Enhanced specificity of the p53 family proteins-based adenoviral gene therapy in uterine cervical cancer cells with E2F1-responsive promoters.

p63 and p73, the p53 family proteins, are similar to p53 in many aspects: structural homology, transactivation of p53-downstream genes, and induction of apoptosis. Interestingly, they also differ from p53; in particular, they are not inhibited by viral oncoproteins such as HPV E6. This feature would be an advantage over p53 in HPV-associated cancers and therefore, we evaluated the therapeutic potentials of various p53 family proteins (p73alpha, p73beta, p63alpha and p63gamma) against HPV-infected cervical cancers. In clonogenic assay, exogenous expression of p73alpha, p73beta and p63gamma inhibited the colony formation of HPV-positive cervical cancer cells under G418- selection while p53 could not. Recombinant adenoviruses Ad/CMVp73alpha, Ad/CMVp73beta and Ad/CMVp63gamma induced potent apoptosis in all infected cervical cancers (CasKi, SiHa, HeLa, C33A, SNU682, SNU17, SNU1005, SNU703), irrespective of their HPV-infection status. Unfortunately however, Ad/CMVp73alpha, Ad/CMVp73beta, and Ad/CMVp63gamma inhibited also normal cells such as endothelial cells, fibroblasts, and keratinocytes thus, tumorspecific promoter was indispensable to the p53 family proteins-based therapy. Here we report a stringent tumor killing by p73beta in combination with ESM6, a synthetic promoter targeting the DNA tumor virusassociated cancers. Recombinant adenoviruses encoding p73beta by ESM6 (Ad/ESM6p73beta and Ad/ESM6p73bENH) expressed p73beta and induced apoptosis only in the cancer cells but not in normal cells. Collectively, we suggest that the p53 family proteins are potent therapeutic agents for HPV-associated uterine cervical cancers and ESM6 mediated expression of the p53 family proteins would be a safe and strong tumor targeting strategy.