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Investigating the potential of conserved inner core oligosaccharide regions of Moraxella catarrhalis lipopolysaccharide as vaccine antigens: accessibility and functional activity of monoclonal antibodies and glycoconjugate derived sera

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Abstract

We investigated the conservation and antibody accessibility of inner core epitopes of Moraxella catarrhalis lipopolysaccharide (LPS) in order to assess their potential as vaccine candidates. Two LPS mutants, a single mutant designated lgt2 and a double mutant termed lgt2/lgt4, elaborating truncated inner core structures were generated in order to preclude expression of host-like outer core structures and to create an inner core structure that was shared by all three serotypes A, B and C of M. catarrhalis. Murine monoclonal antibodies (mAbs), designated MC2-1 and MC2-10 were obtained by immunising mice with the lgt2 mutant of M. catarrhalis serotype A strain. We showed that mAb MC2-1 can bind to the core LPS of wild-type (wt) serotype A, B and C organisms and concluded that mAb MC2-1 defines an immunogenic inner core epitope of M. catarrhalis LPS. We were unsuccessful in obtaining mAbs to the lgt2/lgt4 mutant. MAb MC2-10 only recognised the lgt2 mutant and the wt serotype A strain, and exhibited a strong requirement for the terminal N-acetyl-glucosamine residue of the lgt2 mutant core oligosaccharide, suggesting that this residue was immunodominant. Subsequently, we showed that both mAbs MC2-1 and MC2-10 could facilitate bactericidal killing of the lgt2 mutant, however neither mAb could facilitate bactericidal killing of the wt serotype A strain. We then confirmed and extended the candidacy of the inner core LPS by demonstrating that it is possible to elicit functional antibodies against M. catarrhalis wt strains following immunisation of rabbits with glycoconjugates elaborating the conserved inner core LPS antigen. The present study describes three conjugation strategies that either uses amidases produced by Dictyostelium discoideum, targeting the amino functionality created by the amidase activity as the attachment point on the LPS molecule, or a strong base treatment to remove all fatty acids from the LPS, thus creating amino functionalities in the lipid A region to conjugate via maleimide-thiol linker strategies targeting the carboxyl residues of the carrier protein and the free amino functionalities of the derived lipid A region of the carbohydrate resulted in a high loading of carbohydrates per carrier protein from these carbohydrate preparations. Immunisation derived antisera from rabbits recognised fully extended M. catarrhalis LPS and whole cells. Moreover, bactericidal activity was demonstrated to both the immunising carbohydrate antigen and importantly to wt cells, thus further supporting the consideration of inner core LPS as a potential vaccine antigen to combat disease caused by M. catarrhalis.

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Acknowledgements

We thank Perry Fleming (core Bacterial Culture Facility) for large scale biomass production, Jacek Stupak for recording CE-ES-MS, Dr. Susan Logan and Annie Aubry for valuable discussions and assistance with developing the bactericidal assay and the NRC-IBS animal facility for animal care. We also thank Dr. Johanna Schwingel and Prof. Tony Campagnari (State University of New York at Buffalo) for providing us with M. catarrhalis mutant strains lgt1 and lgt3. Heather Horan was funded by an Irish Scholarship from the Ireland Canada University Foundation. We are grateful to Novartis Vaccines for providing us with CRM197.

Dedication

The authors would like to dedicate this manuscript to honour the scientific career of Dr. Malcolm Perry on the occasion of his retirement after more than 50 years of contributions to the field of glycobiology. We are all very grateful for having had the opportunity to have worked with him for some of these years.

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  1. Institute for Biological Sciences, National Research Council, 100, Sussex Drive, Ottawa, ON, K1A 0R6, Canada

    Andrew D. Cox, Frank St. Michael, Chantelle M. Cairns, Suzanne Lacelle, Amy Lea Filion, Dhamodharan Neelamegan, Cory Q. Wenzel, Heather Horan & James C. Richards

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  1. Andrew D. Cox
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Correspondence to Andrew D. Cox.

Additional information

Since acceptance of this manuscript, the authors have become aware of a paper from the Gu laboratory [38], that details a somewhat related approach to that described here, wherein O‐deacylated LPS from two different mutants of a serotype A strain were utilised to prepare conjugates with tetanus toxoid as the carrier protein and utilising adipic acid dihydrazide as the linker. Sera derived from immunisation of rabbits with a lgt5 mutant LPS derived conjugate were able to kill all three serotypes albeit at titers lower than we observed in this study. This paper therefore also supports the strategy of truncated LPS‐based vaccines to combat disease caused by Moraxella catarrhalis.

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Cox, A.D., St. Michael, F., Cairns, C.M. et al. Investigating the potential of conserved inner core oligosaccharide regions of Moraxella catarrhalis lipopolysaccharide as vaccine antigens: accessibility and functional activity of monoclonal antibodies and glycoconjugate derived sera. Glycoconj J 28, 165–182 (2011). https://doi.org/10.1007/s10719-011-9332-7

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