Manufacture of vaccines and compositions for the prevention of Salmonella infections
Inventors
Ella, Krishna Murthy • Ramasamy, Venkatesan • Naidu, Mandalapu Gangadhara
Assignees
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Publication Number
US-10702595-B2
Publication Date
2020-07-07
Expiration Date
Abstract
Disclosed are stable conjugate vaccine formulations for protection against Salmonella typhi, and methods of conjugation between Vi-polysaccharide of S. typhi to tetanus toxoid as the carrier protein, responsible for producing improved T-dependent immune response against Typhoid fever caused by Salmonella typhi. The methods disclosed in this invention and the resulting formulations are capable of inducing immunity against typhoid fever including in children below 2 years of age, through only a single injection to comprise a complete vaccination schedule.
Core Innovation
The invention relates to a method of manufacture of a ViP-carrier protein conjugate. The method comprises purifying capsular Vi-polysaccharide antigens (VIPs) of Salmonella, treating the purified VIPs with sodium bicarbonate to form partially de-O-acetylated ViPs, treating the partially de-O-acetylated ViPs with a carbodiimide cross-linking agent, and linking the carbodiimide treated partially de-O-acetylated ViPs with a linker molecule comprising adipic acid dihydrazide.
The linked partially de-O-acetylated ViPs are isolated and then contacted with a carrier protein in the presence of EDAC to form the ViP-carrier protein conjugate. In particular embodiments, capsular Vi-polysaccharide antigens (VIPs) of Salmonella typhi are treated with sodium bicarbonate to form partially de-O-acetylated ViPs having a molecular weight of from 250-300 kDa.
The conjugate is further associated with formulation aspects, including contacting the ViP-carrier protein conjugate with a preservative. The preservative comprises 2-phenoxy ethanol that is free of mercury chloride and/or thiomersal, and the VIP contains no free linker molecules.
Claims Coverage
The partial content provides four independent claims that cover manufacture of a ViP-carrier protein conjugate using partially de-O-acetylated Salmonella Vi-polysaccharides, carbodiimide cross-linking with EDAC, and linking with a linker comprising adipic acid dihydrazide, with additional product-quality and formulation constraints.
Purifying VIPs and forming partially de-O-acetylated ViPs
Purifying capsular Vi-polysaccharide antigens (VIPs) of Salmonella; treating the purified VIPs with sodium bicarbonate forming partially de-O-acetylated ViPs.
Carbodiimide cross-linking and adipic acid dihydrazide linking
Treating the partially de-O-acetylated ViPs with a carbodiimide cross-linking agent; linking the carbodiimide treated partially de-O-acetylated ViPs with a linker molecule, wherein the linker comprises adipic acid dihydrazide.
EDAC-mediated conjugation to carrier protein
Contacting the linked partially de-O-acetylated ViPs with a carrier protein in presence of EDAC to form the ViP-carrier protein conjugate.
EDAC cross-linking with EDAC-containing carbodiimide cross-linking agent
Treating the partially de-O-acetylated ViPs with a carbodiimide cross-linking agent, wherein the cross-linking agent comprises 1-ethyl-3(3-dimethylaminopropyl) carbodiimide (EDAC).
Formulating with preservative comprising 2-phenoxy ethanol free of mercury chloride and/or thiomersal
Contacting the ViP-carrier protein conjugate with a preservative, wherein the preservative comprises 2-phenoxy ethanol which is free of mercury chloride and/or thiomersal.
No free linker molecules and defined ViPs molecular weight range
Treating capsular Vi-polysaccharide antigens (VIPs) of Salmonella typhi with sodium bicarbonate forming partially de-O-acetylated ViPs and linking the partially de-O-acetylated ViPs with a linker comprised of adipic acid dihydrazide, wherein the VIP contains no free linker molecules and has an approximate molecular weight of from 250-300 kDa.
Across the independent claims, the core coverage is manufacturing a ViP-carrier protein conjugate by converting Salmonella Vi-polysaccharides to partially de-O-acetylated ViPs, applying carbodiimide cross-linking including EDAC, coupling via an adipic acid dihydrazide linker, and conjugating with a carrier protein in the presence of EDAC, with additional constraints relating to defined molecular weight, absence of free linker molecules, and optional formulation with 2-phenoxy ethanol free of mercury chloride and/or thiomersal.
Stated Advantages
Not explicitly described in patent.
Documented Applications
Not explicitly described in patent.
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