Gene Therapy-Intoduction to molecular medicine

Question 1

what is gene therapy?

Answer 1

a novel approach to treating diseases based on modifying the expressionof a person’s genes tworad a therapeutic goal

Question 2

what are the types of glcassical gene therapy?

Answer 2

1) somatic gene therapy

2) germ line gene therapy

Question 3

what is somatic gene therapy?

Answer 3

involves the manipulation of gene expressin in cells so as to be corrective for hte patient, but his correction in not inherited by the next generation

Question 4

what is germ line therapy?

Answer 4

involves teh genetic modficication of germ cells that will pass the selected cahnge onto the next generation (limited to animal models)

Question 5

What accidentally occured with the blood transfusion in an HIV positive patient?

Answer 5

the patient was given blood from a donor who had a mutation for CCR5 deletion and now the patient is essentially HIV free

Question 6

what types of diseases are being treated with gene therapy?

Answer 6

genetic diseases; viral infections; autoimmunity; cancer; diseseases in which several genes and they enviornment interact

Question 7

what are the two types of intervetnion strategies for gene therapy applications?

Answer 7

therapeutic stratigies

cytosolic strategies

Question 8

what is the therapeutic stratigy?

Answer 8

vectors carry a gene that encode a protein that is either defective or that is not present due to mutations in the patients’s endogenous genes

Question 9

what is the cytosolic strategy?

Answer 9

vector is designed to destroy or eliminate a diseased cell or tissues

Question 10

what are the requirments and challenges to develope a genetic therapy?

Answer 10

A) gene identification adn cloning
B) Delivery
C) elements to ensure controlled gene expression
D) understanding of the host immune response

Question 11

What are the limitations of genetic therapy delivery?

Answer 11

the delivery must be specific to the cell type and efficent and safe

Question 12

What are the delivery vehicles?

Answer 12

Vectors; most are based on attenuated or modified versions of viruses, some use non-viral vectors (plasmid DNA)

Question 13

What are the challanges of attenuated or modified viruses in gene therapy?

Answer 13

the challange is to remove the disease cauzeing compontes of the virus and insert recombinant genes that will be therapeutic to the patient

Question 14

What are the stratageis for non-viral vectors?

Answer 14

plasmid DNA, liposome transfection (complexes of DNA adn lipids) or use gene gun technology

Question 15

what is gene gun technology?

Answer 15

delivery of DNA on gold particles (primarily for DNA vacines)

Question 16

why is it important to control gene expression?

Answer 16

to make the correct amount of therapeutic proteins at the right time (not cause other issues for the person)
and is essential for maintaining long-term expression of gene in the correct cells or tissue

Question 17

What are the properties of Adenoviruses?

Answer 17

• episomal
• hight transduction efficiency
• infects replicating and non-replicating cells
• elicits an immune response
• insert capacity 8-36Kb

Question 18

What are the properties of Adeno-associated virus?

Answer 18

• integrates genenome into specific region on human chr 19
• low immunogenicity
• no associated disease
• infects both dividing and non-dividing cells
• limited insert capacity ~5Kb

Question 19

What are the properties of herpesviruses?

Answer 19

• large insert capacity
• broad host range
• infects dividing and nondividing cells

Question 20

What are the properties of liposomes/ naked DNA?

Answer 20

• no limit to the size of the genes that can be delivered
• low immunogenicity
• poor levels of gene transfer

Question 21

What are the properties of retroviruses?

Answer 21

• non pathogenic in humans
• stably transduces dividing but non non-dividing
• inserts genome into host cell’s DNA
• longterm expression
• insert capacity of 8Kb
• inactivated by human complement

Question 22

how is a retroviral vector made?

Answer 22

WT gene b/n packign signals is a survival gene, insert to a cell to make a packaging cell, place the therapeutic gene also b/n the packaging signals and insert into the producer cell and get a gene therapy vector (a little confusing sorry)

Question 23

what are some important components to initiate a gene therapy trial?

Answer 23

invovles both internal review boards (IRBs) and ferderal apporval from the FDA adn the recombinant DNA advisory committee (RAC)
can take 10-20 yrs
majority of clinical trials are being done is us (63%)
~1579 trials on going

Question 24

waht is important about AAV? (adeno-associated virus vectors)

Answer 24

• do not stimulate inflammation in the host
• does not elicit antibodies against itself
• can enter non-dividing cells
• integrates successfully into 1 spot int eh genome of its host (chr 19)

Question 25

What was the SCID-X1 success?

Answer 25

an x-linked disorder; mutation in the gamma-subunit of IL (2,4,7,9,&15) receptors
causes early block in T and NK cell differntiation
used ex vivo vectors endoding for the gamma-subunit *hematopeotic stem cell infection) CD 34+ cells
all pts were infants
10/11 treated had a full immune system reconsitituion

Question 26

What occured in the ADA-SCID therapy?

Answer 26

adenosine deaminase deficency, a purine metabolic defect which causes impared lymphocyte development and function
-used retrovirus derived vector containing the ADA cDNA and ex vivo infecton of CD34+ stem cells
therapy was successful

Question 27

Waht occured int eh LCA gene therapy? (leber congenital amaurosis)

Answer 27

a group of AR blinding retinal diseases

• mutation in RPE65 gene
• txed with rAAVV2 carrying human RP65 gene
• pts responed well to txment, younger the pt the better they responded

Question 28

What are the set backs to these treatments?

Answer 28

• death of hte 1st pt in a gene therapy trail (adenovirus ARDs and SIRS)
• serious side effects!!! (many of the SCID pts developed leukemia, insertional ocogenesis next to LMO-2 gene)

Question 29

What are the factors influencign the futrue of the development of gene therapy?

Answer 29

• optimizing vecotr saftey to decrease adverse events
• overcoming the technological obstacles (cell specific targeting and regulated gene expression)
• analysis of human genome
• explanantion of diagnositic industry
• public acceptance of gene therapy