All adaptations of parasites to the host environment reflect molecular responses to external chemical stimuli. These stimuli consist primarily of the various host nutritional, hormonal and immunological factors that have contributed to the evolution of compatible parasite-host associations. But with the advent of chemotherapy, parasites have been exposed to a new form of selective pressure, the introduction of drugs into their environment. During such chemical insult, parasite survival is determined by the capacity to respond to an abrupt environmental change. Some antiparasitic drugs are prodrugs that require enzymatic activation within the target organism to be efficacious. Genetic mutations resulting in the absence or alteration of the catalytic activity of prodrug activating enzyme represents one type of adaptation to an external chemical stimulus that imparts a survival advantage to the parasite. This component of the program project is designed to test the hypothesis that the first step in the mechanism of action of oxamniquine and hycanthone involves enzyme-catalyzed drug esterification by Schistosoma mansoni and that resistance to these two structurally related schistosomicides is a consequence of mutation in the drug activating enzyme gene. Although this mechanism is supported by indirect evidence, the identity of the putative enzyme remains speculative. A number of biochemical and molecular genetic techniques will be used to achieve the following objectives: 1) Elucidate the proximal step of hycanthone and oxamniquine methabolism using cell-free preparations of a susceptible S. mansoni strain; 2) Determine whether the enzyme activity is absent or altered in drug-resistant strains of S. mansoni and in the refractory species S. japonicum; 3) Isolate and characterize the drug activating enzyme from a susceptible strain; and 4) Explore the molecular basis of drug resistance by investigating oxamniquine/hycanthone activating enzyme gene expression in susceptible and resistant S. mansoni strains. In addition to delineating the role of schistosome enzymes in prodrug activation, the results of these studies should contribute to a molecular description of parasite adaptive responses to a chemically-defined stimulus.
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