Gene Therapy Vasileios Antonopoulos (Teacher) Evangelos Tsouramanis (Teacher) Eleni Loukopoulou (Student) Vasileios Vagenas (Student)

Genes Are carried on a chromosome Are composed of strands of a molecule called DNA Hold the instructions how to make a protein Proteins are chemicals that enable the body to work and grow

Picture of a Chromosome

What are Genetic Disorders Every human has about 25,000 genes per cell A mutation, or change, in a gene can cause the gene not to make a normal copy of the protein that it encodes A mutation can result in a disease, physical disability, or shortened life span The above problems caused by mutations are called Genetic Disorders Mutations can be inherited, or can occur spontaneously in some cases

What is Gene Therapy It is a technique for correcting defective genes that are responsible for disease development Approaches to gene therapy that are being tested : Replacing a mutated gene that causes disease with a healthy copy of the gene Inactivating, or “knocking out”, a mutated gene that is functioning improperly Introducing a new gene into the body to help fight a disease

Types of Gene Therapy Somatic Gene Therapy involves introducing a “good” gene into targeted cells to treat the patient – but not the patient’s future children because these genes do not get passed to offspring Germline Gene Therapy involves modifying the genes in egg or sperm cells, which then pass any genetic changes to future generations. This type of therapy is controversial and very little research is being done in this area, both for technical and ethical reasons

Vector Systems A gene cannot be directly inserted into a person’s cell. It must be delivered to the cell using a carrier, or vector Vector systems can be divided into: Viral Vectors Non-viral Vectors Currently, the most common type of vectors are viruses

Delivery techniques Ex vivo = delivery of genes takes place out of the body, and then cells are placed back into the body In vivo = delivery of genes takes place in the body

Picture of Delivery techniques

How Gene Therapy Works A vector delivers the therapeutic gene into a patient’s target cell The target cells become infected with the vector The vector’s genetic material is inserted into the target cell Functional proteins are created from the therapeutic gene causing the cell to return to a normal state

Viruses Scientists manipulate the genome of the virus by removing the disease-causing genes and inserting the therapeutic genes Four different types Retroviruses Adenoviruses Adeno-associated Herpes Simplex

Non-viral Options Direct introduction of therapeutic DNA into target cells. This approach is limited in its application because it can be used only with certain tissues and requires large amounts of DNA Creation of artificial liposomes, which carry the therapeutic DNA and are capable of passing the DNA through the target cell’s membrane

Non-viral Options cont. Chemically linking DNA to molecule that will bind to special cell receptors. Once bound to these receptors, the therapeutic DNA constructs are engulfed by the cell membrane and passed into the interior of the target cell. This delivery system tends to be less effective than other options. A 47 th artificial chromosome. This chromosome would exist autonomously alongside the standard 46 -not affecting their workings or causing any mutations. It would be a large vector capable of carrying substantial amounts of genetic code, and scientists anticipate that, because of its construction and autonomy, the body's immune systems would not attack it. A problem with this potential method is the difficulty in delivering such a large molecule to the nucleus of a target cell.

Problems with Gene Therapy Short-lived nature of gene therapy - Before gene therapy can become a permanent cure for any condition, the therapeutic DNA introduced into target cells must remain functional and the cells containing the therapeutic DNA must be long-lived and stable. Problems with integrating therapeutic DNA into the genome and the rapidly dividing nature of many cells prevent gene therapy from achieving any long-term benefits. Patients will have to undergo multiple rounds of gene therapy. Immune response - Anytime a foreign object is introduced into human tissues, the immune system is designed to attack the invader. The risk of stimulating the immune system in a way that reduces gene therapy effectiveness is always a potential risk. Furthermore, the immune system's enhanced response to invaders, it has seen before, makes it difficult for gene therapy to be repeated in patients.

