Chapter 1
Introduction
Carbohydrate-Based Vaccines: From Concept to Clinic Downloaded from pubs.acs.org by 5.8.47.86 on 08/31/18. For personal use only.
A. Krishna Prasad* Pfizer Vaccines Research and Development, 401 N. Middletown Rd., Pearl River, New York 10965, United States *E-mail:
[email protected].
A vast body of clinical experience has supported the introduction of several conjugate vaccines and related multi-antigen vaccines into routine use across the globe resulting in a dramatic impact on public health. Research and development efforts continue to translate this impact and find scientific and novel technological innovations to target other infectious diseases and address unmet medical needs. The primary motivation of this book is to provide a comprehensive forum for the presentation of various components that define the development and control strategies to produce safe and stable glycoconjugate vaccines that elicit consistent and robust immunogenic responses. The book aims to cover the complete landscape comprising immunological mechanisms; pre-clinical considerations; clinical history; design; development; characterization; conformational modeling; formulation; and chemistry, manufacturing and control (CMC) of glycoconjugate vaccines. These disciplines are closely tied to regulatory aspects leading to the commercial licensure of vaccines with a final chapter providing guidance on “lessons learned” to achieve this goal. All the chapters have been put together by leaders in industry, academia, government and non-profit organizations, with decades of experience in vaccine development and licensure.
© 2018 American Chemical Society
All approaches at a higher level are suspect until confirmed at the molecular level.” Francis Crick in “What Mad Pursuit” (1988)
“The ultimate aim of the modern movement in biology is, in fact, to explain all biology in terms of physics and chemistry.” Francis Crick in “Of Molecules and Men” (1966)
The past three decades have witnessed the development and regulatory approval of glycoconjugate vaccines against several medically important bacterial pathogens, including Haemophilus influenzae type b, Streptococcus pneumoniae, Neisseria meningitides and Salmonella typhi. Immunologic protection against these and many other bacterial diseases is mediated through opsonophagocytic antibodies directed against the surface carbohydrates that define the bacterial serogroup or serotype and serve as virulence factors. These vaccines are composed of bacterial capsular polysaccharides chemically conjugated to immunogenic carrier proteins. Given that the diseases caused by these bacterial pathogens are most pronounced in infants and young children, the development of the glycoconjugate vaccine technology has had a considerable impact on public health. Many of the chapters in this volume were assembled as a follow-up from a symposium entitled “Carbohydrate-based vaccines and adjuvants” which took place at the 254th American Chemical Society National Meeting held in Washington, DC (August 2017). The symposium was sponsored by Pfizer and the Carbohydrate (CARB) and Biotechnology (BIOT) divisions. This book, therefore, reflects the importance of this field toward design, development, manufacture and licensure of the complex carbohydrate-based (glycoconjugate) vaccines. The book has been organized into thirteen chapters, which cover a comprehensive landscape including the clinical history, design, development, chemistry, manufacturing and control (CMC) aspects, pre-clinical assays, adjuvants and the various approaches used to develop carbohydrate-based vaccines. Chapter 2 reviews the clinical experience of glycoconjugates from the ground-breaking monovalent Haemophilus influenzae type b vaccine to contemporary multi-valent (pneumococcal and meningococcal) and multi-component (glycoconjugate vaccines containing protein antigens) products and their positive public health impact. The chapter provides a chronological as well as comprehensive narrative of novel and ground breaking clinical trials, particularly those leading to marketing approvals for new categories of vaccines. Multi-valent pneumococcal polysaccharide conjugate vaccines have been demonstrated to be effective in preventing pneumococcal pneumonia, invasive pneumococcal disease and otitis media in children. A 7-valent conjugate vaccine, 2
including pneumococcal polysaccharides from seven serotypes individually conjugated to CRM197 carrier protein was licensed for use in infants and children < 5 years of age in 2000. The introduction of this first generation pneumococcal conjugate vaccine resulted in a substantial decrease in invasive pneumococcal disease (IPD) in children, less than 5 years of age, in the USA. A 10-valent pneumococcal conjugate (Synflorix®), using non-typable Haemophilus influenzae protein D, tetanus toxoid and diphtheria toxoid as carrier proteins, was later approved for immunization in infants and young children, outside the United States in 2009. This was followed by the regulatory approval of a 13-valent pneumococcal conjugate vaccine in 2009 in European Union and in early 2010, in the USA. In addition, as a result of a vaccine “herd immunity” effect, fewer pneumococcal infections have been observed in older adults more than 65 years of age. The vaccine has also demonstrated direct protective effects in adults against community-acquired pneumonia, resulting in recommended routine use of the vaccine in the United States, for both infants and older adults. A meningococcal glycoconjugate vaccine against meningococcal serogroup C bacterial meningitis was introduced first in the UK in 1999, followed subsequently in many countries around the world. The impact against serogroup C disease has been notable. Similarly, the introduction of a serogroup A meningococcal polysaccharide-tetanus toxoid conjugate vaccine (MenAfriVac®) in the “meningitis belt” of sub-Saharan Africa resulted in the near elimination of the group A