Polysaccharide derivatives and uses in induction of an immune response

Abstract

The present invention generally provides compositions comprising a polysaccharide derivative, and methods of their preparation and use for the prevention or treatment of diseases caused by Neisseria meningitidis bacteria, particularly group B (NmB) strains, and by E. coli K1. The invention provides a de-N-acetylated PS derivative in which one or more residues of the PS has been modified by de-N-acetylation. The invention also includes derivatives in which one or more of the N-acetyl groups of PS containing de-N-acetylated PS are replaced with other N-acyl groups, usually a lower acyl group of C 2 -C 3 . Further, the invention includes de-N-acetylated PS derivatives containing long chain hydrocarbons, as well as conjugates in which the de-N-acetylated PS derivative is linked to a carrier, e.g., a carrier protein.

1 .- 18 . (canceled) 
   
   
       19 . A method of producing a polysaccharide (PS) derivative, the method comprising:
 reacting an at least partially de-N-acetylated PS molecule with a mixture comprising an amine protecting reagent and an acylating reagent in the presence of an organic solvent;   wherein said reacting produces a protected PS derivative having an amine protecting group and an N-acylated group.   
   
   
       20 . A protected polysaccharide (PS) derivative produced by the method of  claim 19 . 
   
   
       21 . The method of  claim 19 , further comprising:
 treating the protected PS derivative under conditions to remove the amine protecting group;   wherein a PS derivative is produced.   
   
   
       22 . The method of  claim 19 , wherein the method further comprises:
 conjugating the protected PS derivative to a carrier protein to produce a conjugated protected PS derivative; and   treating the conjugated protected PS derivative under conditions to remove the amine protecting group;   wherein a conjugated PS derivative is produced.   
   
   
       23 . The method of  claim 19 , wherein the method further comprises:
 reacting the protected PS derivative with an N-acyl amine reagent to produce an amidated protected PS derivative having an amidated sialic acid residue having an alkyl secondary amine; and   treating the amidated protected PS derivative under conditions to remove the removing the amine protecting group;   wherein a PS derivative having an amidated sialic acid residue having an alkyl secondary amine at is produced.   
   
   
       24 . A method of producing a polysaccharide (PS) derivative, the method comprising:
 culturing a  Neisseria meningitidis  Group B bacterium in a growth medium comprising an N-acyl-mannosamine and an amine-protected mannosamine, wherein the bacterium is deficient in production of capsule polysaccharide in the absence of supplemental mannosamine;   wherein said culturing provides for production of a protected PS derivative having an amine protecting group and an N-acylated group.   
   
   
       25 . The method of  24 , wherein the method further comprises:
 conjugating the protected PS derivative to a carrier protein to produce a conjugated protected PS derivative; and   treating the conjugated protected PS derivative under conditions to remove the amine protecting group;   wherein a conjugated PS derivative is produced.   
   
   
       26 . The method of  claim 24 , wherein the method further comprises:
 reacting the protected PS derivative with an N-acyl amine reagent to produce an amidated protected PS derivative having an amidated sialic acid residue having an alkyl secondary amine; and   treating the amidated protected PS derivative under conditions to remove the removing the amine protecting group;   wherein a PS derivative having an amidated sialic acid residue having an alkyl secondary amine at is produced.   
   
   
       27 . A method for identifying a polysialic acid (PSA) epitope bound by an antibody, the method comprising:
 contacting a PSA derivative with an antibody, said contacting being under conditions suitable for formation of antigen-antibody complexes of the antibody and the PS derivative; and   contacting the antigen-antibody complex with a sialidase under conditions suitable for removal of sialidase-accessible PSA derivative residues of the antigen-antibody complex;   wherein PSA derivative residues remaining in the antigen-antibody complex following sialidase treatment define an epitope bound by the antibody.