Generation of neural precursor cell lines

• US 5,580,777 A
• Filed: 10/27/1994
• Issued: 12/03/1996
• Est. Priority Date: 10/29/1987
• Status: Expired due to Term

• Current Assignee: Zenyth Therapeutics Limited (CSL Limited)
• Original Assignee: Zenyth Therapeutics Limited (CSL Limited)
• Inventors: Bartlett, Perry F., Bernard, Ora
• Primary Examiner(s): Ziska, Suzanne E.
• Application Number:
  US08/330,114
• Time in Patent Office: 768 Days
• Field of Search: 435/172.3, 435/320.1, 435/240.2
• US Class Current: 435/456
• CPC Class Codes: C07K 14/82 Translation products from oncogenes
• CPC Class Codes: C12N 15/86 Viral vectors
• CPC Class Codes: C12N 2510/04 Immortalised cells
• CPC Class Codes: C12N 2740/10043 viral genome or elements thereof as genetic vector
• CPC Class Codes: C12N 2740/13043 viral genome or elements thereof as genetic vector
• CPC Class Codes: C12N 5/0623 Stem cells

Abstract

A method for the in vitro production of lines of immortalized neural precursor cells, including cell lines having neuronal and/or glial cell characteristics, comprises the step of infecting neuroepithelium or neural crest cells with a retroviral vector carrying a member of the myc family of oncogenes.

Claims

1. An cell line comprising mammalian neuroepithelial or neural crest cells which have been infected with a retroviral vector carrying a c-myc or N-myc oncogene and in which said c-myc or N-myc gene is expressed.

10. The cell line according to claim 1, wherein said retrovital vector comprises the vector pDol carrying the c-myc oncogene.

11. A cell line according to claim 1, wherein said retroviral vector comprises a vector selected from the group consisting of pZen, pZenSVNeo, pMPZen and pMPZenSVNeo, carrying either the c-myc or the N-myc oncogene.

2. A method for the in vitro production of immortalized mammalian neural precursor cells which comprises infecting mammalian neuroepithelium or neural crest cells with a retroviral vector carrying a c-myc or N-myc oncogene to provide infected cells, wherein said neuroepithelium or neural crest cells are capable of being infected by said retroviral vector, and under conditions whereby the c-myc or N-myc gene is expressed in said infected cells.

3. A method according to claim 2 wherein said retroviral vector comprises the vector pDol carrying the c-myc oncogene.

4. The method according to claim 3 wherein infection of mammalian neuroepithelium or neural crest cells with said retroviral vector is accomplished by:

(a) transfecting fibroblast cells with said retroviral vector to produce virus-producing fibroblast cells, wherein said fibroblast cells are capable of being transfected by said retroviral vector, expressing retrovirus genes borne by said vector, and producing and packaging retroviral particles; and

(b) co-cultivating said virus-producing fibroblast cells with said neuroepithelial or neural crest cells to infect said neuroepithelial or neural crest cells with said retroviral vector.

5. A method according to claim 2, wherein said immortalised neural precursor cells are induced with acidic or basic fibroblast growth factor to differentiate to form at least one of neuronal cells and glial cells.

6. The method according to claim 2 wherein said immortalised neural precursor cells do not differentiate spontaneously in vitro.

7. The method according to claim 2 wherein said retroviral vector comprises a vector selected from the group consisting of pZen, pZenSVNeo, pMPZen and pMPZenSVNeo.

8. The method according to claim 7 wherein infection of mammalian neuroepithelium or neural crest cells with said retroviral vector is accomplished by:

(a) transfecting fibroblast cells with said retroviral vector to produce virus-producing fibroblast cells, wherein said fibroblast cells are capable of being transfected by said retroviral vector, expressing retrovirus genes borne by said vector, and producing and packaging retroviral particles; and

(b) co-cultivating said virus-producing fibroblast cells with said neuroepithelial or neural crest cells to infect said neuroepithelial or neural crest cells with said retroviral vector.

9. The method according to claim 8, wherein said imortalised neural cells are induced with acidic or basic fibroblast growth factor to differentiate to form at least one of neuronal cells and glial cells.

12. The method of claim 4 wherein the fibroblast cells are psi-2 cells.