Peptide mimics of the group B meningococcal capsule induce bactericidal and protective antibodies after immunization

Neisseria meningitidis serogroup B (MenB) is a leading cause of sepsis and meningitis in children. No vaccine is available for the prevention of these infections because the group B capsular polysaccharide (CP) (MenB CP) is unable to stimulate an immune response, due to its similarity with human polysialic acid. Because the MenB CP bears both human cross-reactive and non-cross-reactive determinants, we developed immunogenic peptide mimics of the latter epitopes. Peptides were selected from phage display libraries for their ability to bind to a protective anti-MenB CP mAb. One of these peptides (designated 9M) induced marked elevations in serum bactericidal activity, but not polysialic acid cross-reacting Abs, after gene priming followed by carrier-conjugate boosting. Moreover, the occurrence of bacteremia was prevented in infant rats by administration of immune sera before MenB challenge. 9M is a promising lead candidate for the development of an effective and affordable anti-MenB vaccine.     

Novel blocking human IgG directed against the pentapeptide repeat motifs of Neisseria meningitidis Lip/H.8 and Laz lipoproteins

Ab-initiated, complement-dependent killing contributes to host defenses against invasive meningococcal disease. Sera from nonimmunized individuals vary widely in their bactericidal activity against group B meningococci. We show that IgG isolated from select individuals can block killing of group B meningococci by human sera that are otherwise bactericidal. This IgG also reduced the bactericidal efficacy of Abs directed against the group B meningococcal protein vaccine candidates factor H-binding protein currently undergoing clinical trials and Neisserial surface protein A. Immunoblots revealed that the blocking IgG was directed against a meningococcal Ag called H.8. Killing of meningococci in reactions containing bactericidal mAbs and human blocking Abs was restored when binding of blocking Ab to meningococci was inhibited using either synthetic peptides corresponding to H.8 or a nonblocking mAb against H.8. Furthermore, genetic deletion of H.8 from target organisms abrogated blocking. The Fc region of the blocking IgG was required for blocking because F(ab')(2) fragments were ineffective. Blocking required IgG glycosylation because deglycosylation with peptide:N-glycanase eliminated blocking. C4b deposition mediated by an anti-factor H-binding protein mAb was reduced by intact blocking IgG, but not by peptide:N-glycanase-treated blocking IgG, suggesting that blocking resulted from inhibition of classical pathway of complement. In conclusion, we have identified H.8 as a meningococcal target for novel blocking Abs in human serum. Such blocking Abs may reduce the efficacy of select antigroup B meningococcal protein vaccines. We also propose that outer membrane vesicle-containing meningococcal vaccines may be more efficacious if purged of subversive immunogens such as H.8.     

A novel mimetic antigen eliciting protective antibody to Neisseria meningitidis.

Molecular mimetic Ags are of considerable interest as vaccine candidates. Yet there are few examples of mimetic Ags that elicit protective Ab against a pathogen, and the functional activity of anti-mimetic Abs has not been studied in detail. As part of the Neisseria meningitidis serogroup B genome sequencing project, a large number of novel proteins were identified. Herein, we provide evidence that genome-derived Ag 33 (GNA33), a lipoprotein with homology to Escherichia coli murein transglycosylase, elicits protective Ab to meningococci as a result of mimicking an epitope on loop 4 of porin A (PorA) in strains with serosubtype P1.2. Epitope mapping of a bactericidal anti-GNA33 mAb using overlapping peptides shows that the mAb recognizes peptides from GNA33 and PorA that share a QTP sequence that is necessary but not sufficient for binding. By flow cytometry, mouse antisera prepared against rGNA33 and the anti-GNA33 mAb bind as well as an anti-PorA P1.2 mAb to the surface of eight of nine N. meningitidis serogroup B strains tested with the P1.2 serosubtype. Anti-GNA33 Abs also are bactericidal for most P1.2 strains and, for susceptible strains, the activity of an anti-GNA33 mAb is similar to that of an anticapsular mAb but less active than an anti-P1.2 mAb. Anti-GNA Abs also confer passive protection against bacteremia in infant rats challenged with P1.2 strains. Thus, GNA33 represents one of the most effective immunogenic mimetics yet described. These results demonstrate that molecular mimetics have potential as meningococcal vaccine candidates.     

