Adenovirus-mediated human Topoisomerase II alpha gene transfer increases the sensitivity of etoposide-resistant human breast cancer cells

Cellular resistance to chemotherapeutic agents is attributable to several m echanisms, including alteration of topoisomerase II alpha (topo II alpha) g ene expression. Etoposide-resistant MDA-VP human breast cancer cells expres s lower amounts of enzymatically active and drug-sensitive topo II alpha th an do MDA parent cells, suggesting that the low level of topo II alpha is t he mechanism of resistance. To determine whether transfer of a normal topo II alpha gene into MDA-VP cells can increase topo II alpha gene expression, topo II alpha protein production, and cell sensitivity to etoposide, a rec ombinant adenovirus, Ad-hTopo II alpha, containing the human topo II alpha gene, was constructed. The shuttle vector pAvCvSv-hTopII alpha was construc ted and cotransfected with the pBHG10 packaging vector into 293 cells. Infe ctious recombinant adenovirus plaques were isolated and purified. Presence of the topo II alpha gene was confirmed by PCR and restriction enzyme diges tion. After infection with Ad-hTopoII alpha, topo II alpha mRNA expression in MDA-VP cells increased 7.4-fold, topo II alpha protein production increa sed 5.9-fold, and sensitivity to etoposide was enhanced 4.5-fold compared w ith control transfected cells. Infection of normal human embryonic lung cel ls and human fibroblast cells with Ad-hTopoII alpha did not enhance the exp ression of topo II alpha or sensitivity to etoposide, Viral uptake was comp arable in the MDA-VP and normal cell lines. These data suggest that topo II alpha gene transfer using an adenoviral vector can selectively increase et oposide sensitivity in drug-resistant tumor cells and may enhance the thera peutic index of etoposide.