Investigation of the protective mechanisms mediated by Neisserial Heparin Binding Antigen (NHBA) induced antibodies
Creators: name: PRINCIPATO, SILVIA
name: BINI, LUCA
name: Brunelli, Brunella
Publisher: name: Universit� degli Studi di Siena
Year: 2021

description: Abstract
Neisserial Heparin Binding Antigen (NHBA) is a surface-exposed lipoprotein ubiquitously expressed by Neisseria meningitidis strains and is one of the three main protein components of the Bexsero vaccine. NHBA binds heparin and heparan sulfates through an arginine-rich region and is cleaved by meningococcal and human proteases. Its expression is upregulated at 32�C [1] and it is sparsely distributed on the bacterial surface. Additionally, recent evidence suggests that NHBA plays a key role in bacterial adherence through its arginine-rich region [2].
NHBA induces bactericidal antibodies in humans and confers protective immunity in the infant rat animal model. Anti-NHBA antibodies (polyclonal and monoclonal) from mice and humans are functional, being able to induce complement-mediated bacterial killing, in the presence of rabbit complement (rSBA). However bactericidal activity is not measurable when human serum is used as the source of complement (hSBA).
The aim of this investigation was to elucidate the functional properties that determine the mechanisms of protection mediated by anti-NHBA antibodies.
Negative regulators of the complement system, such as factor H and vitronectin, have been investigated. The effects of antigen density have been explored using a genetically engineered NHBA overexpressing strain to characterize a panel of anti-NHBA monoclonal antibodies isolated from Bexsero immunized adults [3]. Monoclonal antibodies with enhanced C1q recruitment ability have been used to investigate the effect of antigen density and distribution. Moreover, the infant rat challenge model has been used to evaluate in vivo anti-NHBA antibodies induced passive protection.
Nonspecific downregulation of complement-mediated killing due to human factor H was observed to cause an underestimation of the anti-NHBA antibodies ability to efficiently kill bacteria in hSBA, despite the strong passive protection observed in the in vivo infection model. An interaction of NHBA with the complement down-regulator vitronectin was also demonstrated. By using a genetically engineered NHBA overexpressing strain, the relevance of antigen density on the bactericidal activity of antibodies was highlighted. A reduced antigen density on the bacterial surface was shown to be overcome by using engineered monoclonal antibodies with enhanced C1q recruitment ability.
It was demonstrated that multiple interactions with complement regulators interfere with the in vitro measurement of the bactericidal activity mediated by anti-NHBA antibodies in the presence of human complement. Interestingly NHBA, as the Neisseria Opc and NhhA, was found to interact with the extracellular matrix component vitronectin, opening the way to future studies to elucidate implications of this interaction for bacterial colonization. Taken together our findings further support the important role played by NHBA in meningococcal pathogenesis and immunity. 
References
[1] M. Lappann et al., Impact of moderate temperature changes on Neisseria meningitidis adhesive phenotypes and proteome. Infection and immunity, 18, 3484-3495, 2016. 
[2] I. Vacca et al., Neisserial heparin binding antigen (NHBA) contributes to the adhesion of Neisseria meningitidis to human epithelial cells. PloS One, 11, e0162878, 2016. 
[3] M. Giuliani et al., Human protective response induced by meningococcus B vaccine is mediated by the synergy of multiple bactericidal epitopes. Scientific Reports, 8, 3700, 2018.

Disclosures
This work was sponsored by GlaxoSmithKline Biologicals SA.
Silvia Principato is a student of the University of Siena (Life Science Department) and participates in a PhD program at GSK. At present, SP is an employee of the GSK group of companies.
Bexsero is a trademark of the GSK group of companies.
Human monoclonal Antibodies were obtained from adults in a Phase I clinical study conducted in Krakow, Poland and sponsored by Novartis Vaccine, now part of the GSK group of Companies, using two doses of multicomponent serogroup B meningococcal vaccines. The Clinical trial protocol was approved by the Bioethics Committee of the District Medical Doctors? Chamber in Krakow and the study was conducted in accordance with the Declaration of Helsinki. Written informed consent was obtained from each of the subjects.
All animal studies were ethically reviewed and carried out in accordance with European Directive 2010/63/EEC and the GSK policy on the Care, Welfare and Treatment of Animals.