Process for preparing polysaccharide-protein conjugate vaccines
AssigneesThe United States of America, as represented by the Secretary, Department of Health and Human Services, Fiocruz
InventorsEllen Jessouroun, Ivna Alana Freitas Brasileiro Da Silveira, Renata Chagas Bastos, Carl E. Frasch, Che-Hung Robert Lee
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Abstract
Methods for the manufacture of polysaccharide-protein conjugate vaccines at high yield are provided. The methods involve reaction of a hydrazide group on one reactant with an aldehyde group on the other reactant. The reaction proceeds rapidly with a high conjugation efficiency. Simplified purification processes can be employed to separate the conjugate product from the unconjugated protein and polysaccharide and other small molecule by-products.
Claims14
A method for preparing a meningococcal-tetanus toxoid conjugate vaccine in commercial volumes, the method comprising:
- reacting a meningococcal polysaccharide with an oxidizing agent, whereby a solution of an aldehyde-activated meningococcal polysaccharide is obtained;
- reacting a tetanus toxoid protein with hydrazine dichloride at an acidic pH, whereby a solution of a hydrazine-activated tetanus toxoid protein is obtained;
- purifying by diafiltration said solution of hydrazine-activated tetanus toxoid protein to process at least five liters of solution;
- reacting the aldehyde-activated meningococcal polysaccharide with the hydrazine- activated tetanus toxoid protein at a pH of from 5 to 7 in the presence of sodium cyanoborohydride, whereby a meningococcal-tetanus toxoid conjugate is obtained;
- neutralizing unreacted aldehyde groups with adipic acid dihydrazide;
- purifying by diafiltration the resulting solution to process a volume of at least two liters, producing a purified meningococcal-tetanus toxoid conjugate vaccine, and concentrating the purified meningococcal-tetanus toxoid conjugate vaccine by tangential flow ultrafiltration, yielding a concentrated purified meningococcal-tetanus toxoid conjugate vaccine, whereby the meningococcal-tetanus toxoid conjugate vaccine capable of stimulating an immune response is obtained in commercial volumes.
A method for preparing a meningococcal-tetanus toxoid conjugate vaccine in commercial volumes, the method comprising:
- reacting a meningococcal polysaccharide with an oxidizing agent, whereby a solution of an aldehyde-activated meningococcal polysaccharide is obtained;
- buffer exchanging the solution of the aldehyde-activated meningococcal polysaccharide to a pH from about 7 to 8;
- reacting a tetanus toxoid protein with hydrazine dichloride at an acidic pH, whereby a solution of a hydrazine-activated tetanus toxoid protein is obtained;
- raising a pH of the solution of the hydrazine-activated tetanus toxoid protein from 7.0 to 11 and thereafter buffer exchanging the solution of the hydrazine-activated tetanus toxoid protein to a pH from 10.0 to 11.0;
- purifying by diafiltration said solution of hydrazine-activated tetanus toxoid protein to process at least five liters of solution;
- reacting the aldehyde-activated meningococcal polysaccharide with the hydrazine-activated tetanus toxoid protein at a pH from 5 to 7 in the presence of sodium cyanoborohydride, whereby a meningococcal-tetanus toxoid conjugate is obtained;
- neutralizing unreacted aldehyde groups with adipic acid dihydrazide;
- purifying by diafiltration the resulting solution to process a volume of at least two liters, producing a purified meningococcal-tetanus toxoid conjugate vaccine, and concentrating the purified meningococcal-tetanus toxoid conjugate vaccine by tangential flow ultrafiltration, yielding a concentrated purified meningococcal-tetanus toxoid conjugate vaccine, whereby a meningococcal-tetanus toxoid conjugate vaccine capable of stimulating an immune response is obtained in commercial volumes.
Dependencies are inferred from the claim text.