Protein and Polysaccharide Conjugate Vaccines to Enteric

Surface polysaccharides of Gram-negative enteric pathogens, capsules or lipopolysaccharides, are both essential virulence factors and protective antigens. The immunogenicity of these polysaccharides is enhanced by their binding to carrier proteins. Viral surface capsid proteins are commonly found to be protective antigens and suitable as vaccine candidate<P>SALMONELLA TYPHI. The capsular polysaccharide of Salmonella typhi, Vi, is an essential virulence factor and a protective antigen. To improve its immunogenicity in young children, Vi was conjugated to a carrier protein, the recombinant exoprotein A of Pseudomonas aeruginosa (rEPA). A Phase 3 trial of the Vi-rEPA conjugate vaccine in about 12,000 2-5 year old children showed an efficacy of 89% after 48 months of surveillance. A clinical trial of Vi-rEPA injected with DTP in Vietnamese infants has been approved and started in March, 2006. 50 infants have been enrolled since August 2006.<P>SALMONELLA PARATYPHI A is the second most common cause of enteric fever in Southeast Asia. A conjugate vaccine of the O-specific polysaccharide (O-SP) of S. paratyphi A and TT was found to be safe and immunogenic in adults, teenagers, and 2-4 year old children. The O-acetyl content of the PS was found to be related directly to the immunogenicity of the conjugate.To enhance the immunogenicity of O-SP by increasing its molecular weight, a mutant S. paratyphi A was constructed by replacing the LPS elongation chain length regulator, wzz gene, with that of E. coli K12. Silver staining of an SDS-PAGE of this product showed a higher molecular weight distribution than of the native LPS. <P>ESCHERICHIA COLI O157 is a major cause of hemolytic uremic syndrome, especially in young children. A phase 1 trial of it?s O-SP-rEPA conjugate demonstrated safety and immunogenicity in adult volunteers. In a phase 2 study of this conjugate, 55 children 2-5 years old were injected once or twice. The vaccine was found to be safe and immunogenic; 98% of children responded with a greater than 4 fold rise of IgG anti-LPS at 6 months after the first injection, with no booster response after the second. <P>ENTEROTOXIGENIC ESCHERICHIA COLI (ETEC): E. coli that secrete cholera-like toxin (ETEC) cause diarrhea in most countries. The incidence, morbidity, and mortality of ETEC diarrhea are highest in developing countries. ETEC diarrhea is caused by two exotoxins: 1, similar to the cholera toxin in structure and action, is known as the heat-labile toxin (LT); and 2, a polypeptide of 19 amino acids called heat stable toxin (ST). About 40% of ETEC in strains from patients secrete LT alone, 20% secrete both LT and ST, and 60% secrete ST alone. The immunogenicity or protective actions of LT and ST have not been studied thus far.Purified LT and a recombinant mutant non-toxic protein (rLT) have been obtained from John Clements, Tulane University. The mutant LT and rLT, treated with low concentrations of formaldehyde, were injected into young outbred mice. Both induced high levels of IgG anti LT. There was a dosage dependent swelling at the injection site. Adsorption of these proteins onto aluminum hydroxide delayed this adverse reaction. <P>VIBRIO CHOLERA O1 AND O139 are the major disease causing serotypes. The capsular polysaccharide of V. cholera O139 or the O-SP of V. cholera O1 conjugated to various carrier proteins elicited vibriocidal antibodies in mice, with higher levels than those induced by the LPS alone. <P>CAMPYLOBACTER JEJUNI infection is one of the most common enteric infections around the world. Most infections are mild, but serious complications, notably Guillain- Barre syndrome , may occur.Campylobacter is microaerophilic and fermentation in liquid media has been difficult. To design an experimental vaccine, based on it?s lipooligosaccharide (LOS), culture conditions were investigated; various media and fermentation conditions were studied and requirements for growth in liquid media were established. Several C. jejuni serotypes were then cultivated. Chemical analysis of the LOS obtained showed several keto groups in both types. Protein conjugates of the LOS or of the de-acylated LOS were immunogenic in mice and the antibodies elicited by these conjugates showed complement dependent bactericidal activity.28 isolates of C. jejuni from pediatric patients in Israel were analyzed for serotype distribution based on the HS (capsule) scheme. A diverse serotype distribution was found. Chemical and serological analysis of the LOS of these isolates revealed no correlation between the HS types and LOS carbohydrate compositions. Furthermore approximately 40% of the 16 isolates chemically analyzed contained sialic acid in their LOS and may bind to antisera of gangaliosides. THE EFFECT OF LOW BIRTH WEIGHT ON THE IMMUNE RESPONSE IN ADULTHOOD was studied using the Vi polysaccharide from S. typhi as a probe. Substantial evidence links low birth weight (not due to prematurity) to susceptibility later in life to chronic disease. As components of the immune system may be programmed early in life, we investigated the potential association between birth weight and response to vaccination with Vi in a cohort of 257 adults (mean age: 29.4 y; 146 men) born in an urban slum in Lahore, Pakistan, during 1964?1978. Primary immunization with Vi showed a correlation between birth weight and IgG anti-Vi response, with the lower birth weight the lower the antibody levels. These adults were re-injected with the same vaccine 2 years later; no change in results obtained. In contrast, no birth weight/antibody response relation was observed in a comparable group of adults injected with T-cell dependent antigens such as Hib-conjugate and rabies vaccines.ROTAVIRUS infection is the most common cause of infantile diarrhea, regardless of economic status. Vaccine development strategies have mostly taken the approach of the reassortant whole cell oral vaccine. One such vaccine, Rotashield, was withdrawn after a suspected increase in the incidence of intusseception following vaccination. We have initiated a study of the surface proteins, capsits VP8 and VP7, as immunogens.