TRANSPLANTATION
OF NORMAL AND GENETICALLY ALTERED MYOBLASTS
The goal of Dr. Tremblay laboratory is to develop a treatment for Duchenne Muscular Dystrophy (DMD). This hereditary disease is characterized by a progressive weakness of the skeletal muscles. The weakness becomes evident at the age of 5 when the patient starts to have difficulties in walking. By the age of 10, the patient is confined to a wheel chair. Death occurs between the ages of 17 to 23 year due to weakening of the respiratory muscles. The gene responsible for this disease was identified in 1987. It encodes for a large protein, called dystrophin, which is located under the membrane of the muscle fiber. The absence of this protein in DMD patients makes the muscle fiber membrane more vulnerable to exercise. The damaged muscle fibers are repaired by the proliferation of small cells, called myoblasts, which are located near each muscle fiber. Unfortunately, the repaired muscle fibers are still void of dystrophin and will be damaged again. The repeated cycles of degeneration and regeneration leads to senescence of the myoblasts and thus to a reduced repair capacity which in turn causes muscle weakness. Since the discovery of the dystrophin mutation, scientists have been seeking ways to restore this protein in the patients muscle fibers. Since 1987, Dr. Tremblay's laboratory has been transplanting normal donor myoblasts into DMD patients msucles. This potential treatment was investigated in patients in 1990-91 but the results only showed small increases of the dystrophin positive fibers and only transient strength increases in a few patients.
Since this early clinical trial, Dr. Tremblay's laboratory has identified the problems responsible for this limited success and is currently looking for solutions. The first problem is a rapid inflammatory reaction following the intramuscular injection of myoblasts. This problem is reduced in animal models by using an antibody against a neutrophil adhesion molecule called LFA-1. The second problem is a limited diffusion of the injected cells due to the presence of an extracellular matrix. Dr. Tremblay is trying to resolve this problem by inserting in the myoblasts, prior to their transplantation, a gene encoding a metalloproteinase (an enzyme which digests the extracellular matrix). Finally the third problem is a specific immune response against the donor cells. This problem is resolved by immunosuppressing the patient with FK506.
The research conducted by Dr. Tremblay's team has achieved very good transplantation results in animal models. In dystrophic mice, the transplantation of myoblasts has restored the expression of dystrophin in 95% of the muscle fibers. In monkeys, more than 10,000 muscle fibers in a cm3 expressed a reporter gene present in the transplanted myoblasts.
Although myoblast transplantation is a potential solution to many myopathies, it has also been observed that myoblasts are excellent secretory cells. The transplantation of genetically modified myoblasts has allowed for the secretion of factor IX and Granulocyte Colony Stimulating Factor (GCSF).
Dr. Tremblay is therefore currently pursuing research aimed at transplanting normal or genetically modified cells for the treatment of hereditary and acquired diseases.
These projects are supported by the Muscular Dystrophy Association of Canada and the Association française contre les myopathies.
TREMBLAY, J.P., MALOUIN, F., ROY, R., HUARD, J., BOUCHARD, J.P., SATOH, A. and RICHARDS, C.L. (1993) Results of a triple blind clinical study of myoblast transplantations without immunosuppresive treatment in young boys with Duchenne muscular dystrophy. Cell Transplantation 2, 99-112.
TREMBLAY, J.P., BOUCHARD, J.P., THÉAU, D., COTTRELL, F., COLLIN, H., TREMBLAY, M., RICHARDS, C.L., MALOUIN, F., TOMÉ, F. and FARDEAU, M. (1993) Myoblast transplantation between monozygotic twin girls which are carriers of Duchenne muscular dystrophy. Neuromus. Disorders 3, 583-593.
KINOSHITA, I., VILQUIN, J.T., GUÉRETTE, B., ASSELIN, I., ROY, R. and TREMBLAY, J.P. (1994) Very efficient myoblast allotransplantation in mice under FK506 immunosuppression. Muscle and Nerve 17, 1407-1415.
VILQUIN, J.T., WAGNER, E., KINOSHITA, I., ROY, R. and TREMBLAY, J.P. (1995) Successful histocompatible myoblast transplantation in dystrophin-deficient mdx mouse despite the production of antibodies against dystrophin. J. Cell Biol. 131:975-988.
KINOSHITA, I., ROY, R., DUGRÉ, F.J., GRAVEL, C., VILQUIN, J.T., ROY, B., GOULET, M., ASSELIN, I. and TREMBLAY, J.P. (1996) Myoblast transplantation in monkeys: control of immune response by FK506. J. Neuropath. and Exp. Neurol. 55:687-697.