epidemics among children and young adults in this part of the world. Several multi-valent glycoconjugate vaccines covering serogroups A, C, Y and W135 (Menactra®, Menveo® and Nimenrix®) have also been developed and approved to date. Salmonella Typhi is the major cause of enteric fever in lower income countries. Recently, WHO announced the prequalification of a typhoid conjugate vaccine (Typbar-TCV) manufactured by the Indian firm Bharat Biotech, initially licenced for use in India and Nepal. In a recent Phase 1 clinical study, a multicomponent 4-antigen Staphylococcus aureus vaccine, composed of capsular polysaccharide conjugates of serotypes 5 and 8 (CP5 and CP8), with two additional recombinant proteins clumping factor (rmClfA), and recombinant manganese transporter protein C (rMntC) were evaluated to confirm safety and immunogenicity of SA4Ag in a surgical population. This vaccine candidate is now in an efficacy study examining its potential to prevent ortheopedic nosocomial infections. Chapter 3, covers the immunological aspects related to glycoconjugate vaccines, with emphasis on T-cell immunity elicited by glycoconjugate vaccines. Purified “free” polysaccharides are T cell-independent immunogens and, as such, are poorly immunogenic in infants. By contrast, the carrier proteins used for conjugation are T-cell dependent immunogens and the covalent linkage of the polysaccharides to these proteins causes the anti-polysaccharide immune response to also become T cell-dependent. This renders the conjugated polysaccharides immunogenic for infants. Chapter 4 describes the strategies to produce a glyconjugate vaccine that can stimulate a potent and specific immune response to a saccharide antigen. This chapter discusses several key elements that need to be considered during 3
the design and development of conjugate vaccines. This includes preservation of critical immunogenic saccharide epitopes and other features such as selection of an effective and well-tolerated carrier protein as well as the stability of the conjugate in the drug substance as well as drug product. The key design features necessary to develop the “optimal conjugate vaccine construct” and the "toolbox" to achieve these ends are also covered in this chapter. Due to the distinguishing, sometimes subtle, structural differences resulting in variations in physicochemical attributes that differ by each saccharide antigen (in multi-valent vaccines), the conjugation to the carrier protein needs to be carried out by the specific customized process chemistry. Keeping in mind this variability in polysaccharide reactivity, the parameters that affect the final conjugate structural attributes also need to be identified based on the specific polysaccharide antigen structure at each stage of the process (fermentation, purification, activation/conjugation). The final optimal conjugate construct for each polysaccharide serotype is determined, therefore, by a number of factors to generate the optimal conjugate construct for robust immunogenicity and stability. Chapter 5 provides the commercial process development and manufacturing considerations of glyconjugate vaccines. It provides comprehensive coverage of the key elements required for process/product control strategy approaches during various clinical phases through to licensure including statistical process control and performance qualification necessary for a full process understanding. Central to this approach is the development of a control strategy to demonstrate consistency, robustness testing and transfer of commercially viable process technologies into manufacturing facilities, along with the production of clinical trial material. Multi-valent carbohydrate-based vaccines to prevent bacterial infection caused by several serogroups of Neisseria meningitidis have been licensed or are currently in clinical development. Chapter 6 discusses the various conjugation routes employed. Other key attributes, such as the chemical structures and functional groups involved in the covalent coupling of polysaccharides to carrier proteins for the preparation of tetravalent meningococcal serogroup A, C, W, Y glycoconjugates vaccines, have been described in depth. Coverage of a vaccine could potentially be expanded when specific antigens have been demonstrated to provide cross-protection against infection by structurally closely related, non-vaccine strains. However, structural similarity between carbohydrate antigens is not a reliable predictor of cross-protection. Conformational analysis can provide a mechanistic insight into clinical observations on cross-protection and may further indicate the importance of specific structural features, such as non-saccharide substituents. This may augment vaccine design and development strategies toward potential vaccine coverage expansion. Chapter 7 describes, in detail, the computational methodologies employed to model carbohydrate antigens and the valuable role that molecular simulations can play in furthering our understanding of carbohydrate immunogenicity and cross-protection. The authors outline molecular modeling case studies of polysaccharide antigens for meningococcal serogroups Y and W and pneumococcal serogroups 6, 19 and 23, as well O-antigens of Salmonella enterica and Shigella flexneri. 4