Problems with Gene Therapy cont. Problems with viral vectors - Viruses, while the carrier of choice in most gene therapy studies, present a variety of potential problems to the patient --toxicity, immune and inflammatory responses, and gene control and targeting issues. In addition, there is always the fear that the viral vector, once inside the patient, may recover its ability to cause disease. Multigene disorders - Conditions or disorders that arise from mutations in a single gene are the best candidates for gene therapy. Unfortunately, some of the most commonly occurring disorders, such as heart disease, high blood pressure, Alzheimer's disease, arthritis, and diabetes, are caused by the combined effects of variations in many genes. Multigene or multifactorial disorders, such as these, would be especially difficult to treat effectively using gene therapy.

The First Case The first gene therapy was performed on September 14 th, 1990 Four-year old Ashanti DeSilva was treated for ADA- SCID (Adenosine Deaminase - Sever Combined Immune Deficiency) She lacked a healthy immune system, and was vulnerable to every passing germ Doctors removed her white blood cells, inserted the missing gene into the cells, and then put them back into her blood stream. This strengthened her immune system Only worked for a few months, and the process must be repeated every few months

The First Case cont.

An Unsuccessful Gene Therapy Jesse Gelsinger case: An 18-year-old with ornithine transcarbamylase (OTC) deficiency participated in a 1999 gene therapy trial at the University of Pennsylvania OTC is one of many enzymes that break down excessive nitrogen in our cells; when OTC is deficient, ammonia builds up and poisons the brain On Monday, September , Gelsinger was injected with adenoviruses carrying functioning OTC genes Adenovirus triggered an overwhelming inflammatory response that caused massive organ failure and finally the death of Gelsinger after three days His death halted all gene therapies in the United States for a time

Recent Gene Therapy for Blindness Leber’s Congenital amaurosis (LCA) is a rare inherited eye disease that appears at or in the first few months of life LCA causes progressive loss of vision, usually leading to complete blindness by age 40 LCA can be caused by mutations in several genes, one of which is called RPE65. This gene is necessary to manufacture an enzyme needed by the retina’s rods and cones

Recent Gene Therapy for Blindness cont. Researchers at Moorfields Eye Hospital and University College London’s institute of Ophthalmology conducted the first gene therapy clinical trial for patients with RPE65 LCA. The first patient was operated upon in early 2007 Researchers at Children’s Hospital of Philadelphia and the University of Pennsylvania have treated six young people via gene therapy They researched the safety of the subretinal delivery of recombinant adeno associated virus (AAV) carrying RPE65 gene, and found it yielded positive results, with patients having modest increase in vision, and, perhaps more importantly, no apparent side-effects

Recent Gene Therapy for Blindness cont.

Gene Therapy for Cancer Multiple gene therapy strategies have been developed to treat a wide variety of cancers. Two-thirds of all gene therapy trials are for cancer and many of these are entering the advanced stage, including a Phase III trial for head and neck cancer and two different Phase III gene vaccine trials for prostate cancer and pancreas cancer. Numerous Phase I and Phase II clinical trials for cancers in the brain, skin, liver, colon, breast and kidney among others, are being conducted in academic medical centers and biotechnology companies, using novel technologies and therapeutics developed on-site.

Some Questions to Consider When should gene therapy be used? Should it be used to treat critically ill patients? Should it be used to treat babies and children? What effect would gene therapy have on future generations if germline (reproductive) cells were genetically altered? How might this alteration affect human variation? Who should decide what are "good" or "bad" uses of genetic modifications? How do you define "normal" with regard to human beings? What if we could alter human traits not associated with disease? Would it be okay to use gene therapy to improve or enhance a person's genetic profile? Who will have access to gene therapy, treatments and long- term follow-ups? Will gene therapy and genetic enhancements create an advantage for those who can afford it?

Some Questions to Consider cont. The questions raised here have no clear right or wrong answer. Your responses will depend on your values, as well as on the opinions of those around you.

References GeneTherapyReview : The American Society of Gene & Cell Therapy : Human Genome Project Information: genomics.energy.gov /genomics.energy.gov Gene Therapy Net : National Human Genome Research Institute: