gene therapy Flashcards
Define Gene Therapy
Any procedure intended to treat or alleviate disease by genetically modifying the cells of the patient
Name 4 types of disorders gene therapy can treat
Inherited diseases
Cancers
Infectious disease
Immune disorders
Name the two broad categories of gene therapy
Germline cell modification
Somatic cell modification
What is Germ-line cell genetic modification?
Permanent modifications transmitted to descendants. Achieved by modifying DNA of a gamete/zygote or early embryo
Is Germ-line cell modifications allowed in humans?
NO
Can somatic cell genetic modification be passed on?
no
What are the two strategies to somatic cell gene therapy?
Modify the disease cell to alleviate the disease
Selectively kill the diseased cell
Name how disease cells can be modified to alleviate the disease
gene augmentation
gene silencing
repair of the mutant gene
What is gene augmentation?
Adding functional copies of the gene back into cells that lack them.
What is gene silencing?
Expression of a gene is selectively inhibited by turning off transcription
Describe how to selectively kill diseased cells
Directly kill with targeting cells eg suicide gene Or indirectly ( assisted killing) by modifying the immune system to enhance response to disease cell and initiate cell death. Uses Prodrug
What is the difference between In-vivo and ex-vivo gene transfer?
In-vivo= direct delivery Ex-vivo= cell based delivery
What are the issues with short lived cells
Cells need to divide to replenish lost cells so gene therapy genetic material might not be passed into newly divided cells
What 4 things does gene transfer need to meet?
Non toxic
Target the DNA to a particular cell
Introduce DNA into a cell in a transcriptionally active form
Express the gene at a reasonable level for sufficient time
Name a non viral delivery vehicle
Liposome
Name 4 viral delivery vehicles
Retrovirus
Lentivirus
Adenovirus
Adeno-associated viruses
Describe how liposomes are delivery vehicles
Liposome contains genetic information
Taken up via endocytosis or clathrin coated pits
Taken into nucleus for gene to be expressed
What is the gene placed in to achieve high levels of expression?
Plasmid expression vector
Name the common expression vector
pJ7 omega
Name the parts of the expression vector
Polyadenylation site Splice donor site Splice Acceptor site CMV promoter cloning sites
What is the role of the polyadenylation site?
Efficient termination of transcription in the expression vector
What is the role of the CMV promoter?
Drives transcription, strongest promoter, works in all eukaryotic cells
What happens to the expression vector in nucleus of patient cell?
RNA polymerase binds CMV and transcribes RNA
RNA cleaved and polyadenylated then spliced
Why is plasmid inefficiently transported into the nucleus?
Due to its large size it cannot always enter through the nuclear pores
What are the two approaches that can get the plasmid DNA into the nucleus
COnjugating specific DNA or protein sequencing
Compact the DNA
How many people does Cystic Fibrosis affect?
1/2500
What causes Cystic Fibrosis?
Defective CFTR gene, which encodes for a Cl- ion trasnporter
Name the main advantage of liposomes?
Non-toxic
What are the disadvantages of lipsomes?
Inefficient transfer of DNA to target cells
Cannot target particular cell types
Poor expression
