1 6688 119 VALPROATE SYNERGIZES WITH PURINE NUCLEOSIDE ANALOGUES TO INDUCE APOPTOSIS OF B-CHRONIC LYMPHOCYTIC LEUKAEMIA CELLS. RESISTANCE TO CHEMOTHERAPY AND DRUG TOXICITY ARE TWO MAJOR CONCERNS OF CHRONIC LYMPHOCYTIC LEUKAEMIA (B-CLL) TREATMENT BY PURINE NUCLEOSIDE ANALOGUES (PNA, I.E. FLUDARABINE AND CLADRIBINE). WE HYPOTHESIZED THAT TARGETING EPIGENETIC CHANGES MIGHT ADDRESS THESE ISSUES AND EVALUATED THE EFFECT OF THE HISTONE DEACETYLASE INHIBITOR VALPROATE (VPA) AT A CLINICALLY RELEVANT CONCENTRATION. VPA ACTED IN A HIGHLY SYNERGISTIC/ADDITIVE MANNER WITH FLUDARABINE AND CLADRIBINE TO INDUCE APOPTOSIS OF B-CLL CELLS. IMPORTANTLY, VPA ALSO RESTORED SENSITIVITY TO FLUDARABINE IN B CELLS FROM POOR PROGNOSIS CLL PATIENTS WHO BECAME RESISTANT TO CHEMOTHERAPY. MECHANISM OF APOPTOSIS INDUCED BY VPA ALONE OR COMBINED WITH FLUDARABINE OR TO CLADRIBINE WAS CASPASE-DEPENDENT AND INVOLVED THE EXTRINSIC PATHWAY. VPA, BUT NEITHER FLUDARABINE NOR CLADRIBINE, ENHANCED THE PRODUCTION OF REACTIVE OXYGEN SPECIES (ROS) AND INHIBITION OF ROS WITH N-ACETYLCYSTEINE DECREASES APOPTOSIS OF CLL CELLS. VPA STIMULATES HYPERPHOSPHORYLATION OF P42/P44 ERK, CYTOCHROME C RELEASE AND OVEREXPRESSION OF BAX AND FAS. TOGETHER, OUR DATA INDICATE THAT VPA MAY AMELIORATE THE OUTCOME OF PNA-BASED THERAPEUTIC PROTOCOLS AND PROVIDE A POTENTIAL ALTERNATIVE TREATMENT IN BOTH THE RELAPSED AND FRONT-LINE RESISTANT PATIENTS AND IN PATIENTS WITH HIGH RISK FEATURES. 2009 2 64 30 A HIGH-THROUGHPUT SCREENING ASSAY FOR SILENCING ESTABLISHED HIV-1 MACROPHAGE INFECTION IDENTIFIES NUCLEOSIDE ANALOGS THAT PERTURB H3K9ME3 ON PROVIRAL GENOMES. HIV-INFECTED MACROPHAGES ARE LONG-LIVED CELLS THAT REPRESENT A BARRIER TO FUNCTIONAL CURE. ADDITIONALLY, LOW-LEVEL VIRAL EXPRESSION BY CENTRAL NERVOUS SYSTEM (CNS) MACROPHAGES CONTRIBUTES TO NEUROCOGNITIVE DEFICITS THAT DEVELOP DESPITE ANTIRETROVIRAL THERAPY (ART). WE RECENTLY IDENTIFIED H3K9ME3 AS AN ATYPICAL EPIGENETIC MARK ASSOCIATED WITH CHRONIC HIV INFECTION IN MACROPHAGES. THUS, STRATEGIES ARE NEEDED TO SUPPRESS HIV-1 EXPRESSION IN MACROPHAGES, BUT THE UNIQUE MYELOID ENVIRONMENT AND THE RESPONSIBLE MACROPHAGE/CNS-TROPIC STRAINS REQUIRE CELL/STRAIN-SPECIFIC APPROACHES. HERE, WE GENERATED AN HIV-1 REPORTER VIRUS FROM A CNS-DERIVED STRAIN WITH INTACT AUXILIARY GENES EXPRESSING DESTABILIZED LUCIFERASE. WE EMPLOYED THIS REPORTER VIRUS IN POLYCLONAL INFECTION OF PRIMARY HUMAN MONOCYTE-DERIVED MACROPHAGES (MDM) FOR A HIGH-THROUGHPUT SCREEN (HTS) TO IDENTIFY COMPOUNDS THAT SUPPRESS VIRUS EXPRESSION FROM ESTABLISHED MACROPHAGE INFECTION. SCREENING ~6,000 KNOWN DRUGS AND COMPOUNDS YIELDED 214 HITS. A SECONDARY SCREEN WITH 10-DOSE TITRATION IDENTIFIED 24 MEETING CRITERIA FOR HIV-SELECTIVE ACTIVITY. USING THREE REPLICATION-COMPETENT CNS-DERIVED MACROPHAGE-TROPIC HIV-1 ISOLATES AND VIRAL GENE EXPRESSION READOUT IN MDM, WE CONFIRMED THE EFFECT OF THREE PURINE ANALOGS, NELARABINE, FLUDARABINE, AND ENTECAVIR, SHOWING THE SUPPRESSION OF HIV-1 EXPRESSION FROM ESTABLISHED MACROPHAGE INFECTION. NELARABINE INHIBITED THE FORMATION OF H3K9ME3 ON HIV GENOMES IN MACROPHAGES. THUS, THIS NOVEL HTS ASSAY CAN IDENTIFY SUPPRESSORS OF HIV-1 TRANSCRIPTION IN ESTABLISHED MACROPHAGE INFECTION, SUCH AS NUCLEOSIDE ANALOGS AND HDAC INHIBITORS, WHICH MAY BE LINKED TO H3K9ME3 MODIFICATION. THIS SCREEN MAY BE USEFUL TO IDENTIFY NEW METABOLIC AND EPIGENETIC AGENTS THAT AMELIORATE HIV-DRIVEN NEUROINFLAMMATION IN PEOPLE ON ART OR PREVENT VIRAL RECRUDESCENCE FROM MACROPHAGE RESERVOIRS IN STRATEGIES TO ACHIEVE ART-FREE REMISSION. IMPORTANCE MACROPHAGES INFECTED BY HIV-1 ARE A LONG-LIVED RESERVOIR AND A BARRIER IN CURRENT EFFORTS TO ACHIEVE HIV CURE AND ALSO CONTRIBUTE TO NEUROCOGNITIVE COMPLICATIONS IN PEOPLE DESPITE ANTIRETROVIRAL THERAPY (ART). SILENCING HIV EXPRESSION IN THESE CELLS WOULD BE OF GREAT VALUE, BUT THE REGULATION OF HIV-1 IN MACROPHAGES DIFFERS FROM T CELLS. WE DEVELOPED A NOVEL HIGH-THROUGHPUT SCREEN FOR COMPOUNDS THAT CAN SILENCE ESTABLISHED INFECTION OF PRIMARY MACROPHAGES, AND IDENTIFIED AGENTS THAT DOWNREGULATE VIRUS EXPRESSION AND ALTER PROVIRUS EPIGENETIC PROFILES. THE SIGNIFICANCE OF THIS ASSAY IS THE POTENTIAL TO IDENTIFY NEW DRUGS THAT ACT IN THE UNIQUE MACROPHAGE