MULTIPOTENT STEM CELLS AND USES THEREOF

• US 20110070205A1
• Filed: 09/24/2010
• Published: 03/24/2011
• Est. Priority Date: 05/03/2007
• Status: Active Grant

• Current Assignee: Brigham and Women's Hospital Incorporated
• Original Assignee: Brigham and Women's Hospital Incorporated
• Inventors: Southgate, Chris, Crawford, Keith D.
• Granted Patent: US 8,574,567 B2
• Time in Patent Office: Days
• US Class Current: 424/93.7
• CPC Class Codes: A61K 2035/124 the cells being hematopoietic, bone marrow derived or blood cells
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 25/16 Anti-Parkinson drugs
• CPC Class Codes: A61P 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 9/00 Drugs for disorders of the cardiovascular system
• CPC Class Codes: A61P 9/04 Inotropic agents, i.e. stimulants of cardiac contraction; Drugs for heart failure
• 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: C12N 5/0653 Adipocytes; Adipose tissue
• CPC Class Codes: C12N 5/0662 Stem cells
• CPC Class Codes: C12N 5/0668 Mesenchymal stem cells from other natural sources

Abstract

The invention provides a quiescent stem cell having the capacity to differentiate into ectoderm, mesoderm and endoderm, and which does not express cell surface markers including MHC class I, MHC class II, CD44, CD45, CD13, CD34, CD49c, CD73, CD105 CD90, CD66A, CD66E, CXCR4, CD133 or an SSEA. The invention further provides a proliferative stem cell, which expresses genes including Oct-4, Nanog, Sox2, GDF3, P16INK4, BMI, Notch, HDAC4, TERT, Rex-1, TWIST, KLF-4 and Stella but does not express cell surface markers including MHC class I, MHC class II, CD44, CD45, CD13, CD34, CD49c, CD73, CD105, CD90, CD66A, CD66E, CXCR4, CD133 or an SSEA. The cells of the invention can be isolated from adult mammals, have embryonic cell characteristics, and can form embryoid bodies. Methods for obtaining the stem cells, as well as methods of treating diseases and differentiated the stem cells, are also provided.

Claims

1. An isolated adult stem cell that is capable of proliferating and differentiating into at least two of ectoderm, mesoderm, or endoderm, expresses at least one of Oct-4, KLF-4, Nanog, Sox-2, Rex-1, GDF-3, and Stella, and does not detectibly express CD13, CD45, CD90, and CD34 and further does not detectibly express at least one of MHC class I, MHC class II, CD44, CD105, CD49c, CD73, CD66A, CD66E, CXCR4, CD133 or an SSEA.

3. The cell of claim 1, wherein the SSEA is SSEA-4.

4. The cell of claim 1, wherein said cell is synovial fluid derived, blood derived or tissue derived.

5. The cell of claim 1, wherein said cell is substantially purified.

6. The cell of claim 1, wherein said cell is isolated from a mammal.

7. The cell of claim 1, wherein said cell is isolated from a human.

8. The cell of claim 1, wherein said cell is isolated from an adult mammal.

9. The cell of claim 1, further comprising a heterologous nucleic acid sequence.

19. A master cell bank comprising a plurality of cryopreserved individually packaged populations of isolated adult stem cells, each population including at least 1×

10² or more of the cells of claim 1.

20. A method of forming an adipocyte, the method comprising culturing a stem cell of claim 1 under adipocyte-differentiating conditions.

21. The method of claim 20, wherein said adipocyte-differentiating conditions include culturing with at least one of dexamethasone, 3-isobutyl-1-methylxanthine (IBMX), insulin, and indomethacin.

22. A method of forming a muscle cell, said method comprising culturing a stem cell of claim 1 under muscle cell differentiating conditions.

23. The method of claim 22, wherein said conditions include culturing said cell with PDGF and TGF-β

1.

24. A method of forming a neural cell, said method comprising the steps of contacting a stem cell of claim 1 under neural cell-forming conditions.

25. The method of claim 24, wherein said conditions include culturing said cell with bFGF, FGF-8, SHH, and BDNF.

26. A method of forming a hepatocyte, said method comprising culturing a stem cell of claim 1 under hepatocyte-forming conditions.

27. The method of claim 26, wherein said conditions include culturing said cell with hepatocyte growth factor (HGF) and FGF-4.

28. A method of forming an endothelial cell, said method comprising culturing a stem cell of claim 1 under endothelial cell-forming conditions.

