Antibody studies in mice of outer membrane antigens for use in an improved meningococcal B and C vaccine

Abstract Since 1988, N. meningitidis , B:4:P1.15, ET-5 complex, has been responsible for an epidemic of meningococcal disease in Greater São Paulo, Brazil. Despite current trials to develop an effective vaccine against group B meningococci, children less than 2 years old have not been protected. It has been suggested that iron-regulated proteins (IRPs) should be considered as potential antigens for meningococcal vaccines. The vaccines under study consisted of outer-membrane vesicles depleted of lipooligosaccharide from three serogroup B strains and one serogroup C strain, IRPs, meningococcal group C polysaccharide and aluminum hydroxide. Four different protein and C polysaccharide concentrations were studied. The ELISA and bactericidal results showed a higher antibody response when 2 injections of 2.0 μg doses were administered. Despite higher IgG reactivity against antigen preparations containing IRPs seen in ELISA, the bactericidal activity was not increased if the target strain was grown in iron-restricted medium. The influence of addition of alkaline-detoxified lipooligosaccharide (dLOS) on immunogenicity of the vaccine was also investigated, and the dLOS provided for a more functionally specific antibody response.     

A comparative study of the immunogenicity of 2 group-B meningococcal vaccines in monkeys

The comparative study of two group B meningococcal vaccines manufactured in the USSR and in Cuba was made. The vaccine manufactured in the USSR contained the noncovalent compound of group B Neisseria meningitidis polysaccharide and outer membrane protein, and the Cuban vaccine contained group B N. meningitidis outer membrane proteins and group C N. meningitidis polysaccharide. The data obtained in this study indicated that both vaccines possessed immunological potency evaluated according to their capacity to stimulate the formation of bactericidal antibodies, whose level was found to increase eightfold after the immunization of monkeys in two injections. Besides, group B meningococcal vaccines did not induce the suppression of nonspecific protective activity characteristics of the body and did not stimulate the formation of autoantibodies to brain and liver tissues, which was indicative of the safety of these vaccines.     

The immunological status indices of monkeys inoculated with a meningococcal B vaccine

The method for the determination of bacterial antibodies to group B meningococci was worked out. The method was used for the determination of antibodies to group B meningococcal vaccine produced in the USSR. The dynamic study of antibodies to protein, polysaccharide and lipopolysaccharide antigens of group B meningococci was made by the method of the enzyme immunoassay (EIA), and the safety of the vaccine was studied by the determination of autoantibodies active against brain tissue antigens. The data thus obtained were indicative of the immunological activity of group B protein-polysaccharide vaccines, manifested by the capacity for stimulating bactericidal antibodies whose level increased 8- to 10-fold after the immunization of monkeys in 2 and 3 injections. Similarity in the dynamics of the formation of bacteriolysins and antibodies to protein antigen, as determined in EIA, was noted. The vaccine was found to stimulate no cytotoxic anticerebral antibodies in the glia migration test, which was indicative of the safety of group B meningococcal vaccine.     

Comparison of meningococcal outer membrane protein vaccines solubilized with detergent or C polysaccharide

Outer membrane proteins (OMPs) were isolated from meningococcal strain H44/76 (B:15:P1.16) by detergent extraction of bacteria. A final product containing class 1 (P1.16), 3(15), 4 OMPs and 5% (w/w) lipooligosaccharide was obtained. Two experimental vaccines were prepared: OMP-detergent and OMP-C polysaccharide. The OMP-detergent vaccine tended to show a better bactericidal: ELISA ratio for the antibodies induced as compared to the OMP-C polysaccharide vaccine. The vaccine induced bactericidal antibodies appeared for the greater part to be directed against the class 1 OMP (P1.16). By comparison of cultures grown in Mueller Hinton Broth with and without 0,25% (w/v) glucose, it was found that monoclonal antibodies against the serotype OMP (class 2 or 3) were not bactericidal against meningococci grown in MHB without glucose. Antibodies against class 1 OMP and lipooligosaccharide were not influenced by this. A new major outer membrane protein (appr. 40kd) is described that may function as a cation-specific porin.     

