Gene therapy Flashcards
Uncommon genetic disorders that cannot be cured
Cystic fibrosis
Sickle cell anaemia
Thalassaemia
Duchenne Muscular Dystrophy
Haemophilia A
Familial hypercholesterolaemia
Phenylketonuria
Tay-Sachs disease
Targets of gene therapy
Single gene, recessive loss of function - Cystic fibrosis, haemophilia - Therapy would be gene addition/replacement
Single gene, haploinsufficiency - Dyschromatosis Symmetrica Hereditaria (DSH) - Gene addition
Single gene, dominant negative - Huntington disease - allele silencing/replacement
Multi-gene or acquired - Cancer, heart disease, rheumatoid arthritis - Addition of therapeutic gene
What rescued chronic non-ischemic heart failure in minipigs?
Cardiac bridging integrator 1 gene therapy
Gene therapy is used to treat…
- Mostly cancer
- Also genetic disease, infection etc
- Treatment been used to treat haemophilia, HIV, multiple myeloma
What types of genetic diseases are treated using gene therapy
Metabolic disease, eye disease, blood coagulation disorders, immunodeficiency, neuromuscular disease, haemoglobinpathy
Types of gene therapy
in vivo:
- Single step process
- Vector administered directly to patient
- Targeted to specific organ/tissue (route of administration or specificity of vector)
ex vivo:
- Two step process
- Cells removed from patient
- Vector added to cells in vitro
- Engineered cells returned to patient
- May be combined with (stem) cell based therapy
Barriers to gene therapy
- Neutralising antibodies bind to antigen on virus
- Uptake, transport and uncoating of virus
3.
Vectors for gene therapy
- Adenovirus
- Adeno-associated virus
- y-Retrovirus (e.g. Moloney Murine Leukaemia Virus-derived)
-Lentivirus (e.g. HIV derived)
- Routine plasmids
- Mini-circles
- Transposons
In vivo gene therapy pros/cons
- Difficulty of delivery
- Accessible organs - lungs, skin, muscles
- Less accessible - liver, retina, brain
- Vector - adenovirus, adeno-associated virus, some use of retroviral vectors
- Treatment of single gene disorders and acquired disease
What are the advantages of adenoviral vectors
- Large capacity - up to 30kb if helper virus provided
- Easily purified
- Infects broad range of cell types
- Efficient transduction
Disadvantages of adenoviral vectors
- Common cold virus - high incidence of neutralising antibodies
- Capsid proteins in highly immunogenic
- Potentially fatal inflammatory response (death of Jesse Gelsinger during OTC trail in 1999)
- Transient expression of transgene
Adeno-associated virus
- Limited capacity (4.7kb ssDNA genome)
- Non-pathogenic, minimal immune response
- rep and cap genes can be replaced with expression cassette
- Can be used in non-dividing cells - maintained as episome
- Different serotypes target different tissues
Successful AAV trails
Glybera (alipogene tiparvovec) - UniQure - EMA approved, 11-2-12 - targets LPL gene - Major indication is LPL deficiency
AAV case study – Leber’s congenital amaurosis (LCA)
Amaurosis (‘darkening’) is vision loss without obvious physical signs
Early-onset blindness
Autosomal recessive (14 genes, including RPE65)
RPE65 codes for retinal pigment epithelium-specific 65kDa protein – required for photoreceptor function
Photoreceptors persist in affected individuals
Vision restored in mouse and dog LCA models using AAV vectors containing RPE65
Successful phase II clinical trials
LCA gene therapy
- Three seperate clinical trials
- AAV2 serotype capsids directly injected beneath retina
- Virus taken up by retinal epithelium
- RPE65 gene expressed from episomal vector
- Light sensitivity restored - maintained for >3 years
- Early intervention required for best results
