focused on the role of differences in hepatic bioactivation and detoxification in determining predisposition to these reactions, it is uncertain and perhaps unlikely that liver-generated reactive metabolites would survive transit to the skin. We have developed a novel hypothesis wherein metabolic activation of drugs in keratinocytes provokes the release of signals that result in the activation of cutaneous dendritic cells, thereby initiating the cascade of events resulting in the manifestations of a CDR. The long term-goal of our project is to elucidate the mechanism of CDR and develop means to predict and/or prevent their occurence. The objective of the present proposal is to test the validity of our hypothesis using sulfamethoxazole (SMX) and dapsone, which are among the most frequent CDR-inducing drugs, as model compounds.
The Specific Aims of this project are to determine: 1) If cyclooxygenase is the enzyme that bioactivates SMX and dapsone in normal human epidermal keratinocytes (NHEK). Preliminary studies have indicated that these compounds can be bioactivated by cyclooxygenase-2. We will identify the enzyme responsible for this bioactivation in NHEK using selective inhibitors and inducers, as well as recombinant enzyme. 2) If cytokines alter the bioactivation or detoxification of SMX and dapsone in NHEK. An inflammatory response has been shown to alter enzymes important in drug bioactivation, an event that appears to be mediated by cytokines. Cytokines may also alter the glutathione content of cells, an alteration that may alter the susceptibility of cells to these hydroxylamine metabolites. We will assess the effects of proinflammatory cytokines on the bioactivation/detoxication of SMX and dapsone in NHEK. 3) If NHEK incubated with hydroxylamine metabolites of SMX and dapsone release signals resulting in the activation of dendritic cells. After determining the mechanism of cell death induced by these metabolites, we will test the hypthothesis that they activate dendritic cells, either directly or indirectly (i.e., via signals released from NHEK). We anticipate that the results of the proposed studies will identify key points of intervention that will permit the prevention or management of these reactions. Moreover, they should enable us to develop in vitro screening tests that will permit the pre-clinical identification of drugs likely to pose a significant risk for the development of such reactions.
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