ENVIRONMENT ON RELEVANT VIRAL STRAINS, WHICH MAY CONTRIBUTE TO ADJUNCTIVE TREATMENT FOR HIV-ASSOCIATED NEUROCOGNITIVE DISORDERS AND/OR PREVENT VIRAL REBOUND IN EFFORTS TO ACHIEVE ART-FREE REMISSION OR CURE. 2023 3 5849 36 SUBEROYLANILIDE HYDROXAMIC ACID (SAHA) AND CLADRIBINE SYNERGISTICALLY INDUCE APOPTOSIS IN NK-LGL LEUKAEMIA. NATURAL KILLER (NK) LARGE GRANULAR LYMPHOCYTE (LGL) LEUKAEMIA FEATURES A CLONAL PROLIFERATION OF CD3(-) NK CELLS THAT CAN BE CLASSIFIED INTO EITHER AGGRESSIVE OR CHRONIC CATEGORIES. THE NKL CELL LINE, DERIVED FROM AN AGGRESSIVE ASIAN NK CELL LEUKAEMIA, AND PATIENT SAMPLES FROM CHRONIC NK-LGL LEUKAEMIA WERE USED IN OUR STUDY TO PROBE FOR SYNERGISTIC EFFICACY OF THE EPIGENETIC DRUGS VORINOSTAT (SAHA) AND CLADRIBINE IN THIS DISEASE. WE DEMONSTRATE THAT HISTONE DEACETYLASES (HDACS) ARE OVER-EXPRESSED IN BOTH AGGRESSIVE AND CHRONIC NK LEUKAEMIA. ADMINISTRATION OF THE HDAC INHIBITOR SAHA REDUCES CLASS I AND II HDAC EXPRESSION AND ENHANCES HISTONE ACETYLATION IN LEUKAEMIC NK CELLS. IN VITRO COMBINATION TREATMENT WITH SAHA AND CLADRIBINE DOSE-DEPENDENTLY EXERTS SYNERGISTIC CYTOTOXIC AND APOPTOTIC EFFECTS ON LEUKAEMIC NK CELLS. EXPRESSION PROFILING OF APOPTOTIC REGULATORY GENES SUGGESTS THAT BOTH COMPOUNDS LED TO CASPASE-DEPENDENT APOPTOSIS THROUGH ACTIVATION OF INTRINSIC MITOCHONDRIAL AND EXTRINSIC DEATH RECEPTOR PATHWAYS. COLLECTIVELY, THESE DATA SHOW THAT COMBINED EPIGENETIC THERAPY, USING HDAC AND DNA METHYLTRANSFERASE INHIBITORS, MAY BE A PROMISING THERAPEUTIC APPROACH FOR NK-LGL LEUKAEMIA. 2015 4 439 35 ANTILEUKEMIC ACTIVITY OF VALPROIC ACID IN CHRONIC LYMPHOCYTIC LEUKEMIA B CELLS DEFINED BY MICROARRAY ANALYSIS. EPIGENETIC CODE MODIFICATIONS BY HISTONE DEACETYLASE INHIBITORS HAVE RECENTLY BEEN PROPOSED AS POTENTIAL NEW THERAPIES FOR HEMATOLOGICAL MALIGNANCIES. CHRONIC LYMPHOCYTIC LEUKEMIA (CLL) REMAINS INCURABLE DESPITE THE INTRODUCTION OF NEW TREATMENTS. CLL B CELLS ARE CHARACTERIZED BY AN APOPTOSIS DEFECT RATHER THAN EXCESSIVE PROLIFERATION, BUT PROLIFERATION CENTERS HAVE BEEN FOUND IN ORGANS SUCH AS THE BONE MARROW AND LYMPH NODES. IN THIS STUDY, WE ANALYZED GENE EXPRESSION MODIFICATIONS IN CLL B CELLS AFTER TREATMENT WITH VALPROIC ACID (VPA), A WELL-TOLERATED ANTI-EPILEPTIC DRUG WITH HDAC INHIBITORY ACTIVITY. CLL B CELLS OBTAINED FROM 14 PATIENTS WERE TREATED IN VITRO WITH A CONCENTRATION OF 1 MM VPA FOR 4 H. VPA EFFECTS ON GENE EXPRESSION WERE THEREAFTER STUDIED USING AFFYMETRIX TECHNOLOGY, AND SOME IDENTIFIED GENES WERE VALIDATED BY REAL-TIME PCR AND WESTERN BLOT. WE OBSERVED THAT VPA INDUCED APOPTOSIS BY DOWNREGULATING SEVERAL ANTI-APOPTOTIC GENES AND BY UPREGULATING PRO-APOPTOTIC GENES. FURTHERMORE, VPA SIGNIFICANTLY INCREASED CHEMOSENSITIVITY TO FLUDARABINE, FLAVOPIRIDOL, BORTEZOMIB, THALIDOMIDE AND LENALIDOMIDE. VPA INHIBITED THE PROLIFERATION OF CPG/IL2-STIMULATED CLL B CELLS AND MODULATED MANY CELL CYCLE MESSENGER RNAS. IN CONCLUSION, EXPOSURE OF CLL B CELLS TO VPA INDUCED APOPTOSIS, POTENTIATED CHEMOTHERAPEUTIC AGENT EFFECTS AND INHIBITED PROLIFERATION. THESE DATA STRONGLY SUGGEST THE USE OF VPA IN CLL TREATMENT, PARTICULARLY IN COMBINATION WITH ANTILEUKEMIA AGENTS. 2009 5 6523 17 TRANSCRIPTIONAL AND EPIGENETIC REGULATORY MECHANISMS AFFECTING HTLV-1 PROVIRUS. HUMAN T-CELL LEUKEMIA VIRUS TYPE 1 (HTLV-1) IS A RETROVIRUS ASSOCIATED WITH HUMAN DISEASES, SUCH AS ADULT T-CELL LEUKEMIA (ATL) AND HTLV-1-ASSOCIATED MYELOPATHY/TROPIC SPASTIC PARAPARESIS (HAM/TSP). AS A RETROVIRUS, ITS LIFE CYCLE INCLUDES A STEP WHERE HTLV-1 IS INTEGRATED INTO THE HOST GENOMIC DNA AND FORMS PROVIRAL DNA. IN THE CHRONIC PHASE OF THE INFECTION, HTLV?1 IS KNOWN TO PROLIFERATE AS A PROVIRUS VIA THE MITOTIC DIVISION OF THE INFECTED HOST CELLS. THERE ARE GENERALLY TENS OF THOUSANDS OF INFECTED CLONES WITHIN AN INFECTED INDIVIDUAL. THEY EXIST NOT ONLY IN PERIPHERAL BLOOD, BUT ALSO IN VARIOUS LYMPHOID ORGANS. VIRAL PROTEINS ENCODED IN HTLV-1 GENOME PLAY A ROLE IN THE PROLIFERATION AND SURVIVAL OF THE INFECTED CELLS. AS IS THE CASE WITH OTHER CHRONIC VIRAL INFECTIONS, HTLV-1 GENE EXPRESSION INDUCES THE ACTIVATION OF THE HOST IMMUNITY AGAINST THE VIRUS. THUS, THE TRANSCRIPTION FROM HTLV-1 PROVIRUS NEEDS TO BE CONTROLLED IN ORDER TO EVADE THE HOST IMMUNE SURVEILLANCE. THERE SHOULD BE A DYNAMIC AND COMPLEX REGULATION IN VIVO, WHERE AN EQUILIBRIUM BETWEEN VIRAL ANTIGEN EXPRESSION AND HOST IMMUNE SURVEILLANCE IS ACHIEVED. THE MECHANISMS REGULATING VIRAL GENE EXPRESSION FROM THE PROVIRUS ARE A KEY TO UNDERSTANDING THE PERSISTENT/LATENT INFECTION WITH HTLV-1 AND ITS PATHOGENESIS. IN THIS ARTICLE, WE WOULD LIKE TO REVIEW OUR CURRENT UNDERSTANDING ON THIS TOPIC. 