1 3744 123 INSIGHTS INTO THE CRYSTAL STRUCTURE OF BRD2-BD2 - PHENANTHRIDINONE COMPLEX AND THEORETICAL STUDIES ON PHENANTHRIDINONE ANALOGS. BROMODOMAIN AND EXTRA-TERMINAL FAMILY PROTEINS RECOGNIZE THE ACETYLATED HISTONE CODE ON CHROMATIN AND PARTICIPATE IN DOWNSTREAM PROCESSES LIKE DNA REPLICATION, MODIFICATION, AND REPAIR. AS PART OF EPIGENETIC APPROACHES, BRD2 AND BRD4 WERE IDENTIFIED AS PUTATIVE TARGETS, FOR THE MANAGEMENT OF CHRONIC DISEASES. WE HAVE RECENTLY REPORTED THE DISCOVERY OF A NEW SCAFFOLD OF THE PHENANTHRIDINONE-BASED INHIBITOR (L10) OF THE SECOND BROMODOMAIN OF BRD2 (BRD2-BD2). HERE, WE PRESENT THE CRYSTAL STRUCTURE OF THE BRD2-BD2, REFINED TO 1.4 A RESOLUTION, IN COMPLEX WITH BETA-MERCAPTOETHANOL (A COMPONENT OF THE PROTEIN BUFFER). THE BETA-MERCAPTOETHANOL COVALENTLY LINKS TO C425 OF BD2 IN THE ACETYL-LYSINE BINDING POCKET, TO FORM A MODIFIED CYSTEINE MERCAPTOETHANOL (CME). THE CME MODIFICATION SIGNIFICANTLY HINDERS THE ENTRY OF LIGANDS INTO THE BD2 BINDING POCKET, SUGGESTING THAT BETA-MERCAPTOETHANOL SHOULD BE REMOVED DURING PROTEIN PRODUCTION PROCESS. NEXT, TO CONFIRM WHETHER PHENANTHRIDIONONE SCAFFOLD IS A NEW INHIBITOR FAMILY OF BRD2-BD2, WE HAVE DETERMINED THE CRYSTAL STRUCTURE OF BD2 IN COMPLEX WITH 6(5H)-PHENANTHRIDINONE (A CORE MOIETY OF L10), REFINED TO 1.28 A RESOLUTION. IT CONFIRMED THAT THE PHENANTHRIDINONE MOLECULE, UNAMBIGUOUSLY, BINDS TO BD2. MOREOVER, WE PERFORMED MOLECULAR DOCKING AND MOLECULAR DYNAMIC STUDIES ON SELECTED PHENANTHRIDINONE ANALOGS. THE PREDICTED L10 ANALOGS ARE STABLE WITH ESSENTIAL HYDROPHOBIC AND HYDROPHILIC INTERACTIONS WITH BD2 DURING MOLECULAR DYNAMIC SIMULATIONS. WE PROPOSE THAT THE PREDICTED PHENANTHRIDINONE ANALOGS MAY BE POTENTIAL MOLECULES FOR INHIBITING THE BD2 FUNCTION OF ACETYLATED HISTONE RECOGNITION. 2018 2 3492 34 IDENTIFICATION OF HISTONE DEACETYLASE 10 (HDAC10) INHIBITORS THAT MODULATE AUTOPHAGY IN TRANSFORMED CELLS. HISTONE DEACETYLASES (HDACS) ARE A FAMILY OF 18 EPIGENETIC MODIFIERS THAT FALL INTO 4 CLASSES. HISTONE DEACETYLASE INHIBITORS (HDACI) ARE VALID TOOLS TO ASSESS HDAC FUNCTIONS. HDAC6 AND HDAC10 BELONG TO THE CLASS IIB SUBGROUP OF THE HDAC FAMILY. THE TARGETS AND BIOLOGICAL FUNCTIONS OF HDAC10 ARE ILL-DEFINED. THIS LACK OF KNOWLEDGE IS DUE TO A LACK OF SPECIFIC AND POTENT HDAC10 INHIBITORS WITH CELLULAR ACTIVITY. HERE, WE HAVE SYNTHESIZED AND CHARACTERIZED PIPERIDINE-4-ACRYLHYDROXAMATES AS POTENT AND HIGHLY SELECTIVE INHIBITORS OF HDAC10. THIS WAS ACHIEVED BY TARGETING THE ACIDIC GATEKEEPER RESIDUE GLU274 OF HDAC10 WITH A BASIC PIPERIDINE MOIETY THAT MIMICS THE INTERACTION OF THE POLYAMINE SUBSTRATE OF HDAC10. WE HAVE CONFIRMED THE BINDING MODES OF SELECTED INHIBITORS USING X-RAY CRYSTALLOGRAPHY. PROMISING CANDIDATES WERE SELECTED BASED ON THEIR SPECIFICITY BY IN VITRO PROFILING USING RECOMBINANT HDACS. THE MOST PROMISING HDAC10 INHIBITORS 10C AND 13B WERE TESTED FOR SPECIFICITY IN ACUTE MYELOID LEUKEMIA (AML) CELLS WITH THE FLT3-ITD ONCOGENE. BY IMMUNOBLOT EXPERIMENTS WE ASSESSED THE HYPERACETYLATION OF HISTONES AND TUBULIN-ALPHA, WHICH ARE CLASS I AND HDAC6 SUBSTRATES, RESPECTIVELY. AS VALIDATED TEST FOR HDAC10 INHIBITION WE USED FLOW CYTOMETRY ASSESSING AUTOLYSOSOME FORMATION IN NEUROBLASTOMA AND AML CELLS. WE DEMONSTRATE THAT 10C AND 13B INHIBIT HDAC10 WITH HIGH SPECIFICITY OVER HDAC6 AND WITH NO SIGNIFICANT IMPACT ON CLASS I HDACS. THE ACCUMULATION OF AUTOLYSOSOMES IS NOT A CONSEQUENCE OF APOPTOSIS AND 10C AND 13B ARE NOT TOXIC FOR NORMAL HUMAN KIDNEY CELLS. THESE DATA SHOW THAT 10C AND 13B ARE NANOMOLAR INHIBITORS OF HDAC10 WITH HIGH SPECIFICITY. THUS, OUR NEW HDAC10 INHIBITORS ARE TOOLS TO IDENTIFY THE DOWNSTREAM TARGETS AND FUNCTIONS OF HDAC10 IN CELLS. 