Bacteria engineered to treat diseases that benefit from reduced gut inflammation and/or tighten gut mucosal barrier

• US 20160206666A1
• Filed: 12/22/2015
• Published: 07/21/2016
• Est. Priority Date: 12/22/2014
• Status: Abandoned Application

• Current Assignee: Synlogic Operating Company, Inc. (Synlogic, Inc.)
• Original Assignee: Synlogic, Inc.
• Inventors: Falb, Dean, Isabella, Vincent M., Kotula, Jonathan W., Miller, Paul F.
• Application Number:
  US14/998,376
• Publication Number:
  US 20160206666A1
• Time in Patent Office: Days
• US Class Current: 1/1
• CPC Class Codes: A61K 2035/115 Probiotics
• CPC Class Codes: A61K 31/19 Carboxylic acids, e.g. valproic acid salicylic acid A61K31/60
• CPC Class Codes: A61K 31/198 Alpha-amino acids, e.g. alanine or edetic acid [EDTA] betaine A61K31/205; proline A61K31/401; tryptophan A61K31/405; histidine A61K31/4172; peptides not degraded to individual amino acids A61K38/00
• CPC Class Codes: A61K 35/741 Probiotics probiotic yeast, e.g. saccharomyces A61K36/06
• CPC Class Codes: A61K 38/20 Interleukins [IL]
• CPC Class Codes: A61K 38/2013 IL-2
• CPC Class Codes: A61K 38/2066 IL-10
• CPC Class Codes: A61K 38/26 Glucagons
• CPC Class Codes: A61K 38/446 Superoxide dismutase (1.15)
• CPC Class Codes: C12N 15/70 Vectors or expression systems specially adapted for E. coli
• CPC Class Codes: C12N 9/001 acting on the CH-CH group of donors (1.3)
• CPC Class Codes: C12N 9/1217 Phosphotransferases with a carboxyl group as acceptor (2.7.2)
• CPC Class Codes: C12Y 103/08001 Butyryl-CoA dehydrogenase (1.3.8.1), i.e. short chain acyl-CoA dehydrogenase
• CPC Class Codes: C12Y 115/01001 Superoxide dismutase (1.15.1.1)
• CPC Class Codes: C12Y 207/02007 Butyrate kinase (2.7.2.7)
• CPC Class Codes: 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

Abstract

Genetically engineered bacteria, pharmaceutical compositions thereof, and methods of treating or preventing autoimmune disorders, inhibiting inflammatory mechanisms in the gut, and/or tightening gut mucosal barrier function are disclosed.

Claims

1. A genetically engineered bacterium comprising:

a) one or more non-native copies of a first gene that encodes a transcription factor protein that is regulated by a reactive nitrogen species (RNS), wherein the expression of the gene is operatively linked to a promoter; and

b) one or more of;

i. a second gene encoding a non-native, anti-inflammation molecule;

ii. a second gene encoding a non-native gut barrier function enhancer molecule; and

iii. a gene cassette encoding a biosynthetic pathway, wherein the final product of the biosynthetic pathway is an anti-inflammation and/or a barrier function enhancer molecule, wherein

the expression of the second gene or gene cassette in b) is controlled by a tunable regulatory region heterologous to the gene or gene cassette, wherein induction of the tunable regulatory region is directly or indirectly controlled by the transcription factor.

2. The bacterium of claim 1, wherein the transcription factor is a transcription activator.

3. The bacterium of claim 2, wherein induction of the tunable regulatory region is directly controlled by the transcription activator.

4. The bacterium of claim 1, wherein the transcription factor is a transcription repressor.

5. The bacterium of claim 4, wherein derepression of the tunable regulatory region is directly controlled by the transcription repressor.

6. The bacterium of claim 1, wherein:

the transcription factor is a first transcription repressor;

the bacterium further comprises a gene encoding a second transcription repressor, wherein expression of the second transcription repressor is repressed by the first repressor; and

the tunable regulatory region is repressed by the second transcription repressor.

7. The bacterium of claim 1, wherein at least one of the one or more non-native copies of the gene that encodes the transcription factor is located on a plasmid in the bacterium.

8. The bacterium of claim 1, wherein at least one of the one or more non-native copies of the gene that encodes the transcription factor is located on a chromosome in the bacterium.

9. The bacterium of claim 1, wherein the promoter that controls expression of at least one of the one or more non-native copies of the gene that encodes the transcription factor is a constitutive promoter.

10. The bacterium of claim 1, wherein the promoter that controls expression of at least one of the one or more non-native copies of the gene that encodes the transcription factor is an inducible promoter.

11. The bacterium of claim 1, wherein the gene encoding the anti-inflammation molecule, the gut barrier enhancer molecule, or the gene cassette encoding the biosynthetic pathway is located on a plasmid in the bacterium.

12. The bacterium of claim 1, wherein the gene encoding the anti-inflammation molecule, the gut barrier enhancer molecule, or the gene cassette encoding the biosynthetic pathway is located on a chromosome in the bacterium.

13. The bacterium of claim 1, wherein the gene that encodes the transcription factor protein is NsrR.

14. The bacterium of claim 1, wherein the regulatory region that controls expression of the anti-inflammation molecule, gut barrier enhancer molecule, or the biosynthetic pathway is selected from a native or a modified functional form of any one of norB, aniA, nsrR, hmpA, ytfE, ygbA, hcp, hcr, nrfA, aox.

15. The bacterium of claim 1, wherein the anti-inflammation and/or gut barrier enhancer molecule is selected from propionate, butyrate, acetate, IL-10, IL-27, TGF-β

2, TGF-β

1, GLP-2, NAPEs, elafin, trefoil factor, and scFv, antisense RNA, siRNA, or shRNA directed against a pro-inflammatory molecule.

16. The bacterium of claim 1, wherein the bacterium is a non-pathogenic bacterium.

18. The bacterium of claim 16, wherein the bacterium is selected from the group consisting of Bacteroides, Bifidobacterium, Clostridium, Escherichia, Lactobacillus, and Lactococcus.

19. The bacterium of claim 18, wherein the bacterium is Escherichia coli strain Nissle.

20. The bacterium of claim 1, wherein the bacterium is an auxotroph in a gene that is complemented when the bacterium is present in a mammalian gut.

21. The bacterium of claim 20, wherein mammalian gut is a human gut.

22. The bacterium of claim 20, wherein the bacterium is an auxotroph in diaminopimelic acid or an enzyme in the thymine biosynthetic pathway.

25. A pharmaceutically acceptable composition comprising the bacterium of claim 1 and a pharmaceutically acceptable carrier.

27. A method of treating or preventing an autoimmune disorder, comprising the step of administering to a patient in need thereof, the composition of claim 25.

28. A method of treating a disease or condition associated with gut inflammation and/or compromised gut barrier function, comprising the step of administering to a patient in need thereof, the composition of claim 25.

30. The method of claim 27, wherein the autoimmune disorder is selected from the group consisting of type 1 diabetes, asthma, multiple sclerosis, lupus, rheumatoid arthritis, ulcerative colitis, juvenile arthritis, psoriasis, psoriatic arthritis, Crohn'"'"'s disease, celiac disease, and ankylosing spondylitis.

31. The method of claim 28, wherein the disease or disorder is selected from an inflammatory bowel disease and a diarrheal disease.

17. (canceled)

23. (canceled)

24. (canceled)

26. (canceled)

29. (canceled)