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1) Indicate which of the following diseases could be corrected by directly infecting the affected cell type with a transgene as opposed to grafting:
a) an individual that lacks expression of an enzyme due to a recessive mutation within the gene.
The affected cell type can serve to express and regulate the transgene.
b) an individual with degenerating dopaminergic neurons.
Most cell bodies would no longer exist to express the transgene. Need grafting of stem cells.
c) an individual with atherosclerosis.
Dependent upon at which step therapeutic intervention is targeted. Can correct individuals with hypercholesterolemia by transferring LDL receptor gene into
hepatocytes. Also can graft engineered endothelial cells to deliver drugs to minimize blood clots.
2) Indicate what viral vector or transfection method can be used for gene transfer into the following primary tissues:
a) Fibroblasts
retrovirus for long term expression.
AAV or liposomes for short term expression.
b) Neurons
herpesvirus, liposomes
c) Hepatocytes
adenovirus or AAV
liposome conjugated to asialoglycoprotein
d) Lymphocytes
retrovirus for long term expression.
AAV or liposomes for short term expression.
3) Why is the Lac Z gene encoding b-galactosidase used in gene transfer studies?
To localize all sites of gene delivery and expression.
To calculate the % efficiency of gene transfer.
4) List 3 ways to limit transgene expression to a specific cell type in gene therapy.
1) Surgically deliver gene to limited area.
2) Use tissue targeted delivery vehicles that bind tissue-specific receptors for entry (ligand conjugated liposomes, viral vectors that infect primarily the tissue of
interest).
3) Use tissue-specific promoter drive expression of transgene only in a certain cell type.
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