gene therapy

Question 1

in vivo vs ex vivo gene therapy

Answer 1

• in vivo:
• direct delivery of genes
• delivery into dividing and non-dividing cells
• episomal = unstable = need for repeated tx
• ex: CF
• ex vivo
• genes introduced into patient’s cells in vitro
• only works for dividing cells (requires cell culture)
• integrated into host genome = stable = lifelong
• ex: SCID

Question 2

naked DNA gene delivery

Answer 2

pros: no side effects
cons: temporary, low efficiency

Question 3

modified RNA gene delivery

Answer 3

• pros: easy

Question 4

liposome gene delivery

Answer 4

• pros: episomal, can target many tissues
• cons: temporary, gene is packaged in an endosome

Question 5

viral vectors for gene delivery

Answer 5

• pros: can be long term or temporary, may be able to package large amounts of material
• cons: side effects, safety concerns, targeted expression due to viral tropism

Question 6

retrovirus vector for gene delivery

Answer 6

cons:

• integration of viral DNA (carrying the GOI) into the host genome requires cell division and most somatic cells do not divide
• max size = 7kb and most genes are larger
• inserted genes eventually become silent due to mutation

Question 7

lentiviral vector for gene delivery

Answer 7

pros:

• similar to retrovirus but allows infection of/delivery to nondividing cells

Question 8

adenovirus vector for gene delivery

Answer 8

pros:

• can get DNA into most types of cells
• can carry up to 35 kb

cons:

• does not integrate into host genome but persists at high levels ~1 month and low levels 6-15 months
• immunogenic (could be avoidable by removing all viral genes = helper dependent adenovirus)

Question 9

adeno-associated viral vector for gene delivery

Answer 9

pros:

• less immunogenic than Ad vectors

cons:

• smaller capacity

Question 10

SCID gene therapy

Answer 10

• removed mononuclear cells from kids with SCID
• cultured with mAb OKT3 and IL-2 to induce T cell proliferation
• then added a retroviral vector expressing WT ADA gene
• cells were reinfused into the children => humoral and cellular immune functions

key points:

• ex vivo
• concurrent with the gold-standard PEG ADA tx

Question 11

CF gene therapy

Answer 11

• delivery of CFTR gene into patient lungs (in vivo) via liposome complex, adenovirus, or adenoassociated vector

Question 12

ALS gene therapy

Answer 12

• ex vivo
• injection of myoblasts infected with adenovirus vectors expressing glial cell derived neurotrophic factor into mice limbs

Question 13

glioblastoma gene therapy

Answer 13

• suicide gene therapy (give gene that activates prodrug) + bystander effect (only requires a few cells expressing suicide gene to eradicate entire tumor)
• cells producing a retrovirus vector expressing HSV TK were injected into tumors
• gancyclovir (prodrug) given
• antitumor effect