# US 10,500,264 B2 · Development of mutations useful for attenuating dengue viruses and chimeric dengue viruses

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- Number: US 10,500,264 B2
- Document ID: US10500264B2
- Status: Granted
- Kind: B2
- Application number: 15/650,482
- Filed: 2017-07-14
- Published: 2018-01-11
- Granted: 2019-12-10
- Earliest claimed: 2001-05-22
- Assignees: The United States of America, as represented by the Secretary, Department of Health and Human Services
- Applicants: The United States of America, as represented by the Secretary, Dept. of Health and Human Services
- Inventors: Stephen S. Whitehead; Brian R. Murphy; Kathryn A. Hanley; Joseph E. Blaney; Ching-Juh Lai
- Web page: https://patentgrep.com/patent/US10500264B2

## CPC

- A61K 39/12: Viral antigens
- A61P 31/14: for RNA viruses
- A61P 37/04: Immunostimulants
- C12N 7/00: Viruses; Bacteriophages; Compositions thereof; Preparation or purification thereof (preparing medicinal viral antigen or antibody compositions, e.g. virus vaccines, A61K39/00)
- C12N 7/04: Inactivation or attenuation; Producing viral sub-units
- A61K 39/295: Polyvalent viral antigens (vaccinia virus or variola virus A61K39/285); Mixtures of viral and bacterial antigens
- A61K 2039/5252: {inactivated (killed)}
- A61K 2039/5254: {avirulent or attenuated}
- A61K 2039/5256: {expressing foreign proteins}
- A61K 2039/54: {characterised by the route of administration}
- A61K 2039/575: {humoral response}
- A61K 2039/70: {Multivalent vaccine}
- C12N 2770/24121: Viruses as such, e.g. new isolates, mutants or their genomic sequences
- C12N 2770/24134: Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
- C12N 2770/24151: Methods of production or purification of viral material
- C12N 2770/24161: Methods of inactivation or attenuation
- C12N 2770/24162: by genetic engineering
- C12N 2770/24163: by chemical treatment
- C12N 2770/24171: Demonstrated in vivo effect
- C12Q 1/701: {Specific hybridization probes}
- C12Q 2600/156: Polymorphic or mutational markers
- G01N 2333/18: Togaviridae; Flaviviridae
- G01N 2500/10: involving cells
- 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
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)
- Y02A: TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE

## Abstract

A menu of mutations was developed that is useful in fine-tuning the attenuation and growth characteristics of dengue virus vaccines.

## Claims

### Claim 1

A dengue virus comprising a U to C mutation at nucleotide 10634 of the 3′ untranslated region, wherein the numbering is based upon the prototypic isolate DEN4 Dominica 1981 and wherein the virus has a phenotype comprising temperature sensitivity in Vero cells or human liver cell line HuH-7 and/or attenuation in mice.

### Claim 2 (depends on 1)

The dengue virus of claim 1, further comprising the 430 mutation.

### Claim 3 (depends on 1)

The dengue virus of claim 1, wherein the dengue virus is a dengue virus type 1, a dengue virus type 2, a dengue virus type 3, or a dengue virus type 4.

### Claim 4 (depends on 1)

The dengue virus of claim 1, wherein the dengue virus is a chimeric virus.

### Claim 5 (depends on 4)

The chimeric virus of claim 4 having a dengue 1 backbone, a dengue 2 backbone, a dengue 3 backbone, or a dengue 4 backbone.

### Claim 6 (depends on 1)

A pharmaceutical composition comprising a pharmacologically acceptable vehicle and a dengue virus according to claim 1.

### Claim 7 (depends on 6)

A kit comprising a pharmaceutical composition according to claim 6 in a pack or dispenser device and instructions for administration.

### Claim 8 (depends on 1)

A method of producing neutralizing antibodies against dengue virus in a subject comprising administering a therapeutically effective amount of a pharmaceutical composition comprising a pharmacologically acceptable vehicle and the dengue virus according to claim 1 to the subject.

### Claim 9 (depends on 8)

The method of claim 8, wherein administration is by subcutaneous, intradermal, or intramuscular injection.

### Claim 10 (depends on 1)

An immunogenic composition comprising a pharmacologically acceptable vehicle and a dengue virus according to claim 1.

### Claim 11 (depends on 10)

The immunogenic composition of claim 10 in unit dosage form having from about 102-109 plaque forming units (PFU)/ml.

### Claim 12 (depends on 10)

The immunogenic composition of claim 10 in unit dosage form having from about 0.1 to 50 μg of E protein/ml.

### Claim 13 (depends on 1)

A method of making a pharmaceutical composition comprising:
isolating the dengue virus according to claim 1; and
combining the isolated dengue virus with a pharmacologically acceptable vehicle.