The molecular study of defects in ammonia metabolism has been generally limited to rare inborn errors of metabolism. Hyperammonemia, however, is much more common and is associated with non-genetic conditions and environmental exposures. These conditions usually involve some degree of physical hepatic damage or disruption. A common example is the frequent hyperammonemia observed in patients following administration of the drug valproic acid. With the increase in bone marrow transplant procedures (BMT), a number of hyperammonemic deaths have been reported. In addition, more subtle toxicities may result from mild increases in urea cycle precursors (glutamine, alanine, glycine, etc.), or decreases in urea cycle intermediates (arginine and citrulline). Whatever the triggering event, disturbances in ammonia clearance reflect physical or biochemical effects on the urea cycle. My research has focused on characterizing the gene encoding the first, rate-limiting step of ureagenesis, carbamyl phosphate synthetase I (CPSI). In patient studies, I have found a number of rare CPSI molecular defects resulting in severe inherited disruptions of the urea cycle. During these studies, I have also identified a common exonic polymorphism which may qualitatively affect CPSI function. This study is designed to examine the clinical and biochemical significance of this polymorphism and identify other potentially relevant changes in the CPSI gene. These changes may play a key role in the susceptibility to urea cycle toxicity secondary to drug or toxin exposure. I will study the functional affects of this change on expressed CPSI protein using site-directed mutants of our CPSI expression clone. In addition to studying the functional characteristics of this polymorphism, I will determine if there is any association between the CPSI genotype and the toxicity observed in patients undergoing BMT. As part of my collaboration with Dr. Brian Christman, we have observed significant changes in urea cycle intermediates following induction chemotherapy for BMT, and have preliminary data showing disequilibrium with the CPSI polymorphism. We will further test the correlation between the CPSI polymorphic genotypes with the presence of observed biochemical/clinical toxicity in patients beginning therapy with valproic acid. We will also screen the CPSI gene for other exonic polymorphisms which can be tested. We will establish the role of these common polymorphisms in the urea-cycle related toxicity seen in these patients. Positive results in this study will lead to further study of other conditions involving the derangement of waste nitrogen disposal.
