Maple Syrup Urine Disease (MSUD) is a genetic disorder of branched- chain amino acid (BCAA) metabolism. The incidence of MSUD in general population is 1:185,000 and in certain population groups, such as the Mennonites of Pennsylvania, the incidence is as high as 1:176. Children with MSUD suffer from profound metabolic complications that culminate in mental retardation and early death. At the present time, no satisfactory treatment for MSUD is available. To facilitate develop of novel treatments, such as gene therapy, we have recently made a murine model of MSUD by gene knockout of the E2 subunit of branched-chain keto acid dehydrogenase (BCKDH), the rate-limiting enzyme for the catabolism of BCAA. However, as expected, all the MSUD pups die within a few days after birth, thus limiting the utility of this mouse. To enhance the utility of the MSUD model, we are embarking on creating a conditional transgenic rescue of the perinatal lethal phenotype. This refinement will significantly enhance the model's usefulness. The specific aims of this SBIR Phase-I project are: (1) to assemble a tetracycline-regulated E2 expression vector, (2) to test the expression vector in vitro for E2 expression and tetracycline regulation, (3) to create tetracycline-regulated E2 transgenic mouse lines by pronuclear microinjection, and (4) to characterize E2 expression and tetracycline regulation in liver of transgenic mice. Subsequently, the transgenic lines will be crossed onto the present MSUD (E2 null) background to produce mice that harbor the tetracycline-regulated E2 transgene and are homozygous for the endogenous E2 knockout. Such mice will overcome the problem of neonatal lethality and will constitute a very useful MSUD model for the development and optimization of novel approaches to cure this devastating disease. PROPOSED COMMERCIAL APPLICATIONS: Testing of novel therapies using the conditional MSUD mouse will lead to development of new. products. These include diagnostic tools and technologies, gene therapy, vectors, candidate drugs, and novel dietary formulas . The potential use of these products and technologies in a clinical setting is expected to be of significant commercial interest and application.
