CG920 Genomics Lesson 11 Genomics – practical applications Markéta Pernisová Functional Genomics and Proteomics of Plants, Mendel Centre for Plant Genomics and Proteomics, CEITEC - Central European Institute of Technology, Masaryk Univerzity, Brno marketa.pernisova@ceitec.muni.cz, www.ceitec.muni.cz 1 Outline 1. Genomics in medicine 2. Biotechnology 3. Genetically modified organisms 4. GMO – pros and cons 5. Discussion 2 GENOMICS IN MEDICINE 3 GENOMICS IN MEDICINE • Biotechnology and gene technology are crucial technologies in medicine • Improvement in practice: • faster identification and analysis of new pathogens • faster development and production of vaccines and reliable diagnostic tools • genetic tests for screening of heritable diseases or severe genetic defects in the embryo or foetus • many drugs produced in laboratories: • bacteria, yeast and mammalian cell cultures produce human proteins available as drugs • developing of e.g. edible vaccines produced by transgenic plants • 1st biotech drug: insulin, 1982 4 GENOMICS IN MEDICINE • Influence of genomics on medicine: • knowledge of the genome can lead to understanding the origin of disease - mutant gene – misfunction of the protein • knowledge of the gene will allow screening for the disease • understanding gene influence on drug effects and origin of side effects • assessing individual genetic predisposition becomes possible • improving the effectiveness of new approaches to cure diseases • knowledge of bacterial and viral genomes helps easier identification of the mechanism of infection and improve prevention, treatment, as well as accelerate vaccine development • genome research is helping us better understand the aging process and improving the quality of life for older people 5 GENOMICS IN MEDICINE • The use of genomics in medicine: • genetic diagnosis • individualized medicine • gene therapy • regenerative medicine • xenotransplantation • molecular in vitro systems to study human diseases 6 GENETIC DIAGNOSIS • test performed to identify specific genetic traits of an individual • do not need to know the sequence of the entire genome • used for: • clarifying whether a disease appeared as a result of changes in specific genetic traits • confirming and refining standard medical diagnosis of an already apparent disease • identification of hereditary diseases before they become evident • pre-implantation screens of embryos fertilized in vitro • assessment of individual predispositions of a person for developing certain diseases later in life • DNA fingerprinting in criminal investigations and paternity disputes 7 PERSONALIZED MEDICINE The medical approach that emphasizes the systematic use of information specific to an individual patient to set up an optimized preventive and therapeutic plan for each patient. • Presumptions: • detailed knowledge of the genome of the patient, ideally the entire genome sequence • knowledge of the function of individual genes, including functional differences of individual alleles represented in the population • correlation of sequences with the prognosis of the disease, and with the success rate of therapeutic procedures ... 8 PERSONALIZED MEDICINE • uses knowledge of the genome for: • prediction of health risks • diagnosis • selection of the most appropriate type of treatment • minimizing the side effects of treatment • prevention http://www.personalizedmedicinecoalition.org/ 9 PERSONALIZED MEDICINE 10 PERSONALIZED MEDICINE 11 PERSONALIZED MEDICINE 12 PERSONALIZED MEDICINE • Problem: • multigene conditionality of most human diseases Goh et al., 2007 13 PERSONALIZED MEDICINE • Problem-solving: • systems biology - uses e.g. gene clustering to identify genes involved in the observed phenomenon Topotecan- resistant Topotecansensitive Dietel and Sers, 2006 14 PERSONALIZED MEDICINE • Problem-solving : • biomarkers • tests The Case for Personalized Medicine, 3rd edition 15 PERSONALIZED MEDICINE • Other problems • Ethical issues: • the condition is genetic testing or knowledge of the genome - easily abused • risk: insufficient data security • in some countries, employers or insurance companies do not have access to such data • High costs - risks: • medicine could be divided into first-class and low-class services • globalization gap could grow even larger - poor countries would not be able to afford this • Privacy: • crucial and complex issue • what information about oneself can/should be considered private? 