Method for transdifferentiation of non pancreatic stem cells to the pancreatic differentiation pathway

• US 20020182728A1
• Filed: 03/29/2002
• Published: 12/05/2002
• Est. Priority Date: 03/29/2001
• Status: Active Grant

• Current Assignee: Ixion Biotechnology Inc. (Galderma Holding SA)
• Original Assignee: Ixion Biotechnology Inc. (Galderma Holding SA)
• Inventors: Clark, Amy, Ramiya, Vijayakumar
• Granted Patent: US 7,202,080 B2
• Time in Patent Office: Days
• US Class Current: 435/366
• 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: A61P 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: C12N 2500/25 Insulin-transferrin; Insulin-transferrin-selenium
• CPC Class Codes: C12N 2501/11 Epidermal growth factor [EGF]
• CPC Class Codes: C12N 2501/12 Hepatocyte growth factor [HGF]
• CPC Class Codes: C12N 2501/125 Stem cell factor [SCF], c-kit ligand [KL]
• CPC Class Codes: C12N 2501/13 Nerve growth factor [NGF]; Brain-derived neurotrophic factor [BDNF]; Cilliary neurotrophic factor [CNTF]; Glial-derived neurotrophic factor [GDNF]; Neurotrophins [NT]; Neuregulins
• CPC Class Codes: C12N 2501/148 Transforming growth factor alpha [TGF-a]
• CPC Class Codes: C12N 2501/315 Prolactin
• CPC Class Codes: C12N 2501/335 Glucagon; Glucagon-like peptide [GLP]; Exendin
• CPC Class Codes: C12N 2501/345 Gastrin; Cholecystokinins [CCK]
• CPC Class Codes: C12N 2506/11 from blood or immune system cells
• CPC Class Codes: C12N 2506/1353 from bone marrow mesenchymal stem cells (BM-MSC)
• CPC Class Codes: C12N 2510/02 Cells for production
• CPC Class Codes: C12N 5/0676 Pancreatic cells

Abstract

The subject invention comprises culture methods for transdifferentiation of non-pancreatic stem cells to the pancreatic differentiation pathway. It also concerns the endocrine hormones that can be produced by such cultures, and the use of the transdifferentiated cells in the treatment of pancreatic diseases.

Claims

1. A method of transdifferentiating mammalian non-pancreatic stem cells to enter the pancreatic differentiation pathway, comprising:

culturing said stem cells in a medium that permits the expression of pancreatic differentiation markers.

2. The method of claim 1 wherein said medium comprises Dulbecco'"'"'s Minimal Essential Medium (DMEM) with high glucose and sodium pyruvate, bovine serum albumin (BSA), 2-mercaptoethanol, fetal calf serum (FCS), penicillin and streptomycin (Pen-Strep), and insulin, transferrin and selenium (ITS) for at least about 10-14 days.

3. The method of claim 1 wherein the pancreatic markers that are expressed are selected from the group consisting of Pdx-1, Isl-1, Pax-4, Pax-6, Glut-2, CK-19 and insulin.

4. The method of claim 1, wherein the medium further contains an effective combination and effective concentration of growth factors selected from the group consisting of stem cell factor (SCF), glucagon-like peptide-1 (GLP-1), activin A, betacellulin, hepatocyte growth factor (HGF), epidermal growth factor (EGF), keratinocyte growth factor (KGF), prolactin (PRL), nerve growth factor (NGF), transforming growth factor-α

(TGF-α

), gastrin, and conditioned medium of a cell line selected from the group consisting of Capan-1, ARIP and AR42J.

5. The method of claim 4 wherein the growth factors comprise SCF, GLP-1, activin A, betacellulin, HGF, EGF, KGF, PRL, NGF, TGF-α

, gastrin and conditioned medium of a cell line selected from the group consisting of Capan-1, ARIP and AR42J.

6. The method of claim 1 wherein said non-pancreatic stem cells are selected from the group consisting of hematopoietic stem cells (HSCs) and mesenchymal stem cells (MSCs).

7. The method of claim 6 wherein the HSCs are CD34+.

8. The method of claim 6 wherein the MSCs are CD105+, CD166+, CD29+ and CD44+.

9. The method of claim 1 wherein the stem cells are human.

10. A method of producing an endocrine hormone comprising the method of claim 1, and further comprising the step of continuing to culture said transdifferentiated cells in said medium, whereby an endocrine hormone may be produced.

11. The endocrine hormones produced by the method of claim 9.

12. A method of treating a mammal with a pancreatic disorder, comprising:

a) culturing non-pancreatic stem cells according to claim 1, whereby said stem cells enter the pancreatic differentiation pathway;

b) using a product of the culture of step (a) to treat said mammal.

13. The method of claim 12 wherein the pancreatic disorder is insulin-dependent diabetes.

14. The method of claim 12 wherein the product in step (b) is an endocrine hormone that is administered to the mammal.

15. The method of claim 12 wherein the product of claim 12, step (b), is transdifferentiated cells, and said method further comprises:

c) implanting said product in said mammal.

16. The method of claim 15, wherein said implantation is in pancreatic, kidney or liver tissue, or in a subcutaneous pocket.

17. The method of claim 15, wherein said transdifferentiated cells are selected from the group consisting of transdifferentiated HSCs and transdifferentiated MSCs.

18. The method of claim 15, wherein said transdifferentiated cells are modified to substantially reduce expression of an antigen selected from the group consisting of insulin dependent diabetes associated autoantigens, GAD, 64 kD islet cell surface antigen and human leukocyte antigens, whereby the modified transdifferentiated cells do not substantially express said antigen.

19. The method of claim 12, wherein the non-pancreatic stem cells originate from an individual that is the same as the treated mammal of step (b).

20. The method of claim 15, wherein said implanted transdifferentiated cells are encapsulated in an endocrine hormone permeable capsule.

21. The method of claim 12, wherein the stem cells are human.

22. The transdifferentiated MSC produced by the method of claim 17.

24. The transdifferentiated HSC produced by the method of claim 17.

26. The transdifferentiated cell produced by the method of claim 1.

23. A transdifferentiated MSC that expresses mRNA for Isl-1, Pax-6, Glut-2, glucagon and optionally Pdx-1 or Pax4.

25. A transdifferentiated HSC that expresses mRNA for Isl-1, Pax-6, CK-19 and optionally insulin.

27. A therapeutic composition comprising a transdifferentiated MSC or transdifferentiated HSC encapsulated in an endocrine hormone permeable capsule.