Autotransplantation in mdx mice of mdx myoblasts genetically corrected by an HSV-1 amplicon vector.
Duchenne muscular dystrophy (DMD) is an X-linked recessive disorder, characterized by a lack of dystrophin. To eliminate the need for immunosuppressive drugs, transplantation of genetically modified autologous myoblasts has been proposed as a possible therapy for this myopathy. An HSV-1 amplicon vector (HSVDGN), containing a 17.3-kb full-length MCK-driven mouse dystrophin cDNA, an eGFP gene, and a neomycin resistance gene driven by CMV or SV40 promoters, respectively, was constructed and used to transduce mdx primary myoblasts. The presence of the eGFP and neomycin resistance genes facilitated the evaluation of the initial transduction efficiency and the permanent transduction frequency. At low multiplicities of infection (MOI 1-5), the majority of myoblasts (60-90%) expressed GFP. The GFP-positive mdx myoblasts were sorted by FACS and selected with neomycin (300 microg/ml) for 2 weeks. Up to 2% of initially infected mdx myoblasts stably expressed the three transgenes without further selection at that time. These altered cells were grafted into the tibialis anterior muscles of 18 mdx mice. Some of the mice were immunosuppressed with FK506 due to the anticipation that eGFP and the product of neomycin resistance gene might be immunogenic. One month after transplantation, numerous muscle fibers expressing mouse dystrophin were detected by immunohistochemistry, in both immunosuppressed (10-50%) and nonimmunosuppressed (5-25%) mdx mice. Our results demonstrated the capability of permanently expressing a full-length dystrophin in dystrophic myoblasts with HSV-1 amplicon vector and raised the possibility of an eventual treatment of DMD based on the transplantation of genetically modified autologous myoblasts.
Bujold
Mathieu
M
Laboratoire de Génétique Humaine, Centre de Recherche du Centre Hospitalier de l'Université Laval, Ste-Foy (Qc), Canada, G1V 4G2.
Caron
Nicolas
N
Camiran
Goeffrey
G
Mukherjee
Santwana
S
Allen
Paul D
PD
Tremblay
Jacques P
JP
Wang
Yaming
Y
D000818:Animals; D018076:DNA, Complementary; D016189:Dystrophin / Q000235:genetics*; D015967:Gene Expression Regulation, Viral; D018014:Gene Transfer Techniques; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D018259:Herpesvirus 1, Human / Q000235:genetics*; D051379:Mice; D018101:Mice, Inbred mdx; D009137:Muscular Dystrophy, Animal / Q000628:therapy*; D020388:Muscular Dystrophy, Duchenne / Q000628:therapy; D032448:Myoblasts, Skeletal / Q000502:physiology* / Q000637:transplantation*; D011401:Promoter Regions, Genetic; D013601:T-Lymphocytes; D014182:Transplantation, Autologous
D018076:DNA, Complementary; D016189:Dystrophin