Connexin 43-enhanced suicide gene therapy using herpesviral vectors

Tumor cell transduction with the herpes simplex virus (HSV) thymidine kinas e (tk) gene and treatment with ganciclovir (GCV) is a widely studied cancer gene therapy. Connexin (Cx)-dependent gap junctions between cells facilita te the intercellular spread of TK-activated GCV, thereby creating a bystand er effect that improves tumor cell killing. However, tumor cells often have reduced connexin expression, thus thwarting bystander killing and the effe ctiveness of TK/GCV gene therapy. To improve the effectiveness of this ther apy, we compared an HSV vector (TOCX) expressing Cx43 in addition to TK wit h an isogenic tk vector (TOZ.1) for their abilities to induce bystander kil ling of Cx-positive U-87 MC human glioblastoma cells and Cx-negative L929 f ibrosarcoma cells in vitro and in vivo. The results showed that low-multipl icity infection of U-87 MG cells with TOCX only minimally increased GCV-med iated cell death compared with infection by TOZ.1, consistent with the endo genous level of Cx in these cells. In contrast, bystander killing of L929 c ells was markedly enhanced by vector-mediated expression of Cx. In vivo exp eriments in which U-87 MC cells were preinfected at low multiplicity and in jected into the flanks of nude mice showed complete cures of all animals in the TOCX group following GCV treatment, whereas untreated animals uniforml y formed fatal tumors. TOCX injection into U-87 MG intradermal and intracra nial tumors resulted in prolonged survival of the host animals in a GCV-dep endent manner. Together; these results suggest that the combination of TK a nd Cx may be beneficial for the treatment of human glioblastoma.