Biotech: Gene Therapy

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Defined as the introduction of corrected genes into patient cells

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gene therapy

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Describe 2 types of gene therapy:

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1) Germ-line
2) Somatic - changes in gene expression in somatic or non-germ cells to restore fcn or target abnormal cellsfor elimination (ex. cancer cells)

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Requirements of Gene Therapy:

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1) Caused by a single gene defect
2) Affects one cell type w/c must be accessible for removal/replacement
3) Disease causing gene must be already identified and cloned
4) Regulated expression of this gene is not required to correct the defect

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Is Somatic Gene Therapy heritable?

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no

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What is the therapeutic agent to modify gene expression in a specific target cell and thus does not alter permanently the heritable genetic material of the individual?

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corrective gene

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What is involved in current Gene Therapy?

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Gene addition whereby an intact version of gene is added to the DNA chromosome of the target cell population

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What are the 2 Gene Therapy Approach:

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1) Ex vivo
2) In vivo

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It involves the transfer of cloned genes into cells grown in culture.

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Ex vivo

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What kind of cell is usually used in ex vivo therapy?

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autologous cells

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What type of cells can be collected from patient and introduced to same patient in utilizing ex vivo therapy?

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cells of hematopoietic system and skin cells

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What are autologous cells?

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involves transplantation of patient's own genetically modified cells

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How does ex vivo therapy correct genetic defect from collected cells?

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gene transfer

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How does in vivo gene and ex vivo transfer differ?

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- IN VIVO cloned genes are transfered directly into patient tissue.
- IN VIVO the therapeutic DNA is injected directly into the body cells, usually via one of two types of viruses. The most frequently used type is the very simple retrovirus.

- EX VIVO involves surgically removing cells from the affected tissue area, injecting or splicing the new DNA (the DNA that will correct the disease) into the cells and letting them divide in cultures. The new tissues are placed back into the affected area of the patient.

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What types of tissues can be used in in vivo gene transfer?

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tissues that cannot be cultured in vitro in sufficient numbers (ex. brain cells)

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in vivo gene therapy requires 2 things before it is delivered to cells of a patient.

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1) promoter - tissue specific
2) remedial gene - encodes a protein that corrects the genetic defect

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What happens following gene transfer?

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inserted gene may integrate into chromosomes genetic elements (episomes)
-episomes independently replicate outside nucleus but not integrate w/ human DNA

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Episomes vs Plasmids

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- An episome is a non-essential genetic element. In addition to its independent existence, an episome can also exist as an integrated part of the host genome of bacteria. It originates outside the host, in a virus or another bacterium. When integrated, a new copy of the episome will be made as the host chromosome undergoes replication. As an autonomous unit, the viral episome genetic material destroys the host cell as it utilizes the cellular replication machinery to make new copies of itself. But, when integrated into the bacterial chromosome they multiply in cell division and are transferred to the daughter cells.
- In contrast to episomes, a plasmid exists only as an independent piece of DNA. It is not capable of integration with the chromosomal DNA; it carries all the information necessary for its own replication.

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What is the target in actively dividing cells?

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stem cells

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Disadvantage of chromosomal integration:

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1) random insertion = location varies
2) inserted gene may not be expressed
3) integration may result in death of host cell