# US 11,439,673 B2 · Bacteria engineered to treat liver disease > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 11,439,673 B2 - Document ID: US11439673B2 - Status: Granted - Kind: B2 - Application number: 17/058,172 - Filed: 2019-05-31 - Published: 2021-05-27 - Granted: 2022-09-13 - Earliest claimed: 2018-06-02 - Assignees: Synlogic Operating Company, Inc. - Applicants: Synlogic Operating Company, Inc. - Inventors: Paul F. Miller; Vincent M. Isabella; Dean Falb; Jonathan W. Kotula; Yossi Dagon - Web page: https://patentgrep.com/patent/US11439673B2 ## CPC - A61K 35/741: Probiotics (probiotic yeast, e.g. saccharomyces A61K36/06) - A61K 35/742: Spore-forming bacteria, e.g. Bacillus coagulans, Bacillus subtilis, clostridium or Lactobacillus sporogenes - A61K 35/744: Lactic acid bacteria, e.g. enterococci, pediococci, lactococci, streptococci or leuconostocs - A61K 35/745: Bifidobacteria - A61P 1/16: for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics - A61P 7/06: Antianaemics - A61P 9/00: Drugs for disorders of the cardiovascular system - A61P 11/00: Drugs for disorders of the respiratory system - A61P 35/00: Antineoplastic agents - 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 ## Abstract The disclosure provides genetically engineered bacteria that are capable of reducing excess ammonia and converting ammonia and/or nitrogen into alternate byproducts. The invention also provides pharmaceutical compositions comprising the genetically engineered bacteria, and methods of modulating and treating disorders associated with excess ammonia and inflammation in the gut and the liver, including, for example, hepatic encephalopathy, NASH/NAFLD, HCV, and Huntington's disease, in a subject, comprising administering the engineered bacterium to the subject. ## Claims ### Claim 1 A method for treating a subject having liver disease, the method comprising: administering an engineered bacterium, or a pharmaceutical composition comprising the engineered bacterium, to the subject, and measuring gene expression in the colon and/or liver before the administering, and/or measuring gene expression in the colon and/or liver after administering, wherein the administering: i) reduces IL-6 gene expression in the liver by at least 10% as compared to IL-6 gene expression in the liver before the administering; ii) reduces TNFα gene expression in the liver by at least 10% as compared to TNFα gene expression in the liver before the administering; iii) reduces TGFβ gene expression in the liver by at least 10% as compared to TGFβ gene expression in the liver before the administering; iv) reduces αSMA gene expression in the liver by at least 10% as compared to αSMA gene expression in the liver before the administering; v) reduces IL-6 gene expression in the colon by at least 10% as compared to IL-6 gene expression in the colon before the administering; and/or vi) wherein blood ammonia levels are decreased by at least 5% after administering as compared to blood ammonia levels before the administering. ### Claim 2 (depends on 1) The method of claim 1, wherein the administering reduces IL-6 gene expression in the liver by at least 15%, 20%, 25%, 30%, 35%, 40%, 45%, or 50% as compared to IL-6 gene expression in the liver before the administering. ### Claim 3 (depends on 1) The method of claim 1, wherein administering reduces TGFβ gene expression in the liver by at least 15% or 20% as compared to TGFβ gene expression in the liver before the administering. ### Claim 4 (depends on 1) The method of claim 1, wherein administering reduces αSMA gene expression in the liver by at least 15%, 20%, 25%, 30%, 35%, 40%, 45%, or 50% as compared to αSMA gene expression in the liver before the administering. ### Claim 5 (depends on 2) The method of claim 2, wherein administering reduces IL-6 gene expression in the colon by at least 15% or 20% as compared to IL-6 gene expression in the colon before the administering. ### Claim 6 (depends on 1) The method of claim 1, wherein blood ammonia levels are decreased by at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, or 50% as compared to blood ammonia levels before the administering. ### Claim 7 (depends on 1) The method of claim 1, further comprising measuring blood ammonia levels before the administering and/or further comprising measuring blood ammonia levels after the administering. ### Claim 8 (depends on 1) The method of claim 1, wherein the engineered bacterium reduces inflammation in the colon of the subject. ### Claim 9 (depends on 1) The method of claim 1, wherein the engineered bacterium reduces inflammation in the liver of the subject. ### Claim 10 (depends on 1) The method of claim 1, wherein the bacterium comprises one or more gene sequences encoding an ammonia consumption circuit. ### Claim 11 (depends on 1) The method of claim 1, wherein the bacterium comprises one or more gene sequences encoding an arginine production circuit. ### Claim 12 (depends on 1) The method of claim 1, wherein the bacterium comprises a gene encoding an arginine feedback resistant N-acetylglutamate synthetase (ArgAfbr), wherein the ArgAfbr has reduced arginine feedback inhibition as compared to a wild-type N-acetylglutamate synthetase from the same bacterial subtype under the same conditions and wherein expression of the gene encoding ArgAfbr is controlled by a promoter that is induced by low-oxygen or anaerobic conditions; and wherein the bacterium has been genetically engineered to lack a functional ArgR. ### Claim 13 (depends on 1) The method of claim 1, wherein each copy of a functional argR gene normally present in a corresponding wild-type bacterium has been deleted. ### Claim 14 (depends on 1) The method of claim 1, wherein under low-oxygen or anaerobic conditions, the transcription of each gene in the engineered bacterium that is present in an operon comprising a functional ARG box and which encodes an arginine biosynthesis enzyme is increased as compared to a corresponding gene in a wild-type bacterium under the same conditions. ### Claim 15 (depends on 1) The method of claim 1, wherein the bacterium comprises a gene sequence encoding a biosynthetic pathway for producing butyrate. ### Claim 16 (depends on 12) The method of claim 12, wherein the promoter that is induced under low-oxygen or anaerobic conditions is an FNR promoter. ### Claim 17 (depends on 1) The method of claim 1, wherein the bacterium is a non-pathogenic bacterium and/or a probiotic bacterium. ### Claim 18 (depends on 1) The method of claim 1, wherein the bacterium is selected from the group consisting of Bacteroides, Bifidobacterium, Clostridium, Escherichia, Lactobacillus, and Lactococcus. ### Claim 19 (depends on 18) The method of claim 18, wherein the bacterium is Escherichia coli strain Nissle. ### Claim 20 (depends on 1) The method of claim 1, wherein the bacterium is a thyA or dapB auxotroph. ### Claim 21 (depends on 1) The method of claim 1, wherein the liver disease is selected from NASH, NAFLD, and hepatic encephalopathy. ### Claim 22 (depends on 1) The method of claim 1, wherein administering reduces TGFα gene expression in the liver by at least 15% or 20% as compared to TGFα gene expression in the liver before the administering.