Nucleic-acid- and cell-based therapeutics - Pharmaceutical Biotechnology: Concepts and Aapplications - page 442

sequence, but also upon detailed regulation of gene expression. Such tight regulation of expression
of transferred genes is beyond the capability of gene therapy technology as it currently stands.
Another early genetic disease for correction by gene therapy was SCID. One form of this disease
is caused by a lack of adenosine deaminase (ADA) activity. ADA is an enzyme that plays a central
role in the degradation of purine nucleosides (it catalyses the removal of ammonia from adenosine,
forming inosine, which, in turn, is usually eventually converted to uric acid). This leads to T- and
B-lymphocyte dysfunction. Lack of an effective immune system means that SCID sufferers must
be kept in an essentially sterile environment.
When compared with treating diseases such as thalassaemia, regulation of the level of expression
of a corrected ADA gene was believed to be less important for a successful therapeutic outcome.
(In most, though not all, metabolic diseases caused by an enzyme defi ciency, it appears that expres-
sion of even a fraction of normal enzyme levels is suffi cient to ameliorate the disease symptoms.)
Gene therapy trials aimed at counteracting ADA defi ciency were initiated in 1990. The fi rst recipi-
ent was a 4-year-old SCID sufferer. The protocol used entailed the isolation of the child's peripheral
lymphocytes, followed by the in vitro introduction of the human ADA gene into these cells, using a
retroviral vector. After a period of expansion (by culture in vitro ), these treated cells were re-injected
into the patient. As the lymphocytes (and, by extension, the corrective gene) had a fi nite life span, the
therapy was repeated every 6-8 weeks. This approach appeared successful, in that it has resulted in a
marked and sustained improvement in the recipient's immune function. Critically, however, interpreta-
tion of this outcome was made more diffi cult owing to the later revelation that the patient also initiated
more conventional SCID therapy just prior to the gene therapy treatment. A second retroviral-based
trial aiming to treat a different form of SCID has also been discussed earlier in this chapter.
Haematopoietic (and indeed other) stem cells are attractive potential gene therapy recipient
cells because they are immortal. Successful introduction of the target gene into these cells should
facilitate ongoing production of the gene product in mature blood cells, which are continually
derived from the stem cell population. This would likely remove the requirement for repeat gene
transfers to the affected individual.
The routine transduction of haematopoietic stem cells has, thus far, proven technically diffi cult.
They are found only in low quantities in the bone marrow, and there is a lack of a suitable assay
for stem cells. However, recent progress has been made in this regard, and routine transduction of
such cells will likely be achievable within the next few years.
Additional genetic diseases for which a gene therapy approach is currently being evaluated include
familial hypercholesterolaemia and cystic fi brosis. Familial hypercholesterolaemia is caused by the ab-
sence (or presence of a defective form of) low-density lipoprotein receptors on the surface of liver cells.
This results in highly elevated serum cholesterol levels, normally accompanied by early onset of seri-
ous vascular disease. The gene therapy approaches that have been attempted thus far to counteract this
condition have entailed the initial removal of a relatively large portion of the liver. Hepatocytes derived
from the liver are then cultured in vitro , with gene transfer being undertaken using retroviral vectors.
The corrected hepatocytes are then usually infused back into the liver via a catheter. Although studies in
animals have been partially successful, transduction of only a small proportion of the hepatocytes is nor-
mally observed. Subsequent expression of the corrective gene can also be variable. In vivo approaches to
hepatic gene correction, using both viral and non-viral approaches, are also currently being assessed.
The cystic fi brosis ( cf ) gene was fi rst identifi ed in 1989. It codes for CFTR, a 170 kDa pro-
tein that serves as a chloride channel in epithelial cells. Inheritance of a mutant cftr gene from
both parents results in the cystic fi brosis phenotype. While various organs are affected, the most
severely affected are the respiratory epithelial cells. These cells have, unsurprisingly, become the