Methods of controlling proliferation and differentiation of stem and progenitor cells

• US 7,855,075 B2
• Filed: 05/19/2008
• Issued: 12/21/2010
• Est. Priority Date: 02/17/1998
• Status: Expired due to Fees

• Current Assignee: Gamida Cell, Ltd., Hadasit Medical Research Services & Development Ltd (Hadassah, The Women's Zionist Organization of America, Inc.)
• Original Assignee: Gamida Cell, Ltd., Hadasit Medical Research Services & Development Ltd (Hadassah, The Women's Zionist Organization of America, Inc.)
• Inventors: Fibach, Eitan, Treves, Avi, Peled, Tony
• Primary Examiner(s): Belyavskyi; Michail A
• Application Number:
  US12/154,058
• Publication Number:
  US 20090004158A1
• Time in Patent Office: 946 Days
• Field of Search: None
• US Class Current: 435/372
• CPC Class Codes: A61K 31/13 Amines A61K31/04 takes precedence
• CPC Class Codes: A61K 31/131 acyclic
• 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 31/395 having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
• CPC Class Codes: A61K 31/401 Proline; Derivatives thereof, e.g. captopril
• CPC Class Codes: A61K 31/495 having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazinesA61K31/48 takes precedence ; with three nitrogen atoms A61K31/53
• CPC Class Codes: A61K 31/4995 Pyrazines or piperazines forming part of bridged ring systems
• CPC Class Codes: A61K 33/30 Zinc; Compounds thereof
• CPC Class Codes: A61K 35/14 Blood; Artificial blood perfluorocarbons A61K31/02; umbilical cord blood A61K35/51; haemoglobin A61K38/42
• 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 38/06 Tripeptides
• CPC Class Codes: A61P 35/02 specific for leukemia
• CPC Class Codes: A61P 7/00 Drugs for disorders of the blood or the extracellular fluid
• CPC Class Codes: C12N 2500/20 Transition metals
• CPC Class Codes: C12N 2500/22 Zinc; Zn chelators insulin-zinc complexes C12N2501/33
• CPC Class Codes: C12N 2500/46 Amines, e.g. putrescine
• CPC Class Codes: C12N 2501/125 Stem cell factor [SCF], c-kit ligand [KL]
• CPC Class Codes: C12N 2501/14 Erythropoietin [EPO]
• CPC Class Codes: C12N 2501/145 Thrombopoietin [TPO]
• CPC Class Codes: C12N 2501/18 Liver cell growth factor (LCGF, Gly-His-Lys)
• CPC Class Codes: C12N 2501/22 : Colony stimulating factors (G-CSF, GM-CSF)
• CPC Class Codes: C12N 2501/23 : Interleukins [IL]
• CPC Class Codes: C12N 2501/26 : Flt-3 ligand (CD135L, flk-2 ligand)
• CPC Class Codes: C12N 5/0641 : Erythrocytes
• CPC Class Codes: C12N 5/0647 : Haematopoietic stem cells; Uncommitted or multipotent progenitors

Abstract

A method of ex-vivo expanding a population of stem cells, while at the same time inhibiting differentiation of the stem cells. The method comprises ex-vivo providing the stem cells with conditions for cell proliferation and with at least one copper chelator in an amount and for a time period for permitting the stem cells to proliferate and, at the same time, for reducing a capacity of the stem cells to differentiate

Claims

1. An ex-vivo expanded cell population comprising cells having reduced available intracellular copper as compared to ex-vivo cells cultured in the absence of a transition metal chelator having affinity for copper and wherein said cells are further characterized by increased proportion of undifferentiated cells, wherein said cells are selected from the group consisting of erythroblast precursors, embryonic stem cells, undifferentiated hepatic cells and hematopoietic cells.

2. The ex-vivo expanded cell population of claim 1, wherein said cells are erythroblast precursors and said undifferentiated cells are non-hemoglobin producing cells.

3. The ex-vivo expanded cell population of claim 1, wherein said cells are myeloid cells and said undifferentiated cells are non-phagocytic cells.

4. The ex-vivo expanded cell population of claim 1, wherein said cells are provided in a culture medium.

5. The ex-vivo expanded cell population of claim 4, wherein said culture medium comprises cytokines and a transition metal chelator having affinity for copper.

6. The ex-vivo expanded cell population of claim 1, wherein said cells are isolated from a culture medium.

7. A pharmaceutical composition comprising the ex-vivo expanded cell population of claim 1.

8. A pharmaceutical composition comprising the ex-vivo expanded cell population of claim 5.

9. The ex-vivo expanded cell population of claim 1, wherein said transition metal chelator is selected from the group consisting of polyamine chelating agents, ethylendiamine, diethylenetriamine, triethylenetetramine, triethylenediamine, tetraethylenepentamine, aminoethylethanolamine, aminoethylpiperazine, pentaethylenehexamine, triethylenetetramine-hydrochloride, tetraethylenepentamine-hydrochloride, pentaethylenehexamine-hydrochloride, tetraethylpentamine, captopril, penicilamine, N,N′

-bis(3-aminopropyl)-1,3-propanediamine, N,N,Bis (2 animoethyl) 1,3 propane diamine, 1,7-dioxa-4,10-diazacyclododecane, 1,4,8,11-tetraaza cyclotetradecane-5,7-dione, 1,4,7-triazacyclononane trihydrochloride, 1-oxa-4,7,10-triazacyclododecane, 1,4,8,12-tetraaza cyclopentadecane or 1,4,7,10-tetraaza cyclododecane, preferably tetraethylpentamine.

10. The ex-vivo expanded cell population of claim 1, wherein said transition metal chelator is tetraethylenepentamine.

11. The ex-vivo expanded cell population of claim 10, wherein said transition metal chelator concentration is about 0.1 μ

M to about 100 μ

M.

12. The ex-vivo expanded cell population of claim 11, wherein said transition metal chelator concentration is about 4 μ

M to about 50 μ

M.

13. The ex-vivo expanded cell population of claim 12, wherein said transition metal chelator concentration is about 5 μ

M to about 40 μ

M.

14. The ex-vivo expanded cell population of claim 5, wherein said cytokines are early acting cytokines.

15. The ex-vivo cell population of claim 1, wherein said cells are genetically modified with an exogene.

16. An ex-vivo expanded cell population comprising ex-vivo expanded cells characterized by:

(a) having been cultured for at least one week in the presence of cytokines and a transition metal chelator having affinity for copper, and(b) having a greater proportion of undifferentiated cells following at least 1 week culture in the presence of said transition metal chelator, as compared to ex vivo cells cultured for the same time in a presence of said cytokines yet in an absence of said transition metal chelator, wherein said cells are selected from the group consisting of erythroblast precursors, embryonic stem cells, undifferentiated hepatic cells and hematopoietic cells.

17. The ex-vivo expanded cell population of claim 16, wherein said transition metal chelator is tetraethylenepentamine.