Tissue specific expression of antibodies in chickens

Abstract

Transgenes encoding exogenous antibodies are stably integrated into donor cells and are present in the somatic tissue of chimeric birds. The transgenes encode exogenous antibodies and are preferably expressed in the oviduct for collection in the egg. Tissue specificity is provided by selecting the content of the transgene accordingly. Birds whose genome is comprised of trangene-derived exogenous antibody-encoding DNA express exogenous antibodies having desirable chemical properties with increased therapeutic utility compared to antibodies derived from bacterial expression systems.

Description

PRIORITY INFORMATION [0001] This application is a continuation in part of a U.S. application filed on Feb. 9, 2005 entitled “Tissue Specific Expression of Exogenous Proteins in Transgenic Chickens,” Ser. No. 11 / ______, which is a national phase filing of PCT / US2003 / 025270, publication number WO 2004 / 015123 A1 filed on Aug. 11, 2003.[0002] An invention described herein may have been made with Government support under USDA SBIR 2003-33610 and NIH 2 R44 GM64261-01 and 2 R44 HD 039583-02. The U.S. Government may have certain rights in this invention.BACKGROUND OF THE INVENTION [0003] Genetically modified animals offer the potential for tremendous advances in the sustainable production of valuable pharmaceutical products, such as antibodies. However, the production of genetically modified animals involves significant technical hurdles that have only been overcome for a few species. The ability to incorporate genetic modifications encoding proteins into the DNA of a species for a specific...

AI Technical Summary

Benefits of technology

[0012] The present invention includes populations of birds exhibiting tissue specific expression of antibodies, transgene constructs that enable exogenous antibody expression, isolated compositions of antibodies produced in chickens and having specially defined chemical properties, and related methods for creation of the birds, production of the antibodies and their therapeutic use in humans. The invention uses long term cell cultures and special techniques to produce chimeric or transgenic birds derived from prolonged cell cultures, wherein the genome of the ES cells have a stably integrated transgene expressing an exogenous protein such that progeny of the cultured cells contain the stably integrated transgene. When introduced to a host avian embryo, by the procedures described below, those modified donor cells produce birds that express the transgene into specific, selected somatic tissue of the resulting animals. These birds exhibit a donor-cell derived phenotype and express the antibody in the oviduct to facilitate concentration and collection of the antibody in egg white.

[0013] This invention also includes compositions of antibodies expressed in the chicken expression system and having certain desirable chemical properties compared to vertebrate, plant, or bacterial cell systems. Specifically, the antibodies have reduced concentrations of fucose, galactose, N-acetyl neuraminic acid, N-glycolylneuraminic acid and elevated concentrations of mannose. Antibodies having some or all of these properties exhibit increased therapeutic utility when administered to a human. Specifically, these antibody compositions exhibit enhanced antibody-dependent cellular cytotoxicity (ADCC). Accordingly, the methods of the invention include enhancing the therapeutic utility, based on the ADCC effect, of compositions of antibodies by expressing them in a chicken. In practical application, the methods comprise administering the antibody composition described herein and detecting cellular cytotoxicity in the patient. The invention also includes chickens expressing exogenous antibody, having the advantageous chemistry defined herein, in the oviduct tissue such that exogenous antibody is concentrated in defined quantities in the egg white. In one preferred embodiment, the exogenous protein is a human sequence monoclonal antibody encoded by the transgene construct incorporated into the genome of a donor cell line and progeny. The human monoclonal antibody encoding polynucleotide sequence is contained within a transgene that is specifically constructed for expression in the oviduct and which contains appropriate promoters and regulatory sequences to facilitate tissue specific expression. In the embodiment of a transgenic or chimeric bird expressing exogenous proteins, the invention includes compositions specific to the animal and the protein, such as egg white albumen containing exogenous antibodies as described herein.

Problems solved by technology

However, the production of genetically modified animals involves significant technical hurdles that have only been overcome for a few species.

However, the collection of a valuable antibody from the tissues of an animal typically requires that the expression be limited to certain specific tissue types that facilitate collection of the expressed protein and convey desirable chemical properties.

Under such circumstances, it is unlikely that a meaningful quantity of the antibody could be separated from the animal, the ubiquitous expression of an exogenous antibody is usually very damaging to the overall health and well being of the animal, and the desirable chemical properties exhibited in the chicken oviduct are not present.

The transgenes that enable tissue specific expression are complex and the genetic manipulations that are necessary to incorporate the transgenes into a recipient cell line require extensive manipulation that can threaten the pluripotency of the cells unless the culture conditions are optimized.

Images