1 535 113 ASXL1 MUTATION CORRECTION BY CