DNA sequences encoding modified retroviral gag polypeptides and vaccines containing them or aggregates thereof.

摘要

Conventional HIV-1 vaccine strategies, based on the external part of the env protein gp160/120, were till now not found to induce protective immunity. In addition, enhancing antibodies may negatively influence disease progression. Due to the early appearence in infection and its particulate nature, the virus core protein p55-GAG seems to be a promising vaccine candidate. p55-core expression was assayed by recombinant vaccinia and baculo viruses. Shedding of 90-110 nm core particles into culture medium was observed in both expression systems and proven by ultrathin-section electron microscopy and sedimentation analysis. Addition of the protease coding sequence resulted in an efficient processing of the p55-precursor molecule - in the Baculo-system only. Significant protease mediated processing of the GAG precursor in the Vaccinia-system was only achievable by addition of the entire POL coding sequence. In contrast to other retroviruses (MMLV), HIV-1 protease mediated processing is not dependent on the myristylation of p55-GAG. As protease mediated processing does not complete the maturation process of the p55-precursor particles, but, in contrast, inhibits particle formation, further investigations focused on the application of premature p55-GAG-particles. To extend their immunological spectrum, a consensus sequence of the HIV-major neutralizing epitope V3 of gp120, designed in our lab, was inserted into different regions of the p55 carrier molecule. The expression of the p55/V3 chimeric proteins in E. coli and the Vaccinia expression system was proven by Western blot analysis using (i) monoclonal antibodies directed to p55 and the inserted V3 region and (ii) polyclonal monospecific anti-V3 peptide sera. p55/V3 hybrid particles will be purified from Baculovirus infected cells. Recombinant p55/V3 Vaccinia viruses are, in addition to antigen production, directly used as a life vaccine in experimental animals. <IMAGE>