2016 6 3877 24 KDM6A PROMOTES IMATINIB RESISTANCE THROUGH YY1-MEDIATED TRANSCRIPTIONAL UPREGULATION OF TRKA INDEPENDENTLY OF ITS DEMETHYLASE ACTIVITY IN CHRONIC MYELOGENOUS LEUKEMIA. RATIONALE: DESPITE LANDMARK THERAPY OF CHRONIC MYELOGENOUS LEUKEMIA (CML) WITH TYROSINE KINASE INHIBITORS (TKIS), DRUG RESISTANCE REMAINS PROBLEMATIC. CANCER PATHOGENESIS INVOLVES EPIGENETIC DYSREGULATION AND IN PARTICULAR, HISTONE LYSINE DEMETHYLASES (KDMS) HAVE BEEN IMPLICATED IN TKI RESISTANCE. WE SOUGHT TO IDENTIFY KDMS WITH ALTERED EXPRESSION IN CML AND DEFINE THEIR CONTRIBUTION TO IMATINIB RESISTANCE. METHODS: BIOINFORMATICS SCREENING COMPARED KDM EXPRESSION IN CML VERSUS NORMAL BONE MARROW WITH SHRNA KNOCKDOWN AND FLOW CYTOMETRY USED TO MEASURE EFFECTS ON IMATINIB-INDUCED APOPTOSIS IN K562 CELLS. TRANSCRIPTOMIC ANALYSES WERE PERFORMED AGAINST KDM6A CRISPR KNOCKOUT/SHRNA KNOCKDOWN K562 CELLS ALONG WITH GENE RESCUE EXPERIMENTS USING WILDTYPE AND MUTANT DEMETHYLASE-DEAD KDM6A CONSTRUCTS. CO-IMMUNOPRECIPITATION, LUCIFERASE REPORTER AND CHIP WERE EMPLOYED TO ELUCIDATE MECHANISMS OF KDM6A-DEPENDENT RESISTANCE. RESULTS: AMONGST FIVE KDMS UPREGULATED IN CML, ONLY KDM6A DEPLETION SENSITIZED CML CELLS TO IMATINIB-INDUCED APOPTOSIS. RE-INTRODUCTION OF DEMETHYLASE-DEAD KDM6A AS WELL AS WILD-TYPE KDM6A RESTORED IMATINIB RESISTANCE. RNA-SEQ IDENTIFIED NTRK1 GENE DOWNREGULATION AFTER DEPLETION OF KDM6A. MOREOVER, NTRK1 EXPRESSION POSITIVELY CORRELATED WITH KDM6A IN A SUBSET OF CLINICAL CML SAMPLES AND KDM6A KNOCKDOWN IN FRESH CML ISOLATES DECREASED NTRK1 ENCODED PROTEIN (TRKA) EXPRESSION. MECHANISTICALLY, KDM6A WAS RECRUITED TO THE NTRK1 PROMOTER BY THE TRANSCRIPTION FACTOR YY1 WITH SUBSEQUENT TRKA UPREGULATION ACTIVATING DOWN-STREAM SURVIVAL PATHWAYS TO INVOKE IMATINIB RESISTANCE. CONCLUSION: CONTRARY TO ITS REPORTED ROLE AS A TUMOR SUPPRESSOR AND INDEPENDENT OF ITS DEMETHYLASE FUNCTION, KDM6A PROMOTES IMATINIB-RESISTANCE IN CML CELLS. THE IDENTIFICATION OF THE KDM6A/YY1/TRKA AXIS AS A NOVEL IMATINIB-RESISTANCE MECHANISM REPRESENTS AN UNEXPLORED AVENUE TO OVERCOME TKI RESISTANCE IN CML. 2021 7 1333 32 DEREGULATION AND EPIGENETIC MODIFICATION OF BCL2-FAMILY GENES CAUSE RESISTANCE TO VENETOCLAX IN HEMATOLOGIC MALIGNANCIES. THE BCL2 INHIBITOR VENETOCLAX HAS BEEN APPROVED TO TREAT DIFFERENT HEMATOLOGICAL MALIGNANCIES. BECAUSE THERE IS NO COMMON GENETIC ALTERATION CAUSING RESISTANCE TO VENETOCLAX IN CHRONIC LYMPHOCYTIC LEUKEMIA (CLL) AND B-CELL LYMPHOMA, WE ASKED IF EPIGENETIC EVENTS MIGHT BE INVOLVED IN VENETOCLAX RESISTANCE. THEREFORE, WE EMPLOYED WHOLE-EXOME SEQUENCING, METHYLATED DNA IMMUNOPRECIPITATION SEQUENCING, AND GENOME-WIDE CLUSTERED REGULARLY INTERSPACED SHORT PALINDROMIC REPEATS (CRISPR)/CRISPR-ASSOCIATED PROTEIN 9 SCREENING TO INVESTIGATE VENETOCLAX RESISTANCE IN AGGRESSIVE LYMPHOMA AND HIGH-RISK CLL PATIENTS. WE IDENTIFIED A REGULATORY CPG ISLAND WITHIN THE PUMA PROMOTER THAT IS METHYLATED UPON VENETOCLAX TREATMENT, MEDIATING PUMA DOWNREGULATION ON TRANSCRIPT AND PROTEIN LEVEL. PUMA EXPRESSION AND SENSITIVITY TOWARD VENETOCLAX CAN BE RESTORED BY INHIBITION OF METHYLTRANSFERASES. WE CAN DEMONSTRATE THAT LOSS OF PUMA RESULTS IN METABOLIC REPROGRAMMING WITH HIGHER OXIDATIVE PHOSPHORYLATION AND ADENOSINE TRIPHOSPHATE PRODUCTION, RESEMBLING THE METABOLIC PHENOTYPE THAT IS SEEN UPON VENETOCLAX RESISTANCE. ALTHOUGH PUMA LOSS IS SPECIFIC FOR ACQUIRED VENETOCLAX RESISTANCE BUT NOT FOR ACQUIRED MCL1 RESISTANCE AND IS NOT SEEN IN CLL PATIENTS AFTER CHEMOTHERAPY-RESISTANCE, BAX IS ESSENTIAL FOR SENSITIVITY TOWARD BOTH VENETOCLAX AND MCL1 INHIBITION. AS WE FOUND LOSS OF BAX IN RICHTER'S SYNDROME PATIENTS AFTER VENETOCLAX FAILURE, WE DEFINED BAX-MEDIATED APOPTOSIS TO BE CRITICAL FOR DRUG RESISTANCE BUT NOT FOR DISEASE PROGRESSION OF CLL INTO AGGRESSIVE DIFFUSE LARGE B-CELL LYMPHOMA IN VIVO. A COMPOUND SCREEN REVEALED TRAIL-MEDIATED APOPTOSIS AS A TARGET TO OVERCOME BAX DEFICIENCY. FURTHERMORE, ANTIBODY OR CAR T CELLS ELIMINATED VENETOCLAX RESISTANT LYMPHOMA CELLS, PAVING A CLINICALLY APPLICABLE WAY TO OVERCOME VENETOCLAX RESISTANCE. 