Methods for inducing differentiation of embryonic stem cells and uses thereof

• US 7,390,659 B2
• Filed: 07/16/2002
• Issued: 06/24/2008
• Est. Priority Date: 07/16/2002
• Status: Expired due to Fees

• Current Assignee: Trustees Of Columbia University In The City Of New York (Columbia University)
• Original Assignee: Trustees Of Columbia University In The City Of New York (Columbia University)
• Inventors: Lieberam, Ivo, Wichterle, Hynek, Jessell, Thomas
• Primary Examiner(s): Rao; Manjunath
• Assistant Examiner(s): Gamett; Daniel C
• Application Number:
  US10/196,882
• Publication Number:
  US 20040014210A1
• Time in Patent Office: 2,170 Days
• Field of Search: 435/6, 435/7.1, 435/325, 435/7.21, 435/368
• US Class Current: 435/377
• 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 21/00 Drugs for disorders of the muscular or neuromuscular system
• 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 25/16 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 31/12 Antivirals
• CPC Class Codes: C12N 2501/385 of the family of the retinoic acid recptor, e.g. RAR, RXR; Peroxisome proliferator-activated receptor [PPAR]
• CPC Class Codes: C12N 2501/41 Hedgehog proteins; Cyclopamine (inhibitor)
• CPC Class Codes: C12N 2506/02 from embryonic cells
• CPC Class Codes: C12N 5/0619 Neurons
• CPC Class Codes: Y02A 50/30 Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Abstract

The present invention provides a method for inducing differentiation of an embryonic stem cell into a differentiated neural cell. The present invention further provides a method for producing differentiated neural cells, and a population of cells comprising the differentiated neural cells. Additionally, the present invention provides a method for repopulating a spinal cord in a subject, and a method for treating nervous tissue degeneration in a subject in need of treatment. The present invention further provides neural progenitor cells, differentiated neural cells, and uses of same. Also provided is a transgenic non-human animal containing the differentiated neural cells. The present invention is further directed to a method for isolating a population of differentiated neural cells. Finally, the present invention provides a method for identifying an agent for use in treating a condition associated with neuron degeneration.

Claims

1. A method for identifying a candidate agent for treating a condition associated with motor neuron degeneration, comprising the steps of:

(a) obtaining or generating a culture of embryonic stem cells, wherein said embryonic stem cells contain a mutation of a gene selected from the group consisting of a superoxide dismutase gene and a survival motor neuron protein gene, said mutation associated with motor neuron degeneration;

(b) contacting the culture of embryonic stem cells with an amount of retinoic acid effective to produce neural progenitor cells;

(c) activating a Hedgehog signaling pathway in the neural progenitor cells to produce cells expressing HB9, wherein the Hedgehog signaling pathway is selected from the group consisting of the Sonic hedgehog, Indian hedgehog, and Desert hedgehog signaling pathways;

(d) contacting the cells with an agent; and

(e) determining if the agent enhances growth of the cells as compared to growth of the cells in the absence of the agent;

wherein the ability of the agent to enhance growth of some or all of the cells indicates that it is a candidate agent for use in treating a condition associated with motor neuron degeneration.

2. The method of claim 1, wherein the cells expressing HB9 express enhanced green fluorescent protein.

3. A method for identifying a candidate agent for treating a condition associated with motor neuron degeneration, comprising the steps of:

(a) obtaining or generating a culture of human embryonic stem cells, wherein said embryonic stem cells contain a mutation of a gene selected from the group consisting of a superoxide dismutase gene and a survival motor neuron protein gene, said mutation associated with motor neuron degeneration;

(b) contacting the culture of human embryonic stem cells with an amount of retinoic acid effective to produce neural progenitor cells;

(c) activating a Hedgehog signaling pathway in the neural progenitor cells to produce cells expressing HB9, wherein the Hedgehog signaling pathway is selected from the group consisting of the Sonic hedgehog, Indian hedgehog, and Desert hedgehog signaling pathways;

(d) contacting the cells with an agent; and

(e) determining if the agent enhances growth of the cells as compared to growth of the cells in the absence of the agent;

wherein the ability of the agent to enhance growth of some or all of the cells indicates that it is a candidate agent for use in treating a condition associated with motor neuron degeneration.

4. The method of claim 3, wherein the cells expressing HB9 express enhanced green fluorescent protein.