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EPIGENETICS(i) Cell Biology(ii) New Platform in Drug DiscoveryDr. Ana M Roa
PlasmamembraneNucleusCytoplasmCell: functional units of all livingorganisms
Cell Membrane
CytoplasmCytoskeletonKeep cellular structureCellular divisionMigration & AdhesionSubcelular OrganulesMultiple FunctionsSub...
Chomosomes: DNA & ProteinsNucleusGene Expression
What is Epigenetics?Epigenetics is the study of heritable changes in gene expression orcellular phenotype, caused by mecha...
Epigenetics: The life cycle of the epigenomeA number of mechanisms controllingchromatin remodelling include DNAmethylation...
Epigenetics and EnvironmentCertain environmental and dietary factors have been linked to abnormalchanges in epigenetic pat...
Food rich on methyl donorsNormal diet
Epigenetics in Microorganisms• The filamentous fungus Neurospora crassa is amodel system for understanding the control and...
Epigenetics in Tuberculosis2 billion people worldwide are infected with tuberculosis, 10 million of whomfall ill each year...
Epigenetics in MalariaMalaria is a major public health problem in many developing countries, withthe malignant parasite Pl...
Epigenetics in Plants
Epigenetic Targets: Histone modificationHistonesKDMsKDMsChromosome 1 in 3-years-oldidentical twins (left) and in 50-years-...
Epigenetic alterations associated with diseaseVol 28 Nº10 OCTOBER 2010 nature biotechnology
Epigenetic alterations associated with diseaseVol 28 Nº10 OCTOBER 2010 nature biotechnology
Epigenetic Inhibitors for Therapies
Therapeutic areas targeting epigenetic enzymesTherapeutic area % R&DOncology 71Metabolic disease/Diabetes 38CNS/Neurodegen...
Chemical structures of selected compounds thattarget epigenetic modifications
Looking for new active molecules in Drug DiscoveryDeveloping robust and uHTS assays for searching small-molecule inhibitor...
Epi-Enzymes Jumonji family (JmjD) of KDMs Able to demethylate 3, 2 &1 methyl groups Largest family of HDMs Uniquely req...
JmjD family HTS steps1. Assay development of a Biochemical Assay2. Testing of pharmacology3. Testing the Robustness of the...
Bromodomains recognise acetylatedlysine residues within histone tails45 human bromodomainsTargets proteins identified as B...
BRDs control gene transcription
Challenges with targeting epigenetic readers
• BET Bromodomain Inhibition as a Therapeutic Strategy to Targetc-MycCell 01 September 2011• RNAi screen identifies Brd4 a...
Future trends…..Nuclear magnetic resonance (NMR)Isothermal Titration Calorimetry (ITC)Dynamic light scatteringSurface plas...
Many thanks foryour attention !
Una revisión de los conocimientos fundamentales de la biología de la célula. La epigenética
Una revisión de los conocimientos fundamentales de la biología de la célula. La epigenética
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Una revisión de los conocimientos fundamentales de la biología de la célula. La epigenética

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Conferencia de la Dra. Ana María Roa, Bióloga Molecular, sobre Epigenética, impartida en la Universidad Popular Carmen de Michelena de Tres Cantos el 1 de marzo de 2013.
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http://www.universidadpopularc3c.es/index.php/actividades/conferencias/details/448-conferencia-una-revision-de-los-conocimientos-fundamentales-de-la-biologia-de-la-celula-la-epigenetica

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  • The emerging field of epigenetics, a new field of science that describes how the selective regulation of chromatin state controls key cellular activities such as gene expression, cell fate decisions, and DNA damage repair. Miscues in these normal events contribute to the development of human diseases such as cancer, inflammation, autoimmune diseases and neurodegenerative diseases. Drugs that reverse these miscues should provide high-impact medical benefit. Constellation Pharmaceuticals is building the premier chromatin therapeutics company through the discovery and development of drugs targeting epigenetic function and associated disease biology.
  • The cytoplasm of a cell is surrounded by a cell membrane or plasma membrane . The plasma membrane in plants and prokaryotes is usually covered by a cell wall. This membrane serves to separate and protect a cell from its surrounding environment and is made mostly from a double layer of lipids (hydrophobic fat-like molecules) and hydrophilic phosphorus molecules. Hence, the layer is called a phospholipid bilayer, or sometimes a fluid mosaic membrane. Embedded within this membrane is a variety of protein molecules that act as channels and pumps that move different molecules into and out of the cell. The membrane is said to be 'semi-permeable', in that it can either let a substance (molecule or ion) pass through freely, pass through to a limited extent or not pass through at all. Cell surface membranes also contain receptor proteins that allow cells to detect external signaling molecules such as hormones.