Appearance of new strains associated with group B meningococcal disease and their use for rapid vaccine development

There has been a decrease in the prevalence of disease in the United States due to meningococcal serotypes 2a and 2b containing class 2 proteins with a concomitant increase in nonserotypable strains containing class 3 major outer membrane proteins. A new disease associated strain was identified using monoclonal antibodies as B:4:P1.15. Serotype 4 strains have been heretofore solated almost only from carriers. This B:4:P1.15 strain predominated among group B disease isolates in Cuba from the late 1970s to the present and among Miami, Florida isolates recovered in 1981 and 1982. To determine whether protein vaccines for new strains or serotypes could be prepared using our present methods, a combined vaccine was prepared from a group B strain (B:8:P1.15) recovered during a recent outbreak in Virginia, and a serotype 2b strain, plus group C polysaccharide. The vaccine was prepared with aluminum hydroxide, or with trehalose dimycolate plus monophosphoryl lipid A, or without adjuvant. Four weeks after immunization antibody levels were much higher in mice that received vaccine containing adjuvant.     

Bordetella pertussis fimbriae are effective carrier proteins in Neisseria meningitidis serogroup C conjugate vaccines

Serogroup C meningococcal conjugate vaccines generally use diphtheria or tetanus toxoids as the protein carriers. The use of alternative carrier proteins may allow multivalent conjugate vaccines to be formulated into a single injection and circumvent potential problems of immune suppression in primed individuals. Bordetella pertussis fimbriae were assessed as carrier proteins for Neisseria meningitidis serogroup C polysaccharide. Fimbriae were conjugated to the polysaccharide using modifications of published methods and characterised by size exclusion chromatography; co-elution of protein and polysaccharide moieties confirmed conjugation. The conjugates elicited boostable IgG responses to fimbriae and serogroup C polysaccharide in mice, and IgG:IgM ratios indicated that the responses were thymus-dependent. High bactericidal antibody titres against a serogroup C strain of N. meningitidis were also observed. In a mouse infection model, the conjugate vaccine protected against lethal infection with N. meningitidis. Therefore, B. pertussis fimbriae are effective carrier proteins for meningococcal serogroup C polysaccharide and could produce a vaccine to protect against meningococcal disease and to augment protection against pertussis.     

Evaluation of the immunological activity and safety of group B meningococcal vaccine prepared from a natural complex of specific polysaccharide and outer membrane proteins

Immunological activity and safety of group B meningococcal vaccine prepared from a natural complex of specific polysaccharide and outer membrane proteins were under study. The immunological safety of the vaccine was evaluated by the absence of antibodies to denaturated and native DNA (d-DNA and n-DNA). As shown with the use of the enzyme immunoassay (EIA), the administration of the vaccine did not induce antibody formation to d-DNA and n-DNA during the observation period. The titer of bactericidal antibodies in the immune bacteriolysis assay (IBA) to the vaccine strain B:2b:P1.2 after immunization increased four-fold and greater in 80% of the vaccinated persons. The significant increase of bactericidal antibodies to heterologous strains B:2a:P1.2 and B:15:P1.7 was registered in 20-30% of the vaccinees, respectively. A month after the repeated vaccination an increase in specific IgG antibodies to the complex antigen was found to occur according to EIA results. The use of RIB made it possible to evaluate the preventive activity of group B meningococcal vaccine as a whole and to suppose that the vaccine induced mainly type-specific response.     

Human bactericidal antibody response to meningococcal outer membrane protein vaccines

Several different meningococcal outer membrane protein vaccines have been prepared and used in human safety and immunogenicity studies. The results of these studies have led to some general conclusions regarding the human antibody response to these vaccines. A review of these conclusions, however, indicates that a number of important questions and problems still need to be addressed. Two of these are the determination of the protective level of bactericidal antibody in human serum and the impact of phase variation of surface antigens on vaccine strategy. Bactericidal assays using intrinsic complement and high concentrations of serum suggest that the level of natural immunity to group B meningococci is quite high, but is increased by vaccination with outer membrane protein vaccine. Phase variation in meningococcal surface antigens including capsule, class 1 protein, class 5 protein, and lipopolysaccharide was demonstrated using colony blotting with monoclonal antibodies. Phase variation resulted in differences in susceptibility to the bactericidal activity of human sera.     

