Document Type

Dissertation

Date of Degree

Spring 2010

Degree Name

PhD (Doctor of Philosophy)

Degree In

Chemical and Biochemical Engineering

First Advisor

Aliasger K. Salem

Abstract

Prostate cancer is the most common non-skin cancer in America, and the most commonly diagnosed cancer among males. When metastatic, the disease can ultimately be incurable. Consequently, alternative strategies to current treatments are sought, especially in the area of immunotherapy. Vaccine immunotherapy using a specific antigen, such as prostate specific antigen (PSA) seeks to stimulate both the innate and adaptive immune system to destroy tumor cells in the body. PSA is an ideal target antigen given that it has a narrow distribution in tissues and is expressed in virtually all prostate cancer cases. An adenovirus encoding for PSA (Ad-PSA) can be used to deliver the genomic data encoding for PSA production and secretion to the target cell. This type of viral gene delivery system has already been shown to have the potential to stimulate anti-tumor activity.

To enhance this activity and increase transfection efficiency, we proposed the combination of a viral system with a non-viral system, in the form of a cationic polymer such as poly(ethyl)enimine (PEI) or chitosan. Cationic polymers complex with the negatively charged adenovirus to form nanoparticles that can be used in gene delivery. Delivery in nanoparticle form can give enhanced uptake by the antigen-presenting cells necessary to initiate the targeted immune response. To further augment this response, previous research has shown that CpG sequences act as an adjuvant to enhance the efficacy of the Ad-PSA vaccines' tumor protection. CpG delivered in particulate form has also been shown to be more effective than delivery in solution. The objective of this proposal was to test the hypothesis that co-delivery of this targeted viral/non-viral gene delivery system will enhance tumor protection in a mouse model of prostate cancer.

Using the OVA model antigen system, we found that the adenovirus encoding OVA (AdOVA), coupled with the polymer PEI, enhanced tumor protection in vivo compared to AdOVA alone. To move towards our therapeutic model, these experiments were repeated using chitosan as the cationic polymer carrier, delivering AdOVA, and incorporating CpG into some particles. In this set of experiments, we found that AdOVA + CpG gave the best tumor protection in challenge studies. AdOVA + chitosan + CpG showed a decrease in protective levels and numbers of antigen-specific immune cells.

Further experiments focused on elucidating the mechanisms by which chitosan and CpG modulate the immune response. Using the therapeutic AdPSA model, chitosan was not found to enhance tumor protection or numbers of antigen-specific immune cells. Additional experiments found that this depression was not due to problems with viral infectivity or secretion due to chitosan complexation. A series of kinetics studies were performed which showed that peak levels of effector T cells were present 14 days later in AdPSA + CpG immunized mice than in AdPSA alone. This delayed effect may explain the increased levels of protection in AdPSA + CpG mice, and be useful in future vaccine design concerning the timing of peak response.

Keywords

adenovirus, chitosan, CpG, gene delivery, PEI, prostate cancer

Pages

xii, 137 pages

Bibliography

Includes bibliographical references (pages 132-137).

Copyright

Copyright 2010 Jessica Beth Graham

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