2022 3 5122 35 POSSIBLE ROLE OF FRUCTOSAMINE 3-KINASE GENOTYPING FOR THE MANAGEMENT OF DIABETIC PATIENTS. DIABETES MELLITUS IS A GLOBAL PANDEMIC AND CONTINUES TO INCREASE IN NUMBERS AND SIGNIFICANCE. SEVERAL PATHOGENIC PROCESSES ARE INVOLVED IN THE DEVELOPMENT OF SUCH DISEASE AND THESE MECHANISMS COULD BE INFLUENCED BY GENETIC, EPIGENETIC AND ENVIRONMENTAL FACTORS. NON-ENZYMATIC GLYCATION REACTIONS OF PROTEINS HAVE BEEN STRONGLY RELATED TO PATHOGENESIS OF CHRONIC DIABETIC COMPLICATIONS. THE IDENTIFICATION OF FRUCTOSAMINE 3-KINASE (FN3K), AN ENZYME INVOLVED IN PROTEIN DEGLYCATION, A NEW FORM OF PROTEIN REPAIR, IS OF GREAT INTEREST. FN3K PHOSPHORYLATES FRUCTOSAMINES ON THE THIRD CARBON OF THEIR SUGAR MOIETY, MAKING THEM UNSTABLE AND CAUSING THEM TO DETACH FROM PROTEINS, SUGGESTING A PROTECTIVE ROLE OF THIS ENZYME. MOREOVER, THE VARIABILITY IN FN3K ACTIVITY HAS BEEN ASSOCIATED WITH SOME POLYMORPHISMS IN THE FN3K GENE. HERE WE ARGUE ABOUT GENETIC STUDIES AND EVIDENCE OF FN3K INVOLVEMENT IN DIABETES, TOGETHER WITH RESULTS OF OUR ANALYSIS OF THE FN3K GENE ON A CAUCASIAN COHORT OF DIABETIC PATIENTS. PRESENT KNOWLEDGE SUGGESTS THAT FN3K COULD ACT IN CONCERT WITH OTHER MOLECULAR MECHANISMS AND MAY IMPACT ON GENE EXPRESSION AND ACTIVITY OF OTHER ENZYMES INVOLVED IN DEGLYCATION PROCESS. 2015 4 1338 32 DESIGN, SYNTHESIS AND BIOLOGICAL EVALUATION OF A PHENYL BUTYRIC ACID DERIVATIVE, N-(4-CHLOROPHENYL)-4-PHENYLBUTANAMIDE: A HDAC6 INHIBITOR WITH ANTI-PROLIFERATIVE ACTIVITY ON CERVIX CANCER AND LEUKEMIA CELLS. BACKGROUND: THE EPIGENETIC REGULATION OF GENES IN CANCER COULD BE TARGETED BY INHIBITING HISTONE DEACETYLASE 6 (HDAC6), AN ENZYME INVOLVED IN SEVERAL TYPES OF CANCER SUCH AS LYMPHOMA, LEUKEMIA, OVARIAN CANCER, ETC. OBJECTIVE: THROUGH IN SILICO METHODS, A SET OF PHENYL BUTYRIC ACID DERIVATIVES WITH POSSIBLE HDAC6 INHIBITORY ACTIVITY WERE DESIGNED, RENDERING MONOPHENYLAMIDES AND BIPHENYLAMIDES USING TUBACIN (HDAC6 SELECTIVE INHIBITOR) AS REFERENCE. METHOD: THE TARGET COMPOUNDS WERE SUBMITTED TO THEORETICAL ADMET ANALYSES AND THEIR BINDING PROPERTIES ON DIFFERENT HDAC6 CONFORMERS WERE EVALUATED THROUGH DOCKING CALCULATIONS. RESULTS: THESE IN SILICO STUDIES ALLOWED US TO IDENTIFY A COMPOUND NAMED B-R2B. IN ORDER TO HAVE MORE INFORMATION ABOUT THE B-R2B BINDING RECOGNITION PROPERTIES ON HDAC6, THE B-R2B-HDAC6 COMPLEX WAS SUBMITTED THROUGH 100 NS-LONG MOLECULAR DYNAMICS (MD) SIMULATION COUPLED TO MMGBSA APPROACH, REVEALING THAT B-R2B IS LOCATED AT THE ENTRANCE OF HDAC6 ACTIVE POCKET, BLOCKING THE PASSAGE OF THE SUBSTRATE WITHOUT REACHING THE HDAC6 BINDING SITE. BASED ON THESE RESULTS, B-R2B WAS SYNTHESIZED, CHARACTERIZED AND BIOLOGICALLY TESTED. THE HDAC6 FLUOROMETRIC DRUG DISCOVERY KIT FLUOR-DE-LYS (ENZO LIFE SCIENCES INC.) WAS USED TO DETERMINE THE HDAC6 HUMAN INHIBITORY ACTIVITY (IC50 VALUE) OF B-R2B COMPOUND. IN ADDITION, B-R2B SHOW IC50 VALUES ON CANCER CELL LINES (HELA; IC50 = 72.6 MICROM), ACUTE MYELOID LEUKEMIA (THP-1; IC50 = 16.5 MICROM), HUMAN MAST LEUKEMIA (HMC; IC50 = 79.29 MICROM) AND CHRONIC MYELOGENOUS LEUKEMIA (KASUMI; IC50 = 101 MICROM). CONCLUSION: THESE RESULTS SHOW THAT B-R2B IS A HDAC6 INHIBITOR, SPECIFICALLY A NON-COMPETITIVE TYPE IN A SIMILAR WAY THAT TUBACIN DOES, ACCORDING TO MD SIMULATIONS. 2017 5 5947 28 TARGETING THE EPIGENOME: SCREENING BIOACTIVE COMPOUNDS THAT REGULATE HISTONE DEACETYLASE ACTIVITY. SCOPE: NUTRIGENOMICS IS A RAPIDLY EXPANDING FIELD THAT ELUCIDATES THE LINK BETWEEN DIET-GENOME INTERACTIONS. RECENT EVIDENCE DEMONSTRATES THAT REGULATION OF THE EPIGENOME, AND IN PARTICULAR INHIBITION OF HISTONE DEACETYLASES (HDACS), IMPACT PATHOGENETIC MECHANISMS INVOLVED IN CHRONIC DISEASE. FEW STUDIES, TO DATE, HAVE SCREENED LIBRARIES OF BIOACTIVE COMPOUNDS THAT ACT AS EPIGENETIC MODIFIERS. THIS STUDY SCREENED A LIBRARY OF 131 NATURAL COMPOUNDS TO DETERMINE BIOACTIVE COMPOUNDS THAT INHIBIT ZN-DEPENDENT HDAC ACTIVITY. METHODS AND RESULTS: USING CLASS-SPECIFIC HDAC SUBSTRATES, WE SCREENED 131 NATURAL COMPOUNDS FOR HDAC ACTIVITY IN BOVINE CARDIAC TISSUE. FROM THIS SCREEN, WE IDENTIFIED 18 BIOACTIVE COMPOUND HDAC INHIBITORS. USING OUR CLASS-SPECIFIC HDAC SUBSTRATES, WE NEXT SCREENED THESE 18 BIOACTIVE COMPOUNDS AGAINST RECOMBINANT HDAC PROTEINS. CONSISTENT WITH INHIBITION OF HDAC ACTIVITY, THESE COMPOUNDS WERE CAPABLE OF INHIBITING ACTIVITY OF INDIVIDUAL HDAC ISOFORMS. LASTLY, WE REPORT THAT TREATMENT OF H9C2 CARDIAC MYOBLASTS WITH BIOACTIVE HDAC INHIBITORS WAS SUFFICIENT TO INCREASE LYSINE ACETYLATION AS ASSESSED VIA IMMUNOBLOT. CONCLUSION: THIS STUDY PROVIDED THE FIRST STEP IN IDENTIFYING MULTIPLE BIOACTIVE COMPOUND HDAC INHIBITORS. TAKEN TOGETHER, THIS REPORT SETS THE STAGE FOR FUTURE EXPLORATION OF THESE BIOACTIVE COMPOUNDS AS EPIGENETIC REGULATORS TO POTENTIALLY AMELIORATE CHRONIC DISEASE. 