B群脑膜炎球菌疫苗的研制策略

脑膜炎奈瑟菌主要引起儿童细菌性脑脊髓膜炎和败血症,有较高的发病率和病死率。现用疫苗能够控制A、C、W135和Y群脑膜炎球菌引起的感染,而由于B群荚膜多糖免疫原性弱,外膜蛋白变异性高等原因,仍无安全和具有广泛保护性的疫苗用于控制B群脑膜炎球菌的感染。目前,B群脑膜炎球菌大多已成为引起发达国家侵袭性脑膜炎疾病的主要病原体。随着研究的不断深入,B群脑膜炎球菌疫苗的研究已经取得了很大的进展,外膜囊(Out membrane vesicles,OMV)疫苗已经在控制特异性菌株爆发流行中取得了成功。然而,人们对具有广泛保护性的B群脑膜炎球菌疫苗的探索仍在继续。本文对近年来B群脑膜炎球菌基于不同型抗原疫苗的各种研制策略及其存在的问题进行了综述。     

The reactogenicity and antigenic activity of a meningococcal group B vaccine made from a natural complex of the specific polysaccharide and outer membrane proteins

Group B meningococcal vaccine consisting of the natural complex of specific polysaccharide and outer membrane protein (OMP) has been shown to be moderately reactogenic, safe with respect to the effect of undermining tolerance to human brain tissue antigens and to produce no allergization of humans. The vaccine under study possesses antigenic activity: (a) immunization with this vaccine ensures the fourfold rise of the level of antibodies to the group-specific polysaccharide of group B meningococcus in about 80% of persons with the initially low level of antibodies, this percentage being retained during the whole period of observation, i. e. 85 days; (b) the vaccine enhances the level of antibodies to meningococcal OMP, determined in the enzyme immunoassay and the passive hemagglutination test; (c) these data are indicative of the expediency of immunizing the risk groups of persons with the initially low level of antibodies.     

Molecular mimetics of polysaccharide epitopes as vaccine candidates for prevention of Neisseria meningitidis serogroup B disease

Neisseria meningitidis is a major cause of meningitis and sepsis. Despite nearly 25 years of work, there is no promising vaccine candidate for prevention of disease caused by meningococcal B strains. This review summarizes newer approaches for eliciting protective meningococcal B immune responses, including the use of molecular mimetics of group B polysaccharide and conserved membrane proteins as immunogens. The capsular polysaccharide of this organism is conserved and serum antibody to this capsule confers protection against disease. However, the immunogenicity of meningococcal B polysaccharide-based vaccines is poor. Further, a portion of the antibody elicited has autoantibody activity. Recently, our laboratory produced a panel of murine monoclonal antibodies (Mabs) that react specifically with capsular polysaccharide epitopes on meningococcal B that are distinct from host polysialic acid. These Mabs elicit complement-mediated bactericidal activity and confer passive protection in animal models. The anti-capsular Mabs were used to identify molecular mimetics from phage display peptide libraries. The resulting peptides were antigenic mimetics as defined by binding to the Mabs used to select them but, to date, are poor immunogenic mimetics in failing to elicit anti-capsular antibodies.     

Fusarium verticillioides (Saccardo) Nirenberg Associated with Hardlock of Cotton

The development of new immune potentiators for human vaccines is an important and expanding field of research. In the present study, the ability of the capsular polysaccharide from Neisseria meningitidis serogroup A (CPS-A), a mannose-containing carbohydrate, to enhance the antibody production against a co-administered model vaccine antigen, is examined. A protein-meningococcal serogroup C capsular polysaccharide (CPS-C) conjugate was selected as the model antigen for this study. After subcutaneous immunization of Balb/C mice, the conjugate mixed with CPS-A induced higher anti-CPS-C IgG and IgG_2a antibody levels and higher anti-meningococcal serogroup C bactericidal titers than the conjugate alone or mixed with CPS-C. The immuno-stimulatory properties exhibited by CPS-A and the fact that vaccines based on purified CPS-A has been safely used during decades to fight the serogroup A meningococcal disease, support the proposal to use CPS-A as immune potentiator for human vaccination studies.     

