ANALYSIS OF TISSUE-SPECIFIC GENE DELIVERY BY RECOMBINANT ADENOVIRUSESCONTAINING CARDIAC-SPECIFIC PROMOTERS

Objective: To approach heart muscle diseases by gene transfer, an aden oviral vector system was intended to be established suitable for gene expression in ventricular and/or atrial myocardium. Methods: Two adeno viral vectors (Ad-mhcLuc, Ad-mlcLuc) were constructed, in which the lu ciferase reporter gene is under control of either the ventricle-specif ic myosin light chain-2 (mlc-2v) or the atrial-and ventricular-specifi c alpha-myosin heavy chain (alpha-mhc) promoter. For controls, a recom binant adenovirus without promoter (Ad-Luc) and one with the Rous sarc oma virus (rsv) promoter (Ad-rsvLuc) were generated. A volume of 20 mu l containing 2x10(9) plaque forming units (pfu) of the recombinant ad enoviruses Ad-mhcLuc, Ad-mlcLuc, Ad-rsvLuc or Ad-Luc was injected into the cardiac cavity or the quadriceps femoris muscle of neonatal rats. After five days animals were sacrificed and nine different tissues we re analyzed for reporter gene expression by detection of light activit y relative to mg of tissue. Results: Injections of recombinant adenovi ruses into the cardiac cavity of neonatal rats resulted in heart-speci fic gene expression of Ad-mlcLuc (20 fold of Ad-Luc; 11% of Ad-rsvLuc) , whereas Ad-mhcLuc gave mainly luciferase activity in the heart (6.5 fold of Ad-Luc; 3% of Ad-rsvLuc) with additional activity in lung and liver (2-4 fold of Ad-Luc). In the ventricular tissue Ad-mlcLuc reveal ed a 35-fold higher luciferase activity, whereas Ad-mhcLuc, Ad-rsvLuc and Ad-Luc showed only 2-fold higher luciferase activities compared to the atrium. Viral DNA in atrial and ventricular tissue was detected b y PCR at approximately the same abundance independent of the injected type of adenovirus. Direct injection of Ad-mhcLuc and Ad-mlcLuc into t he thigh muscle revealed only background luciferase activities, Conclu sions: In the adenoviral system only the mlc-2v promoter may fulfil th e safety requirements for a myocardial specific gene expression with a high selectivity for the ventricular myocardium, thus providing a pro mising tool for future gene therapy of cardiomyopathies. (C) 1997 Else vier Science B.V.