2017 6 698 35 BROMODOMAIN CONTAINING PROTEIN 4 (BRD4) REGULATES EXPRESSION OF ITS INTERACTING COACTIVATORS IN THE INNATE RESPONSE TO RESPIRATORY SYNCYTIAL VIRUS. BROMODOMAIN-CONTAINING PROTEIN 4 PLAYS A CENTRAL ROLE IN COORDINATING THE COMPLEX EPIGENETIC COMPONENT OF THE INNATE IMMUNE RESPONSE. PREVIOUS STUDIES IMPLICATED BRD4 AS A COMPONENT OF A CHROMATIN-MODIFYING COMPLEX THAT IS DYNAMICALLY RECRUITED TO A NETWORK OF PROTECTIVE CYTOKINES BY BINDING ACTIVATED TRANSCRIPTION FACTORS, POLYMERASES, AND HISTONES TO TRIGGER THEIR RAPID EXPRESSION VIA TRANSCRIPTIONAL ELONGATION. OUR PREVIOUS STUDY EXTENDED OUR UNDERSTANDING OF THE AIRWAY EPITHELIAL BRD4 INTERACTOME BY IDENTIFYING OVER 100 FUNCTIONALLY IMPORTANT COACTIVATORS AND TRANSCRIPTION FACTORS, WHOSE ASSOCIATION IS INDUCED BY RESPIRATORY SYNCYTIAL VIRUS (RSV) INFECTION. RSV IS AN ETIOLOGICAL AGENT OF RECURRENT RESPIRATORY TRACT INFECTIONS ASSOCIATED WITH EXACERBATIONS OF CHRONIC OBSTRUCTIVE PULMONARY DISEASE. USING A HIGHLY SELECTIVE SMALL-MOLECULE BRD4 INHIBITOR (ZL0454) DEVELOPED BY US, WE EXTEND THESE FINDINGS TO IDENTIFY THE GENE REGULATORY NETWORK DEPENDENT ON BRD4 BROMODOMAIN (BD) INTERACTIONS. HUMAN SMALL AIRWAY EPITHELIAL CELLS WERE INFECTED IN THE ABSENCE OR PRESENCE OF ZL0454, AND GENE EXPRESSION PROFILING WAS PERFORMED. A HIGHLY REPRODUCIBLE DATASET WAS OBTAINED WHICH INDICATED THAT BRD4 MEDIATES BOTH ACTIVATION AND REPRESSION OF RSV-INDUCIBLE GENE REGULATORY NETWORKS CONTROLLING CYTOKINE EXPRESSION, INTERFERON (IFN) PRODUCTION, AND EXTRACELLULAR MATRIX REMODELING. INDEX GENES OF FUNCTIONALLY SIGNIFICANT CLUSTERS WERE VALIDATED INDEPENDENTLY. WE DISCOVER THAT BRD4 REGULATES THE EXPRESSION OF ITS OWN GENE DURING THE INNATE IMMUNE RESPONSE. INTERESTINGLY, BRD4 ACTIVATES THE EXPRESSION OF NFKAPPAB/RELA, A COACTIVATOR THAT BINDS TO BRD4 IN A BD-DEPENDENT MANNER. WE EXTEND THIS FINDING TO SHOW THAT BRD4 ALSO REGULATES OTHER COMPONENTS OF ITS FUNCTIONAL INTERACTOME, INCLUDING THE MEDIATOR (MED) COACTIVATOR COMPLEX AND THE SWI/SNF-RELATED, MATRIX-ASSOCIATED, ACTIN-DEPENDENT REGULATOR OF CHROMATIN (SMARC) SUBUNITS. TO PROVIDE FURTHER INSIGHT INTO MECHANISMS FOR BRD4 IN RSV EXPRESSION, WE MAPPED 7,845 RSV-INDUCIBLE TN5 TRANSPOSASE PEAKS ONTO THE BRD4-DEPENDENT GENE BODIES. THESE WERE LOCATED IN PROMOTERS AND INTRONS OF CYTOSTRUCTURAL AND EXTRACELLULAR MATRIX (ECM) FORMATION GENES. THESE DATA INDICATE THAT BRD4 MEDIATES THE DYNAMIC RESPONSE OF AIRWAY EPITHELIAL CELLS TO RNA INFECTION BY MODULATING THE EXPRESSION OF ITS COACTIVATORS, CONTROLLING THE EXPRESSION OF HOST DEFENSE MECHANISMS AND REMODELING GENES THROUGH CHANGES IN PROMOTER ACCESSIBILITY. 2021 7 4741 28 NOVEL HDAC INHIBITOR MAKV-8 AND IMATINIB SYNERGISTICALLY KILL CHRONIC MYELOID LEUKEMIA CELLS VIA INHIBITION OF BCR-ABL/MYC-SIGNALING: EFFECT ON IMATINIB RESISTANCE AND STEM CELLS. BACKGROUND: CHRONIC MYELOID LEUKEMIA (CML) PATHOGENESIS IS MAINLY DRIVEN BY THE ONCOGENIC BREAKPOINT CLUSTER REGION-ABELSON MURINE LEUKEMIA VIRAL ONCOGENE HOMOLOG 1 (BCR-ABL) FUSION PROTEIN. SINCE BCR-ABL DISPLAYS ABNORMAL CONSTITUTIVE TYROSINE KINASE ACTIVITY, THERAPIES USING TYROSINE KINASE INHIBITORS (TKIS) SUCH AS IMATINIB REPRESENT A MAJOR BREAKTHROUGH FOR THE OUTCOME OF CML PATIENTS. NEVERTHELESS, THE DEVELOPMENT OF TKI RESISTANCE AND THE PERSISTENCE OF LEUKEMIA STEM CELLS (LSCS) REMAIN BARRIERS TO CURE THE DISEASE, JUSTIFYING THE DEVELOPMENT OF NOVEL THERAPEUTIC APPROACHES. SINCE THE ACTIVITY OF HISTONE DEACETYLASE (HDAC) IS DEREGULATED IN NUMEROUS CANCERS INCLUDING CML, PAN-HDAC INHIBITORS MAY REPRESENT PROMISING THERAPEUTIC REGIMENS FOR THE TREATMENT OF CML CELLS IN COMBINATION WITH TKI. RESULTS: WE ASSESSED THE ANTI-LEUKEMIC ACTIVITY OF A NOVEL HYDROXAMATE-BASED PAN-HDAC INHIBITOR MAKV-8, WHICH COMPLIED WITH THE LIPINSKI'S "RULE OF FIVE," IN VARIOUS CML CELLS ALONE OR IN COMBINATION WITH IMATINIB. WE VALIDATED THE IN VITRO HDAC-INHIBITORY POTENTIAL OF MAKV-8 AND DEMONSTRATED EFFICIENT BINDING TO THE LIGAND-BINDING POCKET OF HDAC ISOENZYMES. IN CELLULO, MAKV-8 SIGNIFICANTLY INDUCED TARGET PROTEIN ACETYLATION, DISPLAYED CYTOSTATIC AND CYTOTOXIC PROPERTIES, AND TRIGGERED CONCOMITANT ER STRESS/PROTECTIVE AUTOPHAGY LEADING TO CANONICAL CASPASE-DEPENDENT APOPTOSIS. CONSIDERING THE SPECIFIC UPREGULATION OF SELECTED HDACS IN LSCS FROM CML PATIENTS, WE INVESTIGATED THE DIFFERENTIAL TOXICITY OF A CO-TREATMENT WITH MAKV-8 AND IMATINIB IN CML VERSUS HEALTHY CELLS. WE ALSO SHOWED THAT BECLIN-1 KNOCKDOWN PREVENTED MAKV-8-IMATINIB COMBINATION-INDUCED APOPTOSIS. MOREOVER, MAKV-8 AND IMATINIB CO-TREATMENT SYNERGISTICALLY REDUCED BCR-ABL-RELATED SIGNALING PATHWAYS INVOLVED IN CML CELL GROWTH AND SURVIVAL. SINCE OUR RESULTS SHOWED THAT LSCS FROM CML PATIENTS OVEREXPRESSED C-MYC, IMPORTANTLY MAKV-8-IMATINIB CO-TREATMENT REDUCED C-MYC LEVELS AND THE LSC POPULATION. IN VIVO, TUMOR GROWTH OF XENOGRAFTED K-562 CELLS IN ZEBRAFISH WAS COMPLETELY ABROGATED UPON COMBINED TREATMENT WITH MAKV-8 AND IMATINIB. CONCLUSIONS: COLLECTIVELY, THE PRESENT FINDINGS SHOW THAT COMBINATIONS HDAC INHIBITOR-IMATINIB ARE LIKELY TO OVERCOME DRUG RESISTANCE IN CML PATHOLOGY. 