Human Disease Gene Identification and Correction 
cells with supernatant containing vector viruses 
(and no cells) , rather than with virus-producing 
cells, because the former method is considered to 
be safer. Supernatant viral infection of bone 
marrow cells resulting in the acquisition of ADA 
activity can be achieved, but high efficiency re- 
quires the presence of other supporting cells, 
which is impractical in the context of an autolo- 
gous bone marrow transplant. Attempts are under 
way to purify bone marrow stem cells, which are 
responsible for the regeneration of the entire he- 
mopoietic system, since targeting this group of 
cells would provide a more efficient approach to 
the permanent repopulation of bone marrow 
with genetically corrected cells. 
Ornithine transcarbamylase (OTC) deficiency, 
the most common urea cycle defect in humans, is 
inherited in an X-linked recessive manner. Coma, 
seizures, and retardation are the result of hyper- 
ammonemia secondary to the enzyme deficiency. 
Liver transplantation provides a once-per-lifetime 
cure, although again with the potential compli- 
cations of a graft-versus-host reaction. This condi- 
tion is poorly managed through dietary protein 
restriction and medical therapy, making the dis- 
order another excellent candidate for gene 
therapy. 
Two mouse models of the human disorder are 
available for developing gene therapy. In this lab- 
oratory, the OTC deficiency in one of the strains 
of mice has been corrected by retroviral delivery 
of the human OTC gene to the small intestine. In 
current studies toward the development of hu- 
man gene therapy for OTC deficiency, retrovi- 
ruses and defective (safe) adenovirus vectors are 
used to transfer the human gene to cultured liver 
cells. 
Duchenne muscular dystrophy (DMD) is a se- 
vere disorder, also inherited in an X-linked reces- 
sive manner. Deficiency of the protein dystro- 
phin leads to multiple muscle abnormalities and 
eventually death. The gene is large and complex. 
Several mouse models of DMD are available for 
study, and the dystrophin deficiency in one of 
these strains has been corrected by introduction 
of a construct expressing mouse dystrophin. 
In collaboration with Helen Blau (Stanford 
University), myoblasts (precursor muscle fiber 
cells) are being isolated from very young patients 
with DMD. Different constructs will be tested for 
the transfer of the dystrophin gene and for creat- 
ing gene-corrected myoblasts that can be re- 
turned to the patient. The gene is so large that 
currently used viral vectors will not accommo- 
date it. Approaches being used to overcome this 
problem include modifying the vectors, truncat- 
ing the gene, and using physical methods (that do 
not present a size limitation) for transferring the 
gene, such as electroporation or ballistic gene 
transfer (using the Du Pont "gene gun"). 
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