Induction of anti-idiotypic humoral and cellular immune responses by a murine monoclonal antibody recognizing the ovarian carcinoma antigen CA125 encapsulated in biodegradable microspheres

The use of biodegradable poly(dl-lactic-co-glycolic acid) microspheres as a cancer vaccine delivery system for induction of anti-idiotypic responses was investigated using a murine monoclonal antibody B43.13 that recognizes the human ovarian cancer antigen CA125. Immunization of mice with mAb B43.13 encapsulated in poly(dl-lactic-co-glycolic acid) microspheres resulted in enhanced humoral and cellular immune responses compared with mAb B43.13 alone or mAb B43.13 mixed with microspheres. The antibody responses could be further enhanced by the co-encapsulation of mAb B43.13 with monophosphoryl lipid A, a non-toxic adjuvant, in microspheres. Anti-idiotypic humoral responses were shown to result in Ab2 antibodies mimicking the nominal antigen CA125 and Ab3 antibodies recognizing CA125. Further, microsphere delivery of mAb B43.13 also resulted in induction of T cell responses involving T2 cells reactive with mAb B43.13 epitopes and T3 cells recognizing CA125. These results indicate that microsphere delivery of Ab1 can induce both humoral and cellular anti-idiotypic responses relevant to cancer antigens. This raises the possibility of the use of such formulations for anti-idiotypic induction immunotherapy for cancer. Received: 27 August 1997 / Accepted: 24 April 1998     

Molecular characterization of the anti-idiotypic immune response of a relapse-free neuroblastoma patient following antibody therapy: a possible vaccine against tumors of neuroectodermal origin?

Neuroblastoma treatment with chimeric antidisialoganglioside GD2 Ab ch14.18 showed objective antitumor responses. Production of anti-idiotypic Abs (Ab2) against ch14.18 (Ab1) in some cases was positively correlated with a more favorable prognosis. According to Jerne's network theory, a subset of anti-idiotypic Abs (Ab2beta) carries an "internal image" of the Ag and induces Abs (Ab3) against the original Ag. The molecular origin of an anti-idiotypic Ab response in tumor patients was not investigated previously. To clone anti-idiotypic Abs, B cells of a ch14.18-treated neuroblastoma patient with Ab2 serum reactivity were used to construct Ab phage display libraries. After repeated biopannings on ch14.18 and its murine relative, anti-GD2 mAb 14G2a, we selected 40 highly specific clones. Sequence analysis revealed at least 10 of 40 clones with different Ig genes. Identities to putative germline genes ranged between 94.90 and 100% for V(H) and between 93.90 and 99.60% for V(L). An overall high rate of replacement mutations suggested a strong Ag-driven maturation of the anti-idiotypic Abs. Two clones that were analyzed further, GK2 and GK8, inhibited binding of ch14.18 to GD2 just as the patient's serum did. GK8 alone inhibited >80% of the patient's anti-idiotypic serum Abs in binding to ch14.18. Rabbits vaccinated with GK8 or GK2 (weaker) produced Ab3 against the original target Ag GD2. GK8 may be useful as a tumor vaccine for GD2-positive [corrected] tumors.     

T cell recognition of Neisseria meningitidis class 1 outer membrane proteins. Identification of T cell epitopes with selected synthetic peptides and determination of HLA restriction elements.

