Originally published in Science Express as 10.1126/science.1078942 on December 19, 2002
Science, Vol. 299, Issue 5605, 411-414, January 17, 2003
Production of 1,3-Galactosyltransferase-Deficient Pigs
Carol J. Phelps,1 Chihiro Koike,34 Todd D. Vaught,1 Jeremy Boone,1 Kevin D. Wells,1 Shu-Hung Chen,1 Suyapa Ball,1 Susan M. Specht,34 Irina A. Polejaeva,1 Jeff A. Monahan,1 Pete M. Jobst,1 Sugandha B. Sharma,34 Ashley E. Lamborn,1 Amy S. Garst,1 Marilyn Moore,2 Anthony J. Demetris,35 William A. Rudert,36 Rita Bottino,36 Suzanne Bertera,36 Massimo Trucco,36 Thomas E. Starzl,34 Yifan Dai,1* David L. Ayares1*
The enzyme 1,3-galactosyltransferase (1,3GT or GGTA1) synthesizes 1,3-galactose (1,3Gal) epitopes (Gal1,3Gal1,4GlcNAc-R), which are the major xenoantigens causing hyperacute rejection in pig-to-human xenotransplantation. Complete removal of 1,3Gal from pig organs is the critical step toward the success of xenotransplantation. We reported earlier the targeted disruption of one allele of the 1,3GT gene in cloned pigs. A selection procedure based on a bacterial toxin was used to select for cells in which the second allele of the gene was knocked out. Sequencing analysis demonstrated that knockout of the second allele of the 1,3GT gene was caused by a T-to-G single point mutation at the second base of exon 9, which resulted in inactivation of the 1,3GT protein. Four healthy 1,3GT double-knockout female piglets were produced by three consecutive rounds of cloning. The piglets carrying a point mutation in the 1,3GT gene hold significant value, as they would allow production of 1,3Gal-deficient pigs free of antibiotic-resistance genes and thus have the potential to make a safer product for human use.
1 PPL Therapeutics Inc., 1700 Kraft Drive, Blacksburg, VA 24060, USA.
2 PPL Therapeutics Ltd., Roslin, Midlothian, EH25 9PP, UK.
3 Thomas E. Starzl Transplantation Institute,
4 Department of Surgery,
5 Department of Pathology, and
6 Department of Pediatrics (Division of Immunogenetics) of University of Pittsburgh Medical Center (UPMC) and Children's Hospital, Pittsburgh, PA 15213, USA.
* To whom correspondence should be addressed. E-mail: ydai@ppl-therapeutics.com; dayares@ppl-therapeutics.com
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