2022 8 3380 23 HIV-1 INFECTION OF GENETICALLY ENGINEERED IPSC-DERIVED CENTRAL NERVOUS SYSTEM-ENGRAFTED MICROGLIA IN A HUMANIZED MOUSE MODEL. THE CENTRAL NERVOUS SYSTEM (CNS) IS A MAJOR HUMAN IMMUNODEFICIENCY VIRUS TYPE 1 RESERVOIR. MICROGLIA ARE THE PRIMARY TARGET CELL OF HIV-1 INFECTION IN THE CNS. CURRENT MODELS HAVE NOT ALLOWED THE PRECISE MOLECULAR PATHWAYS OF ACUTE AND CHRONIC CNS MICROGLIAL INFECTION TO BE TESTED WITH IN VIVO GENETIC METHODS. HERE, WE DESCRIBE A NOVEL HUMANIZED MOUSE MODEL UTILIZING HUMAN-INDUCED PLURIPOTENT STEM CELL-DERIVED MICROGLIA TO XENOGRAFT INTO MURINE HOSTS. THESE MICE ARE ADDITIONALLY ENGRAFTED WITH HUMAN PERIPHERAL BLOOD MONONUCLEAR CELLS THAT SERVED AS A MEDIUM TO ESTABLISH A PERIPHERAL INFECTION THAT THEN SPREAD TO THE CNS MICROGLIA XENOGRAFT, MODELING A TRANS-BLOOD-BRAIN BARRIER ROUTE OF ACUTE CNS HIV-1 INFECTION WITH HUMAN TARGET CELLS. THE APPROACH IS COMPATIBLE WITH IPSC GENETIC ENGINEERING, INCLUDING INSERTING TARGETED TRANSGENIC REPORTER CASSETTES TO TRACK THE XENOGRAFTED HUMAN CELLS, ENABLING THE TESTING OF NOVEL TREATMENT AND VIRAL TRACKING STRATEGIES IN A COMPARATIVELY SIMPLE AND COST-EFFECTIVE WAY VIVO MODEL FOR NEUROHIV. IMPORTANCE: OUR MOUSE MODEL IS A POWERFUL TOOL FOR INVESTIGATING THE GENETIC MECHANISMS GOVERNING CNS HIV-1 INFECTION AND LATENCY IN THE CNS AT A SINGLE-CELL LEVEL. A MAJOR ADVANTAGE OF OUR MODEL IS THAT IT USES IPSC-DERIVED MICROGLIA, WHICH ENABLES HUMAN GENETICS, INCLUDING GENE FUNCTION AND THERAPEUTIC GENE MANIPULATION, TO BE EXPLORED IN VIVO , WHICH IS MORE CHALLENGING TO STUDY WITH CURRENT HEMATOPOIETIC STEM CELL-BASED MODELS FOR NEUROHIV. OUR TRANSGENIC TRACING OF XENOGRAFTED HUMAN CELLS WILL PROVIDE A QUANTITATIVE MEDIUM TO DEVELOP NEW MOLECULAR AND EPIGENETIC STRATEGIES FOR REDUCING THE HIV-1 LATENT RESERVOIR AND TO TEST THE IMPACT OF THERAPEUTIC INFLAMMATION-TARGETING DRUG INTERVENTIONS ON CNS HIV-1 LATENCY. 2023 9 826 27 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 10 709 31 C-MYC ONCOPROTEIN DICTATES TRANSCRIPTIONAL PROFILES OF ATP-BINDING CASSETTE TRANSPORTER GENES IN CHRONIC MYELOGENOUS LEUKEMIA CD34+ HEMATOPOIETIC PROGENITOR CELLS. RESISTANCE TO CHEMOTHERAPEUTIC AGENTS REMAINS ONE OF THE MAJOR IMPEDIMENTS TO A SUCCESSFUL TREATMENT OF CHRONIC MYELOID LEUKEMIA (CML). MISREGULATION OF THE ACTIVITY OF A SPECIFIC GROUP OF ATP-BINDING CASSETTE TRANSPORTERS (ABC) IS RESPONSIBLE FOR REDUCING THE INTRACELLULAR CONCENTRATION OF DRUGS IN LEUKEMIC CELLS. MOREOVER, A CONSISTENT BODY OF EVIDENCE ALSO SUGGESTS THAT ABC TRANSPORTERS PLAY A ROLE IN CANCER PROGRESSION BEYOND THE EFFLUX OF CYTOTOXIC DRUGS. DESPITE A LARGE NUMBER OF STUDIES THAT INVESTIGATED THE FUNCTION OF THE ABC TRANSPORTERS, LITTLE IS KNOWN ABOUT THE TRANSCRIPTIONAL REGULATION OF THE ABC GENES. HERE, WE PRESENT DATA SHOWING THAT THE ONCOPROTEIN C-MYC IS A DIRECT TRANSCRIPTIONAL REGULATOR OF A LARGE SET OF ABC TRANSPORTERS IN CML. FURTHERMORE, MOLECULAR ANALYSIS CARRIED OUT IN CD34+ HEMATOPOIETIC CELL PRECURSORS OF 21 CML PATIENTS REVEALS THAT THE OVEREXPRESSION OF ABC TRANSPORTERS DRIVEN BY C-MYC IS A PECULIAR CHARACTERISTIC OF THE CD34+ POPULATION IN CML AND WAS NOT FOUND EITHER IN THE POPULATION OF MONONUCLEAR CELLS FROM WHICH THEY HAD BEEN PURIFIED NOR IN CD34+ CELLS ISOLATED FROM HEALTHY DONORS. FINALLY, WE DESCRIBE HOW THE METHYLATION STATE OF CPG ISLANDS MAY REGULATE THE ACCESS OF C-MYC TO ABCG2 GENE PROMOTER, A WELL-STUDIED GENE ASSOCIATED WITH MULTIDRUG RESISTANCE IN CML, HENCE, AFFECTING ITS EXPRESSION. TAKEN TOGETHER, OUR FINDINGS SUPPORT A MODEL IN WHICH C-MYC-DRIVEN TRANSCRIPTIONAL EVENTS, COMBINED WITH EPIGENETIC MECHANISMS, DIRECT AND REGULATE THE EXPRESSION OF ABC GENES WITH POSSIBLE IMPLICATIONS IN TUMOR MALIGNANCY AND DRUG EFFLUX IN CML. 2011 11 2143 27 EPIGENETIC LANDSCAPE OF HIV-1 INFECTION IN PRIMARY HUMAN MACROPHAGE. HUMAN IMMUNODEFICIENCY VIRUS (HIV)-INFECTED MACROPHAGES ARE LONG-LIVED CELLS THAT SUSTAIN PERSISTENT VIRUS