No vaccine is yet available against serogroup B meningococci, which are a common cause of bacterial meningitis. Some outer membrane proteins (OMP), LPS, and capsular polysaccharides have been identified as protective Ag. The amino acid sequence of the protective B cell epitopes present within the class 1 OMP has been described recently. Synthetic peptides containing OMP B cell epitopes as well as capsular polysaccharides or LPS protective B cell epitopes have to be presented to the immune system in association with T cell epitopes to achieve an optimal Ir. The use of homologous, i.e., meningococcal, T cell epitopes has many advantages. We therefore investigated recognition sites for human T cells within the meningococcal class 1 OMP. We have synthesized 16 class 1 OMP-derived peptides encompassing predicted T cell epitopes. Peptides corresponding to both surface loops and trans-membrane regions (some of which occur as amphipathic beta-sheets) of the class 1 OMP were found to be recognized by T cells. In addition, 10 of 11 peptides containing predicted amphipathic alpha-helices and four of five peptides containing T cell epitope motifs according to Rothbard and Taylor (Rothbard, J. B., and W. R. Taylor. 1988. EMBO J 7:93) were recognized by lymphocytes from one or more volunteers. Some of the T and B cell epitopes were shown to map to identical regions of the protein. At least six of the peptides that were found to contain T cell epitopes show homology to constant regions of the meningococcal class 3 OMP and the gonococcal porins PIA and PIB. Peptide-specific T cell lines and T cell clones were established to investigate peptide recognition in more detail. The use of a panel of HLA-typed APC revealed clear HLA-DR restriction patterns. It seems possible now to develop a (semi-) synthetic meningococcal vaccine with a limited number of constant T cell epitopes that cover all HLA-DR locus products.     

High affinity mimotope of the polysaccharide capsule of Cryptococcus neoformans identified from an evolutionary phage peptide library

Cryptococcus neoformans causes a life-threatening meningoencephalitis in a significant percentage of AIDS patients. Mice immunized with a glycoconjugate vaccine composed of the glucuronoxylomannan (GXM) component of the cryptococcal capsular polysaccharide conjugated to tetanus toxoid (TT) produce Abs that, based on the epitope recognized, can be either protective or nonprotective. Since nonprotective Abs block the efficacy of protective Abs, we are interested in developing a vaccine that would focus the immune response specifically to protective epitopes. Previously, we screened a phage display library with 2H1, a protective anti-GXM mAb, and isolated PA1, a representative peptide that had a K(d) of 295 nM for 2H1. Mice immunized with PA1 conjugated to keyhole limpet hemocyanin developed high anti-peptide (1/13,000), but low anti-GXM (maximum, 1/200) titers. We now report our efforts to improve this vaccine by screening a sublibrary with six random amino acids added to either end of the PA1 motif to identify higher affinity peptides. P206.1, a peptide isolated from this sublibrary, had 80-fold higher affinity for 2H1 (K(d) = 3.7 nM) than PA1. P206.1 bound protective, but not nonprotective, anti-GXM Abs. Mice immunized with P206.1 conjugated to various carriers did not mount an Ab response to GXM despite developing high anti-peptide titers. However, mice primed with GXM-TT and boosted with P206.1-TT developed significant anti-GXM titers (maximum, 1/180,000). This latter immunization scheme focused the immune response on protective epitopes, since only 2-5% of these titers were directed against nonprotective de-O-acetylated GXM epitopes compared with 20-60% in animals primed and boosted with GXM-TT.     

A meningococcal factor H binding protein mutant that eliminates factor H binding enhances protective antibody responses to vaccination

Certain pathogens recruit host complement inhibitors such as factor H (fH) to evade the immune system. Microbial complement inhibitor-binding molecules can be promising vaccine targets by eliciting Abs that neutralize this microbial defense mechanism. One such Ag, meningococcal factor H-binding protein (fHbp), was used in clinical trials before the protein was discovered to bind fH. The potential effect of fH binding on vaccine immunogenicity had not been assessed in experimental animals because fHbp binds human fH specifically. In this study, we developed a human fH transgenic mouse model. Transgenic mice immunized with fHbp vaccine had 4- to 8-fold lower serum bactericidal Ab responses than those of control mice whose native fH did not bind the vaccine. In contrast, Ab responses were unimpaired in transgenic mice immunized with a control meningococcal group C polysaccharide-protein conjugate vaccine. In transgenic mice, immunization with an fH nonbinding mutant of fHbp elicited Abs with higher bactericidal activity than that of fHbp vaccination itself. Abs elicited by the mutant fHbp more effectively blocked fH binding to wild-type fHbp than Abs elicited by fHbp that bound fH. Thus, a mutant fHbp vaccine that does not bind fH but that retains immunogenicity is predicted to be superior in humans to an fHbp vaccine that binds human fH. In the case of mutant fHbp vaccination, the resultant Ab responses may be directed more at epitopes in or near the fH binding site, which result in greater complement-mediated serum bactericidal activity; these epitopes may be obscured when human fH is bound to the wild-type fHbp vaccine.     

