Material and methods for the growth of hematopoietic stem cells

• US 7,790,458 B2
• Filed: 05/13/2005
• Issued: 09/07/2010
• Est. Priority Date: 05/14/2004
• Status: Expired due to Fees

• Current Assignee: Becton, Dickinson & Co
• Original Assignee: Becton, Dickinson & Co
• Inventors: DeLuca, Keith, Xu, Ruiling, Heidaran, Mohammad, Liebmann-Vinson, Andrea
• Primary Examiner(s): Crouch; Deborah
• Assistant Examiner(s): SINGH, ANOOP KUMAR
• Application Number:
  US11/128,950
• Publication Number:
  US 20050272152A1
• Time in Patent Office: 1,943 Days
• Field of Search: 435/325, 435/372, 435/404, 435/386, 435/405, 435/406, 435/384
• US Class Current: 435/406
• CPC Class Codes: A61P 19/08 for bone diseases, e.g. rachitism, Paget's disease
• CPC Class Codes: A61P 9/00 Drugs for disorders of the cardiovascular system
• CPC Class Codes: C12N 2500/25 Insulin-transferrin; Insulin-transferrin-selenium
• CPC Class Codes: C12N 2500/36 Lipids
• CPC Class Codes: C12N 2500/42 Organic phosphate, e.g. beta glycerophosphate
• CPC Class Codes: C12N 2500/90 Serum-free medium, which may still contain naturally-sourced components
• CPC Class Codes: C12N 2501/10 Growth factors
• CPC Class Codes: C12N 2501/105 Insulin-like growth factors [IGF]
• CPC Class Codes: C12N 2501/11 Epidermal growth factor [EGF]
• CPC Class Codes: C12N 2501/115 Basic fibroblast growth factor (bFGF, FGF-2)
• 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 2501/135 Platelet-derived growth factor [PDGF]
• CPC Class Codes: C12N 2501/14 Erythropoietin [EPO]
• CPC Class Codes: C12N 2501/145 Thrombopoietin [TPO]
• CPC Class Codes: C12N 2501/15 Transforming growth factor beta (TGF-β)
• CPC Class Codes: C12N 2501/155 Bone morphogenic proteins [BMP]; Osteogenins; Osteogenic factor; Bone inducing factor
• CPC Class Codes: C12N 2501/165 Vascular endothelial growth factor [VEGF]
• CPC Class Codes: C12N 2501/21 Chemokines, e.g. MIP-1, MIP-2, RANTES, MCP, PF-4
• 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 2501/2303 : Interleukin-3 (IL-3)
• CPC Class Codes: C12N 2501/2305 : Interleukin-5 (IL-5)
• CPC Class Codes: C12N 2501/2306 : Interleukin-6 (IL-6)
• CPC Class Codes: C12N 2501/2311 : Interleukin-11 (IL-11)
• CPC Class Codes: C12N 2501/235 : Leukemia inhibitory factor [LIF]
• CPC Class Codes: C12N 2501/26 : Flt-3 ligand (CD135L, flk-2 ligand)
• CPC Class Codes: C12N 2501/32 : Angiotensins [AT], angiotensinogen
• CPC Class Codes: C12N 2501/39 : Steroid hormones
• CPC Class Codes: C12N 2501/415 : Wnt; Frizzeled
• CPC Class Codes: C12N 2501/58 : Adhesion molecules, e.g. ICAM, VCAM, CD18 (ligand), CD11 (ligand), CD49 (ligand)
• CPC Class Codes: C12N 2501/60 : Transcription factors
• CPC Class Codes: C12N 2501/70 : Enzymes
• CPC Class Codes: C12N 2501/999 : Small molecules not provided for elsewhere
• CPC Class Codes: C12N 2533/52 : Fibronectin; Laminin
• CPC Class Codes: C12N 2533/54 : Collagen; Gelatin
• CPC Class Codes: C12N 2533/70 : Polysaccharides
• CPC Class Codes: C12N 5/0647 : Haematopoietic stem cells; Uncommitted or multipotent progenitors
• CPC Class Codes: C12N 5/0663 : Bone marrow mesenchymal stem cells (BM-MSC)

Abstract

The invention provides populations of expanded CD34-expressing cells and methods of use. Particular embodiments provide for defined culture media useful for growing these cells, and grafts comprising these cells. The invention finds use in methods for reconstituting, repairing, and regenerating tissue damage.

Claims

1. A cell culture medium for expanding a population of stem and progenitor cells comprising basic fibroblast growth factor (bFGF;

1-5 ng/ml), erythropoietin (EPO;

1-6 units/ml), Flt-3/Flk-2 ligand (2-10 ng/ml), interleukin-11 (IL-11;

0.05-0.2 ng/ml), interleukin-6 (IL-6;

0.1-0.5 ng/ml), platelet-derived endothelial cell growth factor (PD-ECGF;

1-4 ng/ml), thrombospondin (5-20 ng/ml), and thrombopoietin (TPO;

0.5-2 ng/ml).

2. The cell culture medium of claim 1, wherein the cell culture medium comprises bFGF (2.5 ng/ml), EPO (3 units/ml), Flt-3/Flk-2 ligand (5 ng/ml), IL-11 (0.1 ng/ml), IL-6 (0.2 ng/ml), PD-ECGF (2 ng/ml), thrombospondin (10 ng/ml), and TPO (1 ng/ml).

3. A method of producing an expanded population of hematopoietic stem and progenitor cells expressing the CD34 cell surface marker, said method comprising culturing a population of CD34⁺ stem and progenitor cells in a cell culture medium comprising bFGF (1-5 ng/ml), EPO (1-6 units/ml), Flt-3/Flk-2 ligand (2-10 ng/ml), IL-11 (0.05-0.2 ng/ml), IL-6 (0.1-0.5 ng/ml), PD-ECGF (1-4 ng/ml), thrombospondin (5-20 ng/ml), and TPO (0.5-2 ng/ml), thereby producing an expanded population of hematopoietic stem and progenitor cells expressing the CD34 cell surface marker.

4. The method of claim 3, wherein the cell culture medium comprises bFGF (2.5 ng/ml), EPO (3 units/ml), Flt-3/Flk-2 ligand (5 ng/ml), IL-11 (0.1 ng/ml), IL-6 (0.2 ng/ml), PD-ECGF (2 ng/ml), thrombospondin (10 ng/ml), and TPO (1 ng/ml).