Correction of a genetic defect by nuclear transplantation and combined cell and gene therapy.
Immune-deficient Rag2(-/-) mice were used as nuclear donors for transfer into enucleated oocytes, and the resulting blastocysts were cultured to isolate an isogenic embryonic stem cell line. One of the mutated alleles in the Rag2(-/-) ES cells was repaired by homologous recombination, thereby restoring normal Rag2 gene structure. Mutant mice were treated with the repaired ES cells in two ways. (1) Immune-competent mice were generated from the repaired ES cells by tetraploid embryo complementation and were used as bone marrow donors for transplantation. (2) Hematopoietic precursors were derived by in vitro differentiation from the repaired ES cells and engrafted into mutant mice. Mature myeloid and lymphoid cells as well as immunoglobulins became detectable 3-4 weeks after transplantation. Our results establish a paradigm for the treatment of a genetic disorder by combining therapeutic cloning with gene therapy.
Rideout
William M
WM
Whitehead Institute for Biomedical Research, 9 Cambridge Center, Cambridge, MA 02142, USA.
Hochedlinger
Konrad
K
Kyba
Michael
M
Daley
George Q
GQ
Jaenisch
Rudolf
R
D000818:Animals; D001755:Blastocyst / Q000166:cytology / Q000276:immunology / Q000378:metabolism; D016026:Bone Marrow Transplantation; D002454:Cell Differentiation / Q000235:genetics; D002467:Cell Nucleus / Q000235:genetics / Q000276:immunology; D019976:Cloning, Organism / Q000379:methods*; D004268:DNA-Binding Proteins / Q000172:deficiency* / Q000235:genetics; D004195:Disease Models, Animal; D004624:Embryo Transfer; D005260:Female; D015316:Genetic Therapy; D006085:Graft Survival / Q000235:genetics; D018380:Hematopoietic Stem Cell Transplantation; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000276:immunology* / Q000378:metabolism; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D016513:Mice, SCID; D009154:Mutation / Q000235:genetics; D053652:Nuclear Transfer Techniques; D009865:Oocytes / Q000166:cytology / Q000276:immunology / Q000378:metabolism; D011123:Polyploidy; D016511:Severe Combined Immunodeficiency / Q000235:genetics / Q000276:immunology / Q000628:therapy*
D004268:DNA-Binding Proteins; C499308:Rag2 protein, mouse; C064655:V(D)J recombination activating protein 2