A Pneumococcal Polysaccharide Conjugate Vaccine With The Phenyl Linker And The Study On Its Immunological Property

Streptococcus pneumonia is a serious Gram-positive pathogen and can lead to invasive pneumococcal diseases with high mortality rate.Capsular polysaccharide(PS)is the major virulence factor of S.pneumoniae.Previously,capsular PS was used for protection against pneumococcal diseases.However,capsular PS is a T-cell independent antigen that induces a poor immune response in infants and young children.In addition,the immune responses to PS are of short duration and cannot be boosted and the affinity does not mature.Thus,pneumococcal capsular PS has been conjugated with a carrier protein possessing T-helper cell epitopes to improve its immunogenicity,which effectively protects infants and young children against S.pneumonia.For instance,a 13-valent pneumococcal conjugate vaccine(Pfizer Inc.,USA)was licensed in 2010 and significantly reduced the incidence of invasive pneumococcal disease via the acquisition of PS-specific antibodies.However,the PS-specific immunogenicity of pneumococcal conjugate vaccines still needs to be improved.Several factors such as the length of PS chain,PS-to-protein ratio,and the conjugation chemistry are important to produce the PS-specific antibodies.Conjugation of the conjugate vaccines with adjuvant has also been used to stimulate the production of PS-specific antibodies.Beyond that,the carrier-induced epitopic suppression(CIES)mechanism has been observed in the study of conjugate vaccine.Through the CIES mechanism,the elicited high level carrier Protein specific antibodies can suppress the immune response to the capsular PS on the conjugate.Thus,the strategy to reduce the CIES effect of carrier proteins on the conjugate by introducing chemical linkers that moderately shield the epitope of the carrier protein is efficent.Recently,the hydrophobic phenyl linker has been found can induce a dense conformation of PEG by interaction with the hydrophilic PEG,which extensively shielded most domains adjacent to the antigen epitopes of SAK.Here,three conjugates,(PS-TT,PS-phe-TT and PS-2phe-TT)were prepared to study whether the phenyl linker can reduce the CIES effect and improve the immunogenicity of the pneumococcal polysaccharide conjugates.The structural property and immunogenicity of the three pneumococcal polysaccharide conjugates were investigated.The details were as following:The three conjugates were purified by size exclusion chromatography and identified by 1H NMR,which indicated the successful conjugation of PS and TT.In addition,DLS analysis showed that the molecular radii of PS-TT,PS-phe-TT and PS-2phe-TT were 11.6±0.3 nm,10.9±0.4 nm and 10.3±0.2 nm,respectively.As compared with the conjugate without the phenyl linker(PS-TT),the conjugate with the phenyl linker(PS-phe-TT)showed lower TT-specific IgG titers and significantly higher PS-specific IgG titers and higher IL-5 level.This indicated that the phenyl linker could alter the PS conformation and efficiently shield the TT epitopes.Thus,the PS-specific immunogenicity of the conjugate vaccine could be improved by decreasing the CIES effect of TT with the phenyl linker.In addition,the damage extent of PS epitopes and TT epitopes were minimized for PS-phe-TT.PS-phe-TT preferentially promoted a robust Th2-type immune response.In summary,PS-phe-TT is expected to act as an effective pneumococcal conjugate vaccine for protection against pneumococcal diseases.