Muscle and Exercise Lecture 2.2 : Gene therapy for DMD: Viral - mediated strategies Flashcards Preview

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Flashcards in Muscle and Exercise Lecture 2.2 : Gene therapy for DMD: Viral - mediated strategies Deck (22):

What is the adenovirus, and what are its characteristics?

Causes mild disease in humans, is icosahedral and non-enveloped.
Medium sized with large carrying capacity.


What kind of cells can adenovirus infect?

Both proliferating and non proliferating cells.


What is meant by first generation adenovirus? Does it infect satellite cells?

Room needs to be made for dystrophin.
8kbp of genes responsible for virality cut out, replaced with a mini dystrophin gene. Doesnt infect satellite cells


Do first generation adenoviruses induce immune response? What is a consequence of this? Why cant this be avoided by systemic injection?

Yes, making it localised. Systemic injection will cause sepsis.


What is meant by second generation adenovirus? Does it elicit an immune response?

Like first generation, but further viral genes cut out. Still elicits an immune response.


What is meant by third generation adenovirus? Can it deliver full length dystrophin gene?

Is a completely gutted adenovirus with no viral genes. Carrying capacity at 30kbp, has very little immune response. Can deliver the full dystrophin gene.


Do third generation adenoviruses improve functionality? What are issues regarding immune response? Why are high doses needed.

Yes they do, but high doses are needed for this, and also to infect satellite cells. However, high doses then cause an immune response. High dose needed due to poor muscle permeability.


Is there dystrophin expression with adenovirus if there is an immune response?

There is no expression.


What is the lentivirus? Can it deliver full length dystrophin?

It is an enveloped retrovirus, retroviruses cause more severe diseases.
Genome is 8kbp, so cant deliver full length dystrophin, but can carry a functional dystrophin gene.


Can lentivirus integrate into the genome? Can it infect satellite cells?

It can integrate into the genome, and infect satellite cells.


Is transfection by lentivirus permanent? What does this suggest?

Yes, if done early enough. Suggests it transfects satellite cells.


What is a risk of using lentivirus, or any retrovirus? How does this change how its administered?

High risk of insertional mutagenesis, where the gene is inserted into an oncogene, leading to possible tumour development.
Can't be delivered systemically therefore.


What is the adeno associated virus (AAV)? What percentage of humans has it infected? Why is it a good thing?

Small virus containing parvovirus, infected 80% of the population. Doesn't cause disease, so can safely be used. Humans are its natural host.


What is the carrying capacity of AAV, and what does it require to replicate?

5kbp, and requries a second helper virus to infect.


When using AAV, how many vectors are needed?

2 needed, one has dystrophin, flanked by terminal repeat sequences, and packaged into a capsid.
Injected with the helper virus to induce transfection.


rAAV is the gene of interest vector for AAV transfection. Which serotypes of rAAV are best for DMD?

rAAV1, 6, 8, 9 are best for muscle and heart cells.


Does AAV transfection improve fibrosis?

No it doesn't, despite the expressed dystrophin. Early action is therefore best.


What is required when injecting AAV regarding the subject's immune system?

It must be supressed or the vector will be broken down.


Why can some people have an immune response to dystrophin itself?

As it has regions normally not expressed, they cause an immune response.


Can AAV be systemically delivered?

Yes, it can.


What is utrophin similar to, and what happens to it in DMD?

Similar to dystrophin, and is upregulated in DMD.


Can AAV be used to carry full length dystrophin despite its small carying capacity?

Yes, two vectors needed. One half of dystrophin is put into each vector, a homologous region needed in between.
Recombination occurs in vivo.

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