A群C群脑膜炎球菌结合疫苗稳定性

目的评价A群C群脑膜炎球菌结合疫苗原液和成品的稳定性。方法分别将A群、C群脑膜炎球菌结合疫苗原液及A群C群脑膜炎球菌结合疫苗各选取连续3批,分别放置于37℃、20~25℃和2~8℃3种温度下,在一定的时间取样进行主要项目测定,在关键时间点进行全面检测。结果 A群结合疫苗原液于2~8℃保存9个月,20~25℃保存4周,37℃保存4 d;C群结合疫苗原液于2~8℃保存9个月,20~25℃保存6个月,37℃保存4周;A群C群脑膜炎球菌结合疫苗于2~8℃保存2年3个月,20~25℃保存6个月,37℃可以保存9周;各项检测指标均符合质量标准的要求。结论在2~8℃条件下,A群、C群脑膜炎球菌结合疫苗原液存放6个月,A群C群脑膜炎球菌结合疫苗存放2年,其质量稳定。     

Induction of group 17 specific antibodies by pneumococcal type 17F and 17A polysaccharide vaccines.

Pneumococcal capsular polysaccharide group 17 contains two distinct serotypes, 17F and 17A. Pneumococcal group 17 is present in the licensed 23 valent polysaccharide vaccines. One such vaccine contains type 17A, while the other vaccine contains type 17F. The purpose of these studies was to determine the extent of cross-protection that could be expected, as both type 17F and 17A cause disease. The antibody responses of one group of adults to a vaccine containing type 17F was compared to that of another group that received a type 17A containing vaccine. By ELISA the 17A vaccine induced more cross-reactive antibodies. Opsonophagocytic antibodies are a good predictor of protection and both vaccines induced antibodies opsonic for both 17F and 17A. We conclude that either 17F or 17A will provide similar protection against group 17 disease.     

WHO/Health Canada meeting on regulatory considerations for evaluation and licensing of new meningococcal Group B vaccines,Ottawa, Canada, 3–4 October 2011

Serogroup B Neisseria meningitides (MenB) is a significant cause of endemic and epidemic outbreaks of the disease worldwide. Although polysaccharide and conjugate vaccines are available against other meningococcal serogroups, the poor immunogenicity of MenB polysaccharide has led to the development of protein-based vaccines. However, the diversity and antigenic variability of MenB strains has been a major challenge. Recently a new generation of MenB vaccines that contain conserved antigens has been developed to provide broader coverage and they are in an advanced stage of development and regulatory consideration. In October 2011, the World Health Organization and Health Canada jointly organized a consultation on regulatory considerations for the evaluation and licensing of new MenB vaccines. The aim was to seek consensus on key regulatory issues relevant to the evaluation of candidate MenB vaccines and on approaches to the standardisation of in vitro assays used in the evaluation process. Participants agreed that functional antibodies as measured in the Serum Bactericidal Activity (SBA) assay could be used to evaluate MenB vaccine efficacy and ways of improving assay standardization proposed. Approaches to bridging SBA data to large collections of strains in order to give an indication of the prospective breadth of vaccine coverage were discussed.     

Evaluation of the level of antibodies to purified meningococcal antigens

Group B meningococcal antigens, such as polysaccharide, lipopolysaccharide, protein preparation, as well as sonicates obtained from meningococcal cells, groups A, B and C, have been isolated. On the basis of these preparations the parameters of an enzyme immunoassay system for the detection of antibodies to individual meningococcal antigens have been established, and the specificity of the system and the possibility of using it for the evaluation of the level of antibodies to meningococci in human sera have been studied.     

Meningococcal polysaccharide vaccines: A review

Polysaccharides produced by Neisseria meningitidis are pharmaceutically important molecules, and are the active components of vaccines against N. meningitidis serogroups A, C, W135 and Y. Effective vaccines based on capsular polysaccharide, polysaccharide conjugates and outer membrane vesicles have been developed for strains expressing capsular polysaccharides that define the sero groups A, C, Y and W135. However, conventional approaches to develop a vaccine for group B strains have been largely unsuccessful. This review focuses on the various aspects of fermentative production of meningococcal polysaccharide from N. meningitidis, methods of conjugation for improving the immunogenicity of polysaccharide vaccine, and efficient and cost effective methods for the purification of N. meningitidis capsular polysaccharide and outer membrane vesicles. In addition, different analytical techniques for the quantitative determination of polysaccharide vaccine and evaluation of structural integrity of conjugate vaccine have been described.     

