Date of Award

2016

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Graduate Group

Cell & Molecular Biology

First Advisor

David B. Weiner

Abstract

Monoclonal antibodies (mAbs) have become important therapeutic and prophylactic agents for a number of indications, including infectious diseases. However, due to many issues, particularly the high cost of antibody production, mAb therapies are limited to the world’s richest populations. Furthermore, lengthy product development programs mean only a small number of mAb products can be produced at any one time. Engineering novel, low-cost, and simple methods of developing and delivering mAbs would be highly advantageous, potentially expanding the utility of antibody approaches into a wider array of applications. Here, we describe an approach to deliver human IgG neutralizing mAbs in vivo using DNA plasmid-mediated antibody gene transfer. This approach, which we term DNA mAb (DMAb) delivery, generates biologically relevant levels of mAbs after a single intramuscular injection of antibody-encoding DNA followed by in vivo electroporation (EP). First, we developed antibody-encoding DNA plasmids that could reproducibly deliver human mAbs to mouse serum. We show that these plasmid-encoded antibodies have similar binding capacity and functionality to in vitro-produced purified antibodies. Then, we use a mouse model to show that intramuscular delivery of pDVSF-3 LALA, which encodes a human anti-Dengue virus (DENV) IgG1 neutralizing antibody modified with a mutation that abrogates Fcγ receptor binding, produces anti-DENV antisera capable of binding to and neutralizing DENV1-3. Importantly, mice receiving pDVSF-3 LALA, but not the unmodified pDVSF-3 WT, were protected from both virus-only disease and antibody-enhanced lethal disease. To build upon these initial findings, we evaluated targeted genetic approaches and alternative delivery regimens in order to increase DMAb expression in vivo. Using DMAbs encoding human IgG1 antibodies against Borrelia burgdorferi (the causative agent of Lyme disease) as a model, we show that specific amino acid modifications to the framework regions of antibody variable domains confer increased in vitro and in vivo DMAb expression levels compared to the original DMAb sequences. Of note, these modifications were found to have no detrimental effect on the antibody’s borreliacidal activity. Lastly, we observed that pre-treatment of the DMAb injection site with hyaluronidase resulted in a 2.4 to 6.4-fold increase in human IgG concentration levels in vivo compared to mice receiving EP-mediated DMAb delivery only. Taken together, these data establish DNA plasmid-based antibody gene transfer as a safe, effective means of delivering tailored, protective monoclonal antibodies to hosts.

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