Gene Therapy eolss net

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PHARMACOLOGY Vol II Gene Therapy K K Jain,4 Concluding remarks. Bibliography,Biographical Sketch, Gene therapy can be broadly defined as the transfer of defined genetic material to. specific target cells of a patient for the ultimate purpose of preventing or altering a. particular disease state Two main approaches to gene therapy include in vivo and ex. vivo Ex vivo gene transfer techniques usually involve the genetic alterations of cells. cell lines or human cells mostly by use of viral vectors prior to implanting these into. the tissues of the living body In vivo gene therapy means direct introduction of genetic. material into the human body It can be accomplished by use of non viral vectors In. addition to gene transfer and in vivo production of therapeutic proteins some techniques. are aimed at repairing defective genes or suppressing the function of genes. This article describes the use of various biotechnologies in development of gene. therapy The most frequent method of gene transfer is by viral vectors but several non. viral vectors are also being developed Several routes of administration have been used. although injection is still the most commonly used method Efforts are also being made. to develop orally administered gene therapy Physical methods of gene transfer include. ultrasound electroporation laser irradiation and gene gun Related methods include use. of genetically modified cells antisense therapy and RNA interference Gene therapy is. an excellent method of delivery of therapeutics and various methods of targeted and. controlled gene therapy are described Gene therapy can be considered for almost any. disease Major applications that are currently under investigation include cancer. cardiovascular diseases neurological disorders and infections Some complications of. gene therapy have been encountered and these are being addressed It is anticipated that. gene therapy will become established as a part of human medicine during the next. decade and will fit in with the concepts of personalized medicine. 1 Introduction, Gene therapy can be broadly defined as the transfer of defined genetic material to. specific target cells of a patient for the ultimate purpose of preventing or altering a. particular disease state Applications of gene therapy would be narrow if one uses the. term as a synonym for transfer of defined genetic material to specific target cells using. carriers or delivery vehicles called vectors Vectors are usually viral but several non. viral techniques are being used as well Genes and DNA are now being introduced. without the use of vectors and various techniques are being used to modify the function. of genes in vivo without gene transfer If one adds to this the cell therapy particularly. with use of genetically modified cells the scope of gene therapy becomes much. broader Gene therapy can now combined with antisense techniques such as RNA. interference RNAi further increasing the therapeutic applications. A study of the historical evolution of gene therapy gives an idea of the time lapsing. between fundamental discoveries animal experiments and human applications of a new. Encyclopedia of Life Support Systems EOLSS,PHARMACOLOGY Vol II Gene Therapy K K Jain. technology Although the capability of viruses to transmit genes was recognized as far. back as 1952 suggestion for their use as vectors for gene transfer was not made until 20. years later Another decade passed before retrovirus vectors were used for gene transfer. and the first authorized gene therapy experiment in humans did not take place until a. further decade later in 1992 In 1999 the death in the US of Jesse Gelsinger following. gene therapy led to a setback and an intense scrutiny of the supervision process in gene. therapy trials In 2001 the US FDA introduced stricter guidelines and regulations for. gene therapy There are several late stage clinical trials in cancer gene therapy The first. product for gene therapy of cancer was approved in China in 2003. 2 Relation of Gene Therapy to Other Biotechnologies. Figure 1 Relation of gene therapy to other biotechnologies. Gene therapy is also related to other biotechnologies particularly molecular diagnostics. and genomics proteomics Molecular diagnosis can be defined as the clinical. application of molecular technologies to elucidate diagnose and monitor human. diseases Molecular technologies include the use of DNA deoxyribonucleic acid and. RNA ribonucleic acid as well as recombinant antibodies Polymerase chain reaction. PCR is the best known of molecular diagnostics and has applications in gene therapy. Genomics is the study of all of the genes in an organism their sequences structure. regulation interaction and products Pharmacogenomics an offshoot of genomics. means the application of genomic technologies to drug discovery and development The. completion of the Human Genome Project and the advent of the postgenomic era have. its impact on gene therapy as well Commercial applications of the knowledge of the. human genome include gene therapy transfer of DNA sequences into cells to correct. genetic defects destroy abnormal cells or regulate cellular function Relation of gene. therapy to other biotechnologies is shown in Figure 1. Encyclopedia of Life Support Systems EOLSS,PHARMACOLOGY Vol II Gene Therapy K K Jain.
3 Gene Therapy Technologies,3 1 Classification of Gene Therapy Techniques. Essentially gene therapy involves three steps administration delivery and expression. Administration means introduction of the gene or a vector containing the gene into. Delivery means the transfer of the gene from the site of the administration to the. nucleus of the target cell, Expression means the production of a therapeutic gene product in the cell. A simplified classification of various methods of gene therapy is shown in Table 1. Gene transfer,Chemical calcium phosphate transfection. Physical ultrasound electroporation laser irradiation gene gun. Transduction with recombinant virus vectors,Adeno associated virus. Adenovirus,Herpes simplex virus,Lentivirus,Moloney murine leukemia virus.
Retroviral vectors,Vaccinia virus,Other viruses,Nonviral vectors for gene therapy. Ligand polylysine DNA complexes,Dendrimers and other polycationic polymers. Synthetic peptide complexes,Artificial viral vectors. Artificial chromosomes, Use of genetically modified microorganisms as oncolytic agents. Genetically modified viruses,Genetically modified bacteria.
Cell therapy,Administration of cells modified ex vivo. Implantation of genetically engineered cells to produce therapeutic substances. Gene DNA administration, Direct injection of naked DNA or genes systemic or at target site. Receptor mediated endocytosis, Use of refined methods of drug delivery e g microspheres. Gene regulation, Regulation of expression of delivered genes in target cells by locus control region. technology,Light Activated Gene Therapy, Molecular switch to control expression of genes in vivo.
