Experimental Molecular Design of Combined Vaccines

A method was elaborated to construct combined artificial immunogens simulating virus particles. The gist was exposing protein antigenic determinants of one virus on the particle surface and delivering plasmids with genes for antigenic proteins of another virus to specialized immune cells. Such immunogens were constructed and shown to induce biosynthesis of specific antibodies against HIV-1 and the tick-borne encephalitis virus. The level and duration of the humoral and cell responses were assayed.     

Comparative immunological properties of enantiomeric peptides

Summary The immunogenicity of a peptide composed of only d-amino acids is compared with that of the corresponding l-peptide enantiomer. Following three administrations of 100 g of individual peptide formulated with different adjuvants (Freund's complete adjuvant, QS21, or alum) to BALB/c mice, guinea pigs and rabbits, the l-peptide elicited strong l-peptide-specific IgG antibody responses in all formulations, whereas the d-peptide-induced d-peptide-specific IgG antibodies in the Freund's complete adjuvant and QS21 formulations, but was nonimmunogenic in the alum formulation. Mouse T-cell lines induced by the d-peptide formulated in Freund's complete adjuvant were found to express significant amounts of IL-2 when they were stimulated by the d-peptide. When an equal amount of both enantiomers was mixed and administered in Freund's complete adjuvant, only an l-peptide-specific IgG antibody response was observed. These results suggest that (i) d-peptide is immunogenic when strong adjuvant is provided; (ii) the immune system has preferential recognition of l-amino acid peptide; and (iii) the d-peptide can elicit d-peptide-specific T-cell responses.     

Synthesis of a neoglycoconjugate containing a Chlamydophila psittaci-specific branched Kdo trisaccharide epitope

The branched Kdo trisaccharide sodium (3-deoxy-α-d-manno-oct-2-ulopyranosyl)onate-(2→8)-[sodium (3-deoxy-α-d-manno-oct-2-ulopyranosyl)onate-(2→4)]-sodium (allyl 3-deoxy-α-d-manno-oct-2-ulopyranosid)onate has been prepared utilizing the regioselective glycosylation of the C-7, C-8 diol entity of a Kdo monosaccharide acceptor with a Kdo bromide donor followed by the attachment of the third Kdo unit to O-4 of the disaccharide intermediate. Deacetylation and hydrolysis of the methyl ester groups furnished the trisaccharide allyl glycoside which was converted into the corresponding 3-(2-aminoethylthio)propyl glycoside. Subsequent covalent attachment to bovine serum albumin furnished a neoglycoconjugate serving as an antigen for the induction of Chlamydophila psittaci-specific monoclonal antibodies.     

Clustered carbohydrates as a target for natural killer cells: a model system

Membrane-associated oligosaccharides are known to take part in interactions between natural killer (NK) cells and their targets and modulate NK cell activity. A model system was therefore developed using synthetic glycoconjugates as tools to modify the carbohydrate pattern on NK target cell surfaces. NK cells were then assessed for function in response to synthetic glycoconjugates, using both cytolysis-associated caspase 6 activation measured by flow cytometry and IFN-γ production. Lipophilic neoglycoconjugates were synthesized to provide their easy incorporation into the target cell membranes and to make carbohydrate residues available for cell–cell interactions. While incorporation was successful based on fluorescence monitoring, glycoconjugate incorporation did not evoke artifactual changes in surface antigen expression, and had no negative effect on cell viability. Glycoconjugates contained Le^x, sulfated Le^x, and Le^y sharing the common structure motif trisaccharide Le^x were revealed to enhance cytotoxicity mediated specifically by CD16 ⁺CD56⁺NK cells. The glycoconjugate effects were dependent on saccharide presentation in a polymeric form. Only polymeric, or clustered, but not monomeric glycoconjugates resulted in alteration of cytotoxicity in our system, suggesting that appropriate presentation is critical for carbohydrate recognition and subsequent biological effects.     

The polymorphic epithelial mucin: potential as an immunogen for a cancer vaccine

 The identification and cloning of several tumour antigens together with an improvement in the understanding of the mechanisms involved in antigen presentation and immune recognition has opened up the possibility of using active specific immunotherapy as a treatment for certain cancers. This review discusses the tumour-associated MUC1 gene product of the polymorphic epithelial mucin (PEM), as a potential target molecule for cancer treatment. PEM is both over-expressed and aberrantly glycosylated in many carcinomas resulting in an antigenically distinct molecule. Furthermore, immune responses specific for PEM have been detected in cancer patients. Both syngeneic and transgenic murine model systems have been developed in order to compare the efficacy and toxicity of various PEM-based immunogens in tumour rejection studies, and to further improve the understanding of antigen presentation and the mechanisms underlying tumour rejection. Such models also allow the examination of MUC1-based immunogens as a treatment for existing tumours. Clinical trials in progress using immunogens based on the MUC1 gene product are briefly discussed. Received: 23 November 1995 / Accepted: 4 January 1996     

The Amadori rearrangement as key reaction for the synthesis of neoglycoconjugates

The Amadori rearrangement was introduced as a key step for the conjugation of carbohydrate moieties with suitable amines such as aliphatic amines and amino acid derivatives. The rearrangement products were further transformed into the corresponding 1-N,2-O cyclic carbamates employing triphosgene to obtain anomerically stable glycoconjugates. The reaction conditions were probed on a model substrate, 3,5-di-O-benzyl-alpha,beta-d-glucofuranose and further applied to d-glycero-d-gulo-heptose, which gave 'd-gluco-conjugates' in the alpha-anomeric form exclusively in high isolated yields.     

