Final - Gene Therapy Flashcards
1
Q
Gene therapy
A
- an experimental proedure aimed at replacing, mainpulation, or supplementing nonfunctional or dysfunctional genes with health genes
2
Q
Goals of Gene Therapy
A
- To compensate for a mutant gene that causes a loss of function in a specific protein
- To replace or inactivate a mutant gene
- To counteract the effects of disease pathogenesis in acquired disorders (e.g. cancer)
3
Q
Criteria for Effective Gene Therapy
A
- must have sufficient knowledge on molecular basis of the disease to be treated
- the gene to be transferred must be identified and cloned
- Must have an effective method of introducing the gene so that it is exposed in a large number of target cells
- Target cells should have a long half-life or good potential to replicate in vivo
- The therapy must not harm the patient
4
Q
Recessive genetic disorder gene therapy
A
- requires goal #1 (To compensate for a mutant gene that causes a loss of function in a specific protein)
- involves the addition of a normal gene into target cells so that the recessive allele is masked
5
Q
Dominant genetic disorder gene therapy
A
- requires goal #2 (To replace or inactivate a mutant gene)
- involves replacement of gene which is harder to do.
6
Q
Basic Principal of Gene Therapy
A
- a therapeutic gene inserted into a vector that carries the gene to specific target cells
- can be performed in vivo or ex vivo
7
Q
Viral Vectors
A
- modified to remove harmful gene and replaced with desired gene
1. Retrovirus
2. Adenovirus
3. Adeno-Associated Virus (AAV)
8
Q
Retrovirus
- advantages
- limitations
A
- simple RNA virus with only 3 structural genes
Advantages:
1. enter target cell efficiently
2. Viral DNA integrates into host genome and is stable
3. Can carry a relatively large piece of DNA (8 kb)
Limitations:
- cannot infect nondividing cells (exception = lentiviruses)
- potential disruption of host genes = side effects
9
Q
Adenoviruses
- advantages
- limitations
A
- cause common cold, natural tropism for respiratory epithelium
Advantages:
1. Can infect dividing and nondividing cells
2. Carry large piece of DNA (30-35 kb)
Limitations:
- Don’t integrate into host genome = only expressed transiently
- induce a strong immune response = destroys virus
10
Q
Adeno-Associated virus (AAV)
- advantages
- limitations
A
- DNA virus that does not cause disease in humans
Advantages:
1. infects dividing and nondividing cells
2. integrates into host genome, producing latent, long-term expression.
3. induces a milder immune response
Limitations:
- can only accommodate small pieces of DNA (5 kb)
- Difficult to produce in large quantities
11
Q
Non-Viral Vectors
- advantages
- limitations
- types
A
Advantages:
- No biological risks
- Do not induce an immune response
- Can carry larger amounts of DNA
Limitations:
- Do not integrate into host genome and therefore, have a poor long-term expression
- May be difficult to target tissue of interest
Types:
A. Naked DNA - cDNA with regulatory elements in a plasmid
B. Liposomes- synthetic lipid vesicles which can transport genes
C. Protein-DNA conjugates - allows binding to a cell surface receptor to facilitate entry into cells
12
Q
Ex vivo gene transfer
A
- target cells are removed from the patient, genetically reconstituted with the appropriate gene in vitro and then returned to the patient
- Ex: Gene therapy for ADA defieciency
13
Q
In vivo gene transfer
A
- genetically modified vector is introduced directly into the body (e.g. inhaled)
- Ex: Gene therapy for Cystic Fibrosis
14
Q
ADA Deficiency
A
- the first disease to be treated with gene therapy
- an autosomal recessive disorder that causes severe immunodeficiency
- example of ex vivo gene transfer: Patients T lymphocytes are removed and incubated with MMLV containing the human ADA gene. Genetically modified T cells are injected IV into patient
15
Q
Severe combined immunodeficiency
A
- ex vivo gene transfer
- bone marrow stem cells are genetically altered to include a gene encoding part of a receptor necessary for lymphocyte development
