GUT FLORA-DERIVED EXTRACELLULAR VESICLES, AND METHOD FOR SEARCHING FOR A DISEASE MODEL, VACCINE, AND CANDIDATE DRUG AND FOR DIAGNOSIS USING THE SAME

• US 20150017664A1
• Filed: 07/02/2014
• Published: 01/15/2015
• Est. Priority Date: 09/01/2009
• Status: Active Grant

• Current Assignee: Aeon Medix Inc.
• Original Assignee: Aeon Medix Inc.
• Inventors: GHO, Yong Song, KIM, Yoon Keun, PARK, Kyoung Su, HONG, Bok Sil, KIM, Ji Hyun, KIM, You Sun, LEE, Won Hee, KIM, Jung Wook, KIM, Dae-Kyum
• Granted Patent: US 9,274,109 B2
• Time in Patent Office: Days
• US Class Current: 435/7.92
• CPC Class Codes: A01K 2207/20 Animals treated with compounds which are neither proteins nor nucleic acids
• CPC Class Codes: A01K 2217/075 inducing loss of function, i.e. knock out
• CPC Class Codes: A01K 2227/105 Murine
• CPC Class Codes: A01K 2267/0337 Animal models for infectious diseases
• CPC Class Codes: A01K 2267/035 Animal model for multifactorial diseases
• CPC Class Codes: A01K 67/027 New or modified breeds of vertebrates
• CPC Class Codes: A61P 1/00 Drugs for disorders of the alimentary tract or the digestive system
• CPC Class Codes: A61P 1/02 Stomatological preparations, e.g. drugs for caries, aphtae, periodontitis
• CPC Class Codes: A61P 1/04 for ulcers, gastritis or reflux esophagitis, e.g. antacids, inhibitors of acid secretion, mucosal protectants
• CPC Class Codes: A61P 1/16 for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
• CPC Class Codes: A61P 1/18 for pancreatic disorders, e.g. pancreatic enzymes
• CPC Class Codes: A61P 11/00 Drugs for disorders of the respiratory system
• CPC Class Codes: A61P 19/02 for joint disorders, e.g. arthritis, arthrosis
• CPC Class Codes: A61P 19/10 for osteoporosis
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 29/00 Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• CPC Class Codes: A61P 3/04 Anorexiants; Antiobesity agents
• CPC Class Codes: A61P 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: A61P 31/00 Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
• CPC Class Codes: A61P 31/04 Antibacterial agents
• CPC Class Codes: A61P 35/00 : Antineoplastic agents
• CPC Class Codes: A61P 43/00 : Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 7/00 : Drugs for disorders of the blood or the extracellular fluid
• CPC Class Codes: A61P 9/10 : for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
• CPC Class Codes: A61P 9/12 : Antihypertensives
• CPC Class Codes: C12Q 1/025 : for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics antimicrobial activity C12Q1/18
• CPC Class Codes: C12Q 1/37 : involving peptidase or proteinase
• CPC Class Codes: G01N 33/56916 : Enterobacteria, e.g. shigella, salmonella, klebsiella, serratia

Abstract

The present application relates to a composition including gut flora-derived extracellular vesicles, and to an animal disease model using same. In addition, the present application relates to a method for using the gut flora-derived extracellular vesicles to efficiently search for a candidate drug which may prevent or treat diseases that occur due to gut flora-derived extracellular vesicles, and to a vaccine which can efficiently prevent or treat infections caused by gut flora or diseases that occur due to gut flora-derived extracellular vesicles. Further, the development of diagnostic technology to discover, using the gut flora-derived extracellular vesicles of the present application, the etiology of diseases that occur due to gut flora-derived extracellular vesicles, can be achieved.

Claims

1-6. -6. (canceled)

7. A method of assessing a subject'"'"'s susceptibility to one or more diseases, the method comprising:

providing a database listing pre-identified pathogenic microorganisms and one or more substances of the pre-identified pathogenic microorganisms;

providing a sample comprising gut flora isolated from a subject;

processing the sample to obtain a concentrated population of extracellular vesicles contained in the gut flora;

processing at least part of the concentrated population to determine if the concentrated population contains at least one substance of a first one of the pre-identified pathogenic microorganisms listed in the database; and

once determined that at least one substance is of the first pre-identified pathogenic microorganism, further determining that the subject is potentially susceptible to one or more diseases that are associated with the first pre-identified pathogenic microorganism.

8. The method of claim 7, wherein the at least one substance comprises at least one genetic substance, wherein the database further comprises sequences of genetic substances of at least part of the pre-identified pathogenic microorganisms,

wherein processing at least part of the concentrated population comprises:

sequencing at least one genetic substance contained in the concentrated population of extracellular vesicles, which provides at least one sequence of the at least one genetic substance; and

computerized searching using the database to determine if the at least one sequence is of the first pre-identified pathogenic microorganism.