Investigation towards bivalent chemically defined glycoconjugate immunogens prepared from acid-detoxified lipopolysaccharide of <Emphasis Type="Italic">Vibrio cholerae</Emphasis> O1, serotype Inaba

A free amino group present on the acid-detoxified lipopolysaccharide (pmLPS) of V. cholerae O1 serotype Inaba was investigated for site-specific conjugation. Chemoselective pmLPS biotinylation afforded the corresponding mono-functionalized derivative, which retained antigenicity. Thus, pmLPS was bound to carrier proteins using thioether conjugation chemistry. Induction of an anti-LPS antibody (Ab) response in BALB/c mice was observed for all conjugates. Interestingly, the sera had vibriocidal activity against both Ogawa and Inaba strains opening the way to a possible bivalent vaccine. However, the level of this Ab response was strongly affected by both the nature of the linker and of the carrier. Furthermore, no switch from IgM to IgG, i.e. from a T cell-independent to a T cell-dependent immune response was detected, a result tentatively explained by the possible presence of free polysaccharide in the formulation. Taken together, these results encourage further investigation towards the development of potent pmLPS-based neoglycoconjugate immunogens, fully aware of the challenge faced in the development of a cholera vaccine that will provide efficient serogroup coverage.     

Immunogenicity of synthetic peptides related to the core peptide sequence encoded by the human MUC1 mucin gene: Effect of immunization on the growth of murine mammary adenocarcinoma cells transfected with the human MUC1 gene

The immune response of CAF1 mice to various synthetic peptides (SP) related to the amino acid sequence (PDTRPAPGSTAPPAHGVTSA) of the tandem repeat of the MUC1 human breast mucin core peptide was evaluated. The most immunogenic preparations of the synthetic peptides were those conjugated to keyhole limpet hemocyanin (KLH) or clustered in a dendritic multiple antigenic peptide (MAP-4) configuration. The mice were immunized subcutaneously with synthetic peptides emulsified in RIBI adjuvant, employing various immunization protocols. Equivalently high IgG responses were induced using SP-KLH conjugates (GVTSAPDTRPAPGSTA-KLH) or an SP — MAP-4 chimeric configuration (SP1-6), which also included a universal malarial CST-3 T-helper epitope (SP1-6 = SAPDTRPAEKKIAKMEKASSVFNVVNS — MAP-4). These IgG antibodies bound both the appropriate MUC1 synthetic peptides and the cell surface expressed MUC1 mucin on murine mammary cells that had been transfected with the human MUC1 gene and a human breast cancer cell line that expresses cell-surface MUC1. A MAP-4 molecule, which included the entire 20-aminoacid sequence of the MUC1 tandem repeat (SP1-5 = PDTRPAPGSTAPPAHGVTSA—MAP-4) induced a poor IgG response. In contrast, all three types of molecule: SP-KLH, SP1-6 and SP1-5, were found to be good immunogens for the induction of specific delayedtype hypersensitivity (DTH) reactions measured using either synthetic peptides or MUC1-transfected cells. In addition, immunization with irradiated MUC1-transfected cells induced strong DTH reactions measured using synthetic peptides that expressed the PDTRP sequence, which has been shown to be, or to overlap, a T cell epitope in humans and a B cell epitope in mice. Finally, it was demonstrated that synthetic MUC1 peptide vaccines could be used both prophylactically and therapeutically to inhibit the growth of MUC1-transfected tumor cells and prolong the survival of tumor-bearing mice.     

重要水产养殖动物病原菌高效多价疫苗保护原的研究

以疫苗为核心的免疫防治是控治水产动物养殖业微生物病害的必由之路。鉴于水产养殖动物病原菌种类繁杂、血清型多样,同时从经济方面考虑也不可能对所有病原菌及其血清型逐一进行疫苗预防接种。因此,研制疫苗必须在思路和技术上进行创新。该文提出了以发展多价疫苗为思路的水产养殖动物细菌性疾病控制的新免疫预防措施,介绍了发现和鉴定多价疫苗候选功能靶位的研究思路和技术路线,展望了基于免疫层次抗原组学的新概念对系统深入研究天然高效多价保护原的良好前景。