Synthetically derived saccharides have garnered significant interest due to their potential use as antigens to generate prophylactic (Neisseria meningitidis serogroup A, Shigella flexneri, etc.) vaccines, with positive results. Significant research efforts were aimed toward therapeutic (tumor associate carbohydrate antigens, TACA) glycoconjugate vaccines. Chapter 8 reviews the recent advances in oligosaccharide synthesis with desired stereo- and regio-selectivity, necessary to access high value targets. However, creating a synthesis platform that accommodates the large variation in oligosaccharide conformation and connectivity is a daunting task. Several challenges remain including poor immunogenicity, cost of goods (due to multi-step synthesis) and commercial viability that currently eludes their vaccine potential, especially for antigens with complex structures in their repeat units. For multi-antigen vaccines, this complexity is amplified. Nonetheless, over the last few decades researchers have made significant great progress toward addressing these shortcomings. Formulation is the key final step to required produce a safe, stable and efficacious vaccine drug product, often involving an adjuvant to boost immunogenicity of the antigen components. Among the currently licensed vaccines, the production of multi-valent glycoconjugate vaccines are the most complex. Besides the complexity in manufacturing, there are several other considerations such as facilities, shipping, storage, and shelf-life considerations that need to be addressed. Chapter 9 discusses strategies involved toward the development, production and transfer of robust processes to developing countries. Post conjugation with the carrier protein, the resulting glycoconjugate vaccine must retain the biological appearance of the pathogen’s saccharide antigen to specifically direct an immune response that can recognize and facilitate killing of the pathogen. A significant structural diversity exists in saccharide antigens which vary across serotypes and even among strains within a serotype. Therefore, there is not a single platform conjugation approach that can be applied to all polysaccharide conjugate vaccines, and the ability of candidate vaccines to elicit functional immune responses is often empirically determined. Chapter 10 describes several glycoconjugate features/epitopes that are critical for eliciting functional immune responses. The chapter identifies and discusses important considerations for pre-clinical glycoconjugate vaccine evaluation including in vivo models to measure immunogenicity and in vitro assays that measure immune responses that facilitate killing of the pathogen. Campylobacter jejuni is one of the most common causes of human diarrheal disease worldwide. In developed countries Campylobacteriosis cases are rare, but in developing countries, the incidence is estimated to be at least 10 times higher, which poses a life-threatening risk toward the pediatric population. Chapter 11 describes the development aspects of a multi-valent Campylobacter jejuni capsule polysaccharide conjugate candidate vaccine prepared for human clinical trials. Quality considerations for consistency in the production of glycoconjugate vaccines is a major area that needs to be addressed by all developers and manufacturers during clinical trials as well as commercial supply. Chapter 12 provides a structured guide to the manufacturing processes, regulatory requirements and analytical methods applied to current products and for future candidates in development stage. 5
Despite the availability of significant information in public domain such as literature and patents, these data provide only a partial glimpse of best practices applicable toward the design and manufacture new vaccines. Therefore, the risk of technical failure resulting in poorly immunogenic products remains high for new entrants to the field. Multinational corporate vaccine players with decades of experience, who went through various clinical phases to licensure, have typically made substantial investments in complex glycoconjugate vaccine development and commercialization. Finally, tying all the chapter pieces together, in Chapter 13 several experts in the field, with decades of experience in vaccine industry, academia, government and non-profit organizations, outline the key concepts that define successful design and manufacture of new glycoconjugate vaccines. The chapter provides general guidance from “lessons learned” with the intent of improving the CMC and chances of success for the glycoconjugate vaccines, still in development. Building on this extensive knowledge base from pneumococcal and meningococcal vaccines, glycoconjugates developers are now on the verge of extending this success to other encapsulated bacteria such as Salmonella typhi, Staphylococcus aureus, Shigella and Escherichia coli. Conjugate vaccine candidates, comprising Group B Streptococcus polysaccharide antigens, are currently undergoing clinical trials and hold significant promise to address morbidity and mortality in neonates. While the chapters were designed with the intent to cover all major aspects of glycoconjugate vaccine development, a book of this size cannot capture information on all supporting topics and allied fields in infinite detail. We hope that this book will serve as a useful reference providing valuable insights into the design, development, manufacture and licensure of these complex products.
Acknowledgments The editor thanks all the speakers at the ACS symposium and especially those who contributed chapters to the book. The authors fully recognize and acknowledge, with a deep sense of gratitude the importance, complexity and sensitive nature of the materials required to assemble the chapters to be shared with a vast audience. The authors and the editor gratefully acknowledge these efforts and the support of their respective managements which culminated in the successful completion of the book. The following organizations, whose support enabled this collaborative project to be realized, are gratefully acknowledged: Pfizer, GSK Vaccines, Sanofi Pasteur, Janssen Vaccines, PATH, University of Georgia (Athens, GA), University of Toledo (Ohio), University of Cape Town (South Africa) and University of Guelph (Canada). The editor thanks Drs. Kathrin Jansen, Annaliesa Anderson, Wendy Watson and Mark Ruppen for their overall support and valuable advice on the chapters contributed by Pfizer colleagues. The editor is grateful to Drs. Emilio Emini and Peter Paradiso for their support and to all the reviewers for their efforts to ensure the high scientific quality of the chapters in this book. 6
