Development of Genetic Medicines for the Treatment and Prevention of Infectious Disease, Cancer, and Aging
Creators: Brown, Douglas Wilson
Publisher: name: University of Alberta Library
Year: 2022

<p>Gene therapy has potential to become the Holy Grail of modern medicine, however, its clinical success has been hindered due to the lack of a safe and effective nucleic acid delivery platform. The recent success of non-viral RNA delivery platforms has revitalized the field and spurred great interest in advancing these technologies to the clinic. In this thesis, I first describe the development and design of a novel non-viral nucleic acid delivery platform that utilizes a unique chimeric fusion protein to facilitate intracellular delivery of encapsulated cargo. I demonstrate that incorporation of this fusion protein into a lipid-based delivery platform enhances transgene expression and enables toxic components to be reduced, which significantly increases the tolerability relative to other clinically approved platforms. After establishing the feasibility of this nucleic acid delivery platform (called Fusogenix proteolipid vehicles or FAST-PLVs), I then explore various treatment indications in which gene therapy might provide some benefit. The first is development of a DNA-based vaccine against SARS-CoV-2, the virus that causes the disease known as COVID-19. We show that this vaccine can induce robust humoral immune responses, comparable to convalescent patients, and is able to protect hamsters from morbidity following viral challenge. Next, I examine the potential to create a genetic medicine for the treatment of one of the most complex genetic diseases, cancer. I first explore the consequence of p53 mutation on its own transcriptional regulation. I find that loss of p53 function is correlated with an upregulation in p53 transcriptional activity. Using this information, we designed a DNA-based suicide gene therapy, where the expression of a late-stage apoptotic protein is under control of the p53 promoter. I show that not only does this treatment selectively target cancer cells, but it also offers an effective means to attenuate tumor growth. Additionally, I demonstrate that the combination of this treatment with immunotherapy is able to achieve tumor control in a poorly immunogenic, highly metastatic tumor model. Finally, I explore the possibility of developing a genetic senolytic treatment to improve lifespan and prevent age-related organ function decline. To achieve this, we mimic the original transgenic mouse models that selectively eliminated senescent cells on the basis of their transcriptional activity. By generating a DNA-based suicide gene therapy where the expression of the same late-stage apoptotic protein is under control of the p16Ink4a promoter, we are able to selectively target and remove senescent cells. Combining this construct with the p53 promoter construct described above offers an effective means to improve lifespan in naturally aged mice possibly due to senescent cell elimination in the kidney. My work can help pave the way for the next generation of genetic medicines, as these Fusogenix proteolipid vehicles are currently the only known non-viral DNA delivery platform and our SARS-CoV-2 vaccine has begun phase II clinical trials. Additionally, this work may stimulate the development of new treatment strategies that target complex diseases based on their transcriptional activity. </p>