Week 1 - Gene Therapy

Front 1

What is gene therapy?

Back 1

An approach to treating disease by adding, augmenting, or repairing genes

Front 2

What are some diseases that may be treated with gene therapy?

Back 2

Cystic fibrosis, sickle cell anemia, muscular dystrophy, cancer, AIDS

Front 3

What are the two modes of gene delivery?

Back 3

Ex vivo - the gene is put into isolated cells
In vivo - the gene is put directly into patients

Gene delivery may also be viral or non viral

Front 4

What are the characteristics of an ideal vector for gene therapy?

Back 4

Safe
Sufficient capacity for size of therapeutic DNA
Non-Immunogenic
Allow re-administration
Ease of manipulation
Efficient introduction into target cells/tissues
Efficient and appropriate regulation of expression
Level, tissue specificity, transient, stable?

Front 5

What are the non viral methods of gene delivery? Why are non viral methods difficult?

Back 5

Microinjection, electroporation, or complexing with a carrier (liposomes, cationic lipids). It's difficult because naked DNA or RNA cant enter cells!

Front 6

What is a viral vector?

Back 6

A virus that has been adapted to serve in gene delivery. They include retroviruses, adenoviruses, lentiviruses, and adeno-associated viruses (AAV)

Front 7

How can you use in vivo viral gene therapy to cure a mouse with muscular dystrophy?

Back 7

Create viral vectors with the dystrophin gene. Inject the virus into the mouse. Muscle cells pick up the DNA, incorporate it into their genome, and can produce dystrophin.

Front 8

How and why did gene therapy for hemophilia B (AAV vector used in liver cells) fail? How will future studies remedy this problem?

Back 8

Some patients treated with gene therapy produced factor IV and normal levels, but only transiently. Eventually, the patient's immune response destroyed the liver cells containing the therapeutic gene. In future studies, patients will be given immunosuppressive treatment to prevent the destruction of these cells.

Front 9

Are there risks associated with viral gene therapy?

Back 9

Yes. One patient received high doses of the virus, had a massive infammatory response, and died a few days later.

Front 10

Are problems in bone marrow treated without gene therapy?

Back 10

You can do a bone marrow transplant with healthy cells, say from a brother or sister.

Front 11

What is a complication of bone marrow transplants from relatives?

Back 11

Graft Versus Host disease, where the body rejects the new tissue

Front 12

How can you restore hematopoietic stem cells without facing the complications of graft versus host disease?

Back 12

Take out the patients own hematopoietic stem cells, fix the genetic defect, then reimplant it in the patient

Front 13

How do retroviral vectors work in gene therapy?

Back 13

Take the retrovirus RNA, replace the proliferation genes with your gene of interest, and let it integrate into your patient's tissues. This way it doesn't spread, but still let's your gene of interest incorporate into your patient's genome.

Front 14

What is SCID? What gene mutation is associated with SCID? Can ADA enzyme function be restored with gene therapy?

Back 14

SCID, severe combined immuno deficiency, or bubble boy disease, is a syndrome where infants are born without a functioning immune system (they lack all lymphocytes and suffer from opportunistic infections). The ADA gene, which codes for a protein required for immune cell survival. It appears that the enzyme can be restored with gene therapy, over time.

Front 15

Was gene therapy for XSCID patients successful? How is the gene therapy technique being modified?

Back 15

Gene therapy treatment for XSCID restored most of the patient's immunes systems. However, 1/4 of patients developed a leukemia-like disorder in 2-3 years. This is due to the potent retrovirus promoter activating oncogenes. Future studies will attempt using a less powerful promoter.

Front 16

What is a current treatment of sickle cell disease? What would gene therapy treatment involve?

Back 16

The current treatment is transplanting HSC from a healthy individual to the patient. Gene therapy would involve inserting an "anti-sickling" gene, as seen in fetal hemoglobin, into patient HSC genome

Front 17

What is the molecular basis of sickle cell disease?

Back 17

An amino acid substitution, caused by a point mutation, resulting in the polymerization of hemoglobin and the cells to develop a "sickle" shape

Front 18

How do sickle cell patients with higher amount of fetal hemoglobin levels compare to those with lower amount of fetal hemoglobin?

Back 18

Patients with fetal hemoglobin live longer and less severe symptoms

Front 19

What genetic factors affect susceptibility to HIV? What happened with the HIV patient who received a bone marrow transplant from a CCR5 mutation donor?

Back 19

The CCR5 receptor protein is an important cellular protein involved in the initial step of HIV infection. When this protein is mutated, susceptibility to HIV is decreased. For the patient who received a bone marrow transplant, the HIV became undetectable

Front 20

What gene therapy treatments are available for HIV patients?

Back 20

Inserting a gene for an inhibitory RNA gene, which interferes with CC35 protein translation.

Introducing a double strand break in DNA using gene-specific zinc finger nucleases, and letting the cell repair the DNA imperfectly, which disrupts the CCR5 gene

Front 21

What gene therapy treatments are available to cancer patients?

Back 21

T cells recognize and attack tumor cells based on recognition of foreign surface proteins. Take the T cell receptor genes from a patient who has beaten cancer and insert them into a cancer patient to re engineer their immune system.

Front 22

What is one mechanism of gene repair?

Back 22

Introducing a double stranded break using zinc finger endonucleases, then relying on homologous recombination to repair the gene

Front 23

What are some advantages gene correction has over gene addition?

Back 23

In gene correction, the therapeutic gene is already in the normal location, so expression should be normal. It also avoids problems of genes inserting semi-randomly throughout the genome, which may lead to activation of nearby genes.

Front 24

How can gene therapy be used to generate healthy patient HSCs from a skin biopsy, per se?

Back 24

Tissue samples, such as a skin biopsy, can be taken from a patient. Gene addition or correction is done to this cell. It is then de-differentiated to a pluripotent stem cell, which can then be differentiated into a HSC. The HSC is now transplanted into the patient.