1. Gene therapy and viral vector
Lecture 3

2. Viral Vectors retrovirus adenovirus adeno-associated virus
herpes virus
pox virus
human foamy virus (HFV)
lentivirus  

3. Adenovirus Medium-sized (90-100nm)
Nonenveloped (without an outer lipid bilayer)
Double stranded DNA
Derived from Human Adenoids (tonsils) in 1953
They have a broad range of vertebrate hosts; in humans, 100 distinct adenoviral serotypes have been found to cause a wide range of illnesses, from mild respiratory infections in young children (known as the common cold) to life-threatening multi- organ disease in people with a weakened immune system.
Serotype or serovar are distinct variations within a species of bacteria or viruses or among immune cells of different individuals.
Different types/serotypes are associated with different conditions:
respiratory disease (mainly species HAdV-B and C)
conjunctivitis (HAdV-B and D)
gastroenteritis (HAdV-F types 40, 41, HAdV-G type 52)
obesity or adipogenesis (HAdV-A type 31, HAdV-C type 5, HAdV-D types 9, 36, 37) 

5. How Adenovirus enters the host cell?
Entry of adenoviruses into the host cell involves two sets of interactions between the virus and the host cell.
Entry into the host cell is initiated by the knob domain of the fiber protein binding to the cell receptor. The two currently established receptors are: CD46 for the group B human adenovirus serotypes and the coxsackievirus adenovirus receptor (CAR) for all other serotypes.
This is followed by a secondary interaction with an integrin molecule. It is the co-receptor interaction that stimulates entry of the adenovirus.
Binding to integrin results in endocytosis of the virus particle via clathrin-coated pits.
Attachment to integrin stimulates cell signaling and thus induces actin polymerization resulting in entry of the virion into the host cell within an endosome.

6. Entry in the nucleus
Once the virus has successfully gained entry into the host cell, the endosome acidifies, which alters virus topology.
These changes, as well as the toxic nature of the pentons, destroy the endosome, resulting in the movement of the virion into the cytoplasm.
With the help of cellular microtubules the virus is transported to the nuclear pore complex, whereby the adenovirus particle disassembles.
Viral DNA is subsequently released, which can enter the nucleus via the nuclear pore.
After this the DNA associateswith histone molecules. Thus, viral gene expression can occur and new virus particles can be generated.

7. Benefits Their basic biology has been studied extensively,
The viral genome can accommodate large heterologous transgene insertions,
They readily infect quiescent and dividing cells,
They can be amplified to high titers and they have previously been shown to be relatively safe for use in humans.
The family Adenoviridae consists of five genera, including genus Mastadenovirus and genus Aviadenovirus, which infect mammals and birds respectively.
The adenovirus vector most commonly used for clinical trials and experimental gene therapy applications is species C adenovirus, HAdV-C5.

8. Drawbacks
Adenovirus delivered genes can be lost due to genetic instability therefore repeated doses are necessary to maintain the expression of transgene.
They would not integrate into the host genome, their gene expression is too short term.
Immunologic responses against adenoviruses have made their clinical application limited to a few tissues, such as  liver, lung (especially for CF(Cystic Fibrosis) treatment), or localized cancer gene therapy.

9. Drawbacks
Although the risk of serious disease following natural adenovirus infection is rare and the viral genome would not integrate into the host genome, gene therapy by adenoviral vectors has caused serious bad side effects and even death of some patients.

10. Adeno-associated vectors
Adeno-associated vectors (AAV) are like adenoviral vectors in their features but because of having some deficiency in their replication and pathogenicity, are safer than adenoviral vectors.
In human, AAVs are not  associated with any disease.
Another special character of AAV is their ability to integrate into a specific site on chromosome 19 with no noticeable effects cause long-term expression in vivo. 
AAVs have been used in the treatment of some diseases, such as CF, hemophilia B, Leber congenital amaurosis, and AAT (Alpha-1 antitrypsine) deficiency.

11. The use of antisense oligodeoxynucleotides targeted to the renin-angiotensin system [hormone system that regulates blood pressure and water (fluid) system] and adeno-associated virus vector delivery of antisense DNA offers a new approach to prolonged hypertension therapy with a single administration.

12. Drawbacks
The major disadvantages of these vectors are complicated process of vector production and the limited transgene capacity of the particles (up to 4.8 kb). 

13. Comparison

14. Lecture prepared from
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