Methods for Cell Expansion and Uses of Cells and Conditioned Media Produced Thereby for Therapy

• US 20110129447A1
• Filed: 03/22/2007
• Published: 06/02/2011
• Est. Priority Date: 03/23/2006
• Status: Abandoned Application

• Current Assignee: Pluristem Ltd. (Pluri Inc.)
• Original Assignee: Pluristem Ltd. (Pluri Inc.)
• Inventors: Meretzki, Shai, Burger, Ora, Aberman, Zami
• Application Number:
  US12/225,478
• Publication Number:
  US 20110129447A1
• Time in Patent Office: Days
• US Class Current: 424/93.7
• CPC Class Codes: A61K 2035/122 for inducing tolerance or supression of immune responses
• CPC Class Codes: A61K 2035/124 the cells being hematopoietic, bone marrow derived or blood cells
• CPC Class Codes: A61K 35/12 Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells vaccines or medicinal preparations containing antigens or antibodies A61K39/00
• CPC Class Codes: A61K 35/28 Bone marrow; Haematopoietic stem cells; Mesenchymal stem cells of any origin, e.g. adipose-derived stem cells
• CPC Class Codes: A61K 35/50 Placenta; Placental stem cells; Amniotic fluid; Amnion; Amniotic stem cells
• CPC Class Codes: A61P 1/00 Drugs for disorders of the alimentary tract or the digestive system
• 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 17/02 for treating wounds, ulcers, burns, scars, keloids, or the like
• CPC Class Codes: A61P 19/02 for joint disorders, e.g. arthritis, arthrosis
• CPC Class Codes: A61P 21/00 Drugs for disorders of the muscular or neuromuscular system
• CPC Class Codes: A61P 21/04 for myasthenia gravis
• 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 25/28 for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
• 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 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 37/02 Immunomodulators
• CPC Class Codes: A61P 37/04 Immunostimulants
• CPC Class Codes: A61P 37/06 Immunosuppressants, e.g. drugs for graft rejection
• CPC Class Codes: A61P 43/00 : Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 5/14 : of the thyroid hormones, e.g. T3, T4
• CPC Class Codes: A61P 7/00 : Drugs for disorders of the blood or the extracellular fluid
• CPC Class Codes: A61P 7/06 : Antianaemics
• CPC Class Codes: A61P 9/00 : Drugs for disorders of the cardiovascular system
• 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 2513/00 : 3D culture
• CPC Class Codes: C12N 2531/00 : Microcarriers
• CPC Class Codes: C12N 2533/30 : Synthetic polymers
• CPC Class Codes: C12N 5/0605 : Cells from extra-embryonic tissues, e.g. placenta, amnion, yolk sac, Wharton's jelly
• CPC Class Codes: C12N 5/0653 : Adipocytes; Adipose tissue
• CPC Class Codes: C12N 5/0663 : Bone marrow mesenchymal stem cells (BM-MSC)
• CPC Class Codes: C12N 5/0667 : Adipose-derived stem cells [ADSC]; Adipose stromal stem cells
• CPC Class Codes: C12N 5/0668 : Mesenchymal stem cells from other natural sources
• CPC Class Codes: C12N 5/0669 : Bone marrow stromal cells; Whole bone marrow isolated stem cells from bone marrow C12N5/0647, C12N5/0663

Abstract

A method of cell expansion is provided. The method comprising culturing adherent cells from placenta or adipose tissue under three-dimensional culturing conditions, which support cell expansion.

Claims

1. A method of cell expansion, the method comprising culturing adherent cells from placenta or adipose tissue under three-dimensional culturing conditions, which support cell expansion.

9. A pharmaceutical composition comprising, as an active ingredient, the population of cells generated according to claim 1.

23. The method of claim 1, wherein said three dimensional culture comprises a 3D bioreactor.

29. The method of claim 1, wherein said culturing is effected until said adherent cells reach at least 60% confluence.

2. A method of producing a conditioned medium, the method comprising

(a) culturing adherent cells from a placenta or adipose tissue in three dimensional culturing conditions which allow cell expansion;

and

(b) collecting a conditioned medium of said expanded adherent cells, thereby producing the conditioned medium.