The Development of an Experimental Multiple Serogroups Vaccine for Neisseria meningitidis

A native outer membrane vesicles (NOMV) vaccine was developed from three antigenically diverse strains of Neisseria meningitidis that express the L1,8, L2, and L3,7 lipooligosaccharide (LOS) immunotypes, and whose synX, and lpxL1 genes were deleted.. Immunogenicity studies in mice showed that the vaccine induced bactericidal antibody against serogroups B, C, W, Y and X N. meningitidis strains. However, this experimental NOMV vaccine was not effective against serogroup A N. meningitidis strains. N. meningitidis capsular polysaccharide (PS) from serogroups A, C, W and Y were effective at inducing bactericidal antibody when conjugated to either tetanus toxoid or the fHbp1-fHbp2 fusion protein fHbp(1+2). The combination of the NOMV vaccine and the N. meningitidis serogroup A capsular polysaccharide (MAPS) protein conjugate was capable of inducing bactericidal antibodies against a limited number of N. meningitidis strains from serogroups A, B, C, W, Y and X tested in this study.     

Seroprotection against serogroup C meningococcal disease in adolescents in the United Kingdom: observational study

Objective To determine the persistence of bactericidal antibody titres following immunisation with serogroup C meningococcal glycoconjugate vaccine at age 6-15 years in order to examine changes in persistence of antibodies with age.Design Observational study.Setting Secondary and tertiary educational institutions in the United Kingdom.Participants Healthy adolescents aged 11-20 years previously immunised between 6 and 15 years of age with one of the three serogroup C meningococcal vaccines.Intervention Serum obtained by venepuncture.Main outcome measures Percentage of participants with (rabbit complement) serum bactericidal antibody titres of at least 1:8; geometric mean titres of serogroup C meningococcal serum bactericidal antibody.Results Five years after immunisation, 84.1% (95% confidence interval 81.6% to 86.3%) of 987 participants had a bactericidal antibody titre of at least 1:8. Geometric mean titres of bactericidal antibody were significantly lower in 11-13 year olds (147, 95% confidence interval 115 to 188) than in 14-16 year olds (300, 237 to 380) and 17-20 year olds (360, 252 to 515) (P<0.0001 for both comparisons). Within these age bands, no significant difference in geometric mean titres of bactericidal antibody between recipients of the different serogroup C meningococcal vaccines was seen. More than 70% of participants had received a vaccine from one manufacturer; in this cohort, geometric mean titres were higher in those immunised at aged 10 years or above than in those immunised before the age of 10.Conclusions Higher concentrations of bactericidal antibody are seen five years after immunisation with serogroup C meningococcal vaccine at age 10 years or above than in younger age groups, possibly owing to immunological maturation. This provides support for adolescent immunisation programmes to generate sustained protection against serogroup C meningococcal disease not only for the vaccine recipients but also, through the maintenance of herd immunity, for younger children.     

Role of Factor H Binding Protein in Neisseria meningitidis Virulence and Its Potential as a Vaccine Candidate To Broadly Protect against Meningococcal Disease

SUMMARY

Neisseria meningitidis is a Gram-negative microorganism that exists exclusively in humans and can cause devastating invasive disease. Although capsular polysaccharide-based vaccines against serogroups A, C, Y, and W135 are widely available, the pathway to a broadly protective vaccine against serogroup B has been more complex. The last 11 years has seen the discovery and development of the N. meningitidis serogroup B (MnB) outer membrane protein factor H binding protein (fHBP) as a vaccine component. Since the initial discovery of fHBP, a tremendous amount of work has accumulated on the diversity, structure, and regulation of this important protein. fHBP has proved to be a virulence factor for N. meningitidis and a target for functional bactericidal antibodies. fHBP is critical for survival of meningococci in the human host, as it is responsible for the primary interaction with human factor H (fH). Binding of hfH by the meningococcus serves to downregulate the host alternative complement pathway and helps the organism evade host innate immunity. Preclinical studies have shown that an fHBP-based vaccine can elicit serum bactericidal antibodies capable of killing MnB, and the vaccine has shown very encouraging results in human clinical trials. This report reviews our current knowledge of fHBP. In particular, we discuss the recent advances in our understanding of fHBP, its importance to N. meningitidis, and its potential role as a vaccine for preventing MnB disease.