# US 11,027,005 B2 · Method for producing Hib conjugate vaccine using PRP with lowered molecular weight

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- Number: US 11,027,005 B2
- Document ID: US11027005B2
- Status: Granted
- Kind: B2
- Application number: 16/340,778
- Filed: 2017-10-11
- Published: 2020-02-13
- Granted: 2021-06-08
- Earliest claimed: 2016-10-20
- Assignees: KM Biologics Co., Ltd.
- Applicants: KM Biologies Co., Ltd.
- Inventors: Makoto Moriyama; Katsuhiko Fukada; Kazuhiko Kimachi; Yuki Mihara; Hiroshi Yonemura; Yoichiro Kino
- Web page: https://patentgrep.com/patent/US11027005B2

## CPC

- A61K 39/102: {Pasteurellales, e.g. Actinobacillus}, Pasteurella; Haemophilus
- A61K 39/08: Clostridium, e.g. Clostridium tetani
- A61K 39/385: Haptens or antigens, bound to carriers
- A61P 31/04: Antibacterial agents
- A61P 31/16: for influenza or rhinoviruses
- A61K 2039/6037: {Bacterial toxins, e.g. diphteria toxoid [DT], tetanus toxoid [TT]}
- Y02A 50/30: Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
- 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
- Y02A: TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE

## Abstract

A method for preparing a PRP conjugate having high storage stability and a method for producing a Hib conjugate vaccine are provided. A method for preparing PRP conjugate in accordance with the embodiments of the present invention is characterized by that the release of PRPs after preparing the PRP conjugate is suppressed by using PRP with a lowered molecular weight than native PRP for a coupling reaction between PRP and a carrier protein. Also, a Hib conjugate vaccine comprising as an antigen the PRP conjugate prepared by said method has excellent storage stability and maintain sufficient immunogenicity.

## Claims

### Claim 1

A method for preparing a polyribosylribitol phosphate (PRP) conjugate via a coupling reaction of PRP and a carrier protein wherein the method is characterized by that the release of PRPs after preparing the PRP conjugate is suppressed by using PRP with a lowered molecular weight than native PRP, wherein the PRP conjugate is stored in a solution of pH 5.4 to 6.3 after preparation of the PRP conjugate.

### Claim 2 (depends on 1)

The method of claim 1 wherein a released amount of PRPs in a stress test in a solution at pH 5.4 to 6.3 at 37° C. for 4 weeks is less than 50%.

### Claim 3 (depends on 1)

The method of claim 1 wherein PRP with a lowered molecular weight is obtained by physical fracture.

### Claim 4 (depends on 1)

The method of claim 1 wherein PRP with a lowered molecular weight is obtained by hydrolysis with an acid or an alkali.

### Claim 5 (depends on 1)

The method of claim 1 wherein PRP with a lowered molecular weight has a molecular weight of 80 to 150 kDa.

### Claim 6 (depends on 1)

The method of claim 1 wherein PRP and the carrier protein is subject to the coupling reaction for preparing the PRP conjugate at a weight ratio of from 2:1 to 4.1.

### Claim 7 (depends on 6)

The method of claim 6 wherein PRP and the carrier protein are subject to the coupling reaction at a weight ratio of 4:1.

### Claim 8 (depends on 1)

The method of claim 1 wherein the method comprises a step of activating PRP with a lowered molecular weight using 1-cyano-4-(dimethylamino) pyridinium tetrafluoroborate (CDAP) before the coupling reaction for preparing the PRP conjugate.

### Claim 9 (depends on 1)

The method of claim 1 wherein the carrier protein is tetanus toxoid.

### Claim 10 (depends on 9)

The method of claim 9 wherein a purity of tetanus toxoid is 2,500 to 3,500 LF/mg PN.

### Claim 11 (depends on 10)

The method of claim 10 wherein a purity of tetanus toxoid is 2,900 to 3,300 LF/mg PN.

### Claim 12 (depends on 1)

A method for producing a Haemophilus influenzae type b (Hib) conjugate vaccine comprising the method as set forth in claim 1.