3. (canceled)

4. An isolated population of cells comprising adherent cells of placenta or adipose tissue, wherein said adherent cells are characterized by at least one of:

(i) secretion of a higher level of at least one factor selected from the group consisting of SCF, IL-6, and Flt-3 than that secreted by adherent cells of placenta or adipose tissue grown in a 2D culture;

(ii) expression of a higher level of at least one protein selected from the group consisting of H2A histone family (H2AF), Aldehyde dehydrogenase X (ALDH X), eukaryotic translation elongation factor 2 (EEEF2), reticulocalbin 3, EF-hand calcium binding domain (RCN2) and calponin 1 basic smooth muscle (CNN1) than that expressed by adherent cells of placenta or adipose tissue grown in a 2D culture;

(iii) expression of a lower level of at least one protein selected from the group consisting of heterogeneous nuclear ribonucleoprotein H1 (Hnrph1), CD44 antigen isoform 2 precursor, 3 phosphoadenosine 5 phosphosulfate synthase 2 isoform a (Papss2) and ribosomal protein L7a (rpL7a) than that expressed by adherent cells of placenta or adipose tissue grown in a 2D culture; and

(iiii) a higher immunosuppressive activity than that of adherent cells of placenta or adipose tissue grown in a 2D culture.

8. The isolated population of cells of claim 4, wherein said immunosuppressive activity comprises reduction in T cell proliferation.

11. A pharmaceutical composition comprising, as an active ingredient, the isolated population of cells claim 4.

5-7. -7. (canceled)

10. (canceled)

12. A method of treating a condition which may benefit from stromal cell transplantation in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of adherent cells of a tissue selected from the group consisting of placenta and adipose tissue, thereby treating the condition which may benefit from stem cell transplantation in the subject.

16. The method of claim 12, further comprising administering stem cells.

17. The method of claim 16, wherein said stem cells comprise hematopoietic stem cells.

20. The method of claims 12, wherein said adherent cells are obtained from a three dimensional culture.

21. The method of claims 12, wherein said adherent cells are obtained from a two dimensional culture.

22. The method of claims 12, wherein said condition is selected from the group consisting of stem cell deficiency, heart disease, Parkinson'"'"'s disease, cancer, Alzheimer'"'"'s disease, stroke, burns, loss of tissue, loss of blood, anemia, autoimmune disorders, diabetes, arthritis, Multiple Sclerosis, graft vs. host disease (GvHD), neurodegenerative disorders, autoimmune encephalomyelitis (EAE), systemic lupus erythematosus (SLE), rheumatoid arthritis, systemic sclerosis, Sjorgen'"'"'s syndrome, multiple sclerosis (MS), Myasthenia Gravis (MG), Guillain-Barré

Syndrome (GBS), Hashimoto'"'"'s Thyroiditis (HT), Graves'"'"'s Disease, Insulin dependent Diabetes Melitus (IDDM) and Inflammatory Bowel Disease.

31. The method of claim 12, wherein said adherent cells comprise a positive marker expression array selected from the group consisting of CD73, CD90, CD29 and CD105.

32. The method of claim 12, wherein said adherent cells comprise a negative marker expression array selected from the group consisting of CD45, CD80, HLA-DR, CD11b, CD14, CD19, CD34 and CD79.

38. The method of claim 12, wherein said cells comprise cells having a stromal stem cell phenotype.

42. The method of claim 12, wherein said adherent cells are characterized by at least one of:

(i) secretion of a higher level of at least one factor selected from the group consisting of SCF, IL-6, and Flt-3 than that secreted by adherent cells of placenta or adipose tissue grown in a 2D culture;

(ii) expression of a higher level of at least one protein selected from the group consisting of H2A histone family (H2AF), Aldehyde dehydrogenase X (ALDH X), eukaryotic translation elongation factor 2 (EEEF2), reticulocalbin 3, EF-hand calcium binding domain (RCN2) and calponin 1 basic smooth muscle (CNN1) than that expressed by adherent cells of placenta or adipose tissue grown in a 2D culture;

(iii) expression of a lower level of at least one protein selected from the group consisting of heterogeneous nuclear ribonucleoprotein H1 (Hnrph1), CD44 antigen isoform 2 precursor, 3 phosphoadenosine 5 phosphosulfate synthase 2 isoform a (Papss2) and ribosomal protein L7a (rpL7a) than that expressed by adherent cells of placenta or adipose tissue grown in a 2D culture; and

(iiii) a higher immunosuppressive activity than that of adherent cells of placenta or adipose tissue grown in a 2D culture.

13. A method of treating a condition which may benefit from stromal cell transplantation in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a conditioned medium of adherent cells derived from a tissue selected from the group consisting of placenta and adipose tissue, thereby treating the condition which may benefit from stem cell transplantation in the subject.

14-15. -15. (canceled)

18-19. -19. (canceled)

24-28. -28. (canceled)

30. (canceled)

33-37. -37. (canceled)

39-41. -41. (canceled)