University of Pennsylvania ScholarlyCommons Publicly Accessible Penn Dissertations 2017 Improving Vaccine Design For Viral Diseases Using Modified Antigens And Vectors Michael John Hogan University of Pennsylvania, [email protected] Follow this and additional works at: https://repository.upenn.edu/edissertations Part of the Virology Commons Recommended Citation Hogan, Michael John, "Improving Vaccine Design For Viral Diseases Using Modified Antigens And Vectors" (2017). Publicly Accessible Penn Dissertations. 2342. https://repository.upenn.edu/edissertations/2342 This paper is posted at ScholarlyCommons. https://repository.upenn.edu/edissertations/2342 For more information, please contact [email protected]. Improving Vaccine Design For Viral Diseases Using Modified Antigens And Vectors Abstract Two of the principal challenges facing vaccine design today are how to generate protective antibody responses against viruses that have evolved sophisticated strategies to evade the humoral immune system and how to more rapidly and effectively produce vaccines to address emerging epidemics. In this regard, we explored multiple strategies to improve vaccine design for HIV-1 and Zika virus. In one approach, we derived CD4-independent variants of HIV-1 envelope (Env) with the hypothesis that such Envs would expose conserved epitopes that may be targets of protective, non-neutralizing antibodies. We characterized the biological and structural properties of two CD4-independent Env clones and found that they exhibited significantly greater exposure of a relatively conserved, linear epitope in the second variable loop (V2) that had previously been associated with decreased risk of infection in a clinical HIV-1 vaccine trial. This epitope was significantly more immunogenic in mice and nonhuman primates and, intriguingly, was associated with more rapid development of antibody-dependent cell-mediated cytotoxicity. In another approach, we designed mutations in the cytoplasmic tail of HIV-1 Env that were predicted to increase its cell surface expression and thus its immunogenicity in a vaccinia prime-protein boost vaccine protocol. We found that the highest level of surface expression was mediated by Envs with truncated cytoplasmic tails, and this was associated with higher levels of binding and neutralizing antibodies after vaccinia primes and protein boosts, respectively. These two studies revealed that modifications ot HIV-1 Env immunogens are able to influence both the quality and magnitude of desirable antibody responses. Finally, we used a newly developed vaccine platform based on nucleoside-modified mRNA to design a vaccine against Zika virus. This vaccine, encoding the surface prM and E proteins, was potently immunogenic and elicited high and sustained titers of neutralizing antibodies in mice and nonhuman primates following a single intradermal immunization. We observed rapid and durable protection from Zika virus infection in mice and a high level of protection in monkeys challenged five weeks after vaccination. This vaccine thus represents a promising candidate for clinical use in controlling the spread of Zika virus. Degree Type Dissertation Degree Name Doctor of Philosophy (PhD) Graduate Group Cell & Molecular Biology First Advisor Drew Weissman Second Advisor James A. Hoxie Keywords Antibody, Envelope, HIV, mRNA, Vaccine, Zika Subject Categories Virology This dissertation is available at ScholarlyCommons: https://repository.upenn.edu/edissertations/2342 IMPROVING VACCINE DESIGN FOR VIRAL DISEASES USING MODIFIED ANTIGENS AND VECTORS Michael J. Hogan A DISSERTATION in Cell and Molecular Biology Presented to the Faculties of the University of Pennsylvania in Partial Fulfillment of the Requirements of the Degree of Doctor of Philosophy 2017 __________________________ __________________________ James A. Hoxie, M.D. Drew Weissman, M.D., Ph.D. Professor of Medicine Professor of Medicine Co-Supervisor of Dissertation Co-Supervisor of Dissertation ___________________________ Daniel S. Kessler, Ph.D. Associate Professor of Cell and Developmental Biology Graduate Group Chairperson Dissertation Committee: Paul F. Bates, Ph.D., Professor of Microbiology Beatrice H. Hahn, M.D., Professor of Medicine Scott E. Hensley, Ph.D., Associate Professor of Microbiology David B. Weiner, Ph.D., Emeritus Professor of Pathology and Laboratory Medicine DEDICATION This dissertation is dedicated to my mother, Mary Ellen Hogan, and my step- father, Robert H. Friel, Esq. Mom and Bob are the epitome of selfless people. Without their support over the years, both emotional and material, I would not have been able to realize the educational path I have chosen, nor would I now be guided by such a clear moral compass. With this dedication, I hope to