# US 11,998,599 B2 · Polypeptide-antigen conjugates with non-natural amino acids

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- Number: US 11,998,599 B2
- Document ID: US11998599B2
- Status: Granted
- Kind: B2
- Application number: 15/859,251
- Filed: 2017-12-29
- Published: 2018-11-22
- Granted: 2024-06-04
- Earliest claimed: 2016-12-30
- Assignees: Vaxcyte, Inc.
- Applicants: Vaxcyte, Inc.
- Inventors: Jeffery Fairman; Jon H. Heinrichs; Wei Chan
- Web page: https://patentgrep.com/patent/US11998599B2

## CPC

- A61K 39/385: Haptens or antigens, bound to carriers
- A61K 39/092: {Streptococcus}
- A61K 47/6415: {Toxins or lectins, e.g. clostridial toxins or Pseudomonas exotoxins}
- A61K 47/646: {the entire peptide or protein drug conjugate elicits an immune response, e.g. conjugate vaccines}
- A61P 29/00: Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
- A61P 31/04: Antibacterial agents
- A61P 37/00: Drugs for immunological or allergic disorders
- A61K 2039/575: {humoral response}
- A61K 2039/6037: {Bacterial toxins, e.g. diphteria toxoid [DT], tetanus toxoid [TT]}
- A61K 2039/6087: {Polysaccharides; Lipopolysaccharides [LPS]}
- A61K 2039/627: {characterised by the linker}
- A61K 2039/70: {Multivalent vaccine}
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS

## Abstract

Methods for the production of immunogenic compositions containing a non-natural amino acid are disclosed. The non-natural amino acid can be a site for attachment of antigens, such as bacterial capsular polysaccharides, to make immunogenic conjugates. Bio-orthogonal attachment chemistry incorporated into the non-natural amino acids allows for more efficient and potent antigen presentation to the immune system, simplified purification, and more well-defined structure of these semi-synthetic immunogens.

## Claims

### Claim 1

A composition comprising at least 21 distinct crosslinked carrier protein-antigen conjugates, wherein each of the distinct crosslinked carrier protein-antigen conjugates comprises:
(i) an antigen, wherein the antigen is a distinct serotype of a capsular polysaccharide antigen from Streptococcus pneumoniae,
wherein there is a distinct crosslinked carrier protein-antigen conjugate for each of the capsular polysaccharide antigens of serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9V, 14, 18C, 19A, 19F and 23F and a distinct crosslinked carrier protein-antigen conjugate for each of at least eight additional capsular polysaccharide antigens of serotypes selected from the group consisting of 2, 6C, 8, 9N, 10A, 11A, 12F, 15A, 15B, 15C, 16F, 17F, 20, 20A, 20B, 22F, 23A, 23B, 24F, 24B, 31, 33F, 34, 35B, 35F and 38; and
(ii) a carrier protein, wherein the carrier protein comprises a polypeptide comprising at least one T-cell activating epitope and at least 95% sequence identity to SEQ ID NO: 1, wherein a 2-amino-3-(4-(azidomethyl)phenyl)propanoic acid (pAMF) non-natural amino acid (nnAA) is substituted for a naturally occurring amino acid at residues 34, 213, 245, 265, 386, and 527 numbered according to SEQ ID NO:1,
wherein the capsular polysaccharide antigen is conjugated to the nnAAs in the carrier protein.

### Claim 2 (depends on 1)

The composition of claim 1, wherein the at least 21 distinct crosslinked carder protein-antigen conjugates comprise
(i) at least 21 distinct crosslinked carrier protein-antigen conjugates wherein there is a distinct crosslinked carrier protein-antigen conjugate comprising a capsular polysaccharide for each of Streptococcus pneumoniae serotypes 1, 3, 4, 5, 6A, 6B, 7F, 8, 9V, 10A, 11A, 12F, 14, 15B, 18C, 19A, 19F, 22F, 23F, and 33F; or
(ii) at least 24 distinct crosslinked carrier protein-antigen conjugates wherein there is a distinct crosslinked carrier protein-antigen conjugate comprising a capsular polysaccharide for each of Streptococcus pneumonia serotypes 1, 2, 3, 4, 5, 6A, 6B, 7F, 8, 9V, 9N, 10A, 11A, 12F, 14, 15B, 17F, 18C, 19A, 19F, 20, 22F, 23F, and 33F.

### Claim 3 (depends on 2)

The composition of claim 2, wherein the antigen is linked to the carrier protein according to formula XI or XIa:
wherein
R1 is H, formyl, or at least one amino acid of the carrier protein;
R2 is OH or at least one amino acid of the carrier protein;
W is C;
y is 1;
n is at least 1; and
X is one monosaccharide within the capsular polysaccharide.

### Claim 4 (depends on 3)

The composition of claim 3, wherein n is 3-16.

### Claim 5 (depends on 2)

The composition according to claim 2, wherein the Streptococcus pneumoniae serotype 20 is serotype 20B.

### Claim 6 (depends on 5)

The composition according to claim 5, wherein the composition additionally comprises a distinct crosslinked carrier protein-antigen conjugate wherein the antigen is a capsular polysaccharide of Streptococcus pneumoniae serotype 6C or 7C.

### Claim 7 (depends on 2)

The composition according to claim 2, wherein the composition additionally comprises a distinct crosslinked carrier protein-antigen conjugate wherein the antigen is a capsular polysaccharide of Streptococcus pneumoniae serotype 6C or 7C.

### Claim 8 (depends on 1)

The composition of claim 1, wherein the overall (weight by weight) ratio of all serotype polysaccharides to carrier protein in the composition is at least 1.1.

### Claim 9 (depends on 1)

The composition of claim 1, wherein the overall (weight by weight) ratio of all serotype polysaccharides to carrier protein in the composition is at least 1.2.

### Claim 10 (depends on 1)

The composition of claim 1, wherein the antigen is linked to the carrier protein according to formula XI or XIa:
wherein
R1 is H, formyl, or at least one amino acid of the carrier protein;
R2 is OH or at least one amino acid of the carrier protein;
W is C;
y is 1;
n is at least 1; and
X is one monosaccharide within the capsular polysaccharide.

### Claim 11 (depends on 10)

The composition of claim 10, wherein n is 3-16.

### Claim 12 (depends on 1)

The composition of claim 1, wherein the Streptococcus pneumoniae capsular polysaccharides are conjugated to the nnAAs via a linking moiety, wherein the Streptococcus pneumoniae capsular polysaccharides comprise a cyclooctyne group and are conjugated to the pAMF nnAAs.

### Claim 13 (depends on 12)

The composition of claim 12, wherein the antigen is linked to the carrier protein according to formula XI or Xia:
wherein
R1 is H, formyl, or at least one amino acid of the carrier protein;
R2 is OH or at least one amino acid of the carrier protein;
W is C;
y is 1;
n is at least 1; and
X is one monosaccharide within the capsular polysaccharide.

### Claim 14 (depends on 13)

The composition of claim 13, wherein n is 3-16.

### Claim 15 (depends on 1)

The composition of claim 1, wherein the overall (weight by weight) ratio of all serotype polysaccharides to carrier protein in the composition is at least 1.0.

### Claim 16 (depends on 1)

The composition of claim 1, wherein there is less than about 25% (by weight) total free capsular polysaccharide present in the composition.

### Claim 17 (depends on 1)

The composition according to claim 1, wherein the composition additionally comprises a distinct crosslinked carrier protein-antigen conjugate wherein the antigen is a capsular polysaccharide of Streptococcus pneumoniae serotype 6C or 7C.