9. The method of claim 8, wherein the at least one substance comprises 16S rRNA.

10. The method of claim 8, wherein the first pre-identified pathogenic microorganism comprises a strain of E. coli or Klebsiella spp. that is known to be pathogenic.

11. The method of claim 7, wherein the at least one substance comprises at least one nucleotide sequence,

wherein processing at least part of the concentrated population comprises:

mixing at least part of the concentrated population with a primer that has a sequence complementary to a first genetic substance of a first one of the pre-identified pathogenic microorganisms such that if the concentrated population contains extracellular vesicles of the first pre-identified pathogenic microorganism, the primer would recognize the first genetic substance of the first pre-identified pathogenic microorganism;

subjecting the mixture to amplification; and

processing the amplification product to determine if any nucleotide sequence recognized by the primer has been amplified therein, wherein if at least one nucleotide sequence recognized by the primer has been amplified, the at least one nucleotide sequence is determined to be the at least one substance is of the first pre-identified pathogenic microorganism.

12. The method of claim 11, wherein the at least one substance comprises 16S rRNA.

13. The method of claim 11, wherein the first pre-identified pathogenic microorganism comprises a strain of E. coli or Klebsiella spp. that is known to be pathogenic.

14. The method of claim 7, wherein the at least one substance comprises at least one protein,

wherein processing at least part of the concentrated population comprises:

mixing at least part of the concentrated population with an antibody that has specificity to a first protein of a first one of the pre-identified pathogenic microorganisms such that if the concentrated population contains extracellular vesicles of the first pre-identified pathogenic microorganism, the antibody would bind with the first protein of the first pre-identified pathogenic microorganism; and

subsequently, processing to determine if the antibody has bound with any protein in the mixture, wherein if determined that the antibody has bound with at least one protein, the at least one protein is determined to be the at least one substance of the first pre-identified pathogenic microorganism.

15. The method of claim 14, wherein the first pre-identified pathogenic microorganism comprises a strain of E. coli or Klebsiella spp. that is known to be pathogenic.

16. The method of claim 7, wherein processing at least part of the concentrated population comprises:

contacting at least part of the concentrated population with animal cells such that if the concentrated population contains extracellular vesicles of a first one of pre-identified pathogenic microorganisms, the extracellular vesicles would induce an immune response in the animal cells, wherein the animal cells are pre-cultured in vitro or part of a live animal; and

subsequently, processing to determine if an immune response has occurred in the animal cells, wherein if determined that an immune response has occurred in the animal cells, it is determined that the concentrated population contains at least one substance of the first pre-identified pathogenic microorganism.

17. The method of claim 16, wherein processing to determine if an immune response has occurred in the animal cells comprises:

determining if Th1 immune response has been induced in the animal cells after contacting the at least part of the concentrated population with the animal cells.

18. The method of claim 16, wherein processing to determine if an immune response has occurred in the animal cells comprises:

determining if Th17 immune response has been induced in the animal cells after contacting the at least part of the concentrated population with the animal cells.

19. The method of claim 16, wherein the first pre-identified pathogenic microorganism comprises a strain of E. coli or Klebsiella spp. that is known to be pathogenic.

20. The method of claim 7, wherein the sample is isolated from at least one selected from the group consisting of feces, large intestinal fluid, gastric fluid, small intestinal fluid, and oral fluid.

21. The method of claim 7, wherein the first pre-identified pathogenic microorganism comprises a strain of E. coli that is known to be pathogenic.

22. The method of claim 7, wherein the first pre-identified pathogenic microorganism comprises a strain of Klebsiella spp. that is known to be pathogenic.

23. The method of claim 7, wherein the first pre-identified pathogenic microorganism comprises a strain of Pseudomonas spp. that is known to be pathogenic.

24. The method of claim 7, wherein the first pre-identified pathogenic microorganism comprises a strain of Bacteoides spp. that is known to be pathogenic.

25. The method of claim 7, wherein the first pre-identified pathogenic microorganisms comprises a strain of at least one microorganism selected from the group consisting of E. coli, Klebsiella spp., Pseudomonas spp. and Bacteoides spp.

26. The method of claim 7, wherein the one or more diseases are selected from the group consisting of sepsis, arteriosclerosis, acute coronary syndrome, stroke, emphysema, acute respiratory distress syndrome, osteoporosis, hypertension, obesity, diabetes, arthritis, a cerebral disease, mouth ulcer, oral cancer, esophagitis, esophageal cancer, gastritis, duodenal ulcer, stomach cancer, inflammatory bowel disease, irritable bowel syndrome, colorectal cancer, cholangitis, cholecystitis, pancreatitis, cholangiocarcinoma, and pancreatic cancer.