Muscle and Exercise Lecture 2.1 : Gene therapy for DMD: Non - viral strategies Flashcards Preview

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Flashcards in Muscle and Exercise Lecture 2.1 : Gene therapy for DMD: Non - viral strategies Deck (30)
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1
Q

Name 6 muscle wasting conditions.

A

-Disuse atrophy, when not in use (limb casting/best rest).
-Denervation - spinal cord injury
-Sepsis - systemic infection (in blood)
-Sarcopenia - age related wasting
-Cancer cachexia - cancer related wasting
Muscular dystropy - genetic cause, genetic theraphy most helpful.

2
Q

What is the best therapy for a disease with genetic causes?

A

Genetic therapy is best.

Pharmacological treatment prevents secondary consequences only.

3
Q

What does genetic therapy do (name 3)?

A

Restores or replaces the defective gene. Can also introduce new gene to help fight diease.

4
Q

What is used in cell therapy?

A

Myoblast transfer.

5
Q

What is non-viral mediated therapy best for?

A

Altering a gene

6
Q

What is viral mediated best for? What is needed to carry it out?

A

Replacing a defective gene. A vector is needed to carry it out.

7
Q

What is a common vector?

A

Retrovirus.

8
Q

What is a gene expression construct? What is it made up of (3)?

A

It is the intervention body.

It needs a promotor, the gene of interest and a stabilising element.

9
Q

What is the promotor in a gene expression construct?

A

Promotor is used to induce high levels of expression, or make it cell specific by making it inducible in only certain cells.

10
Q

In making a promotor for DMD, it is best to make it specific to which tissue?

A

Muscle tissue (including cardiac).

11
Q

Aside from inducing high expression levels, what else can a promotor do?

A

Allows you to induce manually, such that one can induce when needed and turn off when uneeded.

12
Q

How can the gene of interest in a gene expression construct be easily identified?

A

Adding a tag.

13
Q

What is the purpose of a stabilising element in a gene expression construct?

A

Prolongs lifespan of the construct, allowing it to enter the cell.

14
Q

How does cell based therapy differ to genetic therapy?

A

The cell is already modified to express the new gene, is injected into the affected area to replace defective cells.

15
Q

In treating DMD, what must the treatment used have to do, and what percentage of muscles must be restored to begin improving function?

A

Must be able to enter both skeletal and cardiac cells, must be specific to them, and must be safe.
Only need 20% restoration to improve quality of life.

16
Q

Whats special about the dystrophin gene? Why is this an issue? How can it be resolved?

A

Its the largest gene in humans. Hard to deliver the entire corrected gene. Can cut out large parts of the gene and still have a functional protein, is truncated.

17
Q

What are the two main non-viral therapy techniques?

A

Unencapsulated plasmid delivery

Dystrophin restoration.

18
Q

What is unencapsulated plasmid delivery, and give a pro/con.

A

Form a plasmid vector with the corrected gene, then inject directly to muscles.
Is safe and simple, but very low efficiency, only 1% of localised cells (injection site, not the whole muscle) epress the gene.

19
Q

How can unencapsulated plasmid delivery be improved (5)?

A

Can combine with other therapies, add a carrier molecule (lipid) and add a DNA stabiliser.
Can use ultrasound to increase permeability.
Also electroporation, but damaging to cells.

20
Q

How can hyaluronidase be used to improve unencapsulated plasmid delivery?

A

It breaks down the extracellular matrix, and improves efficiency when paired with electroporation, not localised to injection site, but the whole muscle.

21
Q

Can unencapsulated plasmid delivery transfer the entire dystrophin gene? Is it permanent?

A

Yes, it can. It is not permanent, is only local to injection site, and repeat injections are needed.

22
Q

Name 2 dystrophin restoration techniques.

A

Stop codon read through

Exon skipping

23
Q

What is stop codon read through? What mutation does it work with?

A

Only works with nonsense mutation, 10-15% of DMD cases.
Is a premature stop codon, truncating the protein.
Stop codon read through uses a drug to allow ribosome to bypass the stop codon and keep reading.

24
Q

What are two drugs in stop codon read through, and what do they interact with? Are they effective?

A

Gentimycin - ribosomal subunit 140s, drops CK levels too, moderately effective.
Ataluren - ribosomal subunit 60s

25
Q

What is exon skipping? What type of mutation does it target? What is a consequence of this?

A

Targets patients with internal deletions.
Restores the mRNA reading frame by skipping an exon downstream of the mutated exon.
ie exon 50 deleted, exon 51 skipped. Will have a missing section, but still gives a usable form of dystrophin.

26
Q

What 3 drugs are used in exon skipping? Are they effective?

A

2MeAON - not very effective
Drisapersen - being reviewed
PMO - most efficient

27
Q

What is a problem with PMO? What was done to resolve it? What new issue arose?

A

It didnt affect the heart much.

PPMO was developed, is PMO with a cell permeable peptide added. Is toxicity associated with it however.

28
Q

Which therapies are patient specific?

A

Stop codon read through

Exon skipping

29
Q

Does exon skipping require repeat injections or is it permanent?

A

Repeat injections needed.

30
Q

Collectively, what percentage of DMD cases can exon skipping and stop codon read through treat and why?

A

Only 30% at best due to the nature of their mutation targets.

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