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The development of herpes simplex virus vectors for cancer and gene therapy

University of British Columbia

The efficient delivery of therapeutic genes and appropriate gene expression are the crucial issues for clinically relevant gene therapy. Viruses are naturally evolved vehicles that efficiently transfer their genes into host cells. This ability made them desirable for engineering virus vector systems for the delivery of therapeutic genes. Among various vector systems, herpes simplex virus (HSV) vectors represent an attractive delivery system, since these vectors have high gene transfer efficiency and mediate high expression of therapeutic genes. Although HSV has been shown to infect most cell types, they were restricted from mature skeletal muscle tissue. As a result, research involving the use of this vector for muscle-directed gene therapy was hampered. Previous studies indicated that the loss of infectivity may be due, at least in part, to the development of the basal lamina throughout the course of muscle maturation. Enzymatic disruptions of the basal lamina showed moderate increases in levels of infection, although marked toxicity with such procedures resulted. To initiate infection, HSV normally attaches to cell surface heparan sulfate, which stabilizes the virus such that it can interact with secondary protein receptors required for entry into host cells. Our studies revealed a downregulation of heparan sulfate biosynthesis during skeletal muscle maturation. Furthermore, infectivity could be restored by exposing mature skeletal myofibers to low concentrations of the glycosaminoglycan analog, dextran sulfate (DS). This molecule appears to act as a surrogate receptor to stabilize the virus at the myofiber surface such that HSV can engage additional receptors. This demonstration that the basal lamina is not an absolute block to HSV infection is remarkable because it allows for the nondestructive targeting of HSV to mature myofibers and greatly expands the usefulness of this vector for the treatment of inherited and acquired diseases. In light of these results, dextran sulfate was further examined for its ability to target cancer cells in a systemic model of delivery. Cancer cells typically display altered glycosaminoglycan profiles, similar to mature skeletal muscle tissue. Vascular delivery of oncolytic HSV and DS significantly delayed tumor growth, with 25% of the animals cured following treatment. Although, DS did not act to stimulate infection of cancer cells, its ability to alter the hemodynamic properties of the animal system in favor of viral accumulation at tumor portals was key. Surprisingly, viral replication was not necessary for antitumor efficacy and relatively low amounts of virus could result in marked oncolysis. Furthermore, immunohistochemistry revealed infection of tumor vasculature alongside very limited infection of surrounding tumor tissue. Taken together, the tumor vasculature is likely the major target for oncolytic HSV in a systemic delivery model of cancer. Thereby efforts to further enhance delivery of oncolytic HSV to tumor vasculature by incorporating targeting peptides and using antiangiogenic viruses have been successful. Understanding the biology of gene therapy systems is crucial to developing the most efficient and specific systems suited for individual disease applications. Increased insights into the entry and trafficking of gene therapy systems in animal models facilitates two approaches to developing appropriate therapies for individual applications. First, understanding more clearly the biology of currently available systems permits a more judicious choice of applications. Second, this also forms the basis for development of advanced delivery systems with increased efficiency, stability and targeting specificity. Therefore studies that provide insight into why biological therapies succeed and fail not only allow for a better understanding of animal and vector systems, but they allow us to exploit this knowledge to improve our arsenal of standard protocols of care for disease.

Thesis/Dissertation

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2009-10-09

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http://hdl.handle.net/2429/13841

Microbiology and Immunology

University of British Columbia

UBCV

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