Polysialic acid derivatives, methods of production, and uses in enhancing cancer antigen production and targeting
View Patent ↗The present invention relates to compositions and methods of their production and use, including use in increasing de-N-acetyl sialic acid antigen of a mammalian cell and methods that exploit the increase in deNAc sialic acid antigen on such cells.
1. A method of producing a substantially unoxidized and isolated polysialic acid derivative having a defined degree of polymerization, a mixture of N-acetyl sialic acid and de-N-acetyl sialic residues, and a non-reducing end de-N-acetyl residue resistant to degradation by exoneuraminidase, the method comprising:
providing a solution comprising a mixture of substantially unoxidized polysialic acid derivatives each having (i) a different degree of polymerization, (ii) a different mixture of N-acetyl residues and de-N-acetyl residues, and (iii) a non-reducing end N-acetyl sialic acid residue;
subjecting said solution to ion exchange chromatography to generate fractions; and
isolating from one or more of said fractions a substantially unoxidized polysialic acid derivative having a defined degree of polymerization, a mixture of N-acetyl sialic acid and de-N-acetyl sialic residues, and a non-reducing end de-N-acetyl residue resistant to degradation by exoneuraminidase, whereby said isolated polysialic acid derivative is produced.
2. The method of claim 1 , wherein said ion exchange chromatography is anion exchange chromatography.
3. The method of claim 1 , wherein said isolated polysialic acid derivative has a degree of polymerization of about 2 to 10.
4. The method of claim 1 , wherein said isolated polysialic acid derivative is substantially unoxidized.
5. An isolated polysialic acid derivative produced according to claim 1 .
6. A pharmaceutical composition comprising the isolated polysialic acid derivative of claim 5 .
7. A composition comprising an isolated polysialic acid derivative, said isolated polysialic acid derivative being substantially unoxidized and comprising (i) mixture of N-acetyl sialic acid and de-N-acetyl sialic residues, and (ii) a non-reducing end de-N-acetyl residue that is resistant to degradation by exoneuraminidase, wherein said composition is substantially free of polysialic acid having a non-reducing end N-acetyl sialic acid residue.
8. The composition of claim 7 , wherein said isolated polysialic derivative comprises at least one dimer of de-N-acetyl sialic acid and N-acetyl sialic acid linked through a glycosidic bond selected from α(2→8) and a(2→9).
9. The composition of claim 7 , wherein said isolated polysialic derivative has a degree of polymerization of about 2-10.
10. The composition of claim 7 , wherein said isolated polysialic derivative has a degree of polymerization of about 2-5.
11. The composition of claim 7 , wherein said isolated polysialic derivative has a degree of polymerization of about 2-4.
12. The composition of claim 7 , wherein said isolated polysialic acid derivative has a degree of polymerization of about 2.
13. The composition of claim 7 , wherein said non-reducing end de-N-acetyl sialic acid residue is linked through a glycosidic bond to an N-acetyl sialic acid residue.
14. The composition of claim 7 , wherein said mixture comprises de-N-acetyl sialic residues in an amount of about 10%-60%.
15. The composition of claim 7 , wherein said isolated polysialic derivative has about 1 de-N-acetyl sialic residue per polysialic acid derivative chain.
16. The composition of claim 7 , wherein said isolated polysialic acid derivative comprises a conjugate.
17. The composition of claim 7 , wherein said de-N-acetyl sialic acid is neuraminic acid, and said N-acetyl sialic acid is N-acetyl neuraminic acid.
18. The composition of claim 7 , wherein at least one of said neuraminic acid and said N-acetyl neuraminic acid comprises at least one O-acetylated group.
19. The composition of claim 18 , wherein said isolated polysialic acid derivative is derivable from a capsular polysaccharide homopolymer of a bacterium selected from the group consisting of Escherichia coli K1 , Neisseria meningitidis Serogroup B, and Neisseria meningitidis Serogroup C.
20. A method of producing an aggregate comprising a polysialic acid derivative, the method comprising:
placing a substantially unoxidized and purified polysialic acid derivative under aggregating conditions so as to form an aggregate, said polysialic acid derivative comprising (i) a mixture of N-acetyl and de-N-acetyl residues, said de-N-acetyl residues comprising about 10%-80% of said mixture, and (ii) a non-reducing end resistant to degradation by exoneuraminidase; and
isolating said aggregate.
21. The method of claim 20 , wherein said aggregating conditions is heating or the addition of an aggregating excipient.
22. The method of claim 21 , where said heating is from about 30° C. to 70° C.
23. The method of claim 21 , wherein said aggregating excipient is aluminum hydroxide.
24. The method of claim 21 , wherein said aggregate is a particle.
25. The method of claim 21 , wherein said particle is microscopic.