EXPRESSION FROM CARDIOMYOCYTE SPECIFIC PROMOTER AFTER ADENOVIRUS-MEDIATED GENE-TRANSFER IN-VITRO AND IN-VIVO

Adenoviruses are very attractive vectors for gene transfer into the ca rdiac muscle; however, their promiscuous tissue tropism, leading to an ectopic expression of the transgene, is a considerable practical limi tation. To restrict expression of a reporter gene in cultured cardiomy ocytes and in the heart of the rat, we have constructed a recombinant adenovirus (Ad-MCL2 beta gal) containing the beta-galactosidase gene u nder the control of the rat ventricle-specific cardiac myosin light ch ain 2 (MCL-2v) promoter. We show in this work that the MLC-2v promoter inside the adenoviral genome retains its cardiac specificity in vitro in culture cardiomyocytes as well as in vivo in the animal heart. Nor thern blot studies after Ad-MLC2 beta gal infection show significant t ranscription only in cells derived from the cardiac muscle and not fro m the skeletal muscle. Quantitative analysis of the beta-galactosidase activity in a number of cell lines also confirms this result. The lev el of beta-galactosidase expression in rat neonatal cardiomyocytes inf ected with Ad-MLC2 beta gal is 8% of that found when primary cells are infected with Ad-RSV beta gal (containing a beta-galactosidase gene u nder the control of the Rous sarcoma virus promoter). The cardiomyocyt es-specific expression is also found after injection of Ad-MLC2 beta g al directly into the rat myocardium, although the viral genome can be detected by polymerase chain reaction (PCR) in other tissues. Lack of expression after direct injection into liver and skeletal muscle confi rms these results. The use of a tissue-specific promoter is a first st ep to restrict transgene expression to a particular cell type of the t argeted tissue.