2020 8 4003 25 LOSS OF PRMT7 REPROGRAMS GLYCINE METABOLISM TO SELECTIVELY ERADICATE LEUKEMIA STEM CELLS IN CML. OUR GROUP HAS REPORTED PREVIOUSLY ON THE ROLE OF VARIOUS MEMBERS OF THE PROTEIN ARGININE METHYLTRANSFERASE (PRMT) FAMILY, WHICH ARE INVOLVED IN EPIGENETIC REGULATION, IN THE PROGRESSION OF LEUKEMIA. HERE, WE EXPLORED THE ROLE OF PRMT7, GIVEN ITS UNIQUE FUNCTION WITHIN THE PRMT FAMILY, IN THE MAINTENANCE OF LEUKEMIA STEM CELLS (LSCS) IN CHRONIC MYELOID LEUKEMIA (CML). GENETIC LOSS OF PRMT7, AND THE DEVELOPMENT AND TESTING OF A SMALL-MOLECULE SPECIFIC INHIBITOR OF PRMT7, SHOWED THAT TARGETING PRMT7 DELAYED LEUKEMIA DEVELOPMENT AND IMPAIRED SELF-RENEWAL OF LSCS IN A CML MOUSE MODEL AND IN PRIMARY CML CD34(+) CELLS FROM HUMANS WITHOUT AFFECTING NORMAL HEMATOPOIESIS. MECHANISTICALLY, LOSS OF PRMT7 RESULTED IN REDUCED EXPRESSIONS OF GLYCINE DECARBOXYLASE, LEADING TO THE REPROGRAMING OF GLYCINE METABOLISM TO GENERATE METHYLGLYOXAL, WHICH IS DETRIMENTAL TO LSCS. THESE FINDINGS LINK HISTONE ARGININE METHYLATION WITH GLYCINE METABOLISM, WHILE SUGGESTING PRMT7 AS A POTENTIAL THERAPEUTIC TARGET FOR THE ERADICATION OF LSCS IN CML. 2022 9 697 35 BROMODOMAIN AND EXTRATERMINAL PROTEINS AS NOVEL EPIGENETIC TARGETS FOR RENAL DISEASES. EPIGENETIC MECHANISMS, ESPECIALLY DNA METHYLATION AND HISTONE MODIFICATIONS, ARE DYNAMIC PROCESSES THAT REGULATE THE GENE EXPRESSION TRANSCRIPTIONAL PROGRAM IN NORMAL AND DISEASED STATES. THE BROMODOMAIN AND EXTRATERMINAL (BET) PROTEIN FAMILY (BRD2, BRD3, BRD4, AND BRDT) ARE EPIGENETIC READERS THAT, VIA BROMODOMAINS, REGULATE GENE TRANSCRIPTION BY BINDING TO ACETYLATED LYSINE RESIDUES ON HISTONES AND MASTER TRANSCRIPTIONAL FACTORS. EXPERIMENTAL DATA HAVE DEMONSTRATED THE INVOLVEMENT OF SOME BET PROTEINS IN MANY PATHOLOGICAL CONDITIONS, INCLUDING TUMOR DEVELOPMENT, INFECTIONS, AUTOIMMUNITY, AND INFLAMMATION. SELECTIVE BROMODOMAIN INHIBITORS ARE EPIGENETIC DRUGS THAT BLOCK THE INTERACTION BETWEEN BET PROTEINS AND ACETYLATED PROTEINS, THUS EXERTING BENEFICIAL EFFECTS. RECENT DATA HAVE DESCRIBED THE BENEFICIAL EFFECT OF BET INHIBITION ON EXPERIMENTAL RENAL DISEASES. EMERGING EVIDENCE UNDERSCORES THE IMPORTANCE OF ENVIRONMENTAL MODIFICATIONS IN THE ORIGIN OF PATHOLOGICAL FEATURES IN CHRONIC KIDNEY DISEASES (CKD). SEVERAL CELLULAR PROCESSES SUCH AS OXIDATION, METABOLIC DISORDERS, CYTOKINES, INFLAMMATION, OR ACCUMULATED UREMIC TOXINS MAY INDUCE EPIGENETIC MODIFICATIONS THAT REGULATE KEY PROCESSES INVOLVED IN RENAL DAMAGE AND IN OTHER PATHOLOGICAL CONDITIONS OBSERVED IN CKD PATIENTS. HERE, WE REVIEW HOW TARGETING BROMODOMAINS IN BET PROTEINS MAY REGULATE ESSENTIAL PROCESSES INVOLVED IN RENAL DISEASES AND IN ASSOCIATED COMPLICATIONS FOUND IN CKD PATIENTS, SUCH AS CARDIOVASCULAR DAMAGE, HIGHLIGHTING THE POTENTIAL OF EPIGENETIC THERAPEUTIC STRATEGIES AGAINST BET PROTEINS FOR CKD TREATMENT AND ASSOCIATED RISKS. 2019 10 913 17 CHRONIC FLT3-ITD SIGNALING IN ACUTE MYELOID LEUKEMIA IS CONNECTED TO A SPECIFIC CHROMATIN SIGNATURE. ACUTE MYELOID LEUKEMIA (AML) IS CHARACTERIZED BY RECURRENT MUTATIONS THAT AFFECT THE EPIGENETIC REGULATORY MACHINERY AND SIGNALING MOLECULES, LEADING TO A BLOCK IN HEMATOPOIETIC DIFFERENTIATION. CONSTITUTIVE SIGNALING FROM MUTATED GROWTH FACTOR RECEPTORS IS A MAJOR DRIVER OF LEUKEMIC GROWTH, BUT HOW ABERRANT SIGNALING AFFECTS THE EPIGENOME IN AML IS LESS UNDERSTOOD. FURTHERMORE, AML CELLS UNDERGO EXTENSIVE CLONAL EVOLUTION, AND THE MUTATIONS IN SIGNALING GENES ARE OFTEN SECONDARY EVENTS. TO ELUCIDATE HOW CHRONIC GROWTH FACTOR SIGNALING ALTERS THE TRANSCRIPTIONAL NETWORK IN AML, WE PERFORMED A SYSTEM-WIDE MULTI-OMICS STUDY OF PRIMARY CELLS FROM PATIENTS SUFFERING FROM AML WITH INTERNAL TANDEM DUPLICATIONS IN THE FLT3 TRANSMEMBRANE DOMAIN (FLT3-ITD). THIS STRATEGY REVEALED COOPERATION