Anti-cancer binary system activated by bacteriophage HK022 integrase.

The binary system presented in this work is based on the bacteriophage HK022 integrase recombinase that activates the expression of a silenced Diphtheria toxin gene, both controlled by the cancer specific hTERT promoter. Using a lung cancer mice model, assays of different apoptotic and anti-apoptotic factors have demonstrated that the Integrase based binary system is highly specific towards cancer cells and more efficient compared to the conventional mono system whose toxin is directly expressed under hTERT. In a mice survival test, this binary system demonstrated longer persistence compared to the untreated and the mono treated ones. The reason underlying the advantage of this binary system over the mono system seems to be an overexpression of various hTERT suppressing factors induced by the mono system.

Follicle activation is a significant and immediate cause of follicle loss after ovarian tissue transplantation.

PURPOSE: Extensive follicle loss has been demonstrated in ovarian grafts post transplantation, reducing their productivity and lifespan. Several mechanisms for this loss have been proposed, and this study aims to clarify when and how the massive follicle loss associated with transplantation of ovarian tissue graft occurs. An understanding of the mechanisms of follicle loss will pinpoint potential new targets for optimization and improvement of this important fertility preservation technique. METHODS: Frozen-thawed marmoset (n = 15), bovine (n = 37), and human (n = 46) ovarian cortical tissue strips were transplanted subcutaneously into immunodeficient castrated male mice for 3 or 7 days. Histological (H&E, Masson's trichrome) analysis and immunostaining (Ki-67, GDF9, cleaved caspase-3) were conducted to assess transplantation-associated follicle dynamics, with untransplanted frozen-thawed tissue serving as a negative control. RESULTS: Evidence of extensive primordial follicle (PMF) activation and loss was observed already 3 days post transplantation in marmoset, bovine, and human tissue grafts, compared to frozen-thawed untransplanted controls (p < 0.001). No significant additional PMF loss was observed 7 days post transplantation. Recovered grafts of all species showed markedly higher rates of proliferative activity and progression from dormant to growing follicles (Ki-67 and GDF9 staining) as well as higher growing/primordial (GF/PMF) ratio (p < 0.02) and higher collagen levels compared with untransplanted controls. CONCLUSIONS: This multi-species study demonstrates that follicle activation plays an important role in transplantation-induced follicle loss, and that it occurs within a very short time frame after grafting. These results underline the need to prevent this activation at the time of transplantation in order to retain the maximal possible follicle reserve and extend graft lifespan.

Cancer-specific binary expression system activated in mice by bacteriophage HK022 Integrase.

Binary systems based on site-specific recombination have been used for tumor specific transcription targeting of suicide genes in animal models. In these binary systems a site specific recombinase or integrase that is expressed from a tumor specific promoter drives tumor specific expression of a cytotoxic gene. In the present study we developed a new cancer specific binary expression system activated by the Integrase (Int) of the lambdoid phage HK022. We demonstrate the validity of this system by the specific expression of a luciferase (luc) reporter in human embryonic kidney 293T (HEK293T) cells and in a lung cancer mouse model. Due to the absence viral vectors and of cytotoxicity the Int based binary system offers advantages over previously described counterparts and may therefore be developed into a safer cancer cell killing system.