Method of Inhibiting the Activity of an Intracellular Constituent
Description of Invention:
Two combinatorial libraries of binding proteins have been engineered. The libraries were designed to genetically shuffle oligonucleotide motifs within the framework of the immunoglobulin heavy chain gene by random mutation of either the CDRI or CDRIII hypervariable regions. The Fd fragment of the heavy chain gene was then reconstructed such that it contained the randomized oligonucleotides in the hypervariable region, resulting in a collection of highly diverse sequences. The libraries of heavy chain proteins encoded by the array of mutated gene sequences potentially have all of the binding characteristics of an immunoglobulin while requiring only the heavy chain Fd protein.
The re-engineered heavy chain gene sequences were ligated into a M13-derived bacteriophage vector that permits expression of the binding proteins as fusion proteins with viral protein 8, which is expressed on the phage surface.
The claims of the patent application provide methods to screen the libraries, to identify the binding protein to a specific antigen and the gene for that specific protein, and to re-engineer the gene for intracellular expression in a eukaryotic cell. Inducible intracellular inactivation of glucose-6-phosphate dehydrogenase (G6PDH) has been demonstrated by in vivo expression of a gene construct encoding a binding protein selected from one of the libraries and specific for G6PDH. Removal of induction restored the enzyme activity.
The libraries of binding proteins, the screening methods, and the methods of inhibiting intracellular components claimed in the patent application provide powerful potential tools for cellular and molecular biology by affording the capability of binding/inactivating any protein of choice.
Inventors:
MJ Mulligan-Kehoe (NCI)
Patent Status:
DHHS Reference No. E-051-1995/0 --
U.S. Patent 5,702,892 issued 30 Dec 1997
U.S. Patent 5,824,520 issued 20 Oct 1998
Portfolios: Cancer
Cancer -Therapeutics-Immunoconjugates-Other Cancer -Therapeutics
For Additional Information Please Contact: Jasbir (Jesse) S. Kindra J.D.
NIH Office of Technology Transfer
6011 Executive Blvd, Suite 325
Rockville, MD 20852-3804
Phone: (301) 435-5559
Email: kindraj@mail.nih.gov
Fax: (301) 402-0220
