Modulation of microbiota function by gene therapy of the microbiome to prevent, treat or cure microbiome-associated diseases or disorders

Patent metadata

Field	Value
Publication number	US-2023330157-A1
Application number	US-202318319158-A
Country	US
Kind code	A1
Filing date	May 17, 2023
Priority date	Apr 8, 2020
Publication date	Oct 19, 2023
Grant date	—

Abstract

The invention encompasses compositions, kits and methods for modifying bacteria, preferably naturally occurring bacteria, in situ. These can be used to treat, prevent or cure microbiome-associated diseases or disorders by modulating the molecules expressed and/or secreted by bacterial populations of the microbiome in a specific manner. The genomic modifications can modify the interactions between part or all of these populations and the host in a way that decreases their deleterious potential on host health. The compositions, kits and methods of the invention do not result in the direct death of these populations or a direct significant inhibition of their growth. The invention further includes methods for screening for genetic modifications in the bacteria, for determining the efficiency of vectors at inducing these genetic mutations, and for determining the effects of these mutations on bacterial growth.

First claim

We claim:

1 . A method of modifying a naturally occurring bacteria in situ in a subject comprising:
genetically modifying a DNA sequence in the naturally occurring bacteria in situ without introducing a double strand break in the DNA sequence, wherein said genetic modification does not lead to the death of bacteria.

2 . The method of claim 1 , comprising contacting said naturally occurring bacteria with a vector.

3 . The method of claim 1 , comprising contacting said naturally occurring bacteria with a vector located inside a delivery vehicle.

4 . The method of claim 3 , wherein said vector located inside a delivery vehicle is a phagemid.

5 . The method of claim 1 , comprising transducing said naturally occurring bacteria with a packaged phagemid.

6 . The method of claim 2 , wherein the vector further comprises a conditional origin of replication which is inactive in the targeted naturally occuring bacteria but is active in a donor bacterial cell.

7 . The method of claim 1 , wherein said genetic modification is a point mutation.

8 . The method of claim 1 , wherein said genetic modification is a point mutation leading to gene disruption.

9 . The method of claim 1 , wherein the bacteria with the genetic modification does not have a reduced in vivo growth rate as compared to the same bacteria without the genetic modification.

10 . The method of claim 1 , wherein the genetic modification is in a bacterial toxin gene.

11 . The method of claim 2 , wherein said vector encodes a gene editing enzyme or system targeting a specific nucleotide sequence in the naturally occurring bacteria.

12 . The method of claim 11 , wherein the gene editing enzyme further comprises one or two uracil DNA glycosylase inhibitor domain(s) (UGI).

13 . The method of claim 11 , wherein the gene editing enzyme further comprises Mu-GAM.

14 . The method of claim 11 , Wherein the gene editing enzyme is a base editor or a prime editor.

15 . The method of claim 11 , wherein the gene editing enzyme is a dual base editor.

16 . The method of claim 11 , wherein the gene editing enzyme further comprises a reverse transcriptase domain.

17 . The method of claim 11 , wherein the gene editing enzyme further comprises an inhibitor of base repair.

18 . The method of claim 11 , wherein the gene editing system is a retron based system.

19 . The method of claim 2 , wherein the vector comprises a nucleic acid sequence encoding a dCas9 (dead-Cas9), an nCas9 (nickase Cas9), a fusion protein of a dCas9 and a deaminase domain, or a fusion protein of the nCas9 and a deaminase domain.

20 . The method of claim 1 , wherein the naturally occurring bacteria are Cutibacterium acnes bacteria, Escherichia coli bacteria, Staphylococcus aureus bacteria, Clostridium difficile bacteria, Porphyromonas gingivalis bacteria, Streptococcus bacteria, or Pseudomonas bacteria.

21 . A method of modulating host-microbiome interaction by genetically modifying naturally occurring bacteria in situ wherein said naturally occurring bacteria is involved in microbiome associated disorder or disease comprising:
genetically modifying a DNA sequence responsible for the microbiome associated disorder or disease in the naturally occurring bacteria in situ without introducing a double strand break in the DNA sequence, wherein said genetic modification reduces the effects of the microbiome associated disorder or disease, and wherein said genetic modification does not lead to the death of bacteria.

Assignees

• Eligo Bioscience

Inventors

• Duportet Xavier
• Decrulle Antoine
• Gil-Cruz Cristina Del Carmen
• Pérez-Shibayama Christian Ivan
• Ludewig Burkhard
• Fernandez Rodriguez Jesus
• Brodel Andreas

Classifications

• A61K35/74 Primary

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

• C12N15/70 Primary

  Vectors or expression systems specially adapted for E. coli · CPC title

• C12N15/74

  Vectors or expression systems specially adapted for prokaryotic hosts other than E. coli, e.g. Lactobacillus, Micromonospora · CPC title

• C12N15/902

  using homologous recombination · CPC title

• C12N15/1079 Primary

  Screening libraries by altering the phenotype or phenotypic trait of the host (reporter assays C12N15/1086) · CPC title