Protective activity of monoclonal antibodies to genome-derived neisserial antigen 1870, a Neisseria meningitidis candidate vaccine

Genome-derived neisserial Ag (GNA) 1870 is a meningococcal vaccine candidate that can be subdivided into three variants based on amino acid sequence variability. Variant group 1 accounts for approximately 60% of disease-producing group B isolates. The Ag went unrecognized until its discovery by genome mining because it is expressed in low copy number by most strains. To investigate the relationship between Ab binding to GNA1870 and complement-mediated protective functions, we prepared a panel of four murine IgG mAbs against rGNA1870 (variant 1) and evaluated their activity against nine genetically diverse encapsulated Neisseria meningitidis strains expressing subvariants of variant 1 GNA1870. Based on flow cytometry with live encapsulated bacteria, surface accessibility of the epitopes recognized by the mAbs appeared to be low in most strains. Yet mAb concentrations <1 to 5 micro g/ml were sufficient to elicit bactericidal activity with human complement and/or activate C3b deposition on the bacterial surface. Certain combinations of mAbs were highly bactericidal against strains that were resistant to bactericidal activity of the respective individual mAbs. The mAbs conferred passive protection against bacteremia in infant rats challenged by strains resistant to bacteriolysis, and the protective activity paralleled the ability of the mAb to activate C3b deposition. Thus, despite low GNA1870 surface exposure, anti-GNA1870 variant 1 Abs are bactericidal and/or elicit C3b deposition and confer protection against bacteremia caused by encapsulated N. meningitidis strains expressing GNA1870 subvariant 1 proteins. The data support GNA1870 as a promising vaccine candidate for prevention of meningococcal group B disease caused by GNA1870 variant 1 strains.     

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.     

Selection of HIV-specific immunogenic epitopes by screening random peptide libraries with HIV-1-positive sera

Efforts to develop a protective HIV-1 vaccine have been hindered by difficulties in identifying epitopes capable of inducing broad neutralizing Ab responses. In fact, the high mutation rate occurring in HIV-1 envelope proteins and the complex structure of gp120 as an oligomer associated with gp41 result in a high degree of antigenic polymorphism. To overcome these obstacles, we screened random peptide libraries using sera from HIV-infected subjects to identify antigenic and immunogenic mimics of HIV-1 epitopes. After extensive counterscreening with HIV-negative sera, we isolated peptides specifically recognized by Abs from HIV-1-infected individuals. These peptides behaved as antigenic mimics of linear or conformational HIV-1 epitopes generated in vivo in infected subjects. Consistent with these findings, sera of simian HIV-infected monkeys also recognized the HIV-specific epitopes. The selected peptides were immunogenic in mice, where they elicited HIV-specific Abs that effectively neutralized HIV-1 isolates. These results demonstrate that pools of HIV-1 mimotopes can be selected from combinatorial peptide libraries by taking advantage of the HIV-specific Ab repertoire induced by the natural infection.     

Antibodies to keyhole limpet hemocyanin cross-react with an epitope on the polysaccharide capsule of Cryptococcus neoformans and other carbohydrates: implications for vaccine development

