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33 Cards in this Set
- Front
- Back
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Why would one use gene therapy?
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To compensate for a loss of function mutation
Replace/inactivate a dominant negative mutation provide a pharmocological effect to counteract the results of a mutation |
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What are the two forms of gene therapy?
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Somatic-manipulating gene expression in cells of patient, but not passing that manipulation on to the next generation
Germline-modifying germline cells, NOT AN OPTION |
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Three approaches to gene therapy?
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Ex Vivo
In Situ In Vivo |
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What is Ex Vivo
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cells removed from body, incubated with vector containing the gene of interest, and cells returned to body
as in stem cells with IL-2 |
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In Situ?
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vector placed directly into affected tissues(a type of in vivo)
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In Vivo?
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vector injected directly to tissue or ECF--taken up selectively by target cells
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Transduction?
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Viral infxn of normal cells
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Nonviral introduction of target gene?
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Plasmid
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What genetic element must the target gene have in order to be produced in the target cell?
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Promoter/Enhancer regions (aka regulatory elements)
confers: amount of expression, tissue specificity, and timing of expression |
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What is necessary for the target gene to be integrated into the host genome?
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Viral DNA that surrounds the target sequence and regulatory elements?
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What part of the virus facilitates uptake?
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The viral coat
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What is necessary for a plasmid induction of target genes?
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Lipid encapsulation or electroporation
(not as effective as viruses) |
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Retrovirus advantages
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Can enter nearly ever cell in target pop.
rendered incapable of replication nontoxic to cells low number of copies integrate stably into host cells |
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Retrovirus disadvantage
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can only infect actively diving cells(tumors)
only hold up to 8 Kb insert potential for insertional mutagenesis |
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Lentivirus adv.?
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Enters all cells via nuclear pore
Neurotropic, and will infect stem cells nontoxic Accepts a large sized insert Can be either integrating or autosomal |
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Lentivirus Disadvantages
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potential for insertional mutagenesis
deemed safe, but only used in one clinical trial for HIV disabled herpes can only transduce 8-10 kb inserts |
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Adenovirus Adv.?
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infect wide variety of cells
8-10 kb size when gutted, can hold up to 22 kb inserts!!!! Episomal, thus no inseritional mutagenesis very efficient, high titre |
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Adenovirus Dis.?
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transient expression in dividing
potential for strong immunological response-->student was killed due to explosive immunological response |
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Adeno-Associated viruses Adv.?
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no known adverse effects in humans, why its the "darling"
infects all cells(dividing and nondividing), thus neurotropic exists primary as an episome long expression time |
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Adeno-Associated disadvantages?
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very small insertion size
host response to viral vector could cause loss of virus producing cells. |
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Name 4 non viral vectors?
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Naked DNA
DNA in liposomes DNA protein conjugates Artificial chromosomes |
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What are advantages and disadvantages of NON viral vectors
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A: no infectious risk, and no limit on insert size
D: low efficiency of infection, degraded by host |
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Risks of Gene therapy?
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To great an immune response to vector
Transferred DNA may integrate into patients DNA, and cause disruption of tumor supressor activites or upregulate protooncogenes...also could disrupt and essential gene |
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Name two cases of gene therapy success?
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Hemophilia B-clotting factor 9 was transduced via adeno-associated virus
Angina-VEGF-2 transduced into patients to stimulate growth of new blood vessels (8/12 experienced decreased drop in angina) |
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What is SCID? Why is it relevant to gene therapy?
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severe combined immunodeficiency
no B or T cell immunity in the past, only able to due bone marrow transplant with gene therapy-->isolated bone marrow stem cells, inserted correct gene, returned stem cells to patient 9/11 developed normal T and B cell fxn 3 patients developed luekemia like symptoms due to insertional mutagenesis |
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What is the basis of gene therapy for cancer?
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Transfer of therapeutic genes to a tumor mass or the peritumoral tissue
--includes transfer of suicide genes to induce cell death following admin. of a prodrug |
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What is Suicide Gene Therapy?
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NOT GENE REPLACEMENT!
introduce a new gene that converts a nontoxic prodrug into a lethal drug compound |
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What are the specific examples mentioned in class regarding suicide genes? Drugs? Enzymes?
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In a tumor cell:
Ganciclovir=prodrug Vector=carries Thymidine Kinase Gene TK phosphoryates Ganc. converting it to its active form, facilitates cell death |
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Parkinson's symptoms?
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tremor of hands, arms, legs or jaw
rigidity of limbs, trunk bradykinesia-slowness of mov't postural instability/impaired balance, coordination classic stooped posture and shuffling gate |
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Parkinson's causes?
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Neuronal death, esp. in Substantia Nigra(lots of dopaminergic receptors)
Substantia Nigra deterioration-->striatum-->Subthalamic nuclei become overactive-->loss of inhibition of motor output |
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What protein is genetically mutated in Parkinson's?
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alpha-Synuclein--leads to aggregate formation known as Lewy bodies
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What environmental factors contribute to Parkinson's?
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Well water, pesticides, herbicides
synthetic narcotic MPTP |
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How can we use gene therapy to treat our future Parkinson's Patients
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Introduce the enzyme GAD-glutamic acid decarboxylase-to synthesize GABA(inhibitory NT).
use adeno associated virus transduced subthalamic nuclei become INHIBITORY vs. excitatory, stimulating motor activity(relieve inhibition of motor output) |