  • El citoplama contiene estructuras subcelulares y líquido citoplasmático Citoesqueleto: Mantiene la estructura celular division migración y adhesion Mitocondrias Retículo endoplásmic, etc Liquido citoplasmático Kinasa, fostatasas,….
  • El nucleo: transmision de información genética durante la división celular Procesos de division celular anómalos pueden dan lugar a crecimientos celulares “descontrolados” (tumores) Cromosomas: ADN y proteinas. Código genético y sitios de unión específicos para factores reguladores de replicación y transcripción
  • The DNA strand in each cell is wound around a protein complex called the histone to compact the DNA into the cell nucleus (chromatin) . Changes in chromatin structure due to either chemical modification of the DNA (methylation) or changes in DNA-histone interactions help control gene expression, silencing or activating genes. Improper gene expression leads to the improper production or non-production of proteins which are the source of most diseases and disorders. Various chemical modifications to the histones cause the DNA to wrap more tightly or loosely around the histones. These chemical modifications are directly responsible for making the DNA accessible or inaccessible to control gene regulation “histone post translational modifications. The post translational modifications which usually take place on the “tails“of histones include methylation, acetylation, phosphorylation and ubiquitination. An increasing number of these newly identified enzymes have been associated with neurodegenerative disorders, metabolic diseases, inflammation, and most notably, cancer. “ The difference between genetics and epigenetics can probably be compared to the difference between writing and reading a book. Once a book is written, the text (the genes or DNA) will be the same in all the copies. However, each individual reader may interpret the story slightly differently. Conrad Waddington (1905-1975) is often credited with coining the term epigenetics in 1942 as “the branch of biology which studies the causal interactions between genes and their products, which bring the phenotype into being”. Epigenetics appears in the literature as far back as the mid 19th century, although the conceptual origins date back to Aristotle (384-322 BC). He believed in epigenesis: the development of individual organic form from the unformed. This controversial view was the main argument against our having developed from miniscule fully-formed bodies. Even today the extent to which we are preprogrammed versus environmentally shaped awaits universal consensus. The field of epigenetics has emerged to bridge the gap between nature and nurture. In the 21st century you will most commonly find epigenetics defined as ‘the study of heritable changes in genome function that occur without a change in DNA sequence‘. But what do the scientists that work in this rapidly expanding research field have to say?
  • The study of the epigenetic mechanism in malaria parasites is still in its infancy, and most knowledge gathered thus far is from focused studies of antigenic variation in P. falciparum. The available information clearly indicates that chromatin-mediated mechanisms underlie many cellular processes in the parasite's development. From an evolutionary point of view, the malaria parasites have a typical nucleosome organization, numerous histone PTMs, and a large catalogue of conserved chromatin modification and remodeling machineries and histone-binding modules, suggesting conserved epigenetic mechanisms in these early-branching protozoan parasites. Therefore, in-depth studies of the epigenome of the malaria parasites will contribute to a better comprehension of how chromatin-mediated mechanisms have evolved. In addition, the malaria parasite also possesses distinct histone PTMs and divergent histone variants, and their combination is expected to generate a complex yet different “histone code.” Besides, among the “writers” and “readers” of the “histone code” in this parasite, many are unique or contain unique domains, but only a countable few have been characterized so far. Thus, elucidation of the epigenetic pathways in malaria parasites not only will help us to understand gene regulation in this unicellular parasite but also will provide insights into many parasite-specific phenomena such as antigenic variation and alternative invasion pathways, which may ultimately lead to the development of novel control measures targeting host-parasite interactions. More importantly, the distinction of the parasite's chromatin modification machinery from those of its mammalian hosts makes it a promising target for antimalarial chemotherapy. For example, HDACs have been explored as potential candidates for antimalarials, and many HDAC inhibitors have potent antimalarial activity (1). With the identification of most of the PTMs, modifiers, and “readers,” future studies will need to characterize and integrate these components and to advance toward a mechanistic understanding of chromatin-mediated gene regulation in the malaria parasite.
