Disorders of Copper Transport

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Menkes disease is an X-linked recessive disorder of copper transport caused by defects in a gene that encodes an evolutionarily conserved copper-transporting ATPase. In mammals, this gene product functions as an intracellular pump to transport copper into trans-Golgi spaces for incorporation into copper-requiring enzymes, and also mediates copper exodus from cells. The disorder presents in infancy with delayed development, failure to thrive, neurodegeneration, and premature death (typically by 3 years of age). Our work on this disorder includes development of rapid and reliable neurochemical and molecular techniques for very early diagnosis, efforts which dovetail with a clinical trial of very early copper histidine treatment for affected infants. We use cell biological, molecular, and biochemical approaches to characterize enrolled patients and to correlate with neurodevelopmental outcomes. Confocal imaging of patient fibroblasts is used to assess quantity and localization of mutant Menkes gene products. The blood-brain barrier poses a challenging treatment obstacle in many Menkes disease patients, and we hypothesized a molecular basis for treatment responsivity in the minority of patients (about 1 in 5) who respond successfully (normal neurodevelopmental outcomes) to early copper histidine. These patients have mutations that enable at least some residual copper transport to the developing brain. Consequently, we are developing alternative therapeutic approaches, including gene therapy, that bypass the blood-brain barrier. As above, although mean survival is significantly enhanced, only a fraction of patients with Menkes disease (~30%) show good or excellent neurological outcomes when treated with copper injections beginning very early in life. The main reason for the disparate outcomes appears to be the amount of residual Atp7a function in these individuals. Those with less Atp7a function are unlikely to respond optimally to copper injection treatment. Therefore, new therapeutic strategies need to be developed for the large percentage of patients who have little or no residual Atp7a function and currently have no ideal treatment options. Toward this goal, the Unit devised an animal study proposal, recently approved, to assess the efficacy of adeno-associated virus (AAV) gene therapy in mouse models of Menkes disease. Gene therapy offers an alternative method for treating genetic disorders, such as Menkes disease, that result from the loss of a protein function. This method provides a way to restore the lost protein function in an affected individual and has been successfully employed to treat a number of genetic disorders in animal models of disease and in at least one human disease. Several mouse models of Menkes disease exist. Like the human patients, the severity of the disease varies substantially from model to model, and only some models respond to injections of copper. Thus, when used in combination, these models provide an effective tool to evaluate the effects of disease severity on the effectiveness of novel treatments. The goal of our study is to evaluate the use of recombinant adeno-associated virus serotype 5 (rAAV5) as a gene therapy vector in two mouse models of Menkes disease, one responsive to early copper therapy and the other which does not respond to early copper therapy. Efficacy will be evaluated by examining life span and several biochemical parameters that are abnormal in Menkes disease mice and Menkes disease patients. If successful, these experiments will lay the groundwork for more effective therapies for a higher percentage of human patients with Menkes disease.