EXPRESSION, WHICH IS BOTH A BARRIER TO VIRAL ERADICATION AND CONTRIBUTOR TO NEUROLOGICAL COMPLICATIONS IN PATIENTS DESPITE ANTIRETROVIRAL THERAPY (ART). TO BETTER UNDERSTAND THE REGULATION OF HIV-1 IN MACROPHAGES, WE COMPARED HIV-INFECTED PRIMARY HUMAN MONOCYTE-DERIVED MACROPHAGES (MDM) TO ACUTELY INFECTED PRIMARY CD4 T CELLS AND JURKAT CELLS LATENTLY INFECTED WITH HIV (JLAT 8.4). HIV GENOMES IN MDM WERE ACTIVELY TRANSCRIBED DESPITE ENRICHMENT WITH HETEROCHROMATIN-ASSOCIATED H3K9ME3 ACROSS THE COMPLETE HIV GENOME IN COMBINATION WITH ELEVATED ACTIVATION MARKS OF H3K9AC AND H3K27AC AT THE LONG TERMINAL REPEAT (LTR). MACROPHAGE PATTERNS CONTRASTED WITH JLAT CELLS, WHICH SHOWED CONVENTIONAL BIVALENT H3K4ME3/H3K27ME3, AND ACUTELY INFECTED CD4 T CELLS, WHICH SHOWED AN INTERMEDIATE EPIGENOTYPE. 5'-METHYLCYTOSINE (5MC) WAS ENRICHED ACROSS THE HIV GENOME IN LATENTLY INFECTED JLAT CELLS, WHILE 5'-HYDROXYMETHYLCYTOSINE (5HMC) WAS ENRICHED IN CD4 CELLS AND MDMS. HIV INFECTION INDUCED MULTINUCLEATION OF MDMS ALONG WITH DNA DAMAGE-ASSOCIATED P53 PHOSPHORYLATION, AS WELL AS LOSS OF TET2 AND THE NUCLEAR REDISTRIBUTION OF 5-HYDOXYMETHYLATION. TAKEN TOGETHER, OUR FINDINGS SUGGEST THAT HIV INDUCES A UNIQUE MACROPHAGE NUCLEAR AND TRANSCRIPTIONAL PROFILE, AND VIRAL GENOMES ARE MAINTAINED IN A NONCANONICAL BIVALENT EPIGENETIC STATE. IMPORTANCE MACROPHAGES SERVE AS A RESERVOIR FOR LONG-TERM PERSISTENCE AND CHRONIC PRODUCTION OF HIV. WE FOUND AN ATYPICAL EPIGENETIC CONTROL OF HIV IN MACROPHAGES MARKED BY HETEROCHROMATIC H3K9ME3 DESPITE ACTIVE VIRAL TRANSCRIPTION. HIV INFECTION INDUCED CHANGES IN MACROPHAGE NUCLEAR MORPHOLOGY AND EPIGENETIC REGULATORY FACTORS. THESE FINDINGS MAY IDENTIFY NEW MECHANISMS TO CONTROL CHRONIC HIV EXPRESSION IN INFECTED MACROPHAGES. 2022 12 6425 25 THE TRANSCRIPTION FACTOR REST UP-REGULATES TYROSINE HYDROXYLASE AND ANTIAPOPTOTIC GENES AND PROTECTS DOPAMINERGIC NEURONS AGAINST MANGANESE TOXICITY. DOPAMINERGIC FUNCTIONS ARE IMPORTANT FOR VARIOUS BIOLOGICAL ACTIVITIES, AND THEIR IMPAIRMENT LEADS TO NEURODEGENERATION, A HALLMARK OF PARKINSON'S DISEASE (PD). CHRONIC MANGANESE (MN) EXPOSURE CAUSES THE NEUROLOGICAL DISORDER MANGANISM, PRESENTING SYMPTOMS SIMILAR TO THOSE OF PD. EMERGING EVIDENCE HAS LINKED THE TRANSCRIPTION FACTOR RE1-SILENCING TRANSCRIPTION FACTOR (REST) TO PD AND ALSO ALZHEIMER'S DISEASE. BUT REST'S ROLE IN DOPAMINERGIC NEURONS IS UNCLEAR. HERE, WE INVESTIGATED WHETHER REST PROTECTS DOPAMINERGIC NEURONS AGAINST MN-INDUCED TOXICITY AND ENHANCES EXPRESSION OF THE DOPAMINE-SYNTHESIZING ENZYME TYROSINE HYDROXYLASE (TH). WE REPORT THAT REST BINDS TO RE1 CONSENSUS SITES IN THE TH GENE PROMOTER, STIMULATES TH TRANSCRIPTION, AND INCREASES TH MRNA AND PROTEIN LEVELS IN DOPAMINERGIC CELLS. REST BINDING TO THE TH PROMOTER RECRUITED THE EPIGENETIC MODIFIER CAMP-RESPONSE ELEMENT-BINDING PROTEIN-BINDING PROTEIN/P300 AND THEREBY UP-REGULATED TH EXPRESSION. REST RELIEVED MN-INDUCED REPRESSION OF TH PROMOTER ACTIVITY, MRNA, AND PROTEIN LEVELS AND ALSO REDUCED MN-INDUCED OXIDATIVE STRESS, INFLAMMATION, AND APOPTOSIS IN DOPAMINERGIC NEURONS. REST REDUCED MN-INDUCED PROINFLAMMATORY CYTOKINES, INCLUDING TUMOR NECROSIS FACTOR ALPHA, INTERLEUKIN 1BETA (IL-1BETA), IL-6, AND INTERFERON GAMMA. MOREOVER, REST INHIBITED THE MN-INDUCED PROAPOPTOTIC PROTEINS BCL-2-ASSOCIATED X PROTEIN (BAX) AND DEATH-ASSOCIATED PROTEIN 6 (DAXX) AND ATTENUATED AN MN-INDUCED DECREASE IN THE ANTIAPOPTOTIC PROTEINS BCL-2 AND BCL-XL. REST ALSO ENHANCED THE EXPRESSION OF ANTIOXIDANT PROTEINS, INCLUDING CATALASE, NF-E2-RELATED FACTOR 2 (NRF2), AND HEME OXYGENASE 1 (HO-1). OUR FINDINGS INDICATE THAT REST ACTIVATES TH EXPRESSION AND THEREBY PROTECTS NEURONS AGAINST MN-INDUCED TOXICITY AND NEUROLOGICAL DISORDERS ASSOCIATED WITH DOPAMINERGIC NEURODEGENERATION. 