# US 10,695,415 B2 · Live attenuated oral vaccine against shigellosis and typhoid fever

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- Number: US 10,695,415 B2
- Document ID: US10695415B2
- Status: Granted
- Kind: B2
- Application number: 15/742,459
- Filed: 2016-07-06
- Published: 2018-09-20
- Granted: 2020-06-30
- Earliest claimed: 2015-07-06
- Assignees: Protein Potential, LLC
- Applicants: Protein Potential, LLC
- Inventors: Yun Wu; Dennis J. Kopecko; B. Kim Lee Sim; Stephen L. Hoffman
- Web page: https://patentgrep.com/patent/US10695415B2

## CPC

- A61K 39/0283: {Shigella}
- A61K 39/0275: {Salmonella}
- A61P 31/04: Antibacterial agents
- A61K 39/116: Polyvalent bacterial antigens
- A61K 2039/522: {avirulent or attenuated}
- A61K 2039/523: {expressing foreign proteins}
- A61K 2039/543: {intranasal}
- A61K 2039/70: {Multivalent vaccine}
- 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

Disclosed is the attenuated Salmonella typhi vaccine Ty21a utilized as a vector for Shigella and/or enterotoxogenic E. coli genes stably integrated in the Ty21a chromosome. These genes include a heterologous Shigella sonnei O-antigen biosynthetic gene region that comprises the wzz gene and expresses Shigella sonnei form 1 O-antigen, as well as a heterologous acid resistance biosynthetic gene system comprising a YbaS gene, which enables increased stability of the Ty21a vector at pH 2.5 relative to Ty21a without the integrated acid resistance biosynthetic gene system.

## Claims

### Claim 1

A transgenic Salmonella typhi Ty21a comprising a heterologous Shigella sonnei O-antigen biosynthetic gene region and a heterologous acid resistance biosynthetic gene system comprising one or more heterologous Shigella acid resistance enzymes,
wherein said heterologous Shigella sonnei O-antigen biosynthetic gene region and said heterologous acid resistance biosynthetic gene system both being integrated into the Salmonella typhi Ty21a chromosome,
wherein said heterologous Shigella sonnei O-antigen biosynthetic gene region comprises a wzz gene, which comprises (i) a DNA sequence that shares at least 95% sequence identity with the DNA sequence of nucleic acids 4,511,904 to 4,513,010 of SEQ ID NO: 4; or (ii) the DNA sequence of nucleic acids 4,511,904 to 4,513,010 of SEQ ID NO: 4, and
wherein:
a. a heterologous Shigella sonnei 0-antigen is stably expressed;
b. one or more heterologous Shigella acid resistance enzymes comprising a YbaS gene are stably expressed;
c. said transgenic Salmonella typhi Ty21a is more acid stable at pH 2.5 than Salmonella typhi Ty21a without the integrated acid resistance biosynthetic gene system;
d. an immune response is elicited against virulent Shigella sonnei challenge; and
e. an immune response is elicited against virulent Salmonella typhi challenge.

### Claim 2 (depends on 1)

The transgenic Ty21a of claim 1, wherein said heterologous Shigella sonnei 0-antigen biosynthetic gene region comprises (i) a DNA sequence that shares at least 95% sequence identity with the DNA sequence of nucleic acids 4,500,076 to 4,513,461 of SEQ ID NO: 4; or (ii) the DNA sequence of nucleic acids 4,500,076 to 4,513,461 of SEQ ID NO: 4.

### Claim 3 (depends on 2)

The transgenic Ty21a of claim 2, wherein the YbaS gene comprises (i) a DNA sequence that shares at least 95% sequence identity with the DNA sequence of nucleic acids 4,503,240 to 4,504,172 of SEQ ID NO: 1; or (ii) the DNA sequence of nucleic acids 4,503,240 to 4,504,172 of SEQ ID NO: 1.

### Claim 4 (depends on 3)

The transgenic Ty21a of claim 3, additionally comprising an O-antigen biosynthetic gene region from a bacterial strain selected from the group consisting of: Shigella dysenteriae, Shigella flexneri, Shigella boydii, Escherichia coli, Salmonela enterica serovars, Vibrio cholera serotypes, Enterobacter species, Yersinia species, and Pseudomonas species.

### Claim 5 (depends on 4)

A composition comprising the transgenic Ty21a of claim 4 and a carrier.

### Claim 6 (depends on 5)

A vaccine comprising the composition of claim 5 which is suitable for oral administration.

### Claim 7 (depends on 6)

The vaccine of claim 6, wherein administration of said vaccine to a human population reduces the incidence of shigellosis in said human population subsequently exposed to pathogenic Shigella sonnei.

### Claim 8 (depends on 6)

The vaccine of claim 6, wherein administration of said vaccine to a human population reduces the incidence of typhoid fever in said human population subsequently exposed to pathogenic S. typhi.

### Claim 9 (depends on 6)

The vaccine of claim 6, wherein administration of said vaccine to a human population reduces the incidence of both shigellosis and typhoid fever in said human population subsequently exposed to both pathogenic Shigella sonnei and pathogenic S. typhi.

### Claim 10 (depends on 1)

The transgenic Ty21a of claim 1, wherein the one or more heterologous Shigella acid resistance enzymes comprise GadA and GadC genes.

### Claim 11 (depends on 1)

The transgenic Ty21a of claim 1, wherein the one or more heterologous Shigella acid resistance enzymes comprise GadB and GadC genes.

### Claim 12 (depends on 1)

The transgenic Ty21a of claim 1, wherein the one or more heterologous Shigella acid resistance enzymes comprise GadA, GadB, and GadC genes.