express my sincere gratefulness for their sacrifice and their example. ii ACKNOWLEDGMENTS None of the projects in this dissertation would have been possible—truly— without the contributions of many wonderful collaborators, so I owe thanks to many people. First, I have benefitted from a very productive co-mentorship under Jim Hoxie and Drew Weissman. I did not necessarily know what I was getting into when I signed up for this, but what I received was an excellent and well-rounded scientific training from two outstanding but, stylistically, very different scientists. Jim has taught me how to be careful and methodical as an investigator and as a presenter. His questions have helped me think more clearly about controls and experimental plans before picking up the pipette. Drew has inspired me to be adventurous as a scientist, to take risks and strike while the iron is hot, but has also shown me how to keep priorities in perspective. What both Jim and Drew have in common is that they have each given me complete flexibility and independence over my projects and have supported me in whatever I chose to do, and I am grateful for that. I am extremely thankful that I have had the opportunity to work with Norbert Pardi as a close collaborator, particularly on the Zika work. Norbi and I match each other well in our zeal for our work, and have made a great scientific team. I am confident that I could not have taken up the project of producing a vaccine for Zika virus with anyone less dependable and dedicated to his work. Outside the lab, I am also grateful for Norbi’s sharing his love of wine. I also worked extremely closely with Angela Conde-Motter for much of the earlier half of my graduate career, and many of the mouse experiments in this dissertation are as much her work as they are mine. The CD4 independence and cytoplasmic tail work would not have been started without her critical contributions. iii Our labs have been fortunate to find ourselves in the midst of excellent col- laborators from other institutions. I am very grateful to Barton Haynes for recognizing early on the potential of the mRNA vaccine platform and providing us access to monkeys for evaluation of the Zika vaccine. Since then, he continues to generously contribute critical reagents and data. I also benefitted from our labs’ membership in a Gates Foundation vaccine discovery consortium. As part of this group, we worked closely with the labs of Shiu-Lok Hu, Kelly Lee, and Shan Lu to advance our vaccines in mice and monkeys. This would not have been possible without the invaluable contributions of Lifei Yang, Brad Cleveland, Wenjin Guo, Tad Davenport, Yu Liang, Shixia Wang, Dong Han, Patricia Firpo, and Deb Diamond, as well as the core facilities of David Montefiori, Georgia Tomaras, Guido Ferrari, and others. All the other members of the Hoxie and Weissman labs have contributed significantly to my success in graduate school. I would like to especially thank Josephine Romano, Beth Haggarty, and Andrea Jordan for taking such good care of me, and Adrienne Swanstrom, Samra Elser, Kitu Kumar, Son Nguyen, and George Leslie for their support, both scientific and otherwise. I cannot imagine entering into a more loving lab family than these wonderful people. I would like to thank my previous mentor, Carthene Bazemore-Walker, for starting me on my research career, my brilliant classmates, Bob Doms for continued guidance, and my support through the NIH T32 training grant in HIV Pathogenesis. Special thanks to my loving grandparents, who taught me enduring life lessons, my father Michael and devoted stepmom Kim, my kind and talented siblings including my older sister and excellent role model, Dr. Katherine Hogan, my Aunt Barbara and many other dear family and friends who have been so generous to my immediate family in recent years, and my partner Joe for his love and support. iv ABSTRACT IMPROVING VACCINE DESIGN FOR VIRAL DISEASES USING MODIFIED ANTIGENS AND VECTORS Michael J. Hogan Drew Weissman, M.D., Ph.D. and James A. Hoxie, M.D. Two of the principal challenges facing vaccine design today are how to generate protective antibody responses against viruses that have evolved sophisticated strategies to evade the humoral immune system and how to more rapidly and effectively produce vaccines to address emerging epidemics. In this regard, we explored multiple strategies to improve vaccine design for HIV-1 and Zika virus. In one approach, we derived CD4-independent variants of HIV-1 envelope (Env) with the hypothesis that such Envs would expose conserved epitopes that may be targets of protective, non-neutralizing antibodies. We characterized the biological and structural properties of two CD4-independent Env clones and found that they exhibited significantly