Enhancement of HSV-TK/GCV suicide gene therapy of cancer

Gene therapy has become a promising alternative treatment form for cancer. Among the broad range of different genetic means to reduce the tumor growth, herpes simplex virus thymidine kinase/ganciclovir (HSV-TK/GCV) suicide gene therapy regimen is the best known approach. In this type of therapy, cancer cells are manipulated to express HSV-TK, followed by administration of the prodrug, the antiviral drug GCV. This prodrug is relatively harmless to normal cells but efficiently kills cells that express HSV-TK. The HSV-TK/GCV suicide gene therapy has been tested extensively, in the laboratory and some recent clinical results have also demonstrated the potential of this treatment form. However, cancer patients still cannot be cured with the method, indicating that this approach needs refinement before true clinical success can be achieved. One way to enhance suicide gene therapy is to increase the number of S phase cells in the tumor, since these cells are undergoing DNA replication and are thus vulnerable to the toxic form of GCV. It is known that alpha-difluoromethylornithine (DFMO), a well-known and well-tolerated polyamine biosynthesis inhibitor, can generate a prominent cell cycle arrest and when DFMO is withdrawn from the cells, they begin to divide again and display an elevated proportion of S phase cells for a certain period of time. This window of increased S phases may be exploited for enhancement of HSV-TK/GCV suicide gene therapy, at least in theory. To verify this hypothesis, the novel combination of polyamine depletion and suicide gene therapy was tested first in cultured cells and thereafter in a mouse tumor model. Furthermore, other types of drugs affecting the polyamine homeostasis or the cell cycle were investigated for this enhancing effect. The combination of HSV-TK/GCV gene therapy and DFMO resulted in an enhanced cytotoxic effect in cultured rat and human tumor cells. This synergistic effect was achieved only when the timing between DFMO treatment and HSV-TK/GCV gene therapy was optimal. A similar effect was also observed in a subcutaneous mouse tumor model, demonstrating the efficacy of this combination in vivo. However, other attemps to manipulate the polyamine homeostasis or cell cycle phase distribution (polyamine catabolism activation with N,N-diethylnorspermine, serum deprivation or treatment with aphidicolin, hydroxyurea, lovastatin, mimosin and resveratrol) did not significantly enhance the efficacy of HSV-TK/GCV gene therapy. Moreover, the duration of the S phase effect needs to be long enough to allow enhancement of HSVTK/GCV –mediated cell killing. In conclusion, our results indicate that correctly timed polyamine depletion with DFMO is an efficient way to enhance the HSV-TK/GCV gene therapy approach in human tumor cells and the effect also appears to be achievable in animal tumor models. Since both DFMO and HSVTK/GCV gene therapy have been extensively tested in clinical trials and their safety profiles have turned out to be excellent, it is realistic to hope that this combination treatment will be successful in clinical studies. However, only further preclinical studies with more relevant (orthotopic) animal models and primary human tumor material will reveal the true utility of the combination. National Library of Medicine Classification: QZ 266, QZ 52, QU 141, QU 61, QU 375 Medical Subject Headings: neoplasms/therapy; gene therapy; genes, transgenic, suicide; cell death; simplexvirus; thymidine kinase; ganciclovir; cell cycle; S phase; polyamines/antagonists & inhibitors; eflornithine; cells, cultured; disease models, animal