BETWEEN THE MAP KINASE (MAPK) INDUCIBLE TRANSCRIPTION FACTOR AP-1 AND RUNX1 AS A MAJOR DRIVER OF A COMMON, FLT3-ITD-SPECIFIC GENE EXPRESSION AND CHROMATIN SIGNATURE, DEMONSTRATING A MAJOR IMPACT OF MAPK SIGNALING PATHWAYS IN SHAPING THE EPIGENOME OF FLT3-ITD AML. 2015 11 6023 28 THE BET PROTAC INHIBITOR DBET6 PROTECTS AGAINST RETINAL DEGENERATION AND INHIBITS THE CGAS-STING IN RESPONSE TO LIGHT DAMAGE. BACKGROUND: CHRONIC INFLAMMATION SIGNIFICANTLY CONTRIBUTES TO PHOTORECEPTOR DEATH IN BLINDING RETINAL DISEASES SUCH AS AGE-RELATED MACULAR DEGENERATION (AMD) AND RETINITIS PIGMENTOSA (RP). BROMODOMAIN AND EXTRATERMINAL DOMAIN (BET) PROTEINS ARE EPIGENETIC READERS THAT ACT AS KEY PROINFLAMMATORY FACTORS. WE RECENTLY FOUND THE FIRST-GENERATION BET INHIBITOR JQ1 ALLEVIATED SODIUM IODATE-INDUCED RETINAL DEGENERATION BY SUPPRESSING CGAS-STING INNATE IMMUNITY. HERE, WE INVESTIGATED THE EFFECTS AND MECHANISM OF DBET6, A PROTEOLYSIS?TARGETING CHIMERA (PROTAC) SMALL MOLECULE THAT SELECTIVELY DEGRADES BET BY THE UBIQUITIN?PROTEASOME SYSTEM, IN LIGHT-INDUCED RETINAL DEGENERATION. METHODS: MICE WERE EXPOSED TO BRIGHT LIGHT TO INDUCE RETINAL DEGENERATION, AND THE ACTIVATION OF CGAS-STING WAS DETERMINED BY RNA-SEQUENCING AND MOLECULAR BIOLOGY. RETINAL FUNCTION, MORPHOLOGY, PHOTORECEPTOR VIABILITY AND RETINAL INFLAMMATION WERE EXAMINED IN THE PRESENCE AND ABSENCE OF DBET6 TREATMENT. RESULTS: INTRAPERITONEAL INJECTION OF DBET6 LED TO THE RAPID DEGRADATION OF BET PROTEIN IN THE RETINA WITHOUT DETECTABLE TOXICITY. DBET6 IMPROVED RETINAL RESPONSIVENESS AND VISUAL ACUITY AFTER LIGHT DAMAGE (LD). DBET6 ALSO REPRESSED LD-INDUCED RETINAL MACROPHAGES/MICROGLIA ACTIVATION, MULLER CELL GLIOSIS, PHOTORECEPTOR DEATH AND RETINAL DEGENERATION. ANALYSIS OF SINGLE-CELL RNA-SEQUENCING RESULTS REVEALED CGAS-STING COMPONENTS WERE EXPRESSED IN RETINAL MICROGLIA. LD LED TO DRAMATIC ACTIVATION OF THE CGAS-STING PATHWAY, WHEREAS DBET6 SUPPRESSED LD-INDUCED STING EXPRESSION IN REACTIVE MACROPHAGES/MICROGLIA AND THE RELATED INFLAMMATORY RESPONSE. CONCLUSIONS: THIS STUDY INDICATES TARGETED DEGRADATION OF BET BY DBET6 EXERTS NEUROPROTECTIVE EFFECTS BY INHIBITING CGAS-STING IN REACTIVE RETINAL MACROPHAGES/MICROGLIA, AND IS EXPECTED TO BECOME A NEW STRATEGY FOR TREATMENT OF RETINAL DEGENERATION. 2023 12 1652 26 DOPAMINE SIGNALING LEADS TO LOSS OF POLYCOMB REPRESSION AND ABERRANT GENE ACTIVATION IN EXPERIMENTAL PARKINSONISM. POLYCOMB GROUP (PCG) PROTEINS BIND TO AND REPRESS GENES IN EMBRYONIC STEM CELLS THROUGH LINEAGE COMMITMENT TO THE TERMINAL DIFFERENTIATED STATE. PCG REPRESSED GENES ARE COMMONLY CHARACTERIZED BY THE PRESENCE OF THE EPIGENETIC HISTONE MARK H3K27ME3, CATALYZED BY THE POLYCOMB REPRESSIVE COMPLEX 2. HERE, WE PRESENT IN VIVO EVIDENCE FOR A PREVIOUSLY UNRECOGNIZED PLASTICITY OF PCG-REPRESSED GENES IN TERMINALLY DIFFERENTIATED BRAIN NEURONS OF PARKISONIAN MICE. WE SHOW THAT ACUTE ADMINISTRATION OF THE DOPAMINE PRECURSOR, L-DOPA, INDUCES A REMARKABLE INCREASE IN H3K27ME3S28 PHOSPHORYLATION. THE INDUCTION OF THE H3K27ME3S28P HISTONE MARK SPECIFICALLY OCCURS IN MEDIUM SPINY NEURONS EXPRESSING DOPAMINE D1 RECEPTORS AND IS DEPENDENT ON MSK1 KINASE ACTIVITY AND DARPP-32-MEDIATED INHIBITION OF PROTEIN PHOSPHATASE-1. CHROMATIN IMMUNOPRECIPITATION (CHIP) EXPERIMENTS SHOWED THAT INCREASED H3K27ME3S28P WAS ACCOMPANIED BY REDUCED PCG BINDING TO REGULATORY REGIONS OF GENES. AN ANALYSIS OF THE GENOME WIDE DISTRIBUTION OF L-DOPA-INDUCED H3K27ME3S28 PHOSPHORYLATION BY CHIP SEQUENCING (CHIP-SEQ) IN COMBINATION WITH EXPRESSION ANALYSIS BY RNA-SEQUENCING (RNA-SEQ) SHOWED THAT THE INDUCTION OF H3K27ME3S28P CORRELATED WITH INCREASED EXPRESSION OF A SUBSET OF PCG REPRESSED GENES. WE FOUND THAT INDUCTION OF H3K27ME3S28P PERSISTED DURING CHRONIC L-DOPA ADMINISTRATION TO PARKISONIAN MICE AND CORRELATED WITH ABERRANT GENE EXPRESSION. WE PROPOSE THAT DOPAMINERGIC TRANSMISSION CAN ACTIVATE PCG REPRESSED GENES IN THE ADULT BRAIN AND THEREBY CONTRIBUTE TO LONG-TERM MALADAPTIVE RESPONSES INCLUDING THE MOTOR COMPLICATIONS, OR DYSKINESIA, CAUSED BY PROLONGED ADMINISTRATION OF L-DOPA IN PARKINSON'S DISEASE. 