Reactions And Immunochemistry Of Carbohydrates

Diabetes, Digestive, Kidney Diseases

Investigators

Linked publications & trials

Abstract

We have three ongoing projects within our continued effort aimed at developing conjugate vaccines for infectious diseases from synthetic fragments of bacterial polysaccharides. Two are concerned with a vaccine for cholera and one with a vaccine for anthrax. Potent vaccines for these diseases are not available. Development of such vaccines is important from both the point of view of public health and of national interest. Development of a potent vaccine for cholera is important because of the involvement of our military in protecting US interests in developing third world countries. While anthrax does not constitute a major health problem in the civilized world, new concerns regarding anthrax have emerged because of potential use of some form of Bacillus anthracis, the etiological cause of anthrax, as a biological weapon. Our work towards a potent conjugate vaccine for cholera involves synthesis of oligosaccharides that mimic the structure of O-specific polysaccharide (O-PS) of Vibrio cholerae in the form suitable for conjugation, conjugation of these antigens to suitable carriers, and serological evaluation of the immunogenicity of the resulting neoglycoconjugates. The approach towards a vaccine for anthrax is based on preparation of a neoglycoconjugate from a suitable carrier and the tetrasaccharide side chain of the major glycoprotein of Bacillus anthracis exosporium. In the cholera project, we are still trying to simplify the synthesis of the hexasaccharide fragment of the O-specific polysaccharide of Vibrio cholerae O1 using glycosyl donors and glycosyl acceptors having the N-(3-deoxy-L-glycero-tetronamido) side chain already in place. In addition, we investigate the role of details in the architecture of neoglycoconjugates upon immunogenicity. We have previously evaluated serological responses of conjugates from the hexasaccharide that mimics the O-PS of Vibrio cholerae O:1. Those conjugates were found potent immunogens, which also elicited in mice bactericidal and protective antibodies for Vibrio cholerae O:1. To determine if there was a minimum saccharide length required for immunogenicity and efficacy, neoglycoconjugates (CHO?BSA) made from shorter saccharides (mono- to pentasaccharide) were tested in mice. Also tested were conjugates that had different length of the linker. The effect of interval between immunizations upon the efficacy these immunogens was also investigated. Immunizations were performed at either a constant mass but differing moles, or equal moles but differing masses. Humoral responses were essentially the same when mice received 9 micrograms of the carbohydrate (0.007 mM with the pentasaccharide) in each of the neoglycoconjugates prepared from mono- through the pentasaccharide, or the same molar amount (0.007 mM), proportionally less by weight when going from the penta- to the monosaccharide. These data showed that, within this dose range, the responses occurred virtually independently of the amount of immunogen. Selected antisera induced by these immunogens were protective, but the ELISA titers of the sera were not predictive of the protective capacity. [unreadable] Regarding conjugates with different length of linker, the majority of mice given immunizations separated by a 14-day gap did not produce vibriocidal or protective antibodies. Mice immunized 28 days apart with immunogens containing the shortest or medium length linker, but not the longest, produced vibriocidal and protective antibodies.[unreadable] Within the anthrax project, our work toward the vaccine is complicated by the fact that the mode of linkage of the tetrasaccharide side chain in the exosporium glycoprotein is not known. We can expect optimum immune response from the neoglycoconjugate only when all epitopes in the antigenic component of the neoglycoconjugate are identical to those in the anthrax spores, including the anomeric configuration at the site of attachment of the tetrasaccharide in the glycoprotein. With that information unknown we will have to synthesize two linker-equipped tetrasaccharide, one with alpha- and one with beta-linked spacer. We have now synthesized the a-linked tetrasaccharide.