Gene therapy refers to the introduction of functional genetic material into target cells to replace or supplement defective genes, or to modify target cells so as to achieve therapeutic goals.
GENE THERAPY
Gene therapy refers to
the introduction of functional genetic material into target cells to replace or
supplement defective genes, or to modify target cells so as to achieve
therapeutic goals. In contrast to all other drugs, this kind of therapy can
impart new functions to a cell. Conceived in the 1960s and started in the 1980s
gene therapy is still experimental, but holds great promise for curing a number
of diseases which at present can, at best, be only palliated or controlled.
Gene defects result in failure to synthesize a functional protein or in the
synthesis of a dysfunctional protein. Equipping the cell (especially the one
which physiologically expresses it) with a normal copy of the defective gene would
overcome the deficiency at the site where it is needed on a long-term (may be
permanent) basis. Apart from inherited genetic disorders with single nucleotide
polymorphism (SNP), the major thrust area of gene therapy are a number of
acquired diseases such as malignancies, immunological disorders, including AIDS,
cardiovascular, neurological and infective diseases, in many of which even
shortterm expression of the introduced gene could be therapeutic.
APPROACHES IN GENE THERAPY
Gene therapy endeavours
to either modify or transfer genes.
1. Gene Modification
This involves correction of the defective portion of a
genomic sequence or removal of the whole defective gene and its replacement by
the normal copy, or alteration of the sequences controlling its expression.
These are technically quite difficult in the in vivo setting.
2. Gene Transfer
This involves introduction of one or more genomic
expression cassettes without removing or altering the existing ones.
Significant progress has been made in this approach. Gene transfer is carried
out by using various physical, chemical or biological delivery methods
(vectors).
Physical Methods: are microinjection in
a single cell, introduction of DNA complexed with gold pCh. No.s/
dextran/lipids, use of microprojectiles or direct parenteral injection of
uncomplexed DNA. However, efficiency of gene transfer is low and duration of
expression is short.
Chemical Methods use liposomes or other
specific cellular ligands. The liposomes can carry genes to the intracellular
location in large amounts, are nonimmunogenic and technically simpler, but the
level of gene transfer achieved is lower than by use of viral vectors.
Liposomal delivery has been used in certain human gene therapy trials.
Biological Methods involve fusion of recipient
cell with bacterial spheroplasts, erythrocyte membrane vesicles, whole cell
fusion, etc, but the most efficient and widely used are the viral vectors.
Adenoviruses, Lenti virus, Herpes simplex virus have been utilized, but
retroviruses are the most popular vectors. For using viral vector, its ‘gag’,
‘pol’, and ‘env’ genes which carry out viral replication are removed and
replaced by the genes to be transferred using recombinant DNA technology. These
viral pCh. No.s are then introduced into a packaging cell line previously
equipped with the ‘gag’, ‘pol’, and ‘env’ genes which then produce multiple
viral pCh. No.s carrying the desired gene. The retroviral vector is then
allowed to infect host cells so that the desired DNA is inserted into a random
site in the host genome which subsequently expresses the transferred gene.
However, the duration of expression of the transferred gene is variable.
Gene transfer could be
carried out into:
· Germline cells The new gene is
introduced into embryonal cells so
that it passes into the next generation. This is ethically not applicable to
humans at present.
· Somatic cells Certain somatic cells
like the bone marrow stem cells,
fibroblasts, hepatocytes or myocytes of the recipient receive the new gene.
This is being extensively pursued.
· Specific organ
directed delivery of genes to the tissues that actually require it is being tried. Success has been achieved
in transferring genes into liver cells, vascular smooth muscle and endothelial
cells, pulmonary cells, etc.
The techniques of gene therapy may utilize either ex vivo or in vivo gene transfer.
Ex vivo gene transfer: The patient’s tissue
cells are isolated and maintained in
tissue culture. These are then transfected with the vector carrying the
relevant gene and injected back into the patient. This method has been widely
utilized in the treatment of single gene defects, metabolic, haematological and
immune disorders.
In vivo gene transfer: The vector, usually a
retrovirus carrying the gene is
injected systemically or directly into the concerned organ. The property of retrovirus
to transduce only dividing cells, such as tumor cells, is utilized for
selective delivery of the gene leaving nondividing cells unaffected. However,
this is a relatively inefficient method of gene therapy.
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