29. The method of claim 28, wherein said conditions include culturing said cell with VEGF.

30. A method of forming a hematopoietic cell, the method comprising culturing a stem cell of claim 1 under hematopoietic cell forming conditions.

31. The method of claim 30, wherein said conditions include culturing said cell with bone morphogenic protein-4 (BMP4), VEGF, bFGF, stem cell factor (SCF), Flt3L, hyper IL6, thrombopoietin (TPO) and erythropoietin (EPO).

32. A method for promoting wound healing in a subject, said method comprising administering a stem cell of claim 1, or a committed or differentiated progeny thereof, to said wound or to a site near said wound in an amount sufficient to promote the healing of said wound.

33. The method of claim 32, wherein said administration of said cells results in reduced scarring at the wound site.

34. A method for treating a cardiovascular disease in a subject, said method comprising administering to said subject a stem cell of claim 1, or a committed or differentiated progeny thereof, in an amount sufficient to treat said disease.

35. The method of claim 34, wherein said cardiovascular disease is myocardial infarction, congestive heart failure, ischemic cardiomyopathy, and coronary artery disease.

36. A method of increasing vascularization in a subject, said method comprising administering to said subject a stem cell of claim 1, or a committed or differentiated progeny thereof, in an amount sufficient to increase vascularization.

37. The method of claim 36, wherein said subject is suffering from type II diabetes.

38. A method for treating a neurological disorder in a subject, said method comprising administering to said subject a stem cell of claim 1, or a committed or differentiated progeny thereof, in an amount sufficient to treat said disease.

39. The method of claim 38, wherein said neurological disorder is a neurodegenerative disease or neurological injury.

40. The method of claim 39, wherein said neurodegenerative disease is Parkinson'"'"'s disease, Alzheimer'"'"'s disease, or Huntington'"'"'s disease.

41. A method for treating an autoimmune disease in a subject, said method comprising administering to said subject a stem cell of claim 1, or a committed or differentiated progeny thereof, in an amount sufficient to treat said disease.

42. A method for reducing or preventing rejection of a transplanted tissue in a subject, said method comprising administering to said subject a stem cell of claim 1, or a committed or differentiated progeny thereof, in an amount sufficient to reduce or prevent said rejection.

45. A process for differentiating the isolated stem cell of claim 1 comprising culturing said isolated stem cell under conditions sufficient to differentiate said isolated stem cell.

2. An isolated quiescent adult stem cell that is capable of proliferating and differentiating into at least two of ectoderm, mesoderm, and endoderm and does not detectibly express Oct-4, CD13, CD45, CD90, CD34 and further does not detectibly express at least one of MHC class I, MHC class II, CD44, CD105, CD49c, CD73, CD66A, CD66E, CXCR4, CD133 or an SSEA.

10. A population of isolated adult stem cells that are capable of proliferating and differentiating into at least two of ectoderm, mesoderm, and endoderm, express at least one of Oct-4, KLF-4, Nanog, Sox-2, Rex-1, GDF-3, and Stella, and does not detectibly express CD13, CD45, CD90, CD34 and further does not detectibly express at least one of MHC class I, MHC class II, CD44, CD105, CD49c, CD73, CD66A, CD66E, CXCR4, CD133 or an SSEA, wherein from about 10% to about 30% of the population of adult stem cells are quiescent.

11. The population of claim 10, wherein said population is a culture expanded population.

12. The population of claim 10, wherein said cells are cryopreserved and wherein said population is included within a container.

13. The population of claim 12, wherein said container is a vial, syringe or other container suitable for local delivery into a site within a human or animal.

14. The population of claim 12, wherein said container is a bag or other container suitable for intravenous delivery of cells within a human or animal.

15. The population of claim 10, wherein said population comprises said stem cells in an amount of at least 1×

10³, at least 1×

10⁶, at least 1×

10⁹, at least 1×

10¹², or at least 1×

10¹⁴.

16. The population of claim 10, wherein said population is contained in a 0.9% NaCL solution.

17. The population of claim 10, further comprising a bioactive compound.

18. The population of claim 10, wherein said bioactive compound is a growth factor, a cytokine, an antibody or fragment thereof, or an organic molecule the molecule having a mass of less than 5,000 daltons.

43. A process for expanding the population of claim 10 comprising passaging the population at least three, at least five, at least ten or at least forty times.

44. A population of stem cells obtained according to the process of claim 43.