  • Epigenetics: the bookmark in the book of life Epigenetics: inheritable changes on gene expression, and hence the proteome and the phenotype, that are not based upon changes in the DNA sequence Redundacy and divergence in enzymes; selectivity: substrate, differential expression Epigenetics allow different interpretations of the fixed genetic code and result in different read-outs, dependent upon the variable conditions” Thomas Jenuwein (Vienna,Austria)
  • In the picture below, certain key functions are selected of the lead identification and lead optimization process. Clinical and development phases are neglected because MDL does not provide solutions in these areas.  The pre-clinical area is important for the near future, because here cheminformatics, genomics, combinatorial chemistry and high throughput screening can make drastic improvements. Such improvements are needed to propel the industry to an seemingly impossible three to eightfold increase in output of new drugs. ( ...some companies are heading for a crash unless they can rethink their approach so completely that R&D costs and lead times plummet, generate additional sales revenues with blockbuster drugs or move into brand new markets.... there could be as few as 13 top companies by the year 2005. Sir Mark Richmond, Glaxo's former Head of Research Worldwide, goes further. "We could see just three...." taken from "Pharma 2005, An Industrial Revolution in R&D from PriceWaterhouseCoopers.) The workflow of the development of a new pharmaceutical entity can be pictured by a huge funnel, which allows the easy input of information that is then compressed to be used to produce a new drug. MDL concentrates on the pre-clinical phase. There you will deal with many chemical structures and any workflow and information system must  be based on handling chemical structures. Handling the chemical structure, also the chemical reaction, was pioneered by MDL
  • Cartoon of the reaction and biological activity, and the assay principle
  • Approx. 110 residue domain; 4 helices, 2 key loops Bromodomain External Terminal
  • Transcript of "Una revisión de los conocimientos fundamentales de la biología de la célula. La epigenética"

    1. 1. EPIGENETICS(i) Cell Biology(ii) New Platform in Drug DiscoveryDr. Ana M Roa
    2. 2. PlasmamembraneNucleusCytoplasmCell: functional units of all livingorganisms
    3. 3. Cell Membrane
    4. 4. CytoplasmCytoskeletonKeep cellular structureCellular divisionMigration & AdhesionSubcelular OrganulesMultiple FunctionsSubcellular organulesMultiple functions(inflammation,cancer)(metabolic disorders,cancer)
    5. 5. Chomosomes: DNA & ProteinsNucleusGene Expression
    6. 6. What is Epigenetics?Epigenetics is the study of heritable changes in gene expression orcellular phenotype, caused by mechanisms other than changes in theunderlying DNA sequence– epi- (Greek: επί- over, above, outer) -genetics.It refers to functionally relevant modifications to the genome that donot involve a change in the nucleotide sequence.– DNA methylation and histone modification, both of which serve toregulate gene expression without altering the underlyingDNA sequence.– These changes may remain through cell divisions for theremainder of the cells life and may also last for multiplegenerations.“The study of the mechanisms of temporal and spatial control of geneactivity during the development of complex organisms.“
    7. 7. Epigenetics: The life cycle of the epigenomeA number of mechanisms controllingchromatin remodelling include DNAmethylation & histone modificationDysregulation of any of them isimplicated in human disease
    8. 8. Epigenetics and EnvironmentCertain environmental and dietary factors have been linked to abnormalchanges in epigenetic pathways in experimental and epidemiological studies.
    9. 9. Food rich on methyl donorsNormal diet
    10. 10. Epigenetics in Microorganisms• The filamentous fungus Neurospora crassa is amodel system for understanding the control andfunction of DNA methylation. In this organisms, DNAmethylation is associated with a genome defensesystem called RIP (repeat-induced point mutation) andsilences gene expression by inhibiting transcriptionelongation.[75]• Bacteria make use of DNA adenine methylation as anepigenetic signal. DNA adenine methylation is important inbacteria virulence in organisms such as Escherichia coli,Salmonella, Vibrio, Yersinia, Haemophilus, and Brucella.
    11. 11. Epigenetics in Tuberculosis2 billion people worldwide are infected with tuberculosis, 10 million of whomfall ill each year. Tuberculosis is the leading cause of death among people co-infected with HIV, the virus that causes AIDS, leading to some half-milliondeaths annually among those co-infected.Genetic and Epigenetic Variation in Mycobacterium tuberculosisdetermine whether M. tuberculosis varies, genetically and/or epigenetically,during the course of single infections and whether this variation is subject toselection by the host immune response.Hypermutability and the Acquisition of Multidrug Resistanceaccount for the emergence of extended drug resistance insome clinical strains.. Current studies on Functional genomics of M.tuberculosis.