2014 13 1036 21 CLASS I HISTONE DEACETYLASES REGULATE P53/NF-KAPPAB CROSSTALK IN CANCER CELLS. THE TRANSCRIPTION FACTORS NF-KAPPAB AND P53 AS WELL AS THEIR CROSSTALK DETERMINE THE FATE OF TUMOR CELLS UPON THERAPEUTIC INTERVENTIONS. REPLICATIVE STRESS AND CYTOKINES PROMOTE SIGNALING CASCADES THAT LEAD TO THE CO-REGULATION OF P53 AND NF-KAPPAB. CONSEQUENTLY, NUCLEAR P53/NF-KAPPAB SIGNALING COMPLEXES ACTIVATE NF-KAPPAB-DEPENDENT SURVIVAL GENES. THE 18 HISTONE DEACETYLASES (HDACS) ARE EPIGENETIC MODULATORS THAT FALL INTO FOUR CLASSES (I-IV). INHIBITORS OF HISTONE DEACETYLASES (HDACI) BECOME INCREASINGLY APPRECIATED AS ANTI-CANCER AGENTS. BASED ON THEIR EFFECTS ON P53 AND NF-KAPPAB, WE ADDRESSED WHETHER CLINICALLY RELEVANT HDACI AFFECT THE NF-KAPPAB/P53 CROSSTALK. THE CHEMOTHERAPEUTICS HYDROXYUREA, ETOPOSIDE, AND FLUDARABINE HALT CELL CYCLE PROGRESSION, INDUCE DNA DAMAGE, AND LEAD TO DNA FRAGMENTATION. THESE AGENTS CO-INDUCE P53 AND NF-KAPPAB-DEPENDENT GENE EXPRESSION IN CELL LINES FROM BREAST AND COLON CANCER AND IN PRIMARY CHRONIC LYMPHATIC LEUKEMIA (CLL) CELLS. USING SPECIFIC HDACI, WE FIND THAT THE CLASS I SUBGROUP OF HDACS, BUT NOT THE CLASS IIB DEACETYLASE HDAC6, ARE REQUIRED FOR THE HYDROXYUREA-INDUCED CROSSTALK BETWEEN P53 AND NF-KAPPAB. HDACI DECREASE THE BASAL AND STRESS-INDUCED EXPRESSION OF P53 AND BLOCK NF-KAPPAB-REGULATED GENE EXPRESSION. WE FURTHER SHOW THAT CLASS I HDACI INDUCE SENESCENCE IN PANCREATIC CANCER CELLS WITH MUTANT P53. 2017 14 826 25 CHARACTERIZATION OF K562 CELLS: UNCOVERING NOVEL CHROMOSOMES, ASSESSING TRANSFERRIN RECEPTOR EXPRESSION, AND PROBING PHARMACOLOGICAL THERAPIES. HUMAN ERYTHROLEUKEMIC K562 CELLS REPRESENT THE PROTOTYPICAL CELL CULTURE MODEL OF CHRONIC MYELOID LEUKEMIA (CML). THE CELLS ARE PSEUDO-TRIPLOID AND POSITIVE FOR THE PHILADELPHIA CHROMOSOME. THEREFORE, K562 CELLS HAVE BEEN WIDELY USED FOR INVESTIGATING THE BCR/ABL1 ONCOGENE AND THE TYROSINE KINASE INHIBITOR, IMATINIB-MESYLATE. FURTHER, K562 CELLS OVEREXPRESS TRANSFERRIN RECEPTORS (TFR) AND HAVE BEEN USED AS A MODEL FOR TARGETING CYTOTOXIC THERAPIES, VIA RECEPTOR-MEDIATED ENDOCYTOSIS. HERE, WE HAVE CHARACTERIZED K562 CELLS FOCUSING ON THE KARYOTYPE OF CELLS IN PROLONGED CULTURE, REGULATION OF EXPRESSION OF TFR IN WILDTYPE (WT) AND DOXORUBICIN-RESISTANT CELLS, AND RESPONSES TO HISTONE DEACETYLASE INHIBITION (HDACI). KARYOTYPE ANALYSIS INDICATES NOVEL CHROMOSOMES AND GENE EXPRESSION ANALYSIS SUGGESTS A SHIFT OF CULTURED K562 CELLS AWAY FROM PATIENT-DERIVED LEUKEMIC CELLS. WE CONFIRM THE HIGH EXPRESSION OF TFR ON K562 CELLS USING IMMUNOFLUORESCENCE AND CELL-SURFACE RECEPTOR BINDING RADIOASSAYS. IMPORTANTLY, HIGH TFR EXPRESSION IS OBSERVED IN PATIENT-DERIVED CELLS, AND WE HIGHLIGHT THE PERSISTENT EXPRESSION OF TFR FOLLOWING DOXORUBICIN ACQUIRED RESISTANCE. EPIGENETIC ANALYSIS INDICATES THAT PERMISSIVE HISTONE ACETYLATION AND METHYLATION AT THE PROMOTER REGION REGULATES THE TRANSCRIPTION OF TFR IN K562 CELLS. FINALLY, WE SHOW RELATIVELY HIGH EXPRESSION OF HDAC ENZYMES IN K562 CELLS AND DEMONSTRATE THE CHEMOTOXIC EFFECTS OF HDACI, USING THE FDA-APPROVED HYDROXAMIC ACID, VORINOSTAT. TOGETHER WITH A DESCRIPTION OF MORPHOLOGY, INFRARED SPECTRAL ANALYSIS, AND EXAMINATION OF METABOLIC PROPERTIES, WE PROVIDE A COMPREHENSIVE CHARACTERIZATION OF K562 CELLS. OVERALL, K562 CELL CULTURE SYSTEMS REMAIN WIDELY USED FOR THE INVESTIGATION OF NOVEL THERAPEUTICS FOR CML, WHICH IS PARTICULARLY IMPORTANT IN CASES OF IMATINIB-MESYLATE RESISTANCE. 2023 15 1455 26 DISCOVERY OF THIENO[2,3-D]PYRIMIDINE-BASED HYDROXAMIC ACID DERIVATIVES AS BROMODOMAIN-CONTAINING PROTEIN 4/HISTONE DEACETYLASE DUAL INHIBITORS INDUCE AUTOPHAGIC CELL DEATH IN COLORECTAL CARCINOMA CELLS. BROMODOMAIN-CONTAINING PROTEIN 4 (BRD4) AND HISTONE DEACETYLASES (HDAC) ARE BOTH ATTRACTIVE EPIGENETIC TARGETS IN CANCER AND OTHER CHRONIC DISEASES. BASED ON THE INTEGRATED FRAGMENT-BASED DRUG DESIGN, SYNTHESIS, AND IN VITRO AND IN VIVO EVALUATIONS, A SERIES OF NOVEL THIENO[2,3-D]PYRIMIDINE-BASED HYDROXAMIC ACID DERIVATIVES ARE DISCOVERED AS SELECTIVE BRD4-HDAC DUAL INHIBITORS. COMPOUND 17C IS THE MOST POTENT INHIBITOR FOR BRD4 AND HDAC WITH IC(50) VALUES AT NANOMOLAR LEVELS, AS WELL AS THE EXPRESSION LEVEL OF C-MYC, AND INCREASES THE ACETYLATION OF HISTONE H3. MOREOVER, 17C PRESENTS INHIBITORY EFFECTS ON THE PROLIFERATION OF COLORECTAL CARCINOMA (CRC) CELLS VIA INDUCING AUTOPHAGIC CELL DEATH. IT ALSO HAS A GOOD PHARMACOKINETIC PROFILE IN RATS AND ORAL BIOAVAILABILITY OF 40.5%. IN THE HCT-116 XENOGRAFT IN VIVO MODELS, 17C DISPLAYS POTENT INHIBITORY EFFICIENCY ON TUMOR