# US 11,078,257 B2 · Recombinant gram negative bacteria and methods of generating and utilizing same

> Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt

- Number: US 11,078,257 B2
- Document ID: US11078257B2
- Status: Granted
- Kind: B2
- Application number: 16/321,093
- Filed: 2017-07-28
- Published: 2020-01-02
- Granted: 2021-08-03
- Earliest claimed: 2016-07-29
- Assignees: The Regents of the University of Michigan
- Applicants: The Regents of the University of Michigan
- Inventors: Ronald W. Woodard; Andrew Pratt
- Web page: https://patentgrep.com/patent/US11078257B2

## CPC

- C07K 16/1232: Escherichia (G)
- A61K 35/74: Bacteria (therapeutic use of a bacterial protein A61K38/00)
- A61K 39/0258: {Escherichia}
- A61K 39/05: {Actinobacteria, e.g. Actinomyces, Streptomyces, Nocardia, Bifidobacterium, Gardnerella}, Corynebacterium; Propionibacterium {(Mycobacterium A61K39/04)}
- A61K 39/095: Neisseria
- A61K 39/102: {Pasteurellales, e.g. Actinobacillus}, Pasteurella; Haemophilus
- A61K 39/105: {Delta proteobacteriales, e.g. Lawsonia; Epsilon proteobacteriales, e.g. campylobacter, helicobacter}
- A61K 39/118: Chlamydiaceae, e.g. Chlamydia trachomatis or Chlamydia psittaci
- C07K 14/22: from Neisseriaceae (F)
- C12N 1/205: Bacterial isolates
- A61K 39/00: Medicinal preparations containing antigens or antibodies (materials for immunoassay G01N33/53)
- A61K 2039/106: {Vibrio; Campylobacter; Not used, see subgroups}
- C12N 9/1048: {Glycosyltransferases (2.4)}
- C12R 2001/15: Corynebacterium
- C12R 2001/19: Escherichia coli
- 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
- C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00)
- 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)
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- C12R: INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- Y02A: TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE

## Abstract

The present invention provides novel, recombinant Gram-negative bacteria. In particular, the invention provides recombinant Gram-negative bacteria (e.g., E. coli) lacking genes involved in lipopolysaccharide (LPS, endotoxin) biosynthesis (e.g., lacking genes required for core oligosaccharide biosynthesis) and also provides recombinant Gram-negative bacteria lacking genes involved in LPS biosynthesis that contain one or more exogenous KDO transferases and/or one or more exogenous heptosyltransferases (e.g., from one or more types and/or strains of bacteria). The invention further provides methods of generating and utilizing (e.g., as or in an immunogenic composition (e.g., as or in an adjuvant and/or vaccine)) the recombinant Gram-negative bacteria therapeutic, preventative, and/or research applications.

## Claims

### Claim 1

A recombinant E. coli cell lacking all endogenous glycosyltransferases involved in the biosynthesis of the core oligosaccharide within lipopolysaccharide (LPS), wherein the cell comprises an exogenous KDO transferase.

### Claim 2 (depends on 1)

The recombinant E. coli cell of claim 1, wherein the cell lacks endogenous waaF, waaC, waaL, waaU, waaZ, waaY, waaJ, waaR, waaB, waaS, waaP, waaG, waaQ, and waaA genes.

### Claim 3 (depends on 1)

The recombinant E. coli cell of claim 1, wherein the cell possesses endogenous hldD and coaD genes.

### Claim 4 (depends on 1)

The recombinant E. coli cell of claim 1, wherein the cell generates sugar precursors of core oligonucleotide biosynthesis but lacks the capability to transfer the sugar precursors to the Lipid IVA molecule.

### Claim 5 (depends on 1)

The recombinant E. coli cell of claim 1, wherein the exogenous KDO transferase is from H. influenzae.

### Claim 6 (depends on 1)

The recombinant E. coli cell of claim 1, wherein the KDO transferase is from C. trachomatis.

### Claim 7 (depends on 1)

The recombinant E. coli cell of claim 1, wherein the KDO transferase is from Chlamydophila psittaci.

### Claim 8 (depends on 1)

The recombinant E. coli cell of claim 1, further comprising an exogenous heptosyltransferase.

### Claim 9 (depends on 8)

The recombinant E. coli cell of claim 8, wherein the heptosyltransferase is from Helicobacter pylori.

### Claim 10 (depends on 8)

The recombinant E. coli cell of claim 8, wherein the heptosyltransferase is from Campylobacter jejuni.

### Claim 11 (depends on 8)

The recombinant E. coli cell of claim 8, wherein the heptosyltransferase is from Aquifex aeolicus.

### Claim 12

A recombinant E. coli cell lacking all endogenous glycosyltransferases involved in the biosynthesis of the core oligosaccharide within lipopolysaccharide (LPS), wherein the cell comprises one or more exogenous KDO transferases.

### Claim 13 (depends on 12)

The recombinant E. coli cell of claim 12, further comprising one or more exogenous heptosyltransferases.