    12. 12. Epigenetics in MalariaMalaria is a major public health problem in many developing countries, withthe malignant parasite Plasmodium falciparum causing the most malaria-associated mortality. The chromatin-mediated mechanisms underlie manycellular processes in the parasites developmentThe distinction of the parasites chromatin modification machinery from those ofits mammalian hosts makes it a promising target for antimalarial chemotherapy. HDACs have been explored as potential candidates for antimalarials. With the identification of most of the PTMs, modifiers, and “readers,”future studies will allow to advance toward a mechanistic understanding ofchromatin-mediated gene regulation in the malaria parasite.Elucidation of the epigenetic pathways in malaria parasites notonly will help us to understand gene regulation in this unicellularparasite but also will provide insights into many parasite-specificphenomena such as antigenic variation and alternativeinvasion pathways, which may ultimately lead to thedevelopment of novel control measures targeting host-parasite interactions.
    13. 13. Epigenetics in Plants
    14. 14. Epigenetic Targets: Histone modificationHistonesKDMsKDMsChromosome 1 in 3-years-oldidentical twins (left) and in 50-years-old (right)P.N.A.S.Divergence with timeWriters, Erasers and Readers in the Book of LifeChromodomainsChromodomainsReadersReadersReadersReadersBromodomainsBromodomainsHDACsHDACsSirtuinsSirtuins
    15. 15. Epigenetic alterations associated with diseaseVol 28 Nº10 OCTOBER 2010 nature biotechnology
    16. 16. Epigenetic alterations associated with diseaseVol 28 Nº10 OCTOBER 2010 nature biotechnology
    17. 17. Epigenetic Inhibitors for Therapies
    18. 18. Therapeutic areas targeting epigenetic enzymesTherapeutic area % R&DOncology 71Metabolic disease/Diabetes 38CNS/Neurodegeneration/Pain 29Inflammatory disease/Autoimmune 29Cardiovascular 28Anti-infectives/Anti-viral 16Others… 15
    19. 19. Chemical structures of selected compounds thattarget epigenetic modifications
    20. 20. Looking for new active molecules in Drug DiscoveryDeveloping robust and uHTS assays for searching small-molecule inhibitors targeting key epigenetic proteins iscrucial in the discovery process of clinically relevantcompounds
    21. 21. Epi-Enzymes Jumonji family (JmjD) of KDMs Able to demethylate 3, 2 &1 methyl groups Largest family of HDMs Uniquely require α-ketoglutarate and Fe(II) At least 30 JmjDs in the human genomewith unique target specificities.Selective inhibition of JmjD may be sufficient formodulation of some diseasesJmjD1aJmjD1bJmjD2aJmjD2bJmjD2cJmjD2d-JmjD3-Gene RepressionHuman H3 N-ARTKQTARKSTGGKAPRKQLAKAARKSAPATGGVKKPHR….4 9 27Background
    22. 22. JmjD family HTS steps1. Assay development of a Biochemical Assay2. Testing of pharmacology3. Testing the Robustness of the assay in the presence ofcompounds4. Determination of the Quality Control parameters and thestatistical expected for the primary screening campaign5. If all OK: Launch the HTS campaign (2M compounds)6. Data analysis with specific software7. Hits selection using chemo-physical properties8. Hits confirmation9. Determination of the potency and selectivity of theconfirmed actives10. Progress the best compounds to the Critical Pathdesigned to determine real potential drugs
    23. 23. Bromodomains recognise acetylatedlysine residues within histone tails45 human bromodomainsTargets proteins identified as BETfamily:– Brd-2, -3, -4Epi-Readers BET family bromodomainsBromodomainsBackgroundDiversity in loop region(selectivity)High species homology at AcKJBC,2007
    24. 24. BRDs control gene transcription
    25. 25. Challenges with targeting epigenetic readers
    26. 26. • BET Bromodomain Inhibition as a Therapeutic Strategy to Targetc-MycCell 01 September 2011• RNAi screen identifies Brd4 as a therapeutic target in acutemyeloid leukemiaNature (2011) 1034• Selective inhibition of BET bromodomainsNature 468, 1067–1073 (23 Dec 2010)
    27. 27. Future trends…..Nuclear magnetic resonance (NMR)Isothermal Titration Calorimetry (ITC)Dynamic light scatteringSurface plasmon resonance (SPR)Dual polarisation interferometryMicroscale thermophoresis (MST)To assess whether (i) the compound binds effectively to thetarget, (ii) the stoïchiometry of binding, (iii) any associatedconformational change and (iv) to identify promiscuousinhibitors.New Technologies for Biophysical testing
    28. 28. Many thanks foryour attention !
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