Use of Human Gamma Satellite Insulator Sequences to Prevent Gene Silencing and Allow for Long Term Expression of Integrated Tran
Posted Jun 30 2008 5:00pm
Description of Invention: The lack of stable expression of transgenes in target cell lines remains a serious problem for gene therapy and cellular reprogramming approaches. Once integrated into chromosomes, the expression of these transgenes may be regulated by epigenetic effects of the surrounding chromatin. These position effects, which include transgene silencing and expression variegation, are often associated with changes in the chromatin structure, and are capable of inhibiting gene expression and neutralizing the intended effect of the inserted transgene.
Experimental results suggest that gene position effects can be partially overcome by flanking the transgene with regulatory elements called chromatin insulators which work by establishing defined domains of transcriptional activity within the eukaryotic genome. These insulators can partially overcome position effects by shielding the promoters from the influence of neighboring regulatory elements, or by preventing the spread of heterochromatin which can lead to subsequent gene silencing.
This invention discloses the use of gamma satellite DNA, residing in the pericentromeric region of human chromosomes, as highly efficient chromatin insulators. These insulators have a remarkable ability to overcome position effects and prevent the silencing of transgenes. When human chromosome 8 gamma satellite sequences were used as flanking DNA for eGFP (enhanced green fluorescent protein) gene expression in mouse erythroleukemia (MEL) cells, stable transgene expression was recorded for well over eight months. Until recently, no chromatin insulator sequences were known to completely prevent gene silencing on a long term basis in transfected cells. The human gamma-satellite sequences demonstrate a higher efficiency than any known chromatin insulator identified so far in intergenic regions, and may have invaluable applications in the fields of gene therapy, protein expression, and cellular reprogramming where adequate expression of the transgene is essential for long term therapeutic or developmental success.
Applications:
Gene therapy
Protein expression
Cellular reprogramming
Development Status: Prolonged transgene expression attained in mouse erythroleukemia (MEL) cells
Inventors: Jung-Hyun Kim (301-480-) Vladimir Larionov (NCI) Tom A Ebersole (NCI)
G Felsenfeld, B Burgess-Beusse, C Farrell, M Gaszner, R Ghirlando, S Huang, C Jin, M Litt, F Magdinier, V Mutskov, Y Nakatani, H Tagami, A West, T Yusufzai. Chromatin boundaries and chromatin domains. Cold Spring Harb Symp Quant Biol. 2004;69:245-250.
T Ebersole, Y Okamoto, VN Noskov, N Kouprina, JH Kim, SH Leem, JC Barrett, H Masumoto, V Larionov. Rapid generation of long synthetic tandem repeats and its application for analysis in human artificial chromosome formation. Nucleic Acids Res. 2005 Sep 1;33(15):e130, doi:10.1093/nar/gni129. [ PubMed abs ]
Licensing Status: Available for licensing.
Collaborative Research Opportunity: The National Cancer Institute Laboratory of Molecular Pharmacology is seeking statements of capability or interest from parties interested in collaborative research to further develop, evaluate, or commercialize gamma-satellite DNA insulators for stable transgene expression in ectopic chromosomal sites and in Human Artificial Chromosomes (HACs). Please contact John D. Hewes, Ph.D. at 301-435-3121 or hewesj@mail.nih.gov for more information.
Portfolios: Gene Based Therapies Gene Based Therapies - Therapeutics Internal Medicine Internal Medicine - Therapeutics
For Additional Information Please Contact: Suryanarayana Vepa Ph.D. NIH Office of Technology Transfer 6011 Executive Blvd. Suite 325, Rockville, MD 20852 United States Email: vepas@mail.nih.gov Phone: 301-435-5020 Fax: 301-402-0220
Description of Invention:
The lack of stable expression of transgenes in target cell lines remains a serious problem for gene therapy and cellular reprogramming approaches. Once integrated into chromosomes, the expression of these transgenes may be regulated by epigenetic effects of the surrounding chromatin. These position effects, which include transgene silencing and expression variegation, are often associated with changes in the chromatin structure, and are capable of inhibiting gene expression and neutralizing the intended effect of the inserted transgene.
Experimental results suggest that gene position effects can be partially overcome by flanking the transgene with regulatory elements called chromatin insulators which work by establishing defined domains of transcriptional activity within the eukaryotic genome. These insulators can partially overcome position effects by shielding the promoters from the influence of neighboring regulatory elements, or by preventing the spread of heterochromatin which can lead to subsequent gene silencing.
This invention discloses the use of gamma satellite DNA, residing in the pericentromeric region of human chromosomes, as highly efficient chromatin insulators. These insulators have a remarkable ability to overcome position effects and prevent the silencing of transgenes. When human chromosome 8 gamma satellite sequences were used as flanking DNA for eGFP (enhanced green fluorescent protein) gene expression in mouse erythroleukemia (MEL) cells, stable transgene expression was recorded for well over eight months. Until recently, no chromatin insulator sequences were known to completely prevent gene silencing on a long term basis in transfected cells. The human gamma-satellite sequences demonstrate a higher efficiency than any known chromatin insulator identified so far in intergenic regions, and may have invaluable applications in the fields of gene therapy, protein expression, and cellular reprogramming where adequate expression of the transgene is essential for long term therapeutic or developmental success.
Applications:
Development Status:
Prolonged transgene expression attained in mouse erythroleukemia (MEL) cells
Inventors:
Jung-Hyun Kim (301-480-)
Vladimir Larionov (NCI)
Tom A Ebersole (NCI)
Patent Status:
HHS, Reference No. E-154-2006/0
US, Application No. 12/527,122 filed 13 Aug 2009
Relevant Publication:
Licensing Status:
Available for licensing.
Collaborative Research Opportunity:
The National Cancer Institute Laboratory of Molecular Pharmacology is seeking statements of capability or interest from parties interested in collaborative research to further develop, evaluate, or commercialize gamma-satellite DNA insulators for stable transgene expression in ectopic chromosomal sites and in Human Artificial Chromosomes (HACs). Please contact John D. Hewes, Ph.D. at 301-435-3121 or hewesj@mail.nih.gov for more information.
Portfolios:
Gene Based Therapies
Gene Based Therapies - Therapeutics
Internal Medicine
Internal Medicine - Therapeutics
For Additional Information Please Contact:
Suryanarayana Vepa Ph.D.
NIH Office of Technology Transfer
6011 Executive Blvd. Suite 325,
Rockville, MD 20852
United States
Email: vepas@mail.nih.gov
Phone: 301-435-5020
Fax: 301-402-0220
Ref No: 1771
Updated: 07/2008