Cryptococcus neoformans causes a life-threatening meningoencephalitis in AIDS patients. Mice immunized with a glycoconjugate vaccine composed of the glucuronoxylomannan (GXM) component of the cryptococcal capsular polysaccharide conjugated to tetanus toxoid produce Abs that can be either protective or nonprotective. Because nonprotective Abs block the efficacy of protective Abs, an effective vaccine must focus the Ab response on a protective epitope. Mice immunized with peptide mimetics of GXM conjugated to keyhole limpet hemocyanin (KLH) with glutaraldehyde developed Abs to GXM. However, control peptides P315 and P24 conjugated to KLH also elicited Abs to GXM. GXM-binding Abs from mice immunized with P315-KLH were inhibited by KLH treated with glutaraldehyde (KLH-g), but not by P315. Furthermore, KLH-g inhibited binding of GXM by serum of mice immunized with GXM-TT, indicating that glutaraldehyde treatment of KLH reveals an epitope(s) that cross-reacts with GXM. Vaccination with KLH-g or unmodified KLH elicited Abs to GXM, but did not confer protection against C. neoformans, suggesting the cross-reactive epitope on KLH was not protective. This was supported by the finding that 4H3, a nonprotective mAb, cross-reacted strongly with KLH-g. Sera from mice immunized with either native KLH or KLH-g cross-reacted with several other carbohydrate Ags, many of which have been conjugated to KLH for vaccine development. This study illustrates how mAbs can be used to determine the efficacy of potential vaccines, in addition to describing the complexity of using KLH and glutaraldehyde in the development of vaccines to carbohydrate Ags.     

Idiotype vaccine for tumor by anti-idiotypic antibody prepared against anti-(bacillus Calmette Guèrin)BCG monoclonal antibody

Summary The anti-idiotypic antibody (Ab2) prepared against the anti-BCG monoclonal antibody (mAb) (Ab1) exhibited potential vaccine activity against Meth A fibrosarcoma that shared a common antigen(s) withMycobacterium bovis strain bacillus Calmette Guèrin (BCG). Mice vaccinated with the anti-idiotypic antibody (Ab2) were protected significantly against growth of the transplanted Meth A tumor (66%), and the presence of anti-(anti-idiotypic antibody) (Ab3) was proved in the Ab2-vaccinated mice by enzyme-linked immunosorbent assay and indirect immunofluorescence analyses using unabsorbed or absorbed sera against the BCG antigen(s) and Meth A tumor cells. This indicated that the anti-idiotypic antibody (Ab2) mimicked the structures of the BCG antigen(s) and behaved as the BCG antigen(s) to induce the Abl-like antibody (Ab3) in vivo. Presumably the Ab2-induced Ab3 plays a significant role in preventing growth of the transplanted tumor in animals. By contrast, the control mice treated with normal mouse serum failed to inhibit the tumor growth. These results suggest the possible development of a tumor vaccine from the anti-idiotypic antibody (Ab2) prepared against the anti-BCG monoclonal antibody, for tumors sharing a common antigen(s) withMycobacterium bovis strain BCG.

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Idiotype vaccine for tumor by anti-idiotypic antibody prepared against anti-(bacillus Calmette Guèrin)BCG monoclonal antibody

     

Peptide mimotopes of pneumococcal capsular polysaccharide of 6B serotype: a peptide mimotope can bind to two unrelated antibodies

Two groups of bacteriophage clones displaying the antigenic properties of serotype 6B pneumococcal capsular polysaccharide (PS) were obtained from different phage libraries expressing random heptameric peptides. One group, biopanned with a mouse mAb (Hyp6BM1), is comprised of 17 phage clones expressing 10 unique sequences of linear peptides. The other group, selected with another mAb (Hyp6BM8), contained six clones, all of which expressed the identical circular peptide. Phage clones expressing the linear peptides (e.g., PhaM1L3) bound only to Hyp6BM1, but not other 6B PS-specific mAb, and their binding could be inhibited with pneumococcal capsular type 6B PS only. In contrast, a phage clone expressing the circular peptide (PhaM8C1) cross-reacted with several other 6B PS-specific mAbs, and their binding could be inhibited with pneumococcal capsular PS of 6A and 6B serotypes. Two short peptides, PepM1L3 and PepM8C1, reflecting the peptide inserts of the corresponding phage clones, could inhibit the binding of the two clones to their respective mAb. Interestingly, the peptide insert in PhaM8C1 was identical to that in PhaB3C4, a previously reported mimotope of alpha(2-->8) polysialic acid, Neisseria meningitidis group B PS. Indeed, PhaM8C1 bound to HmenB3 (a meningococcal Ab), and their association could be inhibited with alpha(2-8) polysialic acid, but not with 6B PS. Conversely, alpha(2-8) polysialic acid could not inhibit the binding of PhaM8C1 to Hyp6BM8. The two-dimensional nuclear magnetic resonance studies indicate that PepM8C1 peptide can assume several conformations in solution. The ability of this peptide to assume multiple conformations might account for its ability to mimic more than one Ag type.     

