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Humoral Responses to Cytomegalovirus Glycoprotein B Vaccine with MF59 Adjuvant

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Humoral Responses to Cytomegalovirus Glycoprotein B Vaccine with MF59 Adjuvant Humoral responses to Cytomegalovirus glycoprotein B vaccine with MF59 adjuvant A thesis submitted to the University College London for the degree of Ph.D. by Ilona Anna Baraniak February 2017 Centre for Virology at Royal Free Campus Division of Infection and Immunity University College London “The only true wisdom is in knowing you know nothing.” ― Socrates Declaration I, Ilona Anna Baraniak, confirm that the work presented in this thesis is my own. Where information has been derived from other sources, I confirm that this has been indicated in the thesis. Abstract Solid organ transplant (SOT) patients are at risk of end-organ diseases such as pneumonitis, hepatitis or enteritis caused by HCMV. HCMV infection can occur via primary infection of a seronegative recipient or upon reinfection or reactivation in a seropositive transplant recipient. Seronegative recipients have the greatest risk of viraemia and disease, showing that pre-existing natural immunity provides substantial protection. This, in turn, underpins vaccination as a viable strategy to control HCMV in the transplant setting. To test this, a clinical trial with a vaccine based on HCMV glycoprotein B (gB) antigen plus MF59 adjuvant was performed in SOT awaiting transplantation. The study showed that antibody titres against the gB antigen were significantly increased in both seropositive and seronegative recipients of the vaccine in comparison to the patients who received placebo, and importantly, higher titres correlated directly with reduced viraemia post-transplant. The aim of my thesis was to identify the component of the specific humoral response responsible for this effect. In comparison with placebo recipients, I could find no evidence for the protection being due to induction of antibodies that mediate neutralisation, antibody dependent cellular cytotoxicity, or prevent cell to cell spread of virus in culture. In contrast, analysis of antigenic domains of gB bound by the antibodies revealed that vaccination of seropositive individuals enhanced antibody responses against antigenic domain 2 and that these correlated with reduced viraemia post-transplant. Antibodies against three other antigenic domains were induced by the vaccine, but did not correlate with protection. These results suggest that antigenic domain 2 should be an important component of future HCMV vaccines to boost pre-existing immune responses that protect from HCMV infection. The protection afforded to seronegatives remains unidentified, but could be explained if another antigenic domain on gB remains to be discovered. Acknowledgements While I alone am responsible for this thesis, it is nonetheless at least as much a product of years of interaction with, and inspiration by, a large number of friends and colleagues as it is my own work. For this reason, I wish to express my warmest gratitude to all those persons whose comments, questions, criticism, support and encouragement, personal and academic, have left a mark on this work. I also wish to thank those institutions which have supported me during the work on this thesis. Regrettably, (but inevitably), the following list of names will be incomplete, and I hope that those who are missing will forgive me and accept my sincere appreciation of their help and encouragement. Foremost, I would like to express my sincere gratitude to my primary advisor Prof Paul Griffiths for the continuous support of my Ph.D. study and research, for his patience, motivation, enthusiasm, and immense knowledge. His guidance was a great help to me during my time of research and writing of this thesis. I would like to express my special appreciation and thanks to my secondary advisor Dr Matthew Reeves, you have been a tremendous mentor for me. I would like to thank you for encouraging my research and for allowing me to grow as a research scientist. Your advice on both research as well as on my career have been invaluable. I really appreciate your commitment and patience you had for such an unruly Ph.D. student as me. I could not have imagined having a better advisor for my Ph.D. study. I would like to thank my lab colleagues, especially to Matthew Murray and Dr Nick Peters for all the help with the lab work; for introducing me to your ‘odd’ British sense of humour; for giving me advice on my personal (turbulent at the times) life and lastly… for all the fun that we have had in the past years. 1 My sincere thanks also go to Lyn Ambrose. Thank you for introducing me to flow cytometry. Thank you for all the assistance you provided, your patience and sharing your knowledge with me. Many thanks to the entire Virology staff at Royal Free Hospital: especially Dr Claire Atkinson for many valuable advices and Emily Rothwell for providing necessary information about the clinical trial and patients; as well as clinical material. Thanks to all members of VacTrain. You have contributed immensely to my personal and professional development. The group has been a source of friendships as well as good advice and collaboration. I am especially grateful to our coordinator Prof Ben van der Zeist and project manager Wendy van Hemmen. It was a real pleasure to be involved in this fantastic European research consortium. Thanks to you I have learned so much though those years, not only about vaccinology, but also about collaborative work. Special thanks to Dr Benedict Halbroth for all the moral support and friendship. It was a great experience to be part of the VacTrain team! Thanks to Dr Leander Grode and the whole Vaccine Project Management team in Hannover for giving me a fantastic opportunity to do an internship at their company. Thank you for all the time and effort to introduce me to the concepts of project management in Vaccine industry; I greatly appreciate this unique possibility to have an insight into the work of a project manager. I would like to thank also Dr Gary McLean and Prof Florian Kern for sharing their expertise and giving me very insightful and valuable advices. I also gratefully acknowledge the funding sources that made my Ph.D. work possible. 2 On personal note: A special thanks to Aonghus D. Thank you for putting up with me during these challenging times, none of this would be possible without you. A big ‘dziękuję’ to my parents. I cannot thank you enough for giving me all the freedom I needed to become who I am now. Thank you for encouraging me to follow my dreams. Also, thank you for being such a good moral support and for all your attempts to cheer me up whenever I needed it. I would also like to thank to my beloved sister Klaudia and brother Łukasz and my best friends: Paula and Marlena for supporting me through all these years. I will always remember and greatly appreciate it. Lastly, I would like express my sincere gratitude to my boyfriend Fabian Lang. Thank you so much for all the support you gave me in those last, challenging months as a Ph.D. student. 3 Funding This work was funded by the European Union under the FP7 Marie Curie Action, Grant number 316655 (VacTrain). Part of the work that is included in chapter 5 was also co-funded by Deutsche Forschungsgemeinschaft MA 929/11-1. 4 Table of contents DECLARATION ..................................................................................................... 0 ABSTRACT ............................................................................................................ 0 ACKNOWLEDGEMENTS ...................................................................................... 1 FUNDING ............................................................................................................... 4 TABLE OF CONTENTS ......................................................................................... 0 LIST OF FIGURES ................................................................................................. 7 LIST OF TABLES ................................................................................................ 13 ABBREVIATIONS ................................................................................................ 14 1. INTRODUCTION. .......................................................................................... 22 1.1. The history of human cytomegalovirus discovery. .................................. 22 1.2. Classification of human cytomegalovirus. ................................................ 23 1.3. Structure of HCMV....................................................................................... 25 1.4. The genetic sequence of the HCMV genome. ........................................... 27 1.5. Genome organization. ................................................................................. 30 1.6. Genetic diversity and antigenic polymorphism in HCMV. ....................... 32 1.6.1. Potential impact of genetic diversity and antigenic polymorphism in HCMV on vaccine development. ....................................................................... 34 1.7. The virus entry process. ............................................................................. 34 0 1.8. HCMV lytic life cycle.................................................................................... 35 1.8.1. Organization of HCMV-expressed genes. .............................................. 36 1.8.2. Replication of the viral genome. ............................................................. 38 1.8.3. DNA packaging and virion egress. ......................................................... 39 1.9. Latency and reactivation. ..........................................................................
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