Adenovirus and Baculovirus Mediated Proangiogenic Gene Therapy in Skeletal Muscle and Uterine Artery Safety Aspects in Peripheral Arterial Disease and Fetal Growth Restriction Therapy

Proangiogenic gene therapy, with viral and non-viral vectors, has been shown to be highly efficient in several animal models. These results have encouraged investigators to proceed to treatment of patients with these new therapies in Phase I to III clinical trials. However, promising results of pre-clinical experiments have not fully translated into clinical success and as such improvements in the vectors and methods are under investigation. In addition, new target diseases are being actively searched for where gene therapy induced angiogenesis might be beneficial. The aims of this study were (1) to test whether a novel gamma secretase inhibitor (Compound X, CX) or baculoviruses are useful in improving skeletal muscle gene therapy efficacy and (2) to investigate the transduction efficiency and safety aspects, in terms of fetal survival, of adenovirus-based gene therapy, when it is injected into the uterine artery during pregnancy. In this study it was found that, CX efficiently improved vascular sprouting in cell cultures and in a mouse eye model. However, when CX was combined with adenoviral vascular endothelial growth factor (VEGF)-A gene transfer, in rabbit skeletal muscle, it actually disrupted the vessel endothelial cell layer integrity. In addition, there was a similar endothelial cell disruption detected in the liver following the combination therapy. Capillary growth was evident in the rabbit skeletal muscles 6 days after the baculovirus mediated VEGF-DΔNΔC gene transfer. When baculovirus was used as a vector to carry VEGF-DΔNΔC gene a significant improvement was seen in different cell lines after addition of VSV-G and WPRE into the vector. In the last study of the dissertation, three different genes were used to test the safety of adenoviral gene transfer to the uterine artery: thymidine kinase (Tk, with Ganciclovir), VEGF-DΔNΔC and LacZ. Adenovirus did not influence fetal survival when injected into the uterine artery at the end of the second trimester of the pregnancy. The transgene was found with PCR in 9 % of the offspring livers but no integration of the transgene was seen. In addition, qPCR analysis did not reveal any expression of the transgene in the fetal livers. In conclusion, we found that the safety profiles of CX or baculoviral gene therapy need to be further improved to become suitable for use in future in vivo studies, at least as proangiogenic therapies. In addition, the safety profile of adenoviral gene therapy was such that further studies are justified. Thus, new vectors and molecules which are highly efficient in cell culture and in rodent models should be evaluated for potency and safety also in larger animal models, such as rabbits or pigs, which better resemble human pharmacological properties before entering clinical trials. National Library of Medicine Classification: QU 107, QU 560, WE 500, WG 500 Medical Subject Headings: Genetic Therapy; Angiogenesis Inducing Agents; Blood Vessels/growth & development; Muscle, Skeletal; Uterine Artery; Amyloid Precursor Protein Secretases/antagonists & inhibitors; Adenoviridae; Vascular Endothelial Growth Factor A; Baculoviridae; Vascular Endothelial Growth Factor D; Cells, Cultured; Endothelial Cells/drug effects; Disease Models, Animal; Safety