Promoter element triggered gene therapy,Encyclopedia of Life Support Systems EOLSS. PHARMACOLOGY Vol II Gene Therapy K K Jain,Repair of defective genes. Involves correction of the gene in situ e g chimeraplasty. Gene repair mediated by single stranded oligonucleotides. Gene replacement, Excision or replacement of the defective gene by a normal gene. Spliceosome mediated RNA trans splicing,Inhibition of gene expression. Antisense oligodeoxynucleotides,Antisense RNA,RNA interference.
Reproduced from Jain KK Gene Therapy technologies markets and companies Jain. PharmaBiotech Publications Basel 2005,Table 1 Classification of methods of gene therapy. 3 2 Physical Methods of Gene Transfer,3 2 1 Electroporation. This is a method of physical transfection for temporarily creating small openings or. pores in cell membranes to introduce macromolecules such as DNA The openings are. created by applying an electrical field to the cell for a few microseconds to a few. milliseconds their size varies according to the amplitude and duration of the pulses. Once the DNA has diffused into the cell the power is discontinued and the pores reseal. themselves, Although DNA can be transported into cells by electroporating their membranes the. molecular basis of this process is controversial Currently there are two major. hypotheses In the first DNA is transported through large stable pores that form without. significant interaction with DNA and then reseal The second more widely supported. hypothesis suggests that DNA is transported into cells by direct interaction with lipids. and intermediates that involve a complex of lipid components and DNA In this case. permeabilization by an electric field might cause a number of smaller pores that by. themselves are not large enough to allow DNA transport but make the membrane. amenable to structural modifications that allow translocation of DNA The simulation of. the large bilayer shows holes that have a diameter of up to 10 nm which would be. sufficiently large to allow the transport of double stranded DNA but and it remains to. be shown that pores of this size are reversible and will reseal when the electric field is. removed Computer simulations of lipid bilayers in the presence of an external electric. field show that pore formation is promoted by an increased likelihood of. transmembrane water defects caused by the interaction of water dipoles with the electric. field gradient at the water lipid or water octane interface. 3 2 2 Particle Bombardment, This method is also known as the gene gun technology or biolistic or ballistic. microprojectile method Gene gun is a convenient hand held device that provides rapid. Encyclopedia of Life Support Systems EOLSS,PHARMACOLOGY Vol II Gene Therapy K K Jain.
and direct gene transfer into a range of targets under in vivo or in situ conditions. Particle mediated gene transfer advanced with the introduction of the helium chamber. in which helium replaced gunpowder as the source of acceleration Modern units. employ an adjustable low pressure helium pulse to sweep DNA or RNA coated gold. microcarriers from the inner wall of a small plastic cartridge directly into the target. Targeted gene delivery is now a reality for in vivo research applications The. microcarriers accelerate for maximum penetration as they move through the barrel. while the helium pulse diffuses outward The spacer maintains the optimal target. distance for in vivo applications and vents the helium gas away from the target to. minimize cell surface impact,3 2 3 Ultrasound mediated Transfection. Several studies during the past decade have suggested that transfection may be. stimulated by ultrasound but the transfection efficiency was low Ultrasound increases. the permeability of the cell membrane and facilitates the passive diffusion of plasmids. into the cells Cavitation microbubbles caused by ultrasound energy produces. breaches in cell membrane and is a key component in ultrasound mediated transfection. Ultrasound is a promising method for introducing foreign DNA or other. macromolecules into cell Advantages of ultrasound mediated transfection are. Ultrasound energy can be transmitted to the walls of containers normally used to. grow cells in vitro, Ultrasound may be used for transfection in vivo because it can be focused into. restricted areas or cavities of the body, Tremendous enhancement in gene expression can be achieved with ultrasound alone. following either intratumoral intramuscular or intravenous injection of a gene product. Ultrasound targeted microbubble destruction can be used to deliver adenoviral or. plasmid DNA to the myocardium This technique holds great promise in applying the. rapidly expanding repertoire of gene therapies being developed for cardiac disease. Cationic acoustic liposomes whose composition and structure enables them to reflect. ultrasound have been used to improve the efficacy of liposomal gene transfer The. ultrasound increases DNA release from liposomes at much lower power than is required. to produce the same effect with standard liposomes Acoustic liposomes conjugated to. antibodies have been shown to target sites of vascular disease This technology will. enable simultaneous identification of the disease site and activation of the therapeutic. agent in situ,3 2 4 Molecular Vibration, Electric field induced molecular vibration enables foreign DNA molecules to penetrate. the plasma membrane and enter the cytoplasm of both primary mesenchymal progenitor. cells and established cell lines of various species at high efficiency and with low cell. mortality This procedure requires no special reagents allows stable expression of. transduced DNA and does not interfere with the normal cellular differentiation. activities of human and chick mesenchymal progenitors. Encyclopedia of Life Support Systems EOLSS,PHARMACOLOGY Vol II Gene Therapy K K Jain.
3 2 5 Gene Transfection using Laser Irradiation, Laser beams have been used for gene transfection The principle of this procedure is. that a small hole is made in a cell membrane by pulse wave Nd YAG. neodymium yttrium aluminum garnet laser irradiation and a gene contained in a. medium is transferred into the cytoplasm through the hole This method enables. transfection of targeted cells i e cells in suspension as well as attached cells This. gene therapy will become established as a part of human medicine during the next decade and will fit in with the concepts of personalized medicine 1 Introduction Gene therapy can be broadly defined as the transfer of defined genetic material to specific target cells of a patient for the ultimate purpose of preventing or altering a particular disease state Applications of gene therapy would

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