SAFETY EVALUATION OF AD5CMV-P53 IN-VITRO AND IN-VIVO

In preparation for a clinical trial of the recombinant p53 adenovirus Ad5CMV-p53 for the treatment of lung cancer, the potential adverse eff ects of Ad5CMV-p53 were assessed in vitro and in vivo. No infectious r eplication of Ad5CMV-p53 was detectable in HeLa cells infected with ex tracts from HeLa cells previously Infected with Ad5CMV-p53. No Ad5CMV- p53 DNA replication was detected by (32)Pi labeling in lung cancer cel ls infected with Ad5CMV-p53 at multiplicities of infection (moi) up to 1,000 pfu/cell (total of 5 x 10(9) pfu viruses). The infectivity and cytotoxicity of Ad5CMV-p53 were examined in vitro in normal human bron chial epithelial (NHBE) cells. At a moi of 50 pfu/cell, Ad5CMV-p53 inf ection and expression were detectable in 80% of the treated cells. The exogenous p53 protein was first detected by western blotting at 8 hr and peaked at 48 hr after infection. Growth of NHBE cells was not affe cted by Ad5CMV-p53 infection at a moi of 100 pfu/cell. The pathogenici ty of Ad5CMV-p53 was assessed in BALB/c mice. The virus was given to f our groups of mice by intratracheal injection at dosages from 10(7) to 10(10) pfu; a fifth group received phosphate-buffered saline alone. N one of the viral injections proved to be lethal; Mild to moderate peri bronchiolar and perivascular infiltration by mononuclear cells and lym phocytes, with patches of pneumonitis, was the most acute toxic effect detected by histologic analysis in the two high-dose groups. Immunohi stochemical analysis of the same paraffin-embedded sections showed tha t infectivity and level of expression of p53 in lung tissue were dose- dependent. Our results demonstrate that Ad5CMV-p53 is a replication-de fective virus that yields a relatively low degree of acute toxicity in mice; these data document a safety profile encouraging for clinical t rials of Ad5CMV-p53 in the therapy of lung cancer.