Gene Therapy: Delivery Flashcards
(22 cards)
What is the Process of Gene Therapy?
What is the Purpose?
- Exogenous genetic material is transferred into somatic cells to correct an inherited or acquired gene defect
- Introduce new function or property to the cell
What are examples of Genetic Diseases that Gene Therapy is used in?
- Cystic fibrosis
- Hemophilia
- Sickle cell anaemia
- Diabetes
What are examples of Acquired Diseases that Gene Therapy is used in?
- Cancer
- Cardiovascular
- Neurological disorders
- Infectious
What is the Concept of Gene Therapy?
- Take piece of genetic material and target the right cells
- Encapsulate it in some sort of delivery system
- Get it into the correct cells in human body and deliver it to cells nucleus
- Integrate DNA into the cell
- Activate the gene - transcribe and translate it to make the protein product it encodes
- Disease treated
RNAi
What is Gene Silencing?
- Cleavage of the targeting mRNA
- Doesn’t change DNA, it just stops it working
What is the process of RNA process in Gene Therapy?
- Get RNA molecule transvected into cell
- Pulled apart into two strands
- One of the strands cleaves the mRNA within the cell
- Gene silenced - no longer creates protein because mRNA molecule has cleaved part of gene that makes the protein
What are the Types of Material Delivered?
- DNA
- RNA
- Oligonucleotides
What is a major challenge of delivery?
- Ribonuclease attack
- Repulsion between negatively charged genetic material and cell wall (also negatively charged)
- As soon as genetic material comes in contact with the atmosphere it undergoes this attack and material is degraded so we need protection against this
What are Vectors used for?
What can vectors be?
- Carriers designed to protect genetic material, enter cells and deposit therapeutic genetic material
- Vectors can be viral or non-viral
What is the function of Viral Vectors?
What are examples of Viral Vectors?
- Replicate by inserting their DNA into a host cell
- e.g. retroviruses, adenovirus, adeno-associated virus
Function of Viral Vectors: What is the Role of Retroviruses (e.g. HIV)?
- Carry genetic material in the form of RNA
- Infect dividing cells only
- Target specific cells through engineering proteins on surface
- May cause an immune resonse
Function of Viral Vectors: What is the Role of Adenovirus (e.g. causes common cold)?
- Carry genetic material in the form of double-stranded DNA
- Infect both dividing and non-dividing cells
- Target specific cells through engineering proteins on surface
- Many patients have developed immunity = problem
Function of Viral Vectors: What is the Role of Adeno-Associated Virus (e.g. don’t cause illness in humans)?
- Carry genetic material in the form of single-stranded DNA
- Infect dividing and non-dividing cells
- Target specific cells through engineering proteins on surface
- Typically, no immune response so it’s useful for delivering genetic material
What are the Limitations of Viral Vectors?
- Complex formulation
- Storage related difficulties
- Off target effects (toxicity, immunogenicity, tumour growth)
What are examples of Non-viral Vectors?
- Naked DNA
- Liposomes (lipofection)
- Dendrimers
Non-viral Vectors: What is the Role of Naked DNA?
- Circular double-stranded DNA
- Forms loop which gives it more stability but still more prone to degradation by enzymes
- DNA transported to cell’s nucleus
- Less efficient compared to viral vectors
- No immune response
Non-viral Vectors: What is the Role of Liposomes (lipofection)?
- Circular double-stranded
- Liposomes interact with cell and deliver DNA
- Interaction into host cell genome has very low efficiency
- Less efficient compared to viral vectors
- No immune response
Non-viral Vectors: What is the Role of Dendrimers?
- Special type of polymer
- Condense DNA/siRNA
- Protect from enzymes, makes genetic material smaller and able to cross cell barrier through endocytosis
What are Other Physical Methods of Gene Therapy Delivery?
- Gene gun
- Sonoporation:
- ultrasonic frequencies to deliver DNA
- Magnetofection
- DNA complexed with magnetic particles
- Electroporation
- short pulses of high voltage, allows DNA to pass through porous cell membrane
What are In-vivo Gene-Therapy Approaches?
- Drug taken into human somehow
- Usually injection
- Can be difficult to target the right place
- If it goes into systemic circulation, there’s a lot of parts of the body that degrade the genetic material before it get’s there
What are Ex-Vivo Gene-Therapy Approaches?
- More common
- Correct cells removed from human
- Treated outside body allowed to grow and put back into body
What are challenges for Gene Therapy Delivery?
- Delivery and activation
- Immune response – can cause serious illness or even death
- Viral vectors – patient may have toxic, immune, inflammatory response
- May induce tumor etc due to insertional mutagenesis
- Multigene Disorders
- Cost
