Neural stem cells

• US 9,309,495 B2
• Filed: 07/21/2014
• Issued: 04/12/2016
• Est. Priority Date: 06/09/2004
• Status: Active Grant

• Current Assignee: University of Edinburgh
• Original Assignee: University of Edinburgh
• Inventors: Conti, Luciano, Pollard, Steven Michael, Smith, Austin Gerard
• Primary Examiner(s): BERTOGLIO, VALARIE E
• Application Number:
  US14/336,513
• Publication Number:
  US 20140370596A1
• Time in Patent Office: 631 Days
• Field of Search: 435/368, 435/375
• US Class Current: 1/1
• 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 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 25/02 for peripheral neuropathies
• CPC Class Codes: A61P 25/14 for treating abnormal movements, e.g. chorea, dyskinesia
• 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 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 2501/11 Epidermal growth factor [EGF]
• CPC Class Codes: C12N 2501/115 Basic fibroblast growth factor (bFGF, FGF-2)
• CPC Class Codes: C12N 2506/02 from embryonic cells
• CPC Class Codes: C12N 2533/52 Fibronectin; Laminin
• CPC Class Codes: C12N 5/0623 Stem cells

Abstract

A homogenous, symmetrically dividing population of adherent neural stem cells is obtained from ES cells or foetal or adult brain isolates, using an activator of a signalling pathway downstream of a receptor of the EGF receptor family, optionally in combination with an activator of a signalling pathway downstream of an FGF receptor. The neural stem cell population is highly pure and retains the ability to differentiate into neurons after in excess of 100 passages.

Claims

1. A composition, comprising (a) neural stem cells;

and either (b) EGF;

or (c) EGF and FGF-2;

wherein at least 80% of said cells are symmetrically dividing neural stem cells and wherein no more than 1% of the cells are positive for the expression of markers from mature astrocytes, neurons and oligodendrocytes, and wherein both EGF and the combination of EGF and FGF-2 promote the symmetrical division of the stem cells.

2. The composition of claim 1, wherein the neural stem cells are in a feeder-free adherent monolayer culture.

3. The composition of claim 1, wherein at least 95% of said neural stem cells are symmetrically-dividing neural stem cells.

4. The composition of claim 1, wherein at least 97% of said neural stem cells are symmetrically-dividing neural stem cells.

5. The composition of claim 1, wherein the neural stem cells are characterized in that they are positive for the expression of RC2, 3CB2, BLBP, and SOX-2.

6. The composition of claim 5, wherein the neural stem cells are further characterized in that they are positive for the expression of at least one of (i) GLAST, (ii) Pax-6, (iii) the neural precursor markers nestin or vimentin, (iv) the LewisX antigen, (v) Musashi-1, and (vi) prominin.

7. The composition of claim 6, wherein the neural stem cells are further characterised in that they are negative for the expression of at least one of Oct4 and Nanog.

8. The composition of claim 6, wherein the neural stem cells are positive for the expression of Sox-2, and negative for the expression of Sox-1.

14. The composition of claim 1, comprising neural stem cells and EGF and FGF-2.

9. A cell line comprising neural stem cells, wherein said neural stem cells are maintained in the presence of either EGF or EGF and FGF-2, wherein at least 80% of said cells are symmetrically-dividing neural stem cells and wherein no more than 1% of the cells are positive for the expression of markers for mature astocytes, neurons, and oligodendrocytes, wherein both EGF and the combination of EGF and FGF-2 promote the symmetrical division of the stem cells.

10. The cell line of claim 9, wherein the cell line is a human neural stem cell line.

11. The cell line of claim 9, wherein the cell line is a mouse neural stem cell line, and wherein at least 80% of said cells are symmetrically-dividing neural stem cells and wherein no more than 1% of cells in the line are positive for expression of GFAP or β

III tubulin.

15. The cell line of claim 9, wherein the neural stem cells are maintained in the presence of EGF and FGF-2.

12. A composition comprising (a) neural stem cells and either (b) EGF or (c) EGF and FGF-2, wherein at least 95% of the cells in the composition are symmetrically-dividing neural stem cells, and wherein both EGF and the combination of EGF and FGF-2 promote the symmetrical division of the stem cells.

13. The composition of claim 12, wherein the neural stem cells are in a feeder-free adherent monolayer culture.

16. The composition of claim 12, wherein the composition comprises neural stem cells and EGF and FGF-2.