Propagator cells and methods for propagating phage, in particular for delivering CRISPR-Cas components via probiotic organisms

Patent metadata

Field	Value
Publication number	US-12516297-B2
Application number	US-202318539066-A
Country	US
Kind code	B2
Filing date	Dec 13, 2023
Priority date	Aug 8, 2017
Publication date	Jan 6, 2026
Grant date	Jan 6, 2026

Abstract

The invention provides propagator cells and methods for propagating phage and transduction particles.

First claim

The invention claimed is:

1 . A method of producing a population of transduction particles comprising nucleic acid packaged by phage coat proteins, wherein the particles are capable of infecting host cells of a first bacterial species or strain by binding a cell-surface receptor comprised by bacteria of the first species or strain, whereby host cells are transduced with the nucleic acid, the method comprising
(a) providing a population of second bacterial cells comprising the receptor on the surface of the second cells, wherein the second cells are of a second species or strain, wherein the second species or strain is different from the first species or strain, and wherein the second cells are capable of producing copies of the nucleic acid;
(b) producing in the second bacterial cells phage coat proteins of a phage that is capable of binding to the receptor comprised by the second bacterial cells; and
(c) allowing the phage coat proteins in the second bacterial cells to package copies of the nucleic acid, thereby producing the population of particles;
wherein the particles are non-replicative transduction particles.

2 . The method of claim 1 , wherein the particles comprise a nucleotide sequence encoding crRNAs that are operable with a Cas in bacteria of the host cell strain or species to form an active CRISPR/Cas system that is capable of targeting one or more protospacer nucleotide sequences, wherein each target sequence is comprised by the genome of the host cells, whereby the crRNAs guide the Cas in host cells to modify the target sequence, thereby killing host cells or reducing host cell population growth.

3 . The method of claim 2 , wherein the genome of each second bacterial cell does not comprise the target sequence.

4 . The method of claim 2 , wherein the crRNAs comprise repeat sequences that are not operable with a Cas of the second cells.

5 . The method of claim 2 , wherein the nucleotide sequence is operably connected with a promoter for transcription of crRNAs in bacteria of the host species or strain, but not in the second species or strain.

6 . The method of claim 2 , wherein the host cells and the second cells are different strains of the same species.

7 . The method of claim 2 , wherein bacteria of the host species or strain comprise an anti-phage toxin or mechanism for reducing the propagation of particles that infect host bacteria, wherein the second bacteria do not comprise the toxin or mechanism.

8 . The method of claim 2 , wherein bacteria of the host species or strain comprise a CRISPR/Cas system that is active for reducing the propagation of particles that infect host bacteria, wherein the second bacteria do not comprise the CRISPR/Cas system.

9 . The method of claim 2 , wherein the second bacterial cells are engineered to produce the receptor, wherein wild-type bacteria of the second species or strain do not produce the receptor.

10 . The method of claim 2 , wherein the second cells are Escherichia coli cells.

11 . The method of claim 2 , wherein the host cells and the second cells are of the same species.

12 . The method of claim 11 , wherein the strain of host cells is a human pathogenic strain and the second cell strain is not a human pathogenic strain.

13 . The method of claim 2 , wherein the receptor is selected from the group consisting of lipopolysaccharides, teichoic acids, proteins and flagella.

14 . The method of claim 2 , wherein the receptor comprises an O-antigen of the host cells.

15 . The method of claim 2 , wherein the particles are operable to express an endolysin or holin in the second cells.

16 . The method of claim 13 , wherein the receptor comprises a teichoic acid moiety that is the product of the action of one or more enzymes in the second cell, wherein the genome of the second cell comprises one or more expressible nucleotide sequences encoding the one or more enzymes, wherein wild-type cells of the second species or strain do not comprise the expressible nucleotide sequences.

17 . The method of claim 16 , wherein the enzymes are selected from the group consisting of TarO, TarA, TarB, TarF, TarK, and TarL.

Assignees

• Snipr Tech Ltd

Inventors

• Clube Jasper

Classifications

• C12N2800/80

  Vectors containing sites for inducing double-stranded breaks, e.g. meganuclease restriction sites · CPC title

• C12N2795/00052

  relating to complementing cells and packaging systems for producing virus or viral particles · CPC title

• C12N2795/00032

  Use of virus as therapeutic agent, other than vaccine, e.g. as cytolytic agent · CPC title

• C12N2795/00023

  Virus like particles [VLP] · CPC title

• A61K35/74

  Bacteria (therapeutic use of a bacterial protein A61K38/00) · CPC title