16 GENE THERAPY Procedure in which the DNA sequence is inserted into the patient genome to replace or supplement an original gene • Options: • replace the mutated gene • repair the mutation • deliver DNA encoding a therapeutic protein • antisense therapy • In the future useful for treating e.g. hereditary diseases • Types: • somatic gene therapy • gene therapy of germ cells 17 GENE THERAPY • Methods: • viral vectors • retroviruses • adenoviruses • herpes simplex virus • vectors capable of replication • non-viral methods • injection of plasmid DNA into muscle • increased efficiency of DNA delivery: • electroporation • sonoporation • „gene gun" • magnetofection • hybrid methods 18 The aim is to recover diseased or injured organs or tissues. • to overcome the problems of transplantation: • shortage of donors • risk of rejection • severe immunosuppresive course • therapeutic cloning - cell therapy that uses stem cells to produce healthy copies of cells/tissues of the patient • potential medical applications: treatment of degenerative diseases such as Parkinson's disease, apoplexy, organ damage, diabetes, burns ... REGENERATIVE MEDICINE 19 REGENERATIVE MEDICINE • embryonic stem cells from animals have been successfully programmed to develop into neural, muscle and other cells - promising results Ethical issues: egg cell is used to form an embryo that does not develop into an organism. Instead, after 5-6 days of development, embryonic stem cells are extracted, which destroys the embryo. Depending on the definition of the point from which you must protect human life, it can be regarded as creating human life to destroy it, and is therefore prohibited. • consequence of ethical debate: increased support for research on adult stem cells 20 GENOMICS IN MEDICINE • The use of genomics in medicine: • genetic diagnosis • individualized medicine • gene therapy • regenerative medicine • xenotransplantation • molecular in vitro systems to study human diseases 21 BIOTECHNOLOGY 22 BIOTECHNOLOGY • It uses living organisms, cells or parts of cells (enzymes) for research, leading to new products and applications in medicine, agriculture, food, environmental protection • Also used in developing better/sustainable production methods for the chemical industry and other industrial processes. • An interdisciplinary approach requiring knowledge of chemistry, biology, physics, material sciences, engineering and informatics. • The origin of biotechnology can be traced back 4,000 years, when the Sumerians (although not knowingly) used microbes for the production of alcoholic beverages. 23 BIOTECHNOLOGY • modern biotechnology often changes the genetic status of cells and organisms to optimize processes, e.g. by chemical or physical treatment, cell fusion or genetic engineering. • genetic engineering modifies isolated nucleic acids • concepts of modern biotechnology and genetic engineering are often used as synonyms • genetic engineering is actually only one branch of the biotechnology industry • genetic engineering - GMO 24 BIOTECHNOLOGY • Examples: • effective utilization of plant biomass for fuel production • acquisition of starting material (monomers) for the production of polymers from living organisms instead of from fossil sources • phytopharmaceutics – using plants to produce new vaccination methods such as expression of antibodies, or antigens suitable for immunization • European Federation of Biotechnology http://www.efb-central.org25 PLANT BIOTECHNOLOGY Agriculture Fibres Organic Acids Polyols Polyurethanes Polyester Nylon Polymers Monomers Biorefining Feed Additives BioFuels Green Solvents (ethyl lactate) Specialty Chemicals High Performance Materials Pharmaceutical Precursors Amino Acids Lignin Ethanol The Biorefinery platform using Agricultural Feed Stocks Marc Van Montagu, ACPD, Praha 2009 26 PLANT BIOTECHNOLOGY • Examples: • production of spider filaments • production of degradable biopolymers • production of elastin-like polypeptides (component of animal tissues) • production of vaccines and antibodies: • human (HIV TNF) • animal (veterinary) • immunomodulation of plant hormones 27 FACTS TO THINK ABOUT • Our civilization is built on farming, the surface area needed for feeding people has decreased by 90% over 10,000 years • To prevent collapse, it is necessary to reduce this area from the current 0.45 ha/person to 0.2 ha/person by the year 2050 • Return to original methods of agriculture would be a return to the original demands on area and therefore would be unsustainable • Intensive farming = conversion of water and oil into food • The goal of plant biotechnology is to use all the available scientific knowledge to breed varieties with higher yield with lower inputs (of land, water, fertilizers, sprays ...) 0 0.05 0.1 0.15 0.2 0.25 0.3 0.35 0.4 0.45 1961 1997 2050 hectarespercapita Source: UN Millennium Ecosystem Assessment 28 GENETICALLY MODIFIED ORGANISMS 29 BREEDING • genetic basis of organisms naturally varies due to mutations • before the era of genetic engineering - question of chance • breeding tools: • selection and crossing • selection: • positive or negative • results were incidental • modern breeder learned to change hereditary information • chemicals, radiation ... Success is not always visible at a glance 30 GENETIC ENGINEERING • Discovery of: • DNA structure (1953) • restriction enzymes • plasmids • targeted change ("targeted breeding") • 1973 - bacteria produce frog protein • recombinant DNA technology = "gene cut" = genetic engineering • ability to transfer genes = transgenosis • according to the law: genetic modification • result: GMO • the first practical application: production of human insulin in bacteria - 1978 31 BREEDING vs. GENETIC ENGINEERING 32 GENETICALLY MODIFIED ORGANISMS (GMOs) • Organisms carrying modified genetic information – either own or foreign (from another organism), enabling targeted changes in the organism and its use for specific purposes • GMO: • plants • bacteria • animals http://www.gmo-compass.org/ 33 GMO PLANTS • Use: • resistance to pests • herbicide resistance • resistance to drought • resistance to cold • resistance to salinity • more efficient nitrogen utilization • increasing nutritional quality http://ipbo.vib-ugent.be/ 34 Bt PLANTS • resistance to insect pests • corn, cotton, rice • genes from Bacillus thuringiensis (Bt) • express delta-endotoxins (Cry proteins) • increasing yields, reducing the amount of chemical sprays 35 Ht PLANTS • resistance to systemic herbicides • glyphosate • interferes in the synthesis of aromatic amino acids; animals without the appropriate enzymatic apparatus = harmless • blocks the enzyme 5-enolpyrovylshikimate-3-phosphate synthase (EPSPS) in chloroplasts – affects green plants • ineffective for bacterial EPSPS - evolutionarily divergent • soya, maize, sugar beet, canola, cotton, alfalfa - added enzyme for tolerance • company Monsanto: Roundup • glufosinate (phosphinothricin) • prevents processing of ammonium - toxic • Streptomyces hygroscopicus synthesizes and transforms it: acetylation by the enzyme phosphinothricin acetyltransferase – coding gene isolated in 1987 - named bar • trade names: Basta, Liberty, Finale, Radical ... 36 MULTIRESISTENT PLANTS • Bt resistance + herbicide • multiresistant corn - the majority of total production in the USA • example of multiresistant corn: • three Bt genes for resistance to air pests • three Bt genes for resistance against soil pests • two genes for herbicide resistance 37 DISEASES TOLERANT PLANTS • viruses - no chemical agents available • gene encoding non-infectious viral envelope protein increases resistance to viral infection • banana; papaya - Hawaii, Southeast Asia • cassava - a basic food ingredient for more than 500 million people + animal feed 38 DROUGHT TOLERANT PLANTS • chickpeas - more resistant to drought, but toxic • corn resistant to drought: commercially utilizable in 2016 Lathyrus sativus Chickpea Cereals Drought in Ethiopia 39 NITROGEN USAGE INCREASE • use of nitrogen from fertilizers • gene from barley - 3x higher nitrogen utilization under oxygen deficiency 40 INCREASING NUTRITIONAL QUALITY • golden rice • several genes from maize encoding enzymes for the biosynthesis of βcarotene (precursor of vitamin A) • avoid problems with your eyes in a large part of the population in India and China • canola and soybean • improved oil properties: stable, resistant to high temperatures, long storage 41 GMO ANIMALS • Transgenic cats • lentiviruses are sensitive to restriction factors • specific restriction factor: rhesus macaque TRIMCyp + eGFP • uniform expression, no mosaicity and no silencing in F1 generation • lymphocytes of transgenic animals resistant to replication of FIV Wongsrikeao et al., 2011, Nature Methods 42 GMO – PROS AND CONS 43 GMO – PROS AND CONS • GMO plants • increased yields without insecticides, pesticides, fungicides • 24% yield gain per acre • 50% yield improvement for small farmers Ohio State University, December 2011 44 GMO – PROS AND CONS • GMO plants • reduce pesticide use - the negative impact of strong chemicals currently used in agriculture • increase nitrogen utilization from mineral fertilizers - the negative impact of excessive fertilizer use on water quality • increase the nutritional quality of crops • reduce cultivation area (increased yield per unit area) • pollen transfer to other species (hybridization) was detected in frequencies between 0.05 to 0.53% 45 GMO – PROS AND CONS • GMO plants • Bt crops: negative impact on other than pest insects ??? • affect some butterflies and beetles • more insects on Bt cotton Lu et al., 2012, Nature berušky pavouci zlatoočka 46 GMO – PROS AND CONS • GMO plants • risk of allergies • GMO plants always contain only one or a few transgenes and are very thoroughly tested • on the other hand, new varieties created using strong mutagens, e.g. X-rays are far less tested. With these methods tens or hundreds of new mutations occur at once • ban on GMO cultivation in Switzerland was decided by referendum • 30% of the EU population believes that the only transgenic plants have genes and refuse to eat them ... 47 GMO – PROS AND CONS Our task: • tirelessly to explain the rational use of scientific knowledge, including genomics, it is necessary both for advances in medicine and for the preservation of our civilization • discussions with the general public about the importance and benefits of GMOs for human society • need to defend GMO plants – crops for 21st century • no technology is without risk, including GMOs, but there is not cause to demonize GMOs and target them - financial benefit of some companies ... 48 LITERATURE • see lecture 49 GMO – PROS AND CONS Discussion 50