2020 13 2326 32 EPIGENETIC REGULATION OF HOTAIR IN ADVANCED CHRONIC MYELOID LEUKEMIA. PURPOSE: CHRONIC MYELOID LEUKEMIA (CML) ACCOUNTS FOR ~10% OF LEUKEMIA CASES, AND ITS PROGRESSION INVOLVES EPIGENETIC GENE REGULATION. THIS STUDY INVESTIGATED EPIGENETIC REGULATION OF HOTAIR AND ITS TARGET MICRORNA, MIR-143, IN ADVANCED CML. PATIENTS AND METHODS: WE FIRST ISOLATED BONE MARROW MONONUCLEAR CELLS FROM 70 PATIENTS WITH DIFFERENT PHASES OF CML AND FROM HEALTHY DONORS AS NORMAL CONTROL; WE ALSO CULTURED K562 AND KCL22 CELLS, TREATED WITH DEMETHYLATION DRUG; MTT ASSAY, FLOW CYTOMETRY, QUANTITATIVE REAL-TIME POLYMERASE CHAIN REACTION (QPCR), METHYLATION-SPECIFIC POLYMERASE CHAIN REACTION (MSP), WESTERN BLOT, LUCIFERASE ASSAY, RNA PULL-DOWN ASSAY AND RNA-BINDING PROTEIN IMMUNOPRECIPITATION (RIP) ASSAY WERE PERFORMED. RESULT: AS MEASURED BY QPCR, HOTAIR EXPRESSION IN K562 CELLS, KCL22 CELLS, AND SAMPLES FROM CASES OF ADVANCED-STAGE CML INCREASED WITH LEVELS OF SEVERAL DNA METHYLTRANSFERASES AND HISTONE DEACETYLATES, INCLUDING DNMT1, DNMT3A, HDAC1, EZH2, AND LSD1, AND MIR-143 LEVELS WERE DECREASED AND HOTAIR LEVELS WERE INCREASED. TREATMENT WITH 5-AZACYTIDINE, A DNA METHYLATION INHIBITOR, DECREASED DNMT1, DNMT3A, HDAC1, EZH2, LSD1 MRNA, PROTEIN LEVELS, AND HOTAIR MRNA LEVELS BUT INCREASED MIR-143 LEVELS. HOTAIR KNOCKDOWN AND MIR-143 OVEREXPRESSION BOTH INHIBITED PROLIFERATION AND PROMOTED APOPTOSIS IN KCL22 AND K562 CELLS THROUGH THE PI3K/AKT PATHWAY. RNA PULL-DOWN, MASS SPECTROMETRY, AND RIP ASSAYS SHOWED THAT HOTAIR INTERACTED WITH EZH2 AND LSD1. A DUAL-LUCIFERASE ASSAY DEMONSTRATED THAT HOTAIR INTERACTED WITH MIR-143. CONCLUSION: OUR FINDINGS DEMONSTRATE THE KEY EPIGENETIC INTERACTIONS OF HOTAIR RELATED TO CML PROGRESSION AND SUGGEST HOTAIR AS A POTENTIAL THERAPEUTIC TARGET FOR ADVANCED CML. FURTHERMORE, OUR RESULTS SUPPORT THE USE OF DEMETHYLATION DRUGS AS A CML TREATMENT STRATEGY. 2018 14 5716 26 SIRT6 PROTECTS VASCULAR SMOOTH MUSCLE CELLS FROM OSTEOGENIC TRANSDIFFERENTIATION VIA RUNX2 IN CHRONIC KIDNEY DISEASE. VASCULAR CALCIFICATION (VC) IS REGARDED AS AN IMPORTANT PATHOLOGICAL CHANGE LACKING EFFECTIVE TREATMENT AND ASSOCIATED WITH HIGH MORTALITY. SIRTUIN 6 (SIRT6) IS A MEMBER OF THE SIRTUIN FAMILY, A CLASS III HISTONE DEACETYLASE AND A KEY EPIGENETIC REGULATOR. SIRT6 HAS A PROTECTIVE ROLE IN PATIENTS WITH CHRONIC KIDNEY DISEASE (CKD). HOWEVER, THE EXACT ROLE AND MOLECULAR MECHANISM OF SIRT6 IN VC IN PATIENTS WITH CKD REMAIN UNCLEAR. HERE, WE DEMONSTRATED THAT SIRT6 WAS MARKEDLY DOWNREGULATED IN PERIPHERAL BLOOD MONONUCLEAR CELLS (PBMCS) AND IN THE RADIAL ARTERY TISSUE OF PATIENTS WITH CKD WITH VC. SIRT6-TRANSGENIC (SIRT6-TG) MICE SHOWED ALLEVIATED VC, WHILE VASCULAR SMOOTH MUSCLE CELL-SPECIFIC (VSMC-SPECIFIC) SIRT6 KNOCKED-DOWN MICE SHOWED SEVERE VC IN CKD. SIRT6 SUPPRESSED THE OSTEOGENIC TRANSDIFFERENTIATION OF VSMCS VIA REGULATION OF RUNT-RELATED TRANSCRIPTION FACTOR 2 (RUNX2). COIMMUNOPRECIPITATION (CO-IP) AND IMMUNOPRECIPITATION (IP) ASSAYS CONFIRMED THAT SIRT6 BOUND TO RUNX2. MOREOVER, RUNX2 WAS DEACETYLATED BY SIRT6 AND FURTHER PROMOTED NUCLEAR EXPORT VIA EXPORTIN 1 (XPO1), WHICH IN TURN CAUSED DEGRADATION OF RUNX2 THROUGH THE UBIQUITIN-PROTEASOME SYSTEM. THESE RESULTS DEMONSTRATED THAT SIRT6 PREVENTED VC BY SUPPRESSING THE OSTEOGENIC TRANSDIFFERENTIATION OF VSMCS, AND AS SUCH TARGETING SIRT6 MAY BE AN APPEALING THERAPEUTIC TARGET FOR VC IN CKD. 2022 15 5703 36 SINGLE-CELL MULTIOMICS REVEAL THE SCALE OF MULTILAYERED ADAPTATIONS ENABLING CLL RELAPSE DURING VENETOCLAX THERAPY. VENETOCLAX (VEN) INHIBITS THE PROSURVIVAL PROTEIN BCL2 TO INDUCE APOPTOSIS AND IS A STANDARD THERAPY FOR CHRONIC LYMPHOCYTIC LEUKEMIA (CLL), DELIVERING HIGH COMPLETE REMISSION RATES AND PROLONGED PROGRESSION-FREE SURVIVAL IN RELAPSED CLL BUT WITH EVENTUAL LOSS OF EFFICACY. A SPECTRUM OF SUBCLONAL GENETIC CHANGES ASSOCIATED WITH VEN RESISTANCE HAS NOW BEEN DESCRIBED. TO FULLY UNDERSTAND CLINICAL RESISTANCE TO VEN, WE COMBINED SINGLE-CELL SHORT- AND LONG-READ RNA-SEQUENCING TO REVEAL THE PREVIOUSLY UNAPPRECIATED SCALE OF GENETIC AND EPIGENETIC CHANGES UNDERPINNING ACQUIRED VEN RESISTANCE. THESE APPEAR TO BE MULTILAYERED. ONE LAYER COMPRISES CHANGES IN THE BCL2 FAMILY OF APOPTOSIS REGULATORS, ESPECIALLY THE PROSURVIVAL FAMILY MEMBERS. THIS INCLUDES PREVIOUSLY DESCRIBED MUTATIONS IN BCL2 AND AMPLIFICATION OF THE MCL1 GENE BUT IS HETEROGENEOUS ACROSS AND WITHIN INDIVIDUAL PATIENT LEUKEMIAS. CHANGES IN THE PROAPOPTOTIC GENES ARE NOTABLY UNCOMMON, EXCEPT FOR SINGLE CASES WITH SUBCLONAL LOSSES OF BAX OR NOXA. MUCH MORE PROMINENT WAS UNIVERSAL MCL1 GENE UPREGULATION. THIS WAS DRIVEN BY AN OVERLYING LAYER OF EMERGENT NF-KAPPAB (NUCLEAR FACTOR KAPPA B) ACTIVATION, WHICH PERSISTED IN CIRCULATING CELLS DURING VEN THERAPY. WE DISCOVERED THAT MCL1 COULD BE A DIRECT TRANSCRIPTIONAL TARGET OF NF-KAPPAB. BOTH THE SWITCH TO ALTERNATIVE PROSURVIVAL FACTORS AND NF-KAPPAB ACTIVATION LARGELY DISSIPATE FOLLOWING VEN DISCONTINUATION. OUR STUDIES REVEAL THE EXTENT OF PLASTICITY OF CLL CELLS IN THEIR ABILITY TO EVADE VEN-INDUCED APOPTOSIS. IMPORTANTLY, THESE FINDINGS PINPOINT NEW APPROACHES TO CIRCUMVENT VEN RESISTANCE AND PROVIDE A SPECIFIC BIOLOGICAL JUSTIFICATION FOR THE STRATEGY OF VEN DISCONTINUATION ONCE A MAXIMAL RESPONSE IS ACHIEVED RATHER THAN MAINTAINING LONG-TERM SELECTIVE PRESSURE WITH THE DRUG. 2022 16 6171 27 THE HDAC INHIBITOR VALPROATE INDUCES A BIVALENT STATUS OF THE CD20 PROMOTER IN CLL PATIENTS SUGGESTING DISTINCT EPIGENETIC REGULATION OF CD20 EXPRESSION IN CLL IN VIVO. TREATMENT WITH ANTI-CD20 ANTIBODIES IS ONLY MODERATELY EFFICIENT IN CHRONIC LYMPHOCYTIC LEUKEMIA (CLL), A FEATURE WHICH HAS BEEN EXPLAINED BY THE INHERENTLY LOW CD20 EXPRESSION IN CLL. IT HAS BEEN SHOWN THAT CD20 IS EPIGENETICALLY REGULATED AND THAT HISTONE DEACETYLASE INHIBITORS (HDACIS) CAN INCREASE CD20 EXPRESSION IN VITRO IN CLL. TO ASSESS WHETHER HDACIS CAN UPREGULATE CD20 ALSO IN VIVO IN CLL, THE HDACI VALPROATE WAS GIVEN TO THREE DEL13Q/NOTCH1WT CLL PATIENTS AND CD20 LEVELS WERE ANALYSED (THE PREVAIL STUDY). VALPROATE TREATMENT RESULTED IN EXPECTED GLOBAL ACTIVATING HISTONE MODIFICATIONS SUGGESTING HDAC INHIBITORY EFFECTS. HOWEVER, ALTHOUGH VALPROATE INDUCED EXPRESSION OF CD20 MRNA AND PROTEIN IN THE DEL13Q/NOTCH1WT I83-E95 CLL CELL LINE, NO SUCH EFFECTS WERE OBSERVED IN THE PATIENTS STUDIED. IN CONTRAST TO THE CELL LINE, IN PATIENTS VALPROATE TREATMENT RESULTED IN TRANSIENT RECRUITMENT OF THE TRANSCRIPTIONAL REPRESSOR EZH2 TO THE CD20 PROMOTER, CORRELATING TO AN INCREASE OF THE REPRESSIVE HISTONE MARK H3K27ME3. THIS SUGGESTS THAT VALPROATE-MEDIATED INDUCTION OF CD20 MAY BE HAMPERED BY EZH2 MEDIATED H3K27ME3 IN VIVO IN CLL. MOREOVER, VALPROATE TREATMENT RESULTED IN INDUCTION OF EZH2 AND GLOBAL H3K27ME3 IN PATIENT CELLS, SUGGESTING TRANSCRIPTIONALLY REPRESSIVE EFFECTS OF VALPROATE IN CLL. OUR RESULTS SUGGEST NEW IN VIVO MECHANISMS OF HDACIS WHICH MAY HAVE IMPLICATIONS ON THE DESIGN OF FUTURE CLINICAL TRIALS IN B-CELL MALIGNANCIES. 2017 17 1287 26 DECITABINE-INDUCED APOPTOSIS IS DERIVED BY PUMA AND NOXA INDUCTION IN CHRONIC MYELOID LEUKEMIA CELL LINE AS WELL AS IN PBL AND IS POTENTIATED BY SAHA. RESTORATION OF CELLULAR APOPTOTIC PATHWAYS PLAYS A CRUCIAL ROLE IN CANCER THERAPY STRATEGIES. IN A BROAD SPECTRUM OF ANTICANCER DRUGS, EPIGENETIC EFFECTORS ARE IN THE CENTER OF INTEREST MOSTLY BECAUSE OF POTENTIAL REVERSIBILITY OF THEIR ACTION. METHYLATION STATUS OF THE CELLS IS INFLUENCED BY METHYLTRANSFERASE INHIBITOR 2-DEOXY-5'-AZACYTIDINE (DECITABINE, DAC), BUT HIGHER CONCENTRATIONS OF THIS AGENT CAUSE A DNA-DAMAGE. IN OUR STUDY, TUMOR SUPRESSOR P53-APOPTOTIC PATHWAY WAS ACTIVATED IN DECITABINE-INDUCED CELL DEATH. EXPRESSION OF P53-INDUCIBLE BH3-ONLY APOPTOTIC PROTEINS PUMA AND NOXA WAS ELEVATED AND LARGE ACTIVATION OF EXECUTIVE CASPASES WAS OBSERVED. THE EXTENT OF ACETYLATION IN THE CELL IS AFFECTED BY HISTONEDEACETYLASE INHIBITOR SUBEROYLANILIDE HYDROXAMIC ACID (SAHA). COMBINATION OF SAHA WITH DECITABINE BROUGHT SYNERGISTIC EFFECT ON APOPTOSIS TRIGGERING IN CML-T1 CELL LINE, BUT APOPTOSIS AS WELL AS NECROSIS OCCURRED ALSO IN NORMAL PERIPHERAL BLOOD LYMPHOCYTES. THEREFORE, PROMISING POTENTIAL OF SUCH COMBINED THERAPY CALLS FOR MORE DETAILED INVESTIGATION OF UNWANTED EFFECTS IN NORMAL CELLS. 2011 18 3202 29 HDAC2 REGULATES ATYPICAL ANTIPSYCHOTIC RESPONSES THROUGH THE MODULATION OF MGLU2 PROMOTER ACTIVITY. HISTONE DEACETYLASES (HDACS) COMPACT CHROMATIN STRUCTURE AND REPRESS GENE TRANSCRIPTION. IN SCHIZOPHRENIA, CLINICAL STUDIES DEMONSTRATE THAT HDAC INHIBITORS ARE EFFICACIOUS WHEN GIVEN IN COMBINATION WITH ATYPICAL ANTIPSYCHOTICS. HOWEVER, THE MOLECULAR MECHANISM THAT INTEGRATES A BETTER RESPONSE TO ANTIPSYCHOTICS WITH CHANGES IN CHROMATIN STRUCTURE REMAINS UNKNOWN. HERE WE FOUND THAT CHRONIC ATYPICAL ANTIPSYCHOTICS DOWNREGULATED THE TRANSCRIPTION OF METABOTROPIC GLUTAMATE 2 RECEPTOR (MGLU2, ALSO KNOWN AS GRM2), AN EFFECT THAT WAS ASSOCIATED WITH DECREASED HISTONE ACETYLATION AT ITS PROMOTER IN MOUSE AND HUMAN FRONTAL CORTEX. THIS EPIGENETIC CHANGE OCCURRED IN CONCERT WITH A SEROTONIN 5-HT(2A) RECEPTOR-DEPENDENT UPREGULATION AND INCREASED BINDING OF HDAC2 TO THE MGLU2 PROMOTER. VIRALLY MEDIATED OVEREXPRESSION OF HDAC2 IN FRONTAL CORTEX DECREASED MGLU2 TRANSCRIPTION AND ITS ELECTROPHYSIOLOGICAL PROPERTIES, THEREBY INCREASING PSYCHOSIS-LIKE BEHAVIOR. CONVERSELY, HDAC INHIBITORS PREVENTED THE REPRESSIVE HISTONE MODIFICATIONS INDUCED AT THE MGLU2 PROMOTER BY ATYPICAL ANTIPSYCHOTICS, AND AUGMENTED THEIR THERAPEUTIC-LIKE EFFECTS. THESE OBSERVATIONS SUPPORT THE VIEW OF HDAC2 AS A PROMISING NEW TARGET FOR SCHIZOPHRENIA TREATMENT. 2012 19 3191 25 HDAC AND HAT INHIBITORS DIFFERENTLY AFFECT ANALGESIA MEDIATED BY GROUP II METABOTROPIC GLUTAMATE RECEPTORS. BACKGROUND: HISTONE DEACETYLASES (HDACS) AND HISTONE ACETYLTRANSFERASES (HATS) ARE KEY PLAYERS IN EPIGENETIC REGULATION OF GENE EXPRESSION. ANALGESIC ACTIVITY BY HDAC INHIBITORS HAS BEEN REPORTED IN DIFFERENT PAIN MODELS INCLUDING INFLAMMATORY AND NEUROPATHIC PAIN. THESE DRUGS INTERFERE WITH GENE EXPRESSION THROUGH DIFFERENT MECHANISMS INCLUDING CHROMATIN REMODELING AND/OR ACTIVATION OF TRANSCRIPTION FACTORS. AMONG OTHER TARGETS, HDAC INHIBITORS REGULATE METABOTROPIC GLUTAMATE RECEPTORS TYPE 2 (MGLU2) EXPRESSION IN CENTRAL AND PERIPHERAL CENTRAL NERVOUS SYSTEM. HOWEVER WHETHER INHIBITION OF HAT ACTIVITY ALSO REGULATES MGLU2 EXPRESSION HAS NOT BEEN REPORTED. FINDINGS: HERE WE REPORT THAT CURCUMIN (CUR), A NATURALLY OCCURRING COMPOUND ENDOWED WITH P300/CREB-BINDING PROTEIN HAT INHIBITORY ACTIVITY, IS ABLE TO INDUCE A DRASTIC DOWN-REGULATION OF THE MGLU2 RECEPTOR IN THE MOUSE SPINAL CORD AFTER SYSTEMIC ADMINISTRATION TOGETHER WITH A MARKED HYPOACETYLATION OF HISTONES H3 AND H4 IN DORSAL ROOT GANGLIA (DRG). FURTHERMORE, THE ANALGESIC ACTIVITY OF THE MGLU2/3 AGONIST, LY379268 IS LOST AFTER A 3-DAY TREATMENT WITH CUR. CONVERSELY THE ANALGESIC ACTIVITY OF LY379268 IS POTENTIATED IN MICE PRETREATED FOR 5 CONSECUTIVE DAYS WITH THE HDAC INHIBITOR, SUBEROYLANILIDE HYDROXAMIC ACID (SAHA), KNOWN TO INDUCE MGLU2-UPREGULATION. CONCLUSIONS: OUR RESULTS DEMONSTRATE THAT SYSTEMICALLY INJECTED CUR IS ABLE TO INHIBIT H3 AND H4 ACETYLATION IN THE DRG AND TO DOWN-REGULATE MGLU2 RECEPTORS IN THE SPINAL CORD. WE ALSO DEMONSTRATE THAT LONG TERM MODIFICATION OF THE MGLU2 EXPRESSION AFFECTS THE ANALGESIC PROPERTIES OF THE ORTHOSTERIC MGLU2/3 AGONIST, LY379268. THESE DATA OPEN UP THE POSSIBILITY THAT EPIGENETIC MODULATORS MIGHT BE GIVEN IN COMBINATION WITH "TRADITIONAL" DRUGS IN A CONTEXT OF A MULTI TARGET APPROACH FOR A BETTER ANALGESIC EFFICACY. 2014 20 5940 30 TARGETING METHYLTRANSFERASE PRMT5 ELIMINATES LEUKEMIA STEM CELLS IN CHRONIC MYELOGENOUS LEUKEMIA. IMATINIB-INSENSITIVE LEUKEMIA STEM CELLS (LSCS) ARE BELIEVED TO BE RESPONSIBLE FOR RESISTANCE TO BCR-ABL TYROSINE KINASE INHIBITORS AND RELAPSE OF CHRONIC MYELOGENOUS LEUKEMIA (CML). IDENTIFYING THERAPEUTIC TARGETS TO ERADICATE CML LSCS MAY BE A STRATEGY TO CURE CML. IN THE PRESENT STUDY, WE DISCOVERED A POSITIVE FEEDBACK LOOP BETWEEN BCR-ABL AND PROTEIN ARGININE METHYLTRANSFERASE 5 (PRMT5) IN CML CELLS. OVEREXPRESSION OF PRMT5 WAS OBSERVED IN HUMAN CML LSCS. SILENCING PRMT5 WITH SHRNA OR BLOCKING PRMT5 METHYLTRANSFERASE ACTIVITY WITH THE SMALL-MOLECULE INHIBITOR PJ-68 REDUCED SURVIVAL, SERIAL REPLATING CAPACITY, AND LONG-TERM CULTURE-INITIATING CELLS (LTC-ICS) IN LSCS FROM CML PATIENTS. FURTHER, PRMT5 KNOCKDOWN OR PJ-68 TREATMENT DRAMATICALLY PROLONGED SURVIVAL IN A MURINE MODEL OF RETROVIRAL BCR-ABL-DRIVEN CML AND IMPAIRED THE IN VIVO SELF-RENEWAL CAPACITY OF TRANSPLANTED CML LSCS. PJ-68 ALSO INHIBITED LONG-TERM ENGRAFTMENT OF HUMAN CML CD34+ CELLS IN IMMUNODEFICIENT MICE. MOREOVER, INHIBITION OF PRMT5 ABROGATED THE WNT/BETA-CATENIN PATHWAY IN CML CD34+ CELLS BY DEPLETING DISHEVELLED HOMOLOG 3 (DVL3). THIS STUDY SUGGESTS THAT EPIGENETIC METHYLATION MODIFICATION ON HISTONE PROTEIN ARGININE RESIDUES IS A REGULATORY MECHANISM TO CONTROL SELF-RENEWAL OF LSCS AND INDICATES THAT PRMT5 MAY REPRESENT A POTENTIAL THERAPEUTIC TARGET AGAINST LSCS. 2016