Lentiviral vectors as tools for gene manipulation
In this thesis the potential for lentiviral vectors as tools for gene manipulation was investigated. This thesis aims to answer the question of whether lentiviral vectors can be used to infect human stem cells with high efficiency without affecting their ability to differentiate and whether these vectors are able to deliver short hairpin RNA interference targeting a clinically relevant gene. The introduction covers first the basics of the areas involved in this thesis, starting with stem cell biology and RNA interference before discussing vector biology and then Kaposi's sarcoma-associated herpesvirus. Human stem cell biology is a novel and rapidly moving field. The applications of lentiviral vectors range from determining mechanisms of cell growth and differentiation to tissue engineering. The first application for these vectors investigated was therefore optimisation of the infection of human stem cells and determining that these cells retain their biological function after infection. Lentiviral vectors to deliver short hairpin RNA were developed and a knockdown in human stem cells demonstrated. Kaposi sarcoma-associated herpesvirus (KSHV), a cause of virally driven malignancies in humans, was chosen as a target to demonstrate the therapeutic potential of these lentiviral vectors. Lentiviral shRNA vectors knocked down latent genes within KSHV in vitro and showed promising therapeutic potential. The final section of this thesis documents the progression of this work to form an in vivo therapeutic against a KSHV-driven malignancy, primary effusion lymphoma (PEL) in a murine model.