Date of Award

2015

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Graduate Group

Cell & Molecular Biology

First Advisor

Valder R. Arruda

Second Advisor

Joel S. Bennett

Abstract

Treatment of hemophilia, which involves infusion of the missing clotting factor, is often hindered by the development of neutralizing antibodies to the replaced clotting factor. We utilized liver-directed AAV gene therapy to tolerize outbred hemophiliac dogs with pre-existing anti-factor VIII and IX antibodies and to treat their underlying hemophilia. Additionally, we sought to shed light on the immunologic mechanisms responsible for this tolerization. Staining for CD4+CD25+FoxP3+ T cells and cytokine profiles of treated dogs suggest that induced Tregs are at least partially responsible for inducing and maintaining tolerance.

The second part of the dissertation attempts to determine the underlying mechanism of the known anti-metastatic effects of activated protein C (aPC). We again utilized liver-directed AAV gene therapy to tease apart the anticoagulation and cytoprotective effects of aPC, as these were the most likely candidates based on available literature, in the B16F10 murine model of metastatic melanoma. Upon finding, however, that neither of these functions were involved in aPC's cancer protection and that zymogen PC is even more protective, we endeavored to find the novel mechanism through which the protein C pathway can modulate tumor progression. While the mechanism has not been found, a number of potential candidates and receptors have been eliminated, including involvement of PAR-4, EPCR, and a potential integrin binding site. We also demonstrate that it is not tumor growth, but rather a step in the metastatic pathway that is inhibited.

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