Ovine adenovirus (OAV) vectors represent a promising tool for human gene therapy since these vectors overcome the problem of pre-existing immunity against human adenovirus vectors. In this report we investigated the in vivo characteristics of this novel vector system with respect to its potential for gene transfer into skeletal muscle. We found that moderate doses of an OAV-derived vector expressing the human alpha1-antitrypsin gene (OAVhaat) infected skeletal muscle in mice very efficiently resulting in high serum hAAT levels. The infection was restricted to skeletal muscle, but gene expression was transient and vector DNA was rapidly cleared. Vector clearance was also observed with a vector that lacked the transgene. The loss of vector DNA was accompanied by a cellular immune response in the infected muscle but was not connected with detectable expression of early or late genes of the viral backbone as analyzed by RT-PCR. A very low dose of OAVhaat (3x 10(7) infectious particles) was sufficient to produce reasonable amounts (>100 ng/ml) of serum hAAT, and this was accompanied by a weak immune response to the vector. Under these conditions, a second intramuscular injection of the same recombinant OAV vector was successful. Our study expands the known tissue tropism of OAV-derived vectors in vivo and points to the possible utility of the vector for muscle gene transfer and vaccination.