Induction of an anti-human type II collagen response by monoclonal anti-idiotypic antibody

Several distinct epitopes on human type II collagen were defined by using mAb. The presence of species-specific and species-nonspecific (common) epitopes was thus clarified. Anti-idiotypic mAb (Ab2) was developed against one of the antibodies (Ab1) reactive with species-specific epitopes. Thus Ab2 was demonstrated to recognize an idiotope expressed on the Ag-binding site (paratope) of Ab1, since the binding of Ab1 to human type II collagen was blocked by Ab2, and the binding of Ab2 to Ab1 was inhibited by soluble human type II collagen, but not by murine and bovine type II collagens. DBA/1 mice immunized with Ab2 coupled to keyhole limpet hemocyanin produced an antibody (Ab3) specifically reactive with human type II collagen. It was also demonstrated that Ab3 expressed an idiotype similar to that of Ab1. These findings indicate that anti-idiotypic antibody directed against mAb to human type II collagen mimics a species-specific epitope on human type II collagen. The anti-idiotypic antibody bearing internal image of type II collagen will open the way to isolation of the arthritogenic epitope on type II collagen.     

A cryptococcal capsular polysaccharide mimotope prolongs the survival of mice with Cryptococcus neoformans infection

Defined Abs to the Cryptococcus neoformans capsular polysaccharide glucuronoxylomannan (GXM) have been shown to be protective against experimental cryptococcosis. This suggests that if a vaccine could induce similar Abs it might protect against infection. However, the potential use of a GXM-based vaccine has been limited by evidence that GXM is a poor immunogen that can induce nonprotective and deleterious, as well as protective, Abs, and that the nature of GXM oligosaccharide epitopes that can elicit a protective response is unknown. In this study, we investigated whether a peptide surrogate for a GXM epitope could induce an Ab response to GXM in mice. The immunogenicity of peptide-protein conjugates produced by linking a peptide mimetic of GXM, P13, to either BSA, P13-BSA, or tetanus toxoid, P13-tetanus toxoid, was examined in BALB/c and CBA/n mice that received four s.c. injections of the conjugates at 14- to 30-day intervals. All mice immunized with conjugate produced IgM and IgG to P13 and GXM. Challenge of conjugate-immunized mice with C. neoformans revealed longer survival and lower serum GXM levels than control mice. These results indicate that 1) P13 is a GXM mimotope and 2) that it induced a protective response against C. neoformans in mice. P13 is the first reported mimotope of a C. neoformans Ag. Therefore, the P13 conjugates are vaccine candidates for C. neoformans and their efficacy in this study suggests that peptide mimotopes selected by protective Abs deserve further consideration as vaccine candidates for encapsulated pathogens.     

An anti-idiotype vaccine elicits a specific response to N-glycolyl sialic acid residues of glycoconjugates in melanoma patients

We generated the 1E10 gamma-type anti-idiotype mAb (Ab2) specific to an Ab1 mAb able to react specifically with N-glycolyl-containing gangliosides and with Ags expressed on human melanoma and breast carcinoma cells. This Ab2 mAb induced an Ab response in animal models sharing immunochemically defined idiotopes with the Ab1. The treatment of tumor-bearing mice with 1E10 mAb induced a strong antitumor activity. A clinical trial was conducted in 20 patients with advanced malignant melanoma. Patients were treated with six intradermal injections of aluminum hydroxide-precipitated 1E10 anti-Id mAb given at 2-wk intervals. Sixteen of the 17 patients who received at least four doses of the anti-Id vaccine develop Ab3 Abs capable of inhibiting Ab2 binding to Ab1 (Ab3Id+). In contrast to the incapacity of 1E10 mAb to generate Ab3 Abs with the same antigenic specificity as the Ab1 mAb in mice, a very specific and strong Ab3 response against N-glycolyl-containing gangliosides was induced in 16 patients (Ab3Ag+). No evidence of serious or unexpected adverse effects has been observed in this clinical trial. 1E10 anti-Id vaccine was safe, well tolerated, and immunologically effective, with most patients being able to generate a specific immune response against 1E10 and Neu-glycolyl-GM(3) ganglioside.     