GROWTH BY INDUCING AUTOPHAGIC CELL DEATH AND SUPPRESSING IL6-JAK-STAT SIGNALING PATHWAYS. OUR RESULTS SUGGEST THAT THE BRD4-HDAC DUAL INHIBITION MIGHT BE AN ATTRACTIVE THERAPEUTIC STRATEGY FOR CRC. 2020 16 2068 22 EPIGENETIC CONTROL OF MICROSOMAL PROSTAGLANDIN E SYNTHASE-1 BY HDAC-MEDIATED RECRUITMENT OF P300. NONSTEROIDAL ANTI-INFLAMMATORY DRUGS ARE THE MOST WIDELY USED MEDICINE TO TREAT PAIN AND INFLAMMATION, AND TO INHIBIT PLATELET FUNCTION. UNDERSTANDING THE EXPRESSION REGULATION OF ENZYMES OF THE PROSTANOID PATHWAY IS OF GREAT MEDICAL RELEVANCE. HISTONE ACETYLATION CRUCIALLY CONTROLS GENE EXPRESSION. WE SET OUT TO IDENTIFY THE IMPACT OF HISTONE DEACETYLASES (HDACS) ON THE GENERATION OF PROSTANOIDS AND EXAMINE THE CONSEQUENCES ON VASCULAR FUNCTION. HDAC INHIBITION (HDACI) WITH THE PAN-HDAC INHIBITOR, VORINOSTAT, ATTENUATED PROSTAGLANDIN (PG)E(2) GENERATION IN THE MURINE VASCULATURE AND IN HUMAN VASCULAR SMOOTH MUSCLE CELLS. IN LINE WITH THIS, THE EXPRESSION OF THE KEY ENZYME FOR PGE(2) SYNTHESIS, MICROSOMAL PGE SYNTHASE-1 (PTGES1), WAS REDUCED BY HDACI. ACCORDINGLY, THE RELAXATION TO ARACHIDONIC ACID WAS DECREASED AFTER EX VIVO INCUBATION OF MURINE VESSELS WITH HDACI. TO IDENTIFY THE UNDERLYING MECHANISM, CHROMATIN IMMUNOPRECIPITATION (CHIP) AND CHIP-SEQUENCING ANALYSIS WERE PERFORMED. THESE RESULTS SUGGEST THAT HDACS ARE INVOLVED IN THE RECRUITMENT OF THE TRANSCRIPTIONAL ACTIVATOR P300 TO THE PTGES1 GENE AND THAT HDACI PREVENTED THIS EFFECT. IN LINE WITH THE ACETYLTRANSFERASE ACTIVITY OF P300, H3K27 ACETYLATION WAS REDUCED AFTER HDACI AND RESULTED IN THE FORMATION OF HETEROCHROMATIN IN THE PTGES1 GENE. IN CONCLUSION, HDAC ACTIVITY MAINTAINS PTGES1 EXPRESSION BY RECRUITING P300 TO ITS GENE. 2017 17 4361 28 MIR-96 ACTS AS A TUMOR SUPPRESSOR VIA TARGETING THE BCR-ABL1 ONCOGENE IN CHRONIC MYELOID LEUKEMIA BLASTIC TRANSFORMATION. MICRORNA-MEDIATED POSTTRANSCRIPTIONAL REGULATION IS AN IMPORTANT EPIGENETIC REGULATORY MECHANISM OF GENE EXPRESSION, AND ITS DYSREGULATION IS INVOLVED IN THE DEVELOPMENT AND PROGRESSION OF A VARIETY OF MALIGNANCIES, INCLUDING CHRONIC MYELOID LEUKEMIA (CML). THE BCR-ABL1 FUSION GENE IS NOT ONLY THE INITIATING FACTOR OF CML, BUT IT IS ALSO AN IMPORTANT DRIVING FACTOR FOR BLASTIC TRANSFORMATION. TYROSINE KINASE INHIBITORS (TKIS) TARGETING BCR-ABL1 TYROSINE KINASE ACTIVITY, REPRESENTED BY IMATINIB, ARE CURRENTLY THE FIRST-LINE TREATMENT FOR CML. HOWEVER, DUE TO PRIMARY RESISTANCE OR SECONDARY RESISTANCE CAUSED BY MUTATIONS IN THE BCR-ABL1 KINASE DOMAIN, TKIS CANNOT COMPLETELY PREVENT THE PROGRESSION OF CML; THUS, THE STUDY OF BCR-ABL1 GENE EXPRESSION REGULATION IS OF GREAT SIGNIFICANCE. IN THIS STUDY, BIOINFORMATICS ANALYSIS AND OUR RESULTS SHOWED THAT MIR-96 COULD DIRECTLY BIND TO THE 3'UTR REGION OF BCR-ABL1 TO REGULATE FUSION PROTEIN EXPRESSION, THEREBY REGULATING ITS DOWNSTREAM SIGNALING PATHWAY ACTIVITY. WE ALSO FOUND THAT MIR-96 WAS DOWNREGULATED DURING THE PROGRESSION FROM THE CHRONIC PHASE (CML-CP) TO THE BLAST CRISIS (CML-BC). DOWNREGULATION OF MIR-96 COULD PROMOTE THE PROLIFERATION AND PARTICIPATE IN THE CELL DIFFERENTIATION OF CML-BC CELLS. ADDITIONALLY, WE FOUND THAT THE NOVEL HISTONE DEACETYLASE DRUG CHIDAMIDE AND THE DNA METHYLTRANSFERASE INHIBITOR DECITABINE COULD RESTORE THE LOW EXPRESSION OF MIR-96 IN CML CELLS, AND THERE WERE TWO ABNORMAL HYPERMETHYLATED SITES IN THE PROMOTER REGION OF MIR-96 IN CML, SUGGESTING THAT ITS LOW EXPRESSION MIGHT BE AT LEAST PARTIALLY REGULATED BY EPIGENETIC MECHANISMS. IN ADDITION, RE-EXPRESSION OF MIR-96 COULD INCREASE THE SENSITIVITY OF CML-BC CELLS TO IMATINIB. THUS, MIR-96 FUNCTIONS AS A TUMOR SUPPRESSOR, AND RE-EXPRESSION OF THIS MICRORNA MIGHT HAVE THERAPEUTIC BENEFITS IN CML BLASTIC TRANSFORMATION. 