RESUMO
The decline in approval of new drugs during the past decade has led to a close analysis of the drug discovery process. One of the main reasons for attrition is preclinical toxicity, frequently attributed to the generation of protein-reactive drug metabolites. In this review, we present a critique of such reactive metabolites and evaluate the evidence linking them to observed toxic effects. Methodology for the characterization of reactive metabolites has advanced greatly in recent years, and is summarized first. Next, we consider the inhibition of key metabolic enzymes by electrophilic metabolites, as well as unfavorable drug-drug interactions that may ensue. One important class of protein-reactive metabolites, not linked conclusively to a toxic event, is acyl glucuronides. Their properties are discussed in light of the safety characteristics of carboxylic acid containing drugs. Many adverse drug reactions (ADRs) are known collectively as idiosyncratic events, that is, not predictable from knowledge of the pharmacology and pharmacokinetics of the parent compound. Observed ADRs may take various forms. Specific organ injury, particularly of the liver, is the most direct: we examine this in some detail. Moving to the cellular level, we also consider the upregulation of induced cellular processes. The related, but distinct, issue of hypersensitivity or allergic reactions to drugs and their metabolites, possibly via the immune system, is considered next. Finally, we discuss the impact of such data on the drug discovery process, both through early detection of reactive metabolites and informed synthetic design, which eliminates unfavorable functionality from drug candidates.
Assuntos
Desenho de Fármacos , Preparações Farmacêuticas/metabolismo , Animais , Pesquisa Biomédica , Sistema Enzimático do Citocromo P-450 , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos , Glucuronídeos/metabolismo , HumanosRESUMO
BACKGROUND: Exposure of patients with cystic fibrosis to sulfonamides is associated with a high incidence of hypersensitivity reactions. OBJECTIVE: To compare mechanisms of antigen presentation and characterize the phenotype and function of T cells from sulfamethoxazole-hypersensitive patients with and without cystic fibrosis. METHODS: T cells were cloned from 6 patients and characterized in terms of phenotype and function. Antigen specificity and mechanisms of antigen presentation to specific clones were then explored. Antigen-presenting cell metabolism of sulfamethoxazole was quantified by ELISA. The involvement of metabolism in antigen presentation was evaluated by using enzyme inhibitors. RESULTS: Enzyme inhibitable sulfamethoxazole-derived protein adducts were detected in antigen-presenting cells from patients with and without cystic fibrosis. A significantly higher quantity of adducts were detected with cells from patients with cystic fibrosis. Over 500 CD4(+) or CD8(+) T-cell clones were generated and shown to proliferate and kill target cells. Three patterns of MHC-restricted reactivity (sulfamethoxazole-responsive, sulfamethoxazole metabolite-responsive, and cross-reactive) were observed with clones from patients without cystic fibrosis. From patients with cystic fibrosis, sulfamethoxazole metabolite-responsive and cross-reactive, but not sulfamethoxazole-responsive, clones were observed. The response of the cross-reactive clones to sulfamethoxazole was dependent on adduct formation and was blocked by glutathione and enzyme inhibitors. Antigen-stimulated clones from patients with cystic fibrosis secreted higher levels of IFN-γ, IL-6, and IL-10, but lower levels of IL-17. CONCLUSION: Sulfamethoxazole metabolism and protein adduct formation is critical for the stimulation of T cells from patients with cystic fibrosis. T cells from patients with cystic fibrosis secrete high levels of IFN-γ, IL-6, and IL-10.
Assuntos
Antibacterianos/efeitos adversos , Antibacterianos/imunologia , Fibrose Cística/tratamento farmacológico , Fibrose Cística/imunologia , Hipersensibilidade a Drogas/imunologia , Sulfametoxazol/efeitos adversos , Sulfametoxazol/imunologia , Antibacterianos/metabolismo , Apresentação de Antígeno , Células Apresentadoras de Antígenos/imunologia , Estudos de Casos e Controles , Proliferação de Células , Células Clonais , Fibrose Cística/complicações , Fibrose Cística/metabolismo , Citocinas/biossíntese , Hipersensibilidade a Drogas/complicações , Hipersensibilidade a Drogas/metabolismo , Humanos , Técnicas In Vitro , Sulfametoxazol/metabolismo , Linfócitos T/imunologia , Linfócitos T/patologiaRESUMO
Sulfamethoxazole (SMX) induces immunoallergic reactions that are thought to be a result of intracellular protein haptenation by its nitroso metabolite (SMX-NO mass, 267 amu). SMX-NO reacts with protein thiols in vitro, but the conjugates have not been defined chemically. The reactions of SMX-NO with glutathione (GSH), a synthetic peptide (DS3), and two model proteins, human GSH S-transferase pi (GSTP) and serum albumin (HSA), were investigated by mass spectrometry. SMX-NO formed a semimercaptal (N-hydroxysulfenamide) conjugate with GSH that rearranged rapidly (1-5 min) to a sulfinamide. Reaction of SMX-NO with DS3 also yielded a sulfinamide adduct (mass increment, 267 amu) on the cysteine residue. GSTP was exclusively modified at the reactive Cys47 by SMX-NO and exhibited mass increments of 267, 283, and 299 amu, indicative of sulfinamide, N-hydroxysulfinamide, and N-hydroxysulfonamide adducts, respectively. HSA was modified at Cys34, forming only the N-hydroxysulfinamide adduct. HSA modification by SMX-NO under these conditions was confirmed with ELISA and immunoblotting with an antisulfonamide antibody. It is proposed that cysteine-linked N-hydroxysulfinamide and N-hydroxysulfonamide adducts of SMX are formed via the reaction of SMX-NO with cysteinyl sulfoxy acids. Evidence for a multistep assembly of model sulfonamide epitopes on GSH and polypeptides via hydrolyzable intermediates is also presented. In summary, novel, complex, and metastable haptenic structures have been identified on proteins exposed in vitro to the nitroso metabolite of SMX.