Study of various presentation forms for a peptide mimetic of Neisseria meningitidis serogroup B capsular polysaccharide

The formulation of a broadly protective vaccine to prevent the serogroup B Neisseria meningitidis (MenB) disease is still an unmet medical need. We have previously reported the induction of bactericidal and protective antibodies against MenB after immunization of mice with a phage-displayed peptide named 4 L-5. This peptide mimics a capsular polysaccharide (CPS) epitope in MenB. With the aim of developing vaccine formulations that could be used in humans, we evaluate in this study various forms of presentation to the immune system of the 4 L-5 sequence, based on synthetic peptides. We synthesized the following: (i) a linear 4 L-5 peptide, (ii) a multiple antigen peptide containing four copies of the 4 L-5 sequence (named MAP), which was then dimerized, and the product named dimeric MAP, and (iii) a second multiple antigen peptide, in this case with two copies of the 4 L-5 sequence and a copy of a T-helper cell epitope of tetanus toxoid, which was then dimerized and the product named MAP-TT. The linear peptide, the MAP, and the dimeric MAP were conjugated to the carrier protein P64K by different conjugation methods. Plain antigens and antigens coupled to P64K were used to immunize BALB/c mice. Of those variants that gave immunogenic results, MAP-TT rendered the highest levels of specific antipeptide IgG antibodies and serum bactericidal activity. These results can find application in the development of meningococcal vaccine candidates and in peptide-based vaccines strategies.     

Induction of neutralizing antibody in mice against poliovirus type II with monoclonal anti-idiotypic antibody

Syngeneic monoclonal anti-idiotope antibody Ab2,2-17C3SCC was raised against an idiotope on a protective monoclonal antibody with specificity for poliovirus type II. Ab2,2-17C3SCC detects a paratope-related interspecies IdX. Ab2,2-17C3SCC purified from supernatant fluids of hybridoma cells by protein A-Sepharose was injected into 4- to 6-wk-old BALB/c mice. The sera of the mice were screened for the expression of antibodies bearing the corresponding idiotope. Immunization of mice with Ab2,2-17C3SCC induced antibodies of complementary specificity. Furthermore, micro VN tests suggest that Ab2,2-17C3SCC can substitute for antigen in the induction of anti-polio neutralizing antibodies, and hence can function as a monoclonal anti-idiotypic vaccine.     

Isotype can affect the fine specificity of an antibody for a polysaccharide antigen

Ab specificity is determined by V region sequence. The murine Mab 18B7 (IgG1) binds to the Cryptococcus neoformans capsular polysaccharide glucuronoxylomannan and produces annular immunofluorescence (IF) on yeast cells. The heavy and light V regions of 18B7 were expressed with the human C regions micro, gamma 1, gamma 2, gamma 3, gamma 4, and alpha1, and the specificity and binding properties of these mouse-human chimeric (ch) Abs was determined. The chIgG1, chIgG2, chIgG4, and the chIgA produced annular IF, whereas the IgM and IgG3 produced punctate IF, despite identical V region sequences. Competition experiments with murine Abs that competed with mAb 18B7 and binding assays to peptide mimetics of glucuronoxylomannan provided additional evidence for altered specificity in some of the ch Abs. Expression of 18B7 heavy V region with murine micro C region produced IgM with a punctate IF, indicating that a change in fine specificity also accompanied the change from murine IgG1 to IgM. Our results show that Ab fine specificity can be a function of isotype. This phenomenon may be most apparent for Abs that bind to Ag with repeating epitopes, such as polysaccharides, where the quarternary structure of the Ag-Ab complex may be influenced by such constraints as Fab-Fab angles, Fc-Fc interactions, Ab size, and solvent accessibility to exposed surfaces. Alterations in Ab fine specificity following isotype change could have important implications for current concepts on the generation of secondary Ab responses to certain Ags and for the isotype preference observed in Abs to polysaccharides.