# US 9,056,899 B2 · Engineered bacteriophages as adjuvants for antimicrobial agents and compositions and methods of use thereof

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- Number: US 9,056,899 B2
- Document ID: US9056899B2
- Status: Granted
- Kind: B2
- Application number: 12/812,212
- Filed: 2009-01-12
- Published: 2010-12-23
- Granted: 2015-06-16
- Earliest claimed: 2008-01-10
- Assignees: Trustees of Boston University; Massachusetts Institute of Technology
- Applicants: James J Collins; Timothy Kuan-Ta Lu
- Inventors: James J Collins; Timothy Kuan-Ta Lu
- Web page: https://patentgrep.com/patent/US9056899B2

## CPC

- C07K 14/005: from viruses
- A61K 35/76: Viruses; Subviral particles; Bacteriophages
- A61P 31/04: Antibacterial agents
- C12N 15/113: Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; {Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing (when used in plants C12N15/8218)}
- C12N 2320/32: Special delivery means, e.g. tissue-specific
- C12N 2330/30: chemically synthesised
- C12N 2795/14122: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
- C12N 2795/14132: Use of virus as therapeutic agent, other than vaccine, e.g. as cytolytic agent
- 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)
- 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)

## Abstract

The present invention relates to the treatment and prevention of bacteria and bacterial infections. In particular, the present invention relates to engineered bacteriophages used in combination with antimicrobial agents to potentiate the antimicrobial effect and bacterial killing by the antimicrobial agent. The present invention generally relates to methods and compositions comprising engineered bacteriophages and antimicrobial agents for the treatment of bacteria, and more particularly to bacteriophages comprising agents that inhibit antibiotic resistance genes and/or cell survival genes, and/or bacteriophages comprising repressors of SOS response genes or inhibitors of antimicrobial defense genes and/or expressing an agent which increases the sensitivity of bacteria to an antimicrobial agent in combination with at least one antimicrobial agent, and their use thereof.

## Claims

### Claim 1

An engineered bacteriophage comprising a nucleic acid operatively linked to a promoter, wherein the nucleic acid encodes:
a bacterial porin or porin-like protein of the OMP superfamily.

### Claim 2 (depends on 1)

The bacteriophage of claim 1, wherein the porin is ompF.

### Claim 3 (depends on 1)

A method to inhibit or eliminate a bacterial infection comprising administering to a surface infected with bacteria, the engineered bacteriophage of claim 1 and at least one antimicrobial agent.

### Claim 4 (depends on 3)

The method of claim 3, wherein the administration of the bacteriophage occurs simultaneously or prior to, or after administration of the antimicrobial agent.

### Claim 5 (depends on 3)

The method of claim 3, wherein the antimicrobial agent is selected from a group consisting of: quinolone, ampicillin, aminoglycoside, ciproflaxacin, levofloxacin, ofloxacin, gatifloxacin, norfloxacin, lomefloxacin, trovafloxacin, moxifloxacin, sparfloxacin, gemifloxacin, pazufloxacin, amikacin, gentamycin, gentamicin, tobramycin, netromycin, streptomycin, kanamycin, paromomycin, neomycin, β-lactam, penicillin, ampicillin, penicillin derivatives, cephalosporins, monobactams, carbapenems, β-lactamase inhibitors and variants or analogues thereof.

### Claim 6 (depends on 3)

The method of claim 3, wherein the bacteria is present in a subject.

### Claim 7 (depends on 6)

The method of claim 6, wherein the subject is a mammal.

### Claim 8 (depends on 7)

The method of claim 7, wherein the mammal is a human.

### Claim 9 (depends on 3)

The method of claim 3, wherein the bacteria is in a biofilm.

### Claim 10 (depends on 1)

A composition comprising the engineered bacteriophage of claim 1 and at least one antimicrobial agent.

### Claim 11 (depends on 1)

A kit comprising an engineered bacteriophage of claim 1, and at least one antimicrobial agent.

### Claim 12 (depends on 10)

The composition of claim 10, wherein the antimicrobial agent is selected from a group consisting of: quinolone, ampicillin, aminoglycoside, ciproflaxacin, levofloxacin, ofloxacin, gatifloxacin, norfloxacin, lomefloxacin, trovafloxacin, moxifloxacin, sparfloxacin, gemifloxacin, pazufloxacin, amikacin, gentamycin, gentamicin, tobramycin, netromycin, streptomycin, kanamycin, paromomycin, neomycin, β-lactam, penicillin, ampicillin, penicillin derivatives, cephalosporins, monobactams, carbapenems, β-lactamase inhibitors and variants or analogues thereof.

### Claim 13 (depends on 11)

The kit of claim 11, wherein the antimicrobial agent is selected from a group consisting of: quinolone, ampicillin, aminoglycoside, ciproflaxacin, levofloxacin, ofloxacin, gatifloxacin, norfloxacin, lomefloxacin, trovafloxacin, moxifloxacin, sparfloxacin, gemifloxacin, pazufloxacin, amikacin, gentamycin, gentamicin, tobramycin, netromycin, streptomycin, kanamycin, paromomycin, neomycin, β-lactam, penicillin, ampicillin, penicillin derivatives, cephalosporins, monobactams, carbapenems, β-lactamase inhibitors and variants or analogues thereof.

### Claim 14 (depends on 1)

The engineered bacteriophage of claim 1, wherein the engineered bacteriophage infects one or more of Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, or Enterococcus faecalis.

### Claim 15 (depends on 1)

The engineered phage of claim 1, wherein the phage is lysogenic.

### Claim 16 (depends on 1)

The engineered phage of claim 1, wherein the phage is lytic.

### Claim 17 (depends on 1)

The engineered phage of claim 1, wherein the phage is an engineered lambda phage, M13 phage, T7 phage, T3 phage, T2 phage, T4 phage, RB69 phage, Pf1 phage, Pf4 phage, phage B40-8, or coliphage MS-2.

### Claim 18 (depends on 1)

The engineered phage of claim 1, wherein the engineered phage increases susceptibility of the bacteria to one or more antibiotic agents selected from a glycopeptide, carbapenum, cephalosporin, fluoroquinolone, quinolone, amino glycoside, β-lactam, sulphonamide, oxazolidinone, and tetracyclines.

### Claim 19 (depends on 18)

The engineered phage of claim 18, wherein the engineered phage increases susceptibility of the bacteria to one or more of an aminoglycoside, quinolone, and β-lactam.

### Claim 20 (depends on 1)

The engineered phage of claim 1, wherein the nucleic acid encodes a bacterial porin or porin-like protein of the OMP superfamily selected from the group consisting of ompA, ompC, ompF, ompG, ompL, ompN, ompW, pgaA, phoE, tolE, tolC, tsx or yncD.