2019 18 84 31 A NOVEL EPIGENETIC MECHANISM OF FXR INHIBITING GLP-1 SECRETION VIA MIR-33 AND ITS DOWNSTREAM TARGETS. TYPE II DIABETES IS A COMPLEX, CHRONIC, AND PROGRESSIVE DISEASE. PREVIOUSLY, WE DEMONSTRATE THAT FXR INHIBITS GLP-1 SECRETION VIA INTERACTING WITH CREB TO INHIBIT THE TRANSCRIPTIONAL ACTIVITY OF CREB, THUS PROMOTING THE DEVELOPMENT OF TYPE II DIABETES. EPIGENETIC MODIFICATIONS, SUCH AS DNA METHYLATION, HISTONE ACETYLATION, AND POST-TRANSCRIPTIONAL RNA REGULATION, ARE ESSENTIAL MEDIATORS CONTRIBUTING TO DIABETES-ASSOCIATED MORBIDITY AND MORTALITY. THUS, WE ATTEMPTED TO INVESTIGATE THE EPIGENETIC MECHANISMS OF FXR MODULATING GLP-1 SECRETION. FIRSTLY, THE INVOLVEMENT OF HISTONE ACETYLATION, DNA METHYLATION, AND POST-TRANSCRIPTIONAL REGULATION IN FXR INHIBITING GLP-1 SECRETION WAS VERIFIED. AS FXR OVEREXPRESSION SIGNIFICANTLY INHIBITED THE ACTIVITY OF GCG 3'-UTR, WE HYPOTHESIZE THAT MIRNA MIGHT PARTICIPATE IN THE MECHANISM. TWO ONLINE TOOLS AND REAL-TIME PCR REVEALED THAT FXR PROMOTED MIR-33 EXPRESSION. MOREOVER, MIR-33 INHIBITED THE EXPRESSION OF GCG AND CREB1 THROUGH DIRECT TARGETING IN STC-1 CELLS. FXR OVEREXPRESSION IN STC-1 CELLS SIGNIFICANTLY REDUCED THE MRNA EXPRESSION AND PROTEIN LEVELS OF BOTH GCG AND CREB1, AS WELL AS THE SECRETION OF GLP-1; MIR-33 INHIBITION EXERTED OPPOSING EFFECTS. MORE IMPORTANTLY, THE EFFECTS OF FXR OVEREXPRESSION WERE SIGNIFICANTLY REVERSED BY MIR-33 INHIBITION, INDICATING THAT FXR INHIBITED GLP-1 SECRETION THROUGH PROMOTING MIR-33 EXPRESSION, THEREFORE INHIBITING THE EXPRESSION OF MIR-33 TARGETS, GCG AND CREB1. IN CONCLUSION, WE PROVIDE A NOVEL EPIGENETIC MECHANISM BY WHICH FXR INHIBITS THE SECRETION OF GLP-1 THROUGH MIR-33 AND ITS TWO DOWNSTREAM TARGETS, GCG AND CREB1. THESE FINDINGS MIGHT PROVIDE INNOVATIVE STRATEGIES FOR IMPROVING TYPE II DIABETES, WHICH NEEDS FURTHER IN VIVO AND CLINICAL INVESTIGATION. 2019 19 3656 29 INDUCIBLE PRMT1 ABLATION IN ADULT VASCULAR SMOOTH MUSCLE LEADS TO CONTRACTILE DYSFUNCTION AND AORTIC DISSECTION. VASCULAR SMOOTH MUSCLE CELLS (VSMCS) HAVE REMARKABLE PLASTICITY IN RESPONSE TO DIVERSE ENVIRONMENTAL CUES. ALTHOUGH THESE CELLS ARE VERSATILE, CHRONIC STRESS CAN TRIGGER VSMC DYSFUNCTION, WHICH ULTIMATELY LEADS TO VASCULAR DISEASES SUCH AS AORTIC ANEURYSM AND ATHEROSCLEROSIS. PROTEIN ARGININE METHYLTRANSFERASE 1 (PRMT1) IS A MAJOR ENZYME CATALYZING ASYMMETRIC ARGININE DIMETHYLATION OF PROTEINS THAT ARE SOURCES OF ASYMMETRIC DIMETHYLARGININE (ADMA), AN ENDOGENOUS INHIBITOR OF NITRIC OXIDE SYNTHASE. ALTHOUGH A POTENTIAL ROLE OF PRMT1 IN VASCULAR PATHOGENESIS HAS BEEN PROPOSED, ITS ROLE IN VASCULAR FUNCTION HAS YET TO BE CLARIFIED. HERE, WE INVESTIGATED THE ROLE AND UNDERLYING MECHANISM OF PRMT1 IN VASCULAR SMOOTH MUSCLE CONTRACTILITY AND FUNCTION. THE EXPRESSION OF PRMT1 AND CONTRACTILE-RELATED GENES WAS SIGNIFICANTLY DECREASED IN THE AORTAS OF ELDERLY HUMANS AND PATIENTS WITH AORTIC ANEURYSMS. MICE WITH VSMC-SPECIFIC PRMT1 ABLATION (SMKO) EXHIBITED PARTIAL LETHALITY, LOW BLOOD PRESSURE AND AORTIC DILATION. THE PRMT1-ABLATED AORTAS SHOWED AORTIC DISSECTION WITH ELASTIC FIBER DEGENERATION AND CELL DEATH. EX VIVO AND IN VITRO ANALYSES INDICATED THAT PRMT1 ABLATION SIGNIFICANTLY DECREASED THE CONTRACTILITY OF THE AORTA AND TRACTION FORCES OF VSMCS. PRMT1 ABLATION DOWNREGULATED THE EXPRESSION OF CONTRACTILE GENES SUCH AS MYOCARDIN WHILE UPREGULATING THE EXPRESSION OF SYNTHETIC GENES, THUS CAUSING THE CONTRACTILE TO SYNTHETIC PHENOTYPIC SWITCH OF VSMCS. IN ADDITION, MECHANISTIC STUDIES DEMONSTRATED THAT PRMT1 DIRECTLY REGULATES MYOCARDIN GENE ACTIVATION BY MODULATING EPIGENETIC HISTONE MODIFICATIONS IN THE MYOCARDIN PROMOTER REGION. THUS, OUR STUDY DEMONSTRATES THAT VSMC PRMT1 IS ESSENTIAL FOR VASCULAR HOMEOSTASIS AND THAT ITS ABLATION CAUSES AORTIC DILATION/DISSECTION THROUGH IMPAIRED MYOCARDIN EXPRESSION. 2021 20 5337 22 QUANTITATIVE ANALYSIS OF HISTONE H3 AND H4 POST-TRANSLATIONAL MODIFICATIONS IN DOXORUBICIN-RESISTANT LEUKEMIA CELLS. THE EPIGENETIC REMODELING OF CHROMATIN THROUGH HISTONE MODIFICATIONS HAS BEEN WIDELY IMPLICATED IN DRUG RESISTANCE OF CANCER CELLS. HOWEVER, WHETHER EPIGENETIC MECHANISMS CONTRIBUTE SPECIFICALLY TO DOXORUBICIN RESISTANCE IN LEUKEMIA HAS NOT BEEN CAREFULLY EXAMINED. USING A STABLE AND SENSITIVE WORKFLOW BASED ON LC-MS, WE QUANTITATIVELY COMPARED THE EXTENTS OF METHYLATION AND ACETYLATION OF HISTONE H3 AND H4 IN ACUTE LEUKEMIA CELL LINE HL60 AND ITS DOXORUBICIN-RESISTANT DERIVATIVE, HL60/ADR, AS WELL AS THE CHRONIC LEUKEMIA CELL LINE K562 AND ITS DOXORUBICIN-RESISTANT DERIVATIVE, K562/ADR. WE FOUND THAT INCREASED LEVELS OF H3K9 METHYLATION, H3K14, H3K18 AND H3K23 ACETYLATION, AND POTENTIALLY H4K20 METHYLATION, ARE ASSOCIATED WITH DRUG RESISTANCE IN BOTH CELLS. OUR RESULTS DEMONSTRATED THAT THE DOXORUBICIN-RESISTANT ACUTE AND CHRONIC LEUKEMIA CELL LINES MAY SHARE A COMMON EPIGENETIC MECHANISM THAT INVOLVES A COMBINATION OF TRANSCRIPTIONAL ACTIVATION AND SILENCING. 2016