Double Suicide Gene Therapy Augments the Antitumor Activity of a Replication-Competent Lytic Adenovirus through Enhanced Cytotoxicity and Radiosensitization

Abstract

Replication-competent adenoviruses may provide a highly efficient means of delivering therapeutic genes to tumors. Previously, we evaluated in vitro a replication-competent adenovirus (Ad5-CD/TKrep) containing a cytosine deaminase (CD)/herpes simplex type 1 thymidine kinase (HSV-1 TK) fusion gene that allows lytic viral therapy to be combined with double suicide gene therapy. Both the CD/5-FC and HSV-1 TK/GCV enzyme/prodrug systems enhanced the tumor cell-specific cytopathic effects of the Ad5-CD/TKrep virus in vitro and sensitized cells to radiation. To extend these in vitro findings in vivo, we evaluated the antitumor activity of the Ad5-CD/TKrep virus in combination with double prodrug therapy and radiation therapy. The Ad5-CD/TKrep virus independently demonstrated significant antitumor activity against C33A cervical carcinoma xenografts. Therapeutic outcome was dramatically improved with systemic administration of double, but not single, prodrug (5-FC + GCV) therapy. When used in a neoadjuvant setting, Ad5-CD/TKrep-mediated double suicide gene therapy dramatically potentiated the effectiveness of radiation therapy. The trimodal approach of Ad5-CD/TKrep viral, double suicide gene, and radiotherapies produced significant tumor regression and ultimately 100% tumor cure. The results demonstrate the high therapeutic potential of the trimodal approach and provide a solid foundation for future clinical trials.

Inclusion of the herpes simplex thymidine kinase gene in a replicating adenovirus does not augment antitumor efficacy

“…Presumably a balance is reached between augmented GCV-induced cell killing due to production of toxic GCV metabolites versus the GCV-induced reduction of cell killing due to impaired viral replication. Our data, and that of Rogulski et al, 24 would suggest that loss of lytic killing by the virus offsets any benefits due to enhanced transgene spread.…”

“…Augmented tumor killing was only seen in those animals treated with both GCV and 5-FC. In this study, the reason for this finding was not determined for certain, however, Rogulski et al 24 showed that the HSVtk/GCV system was effective in inhibiting viral replication. Presumably a balance is reached between augmented GCV-induced cell killing due to production of toxic GCV metabolites versus the GCV-induced reduction of cell killing due to impaired viral replication.…”

“…However, in more recent publications by Morris and Wildner 21,22 using wild-type replicating adenoviruses containing an HSVtk transgene, but with no E1B deletion (Ad.OW34 and Ad.OW37), no augmentation of killing was observed after the addition of GCV. Similar to these latter findings, Freytag et al 23 and Rogulski et al 24 designed a virus that also lacked E1B, but contained a cytosine-deaminase/herpes simplex thymidine kinase fusion gene (FGR virus). The FGR virus had good antitumor effects after injection into cervical carcinoma xenografts, however, addition of GCV did not augment the efficacy of the virus.…”

Enhanced suicide gene effect by adenoviral transduction of a VP22-cytosine deaminase (CD) fusion gene

“…Especially concomitant stimulation of the immune system by chemokines 46 or incorporation of two suicide genes, HSV-1 TK and E. coli CD, into a replicating adenovirus 47 have shown compelling success in animal models. When additionally improving the limited in vivo transduction efficiency with the help of intercellular cargo proteins, successful gene therapy of solid tumours seems feasible.…”

Heat-directed suicide gene therapy for breast cancer

“…Conditionally replicating adenoviral vectors have already been shown to enhance significantly the therapeutic impact of cancer gene therapy strategies employing suicide genes, including the CDTK fusion gene used in this study. 41,42 In addition, Lee et al 35 have described an E1b-attenuated conditionally replicating adenovirus in which CDTK gene expression is under the control of the HSP70b promoter. Infection of DU145 prostatic carcinoma and CX-1 colorectal carcinoma cells with this adenovirus resulted in greater than 1000-fold reductions in survival following treatment with drugs and mild hyperthermia in vitro.…”