US4356170A
Expired / Lapsed
US
⏱️ 0.0 years remaining
Immunogenic polysaccharide-protein conjugates
Assignee: National Research Council Canada
•
Filing: 1981-05-27
•
Granted: 1982-10-26
•
Expires: 2001-05-27
Abstract
Abstract Antigenic polysaccharides are modified to generate a terminally-located aldehyde group by controlled oxidation of vicinal hydroxyl groups, e.g. of unlinked terminal non-reducing sialic acid residues. In some cases where there is a reducing end group, e.g. of the type N-acetylmannosamine residue, it can be made into the most susceptible site for oxidation by initially reducing it to its open chain hydroxyl form, e.g. N-acetylmannosaminitol. The vicinal hydroxyl oxidation is controlled to yield a reactive aldehyde group which is then covalently linked to a free amino group of a selected protein by reductive amination. The resulting polysaccharide-protein conjugates are soluble and have been found to have enhanced antigenicity compared to the polysaccharide alone. This terminal aldehyde:free amine group reductive amination can be applied to various polysaccharide antigens and various well-tolerated proteins, preferably protein immunogens. For example, meningococcal group A, B and C polysaccharides have been linked to tetanus toxoid to give soluble conjugates which have been found to have advantageous immunogenic properties.
Inventors
- 👤 Harold J. Jennings
- 👤 Czeslaw Lugowski
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-23 09:44
CPC Classifications
A; A61; A61K; A61K39/00; A61K39/385; A61K2039/60; A61K2039/6031; A61K2039/6037; Y; Y10; Y10S; Y10S530/00; Y10S530/806; Y10S530/82; Y10S530/825
Claims Analysis
Total Claims: 28
Claim 1
Independent Claim
99 words
A method of preparing antigenic polysaccharide: protein conjugates, comprising: (a) providing an antigenic polysaccharide which has reactive vicinal hydroxyl groups in a terminal portion of the molecule and has a MW above about 2000; (b) subjecting said vicinal hydroxyl groups at a terminal location to controlled oxidation only sufficient to generate a terminal reactive aldehyde group therefrom on the polysaccharide; (c) selecting a physiologically-tolerated protein having a free amino group and reacting said amino group with said aldehyde group by reductive amination, to covalently link said polysaccharide and protein; and (d) recovering a substantially non-crosslinked, soluble, antigenic polysaccharide:protein conjugate.
Claim 2
Independent Claim
99 words
A method of preparing antigenic polysaccharide: protein conjugates, comprising: (a) providing an antigenic polysaccharide which has reactive vicinal hydroxyl groups in a terminal portion of the molecule and has a MW above about 2000; (b) subjecting said vicinal hydroxyl groups at a terminal location to controlled oxidation only sufficient to generate a terminal reactive aldehyde group therefrom on the polysaccharide; (c) selecting a physiologically-tolerated protein having a free amino group and reacting said amino group with said aldehyde group by reductive amination, to covalently link said polysaccharide and protein; and (d) recovering a substantially non-crosslinked, soluble, antigenic polysaccharide:protein conjugate.
Claim 3
Dependent Claim
25 words
The method of claim 1 wherein the antigenic polysaccharide is selected from the group derived from meningococci, Haemophilus influenza, pneumococci, β-hemolytic streptococci, and E. coli.
Claim 4
Dependent Claim
36 words
The method of claim 1 wherein the protein is selected from the group consisting of tetanus toxoid, diphtheria toxoid, and immunogenic proteins derived from bacteria selected from β-hemolytic streptococci, Haemophilus influenza, meningococci, pneumococci, and E. coli.
Claim 5
Dependent Claim
33 words
The method of claim 1 wherein the polysaccharide initially has no terminal vicinal hydroxyl groups but has a terminal reducing sugar residue which is reduced to form reactive vicinal hydroxyl groups in (a).
Claim 6
Dependent Claim
12 words
The method of claim 4 wherein the sugar residue is N-acetylmannosamine residue.
Claim 7
Dependent Claim
19 words
The method of claim 4 wherein the reduction is effected using sodium borohydride at a pH of about 7.5-10.
Claim 8
Dependent Claim
11 words
The method of claim 6 wherein the pH is about 8-9.
Claim 9
Dependent Claim
20 words
The method of claim 1 wherein the controlled oxidation in (b) is effected using periodate reagent for a limited time.
Claim 10
Dependent Claim
22 words
The method of claim 8 wherein the polysaccharide is derived from meningococci, and the time for controlled oxidation is within 10-15 minutes.
Claim 11
Dependent Claim
68 words
The method of claim 1 wherein meningococcal group A, B or C polysaccharides having terminal vicinal hydroxyl groups are utilized and are oxidized sufficiently to convert only the terminal vicinal hydroxyl groups to a terminal aldehyde group, reductive amination carried out to link covalently the terminal aldehyde group to an amino group on the selected protein tetanus toxoid molecule, and recovering a soluble conjugate thereof of enhanced antigenicity.
Claim 12
Dependent Claim
34 words
An antigenic-polysaccharide:protein conjugate wherein the polysaccharide and protein are covalently linked through a ##STR3## linkage to a terminal portion of the polysaccharide without significant cross-linking, said antigenic polysaccharide having a MW above about 2000.
Claim 13
Dependent Claim
34 words
An antigenic-polysaccharide:protein conjugate wherein the polysaccharide and protein are covalently linked through a ##STR3## linkage to a terminal portion of the polysaccharide without significant cross-linking, said antigenic polysaccharide having a MW above about 2000.
Claim 14
Dependent Claim
25 words
The conjugate of claim 11 wherein the antigenic polysaccharide is selected from the group derived from meningococci, Haemophilus influenza, pneumococci, β-hemolytic streptococci, and E. coli.
Claim 15
Dependent Claim
36 words
The conjugate of claim 11 wherein the protein is selected from the group consisting of tetanus toxoid, diphtheria toxoid, and immunogenic proteins derived from bacteria selected from β-hemolytic streptococci, Haemophilus influenza, meningococci, pneumococci and E. coli.
Claim 16
Dependent Claim
25 words
The conjugate of claim 11 wherein the antigenic polysaccharide is selected from meningococcal Group A, B or C polysaccharides and the protein is tetanus toxoid.
Claim 17
Dependent Claim
20 words
The conjugate of claim 11 wherein the antigenic polysaccharide is derived from Haemophilus influenza and the protein is tetanus toxoid.
Claim 18
Dependent Claim
19 words
The conjugate of claim 11 wherein said linkage is through an end-group terminal reducing sugar moiety on the polysaccharide.
Claim 19
Dependent Claim
12 words
The conjugate of claim 16 wherein said reducing sugar moiety is N-acetylmannosaminitol.
Claim 20
Dependent Claim
17 words
The conjugate of claim 11 wherein said linkage is through a lysine amino group on the protein.
Claim 21
Dependent Claim
16 words
The conjugate of claim 11 wherein the polysaccharide and protein are derived from the same bacteria.
Claim 22
Dependent Claim
29 words
A method of immunizing against infection susceptible humans or animals comprising, administering a vaccine comprising the conjugate of claim 11 in an immunogenic amount by intramuscular or subcutaneous injection.
Claim 23
Dependent Claim
25 words
The method of claim 20 wherein the dosage of the conjugate is equivalent to from about 5 to about 25 micrograms for the human infant.
Claim 24
Dependent Claim
32 words
The method of claim 21 wherein human infants are immunized with a vaccine comprising at least one of (a) the conjugate of meningococcal polysaccharide, and (b) the conjugate of H. influenza polysaccharide.
Claim 25
Dependent Claim
12 words
A vaccine comprising at least one conjugate as defined in claim 11.
Claim 26
Dependent Claim
26 words
A vaccine for infants comprising the conjugate of claim 11 wherein the initial non-conjugated polysaccharide is of the type for which the immune response is non-thymus-controlled.
Claim 27
Dependent Claim
24 words
A human infant vaccine comprising the conjugate of claim 11 wherein the polysaccharide comprises at least one of meningococcal polysaccharide and Haemophilus influenza polysaccharide.
Claim 28
Dependent Claim
32 words
A vaccine as in claim 24 in a dosage unit form wherein the conjugate is present in the equivalent of from about 5 to about 25 micrograms based on the human infant.
References Cited (Backward Citations)
US4003792A
Cited
US3847890A
Cited
US4185090A
Cited
US3947352A
Cited
US4125492A
Cited
US4093607A
Cited
US4203893A
Cited
US4303638A
Cited
US4308254A
Cited
US4259232A
Cited
US4310514A
Cited
US4315913A
Cited
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1985-10-25 | 1986-04-26 | 1986-10-26 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1989-10-25 | 1990-04-26 | 1990-10-26 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1993-10-25 | 1994-04-26 | 1994-10-26 | $7700.00 | Window Closed / Lapsed |