Chimeric receptor binding proteins for use in bacterial delivery vehicles

Patent metadata

Field	Value
Publication number	US-11512116-B2
Application number	US-202117527754-A
Country	US
Kind code	B2
Filing date	Nov 16, 2021
Priority date	Nov 27, 2018
Publication date	Nov 29, 2022
Grant date	Nov 29, 2022

Abstract

The present disclosure relates generally to bacterial delivery vehicles for use in efficient transfer of a desired payload into a target bacterial cell. More specifically, the present disclosure relates to bacterial delivery vehicles with desired host ranges based on the presence of a chimeric receptor binding protein (RBP) composed of a fusion between the N-terminal region of a RBP derived from a lambda-like bacteriophage and the C-terminal region of a different RBP.

First claim

What is claimed is:

1. A method for treating a bacterial infection comprising administering to a subject having a bacterial infection in need of treatment the pharmaceutical or veterinary composition comprising a pharmaceutically acceptable carrier and one or more bacterial delivery vehicles comprising a chimeric receptor binding protein (RBP)
wherein the chimeric RBP comprises a fusion between the N-terminal domain of a RBP from a lambda-like or lambda bacteriophage, wherein said lambda-like bacteriophage comprises amino acid sequence homology of around 35% identity for 45 amino acids or more, around 50% identity for 30 amino acids or more, or around 90% identity for 18 amino acids or more in one or more of three amino acid regions ranging from positions 1-150, 320-460, and 495-560 with reference to the lambda bacteriophage stf sequence of SEQ ID NO: 1, and the C-terminal domain of a different RBP, and
wherein said N-terminal domain is fused to said C-terminal domain within one of the amino acids acid regions selected from positions 1-150, 320-460 or 495-560 of the N-terminal RBP with reference to the lambda stf sequence (SEQ ID NO: 1), wherein said region of fusion within the N-terminal RBP from positions 1-150, 320-460 or 495-560 comprises amino acid sequence homology of around 35% identity for 45 amino acids or more, around 50% identity for 30 amino acids or more, or around 90% identity for 18 amino acids or more with reference to the lambda bacteriophage stf sequence of SEQ ID NO: 1.

2. The method of claim 1 , wherein said different RBP is derived from any bacteriophage or bacteriocin.

3. The method of claim 1 , wherein said N-terminal domain of the chimeric RBP is fused to said C-terminal domain within one of the amino acid regions selected from positions 80-150, 320-460, or 495-560 of the N-terminal RBP.

4. The method of claim 1 , wherein the N-terminal domain and the C-terminal domain are fused within said region at an insertion site having at least 80% identity with insertion site selected from the group consisting of amino acids SAGDAS (SEQ ID NO:248), ADAKKS (SEQ ID NO: 249), MDETNR (SEQ ID NO: 250), SASAAA (SEQ ID NO: 251), and GAGENS (SEQ ID NO: 252).

5. The method of claim 1 , wherein the chimeric RBP comprises the amino acid sequence of SEQ ID NO: 2, 4, 7, 9, 12, 15, 17, 20, 23, 24, 25, 27, 29, 31, 33, 35, 37, 39, 41, 42, 44, 46, 47, 48, 49, 50, 51, 52, 53, 56, 59, 123-129, 130, 131, 132, 135, 138, 139, 142, 145, 148, 151, 216, 219, 221, 223, 227, 230, 232, 234, 236, 238, 240, 243, 245 or 246.

6. The method of claim 1 , wherein the C-terminal domain of the different RBP has a depolymerase activity against an encapsulated bacterial strain.

7. The method of claim 1 , wherein the bacterial delivery vehicle further comprises a nucleic acid payload encoding a protein of interest or a nucleic acid of interest.

8. The method of claim 7 , wherein the nucleic acid of interest is selected from the group consisting of Cas nuclease gene, a Cas9 nuclease gene, a guide RNA, a CRISPR locus, a toxin gene, a gene expressing an enzyme, a TALEN, a ZFN, a meganuclease, a recombinase, a bacterial receptor gene, a membrane protein gene, a structural protein gene, a secreted protein gene, a gene expressing resistance to an antibiotic or to a drug in general, a gene expressing a toxic protein or a toxic factor, and a gene expressing a virulence protein or a virulence factor, or any combination thereof.

9. The method of claim 8 , wherein the enzyme is a nuclease or a kinase.

10. The method of claim 7 , wherein the protein of interest is a nuclease that targets cleavage of a host bacterial cell genome or a host bacterial cell plasmid.

11. The method of claim 10 , wherein the cleavage occurs in an antibiotic resistant gene.

12. The method of claim 7 , wherein the nucleic acid payload encodes a therapeutic protein.

13. The method of claim 7 , wherein the nucleic acid payload encodes an anti-sense nucleic acid molecule.

14. A method for reducing the amount of virulent and/or antibiotic resistant bacteria in a bacterial population comprising contacting the bacterial population with a bacterial delivery vehicle comprising a chimeric receptor binding protein (RBP), wherein the chimeric RBP comprises a fusion between the N-terminal domain of a RBP from a lambda-like or lambda bacteriophage, wherein said lambda-like bacteriophage comprises amino acid sequence homology of around 35% identity for 45 amino acids or more, around 50% identity for 30 amino acids or more, or around 90% identity for 18 amino acids or more in one or more of three amino acid regions ranging from positions 1-150, 320-460, and 495-560 with reference to the lambda bacteriophage stf sequence of SEQ ID NO: 1, and the C-terminal domain of a different RBP, and
wherein said N-terminal domain is fused to said C-terminal domain within one of the amino acids acid regions selected from positions 1-150, 320-460 or 495-560 of the N-terminal RBP with reference to the lambda stf sequence (SEQ ID NO: 1), wherein said region of fusion within the N-terminal RBP from positions 1-150, 320-460 or 495-560 comprises amino acid sequence homology of around 35% identity for 45 amino acids or more, around 50% identity for 30 amino acids or more, or around 90% identity for 18 amino acids or more with reference to the lambda bacteriophage stf sequence of SEQ ID NO: 1.

15. The method of claim 14 , wherein said different RBP is derived from any bacteriophage or bacteriocin.

16. The method of claim 14 , wherein said N-terminal domain of the chimeric RBP is fused to said C-terminal domain within one of the amino acid regions selected from positions 80-150, 320-460, or 495-560 of the N-terminal RBP.

17. The method of claim 14 , wherein the N-terminal domain and the C-terminal domain are fused within said region at an insertion site having at least 80% identity with insertion site selected from the group consisting of amino acids SAGDAS (SEQ ID NO:248), ADAKKS (SEQ ID NO: 249), MDETNR (SEQ ID NO: 250), SASAAA (SEQ ID NO: 251), and GAGENS (SEQ ID NO: 252).

18. The method of claim 14 , wherein the chimeric RBP comprises the amino acid sequence of SEQ ID NO: 2, 4, 7, 9, 12, 15, 17, 20, 23, 24, 25, 27, 29, 31, 33, 35, 37, 39, 41, 42, 44, 46, 47, 48, 49, 50, 51, 52, 53, 56, 59, 123-129, 130, 131, 132, 135, 138, 139, 142, 145, 148, 151, 216, 219, 221, 223, 227, 230, 232, 234, 236, 238, 240, 243, 245 or 246.

19. The method of claim 14 , wherein the C-terminal domain of the different RBP has a depolymerase activity against an encapsulated bacterial strain.

20. The method of claim 14 , wherein the bacterial delivery vehicle further comprises a nucleic acid payload encoding a protein of interest or a nucleic acid of interest.

21. The method of claim 20 , wherein the nucleic acid of interest is selected from the group consisting of Cas nuclease gene, a Cas9 nuclease gene, a guide RNA, a CRISPR locus, a toxin gene, a gene expressing an enzyme, a TALEN, a ZFN, a meganuclease, a recombinase, a bacterial receptor gene, a membrane protein gene, a structural protein gene, a secreted protein gene, a gene expressing resistance to an antibiotic or to a drug in general, a gene expressing a toxic protein or a toxic factor, and a gene expressing a virulence protein or a virulence factor, or any combination thereof.

22. The method of claim 21 , wherein the enzyme is a nuclease or a kinase.

23. The method of claim 20 , wherein the protein of interest is a nuclease that targets cl

Assignees

• Eligo Bioscience

Inventors

• Fernandez Rodriguez Jesus

Classifications

• C12N2795/10343

  viral genome or elements thereof as genetic vector · CPC title

• A61P31/00

  Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics · CPC title

• C12N2795/10122

  New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes · CPC title

• A61K48/00

  Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy · CPC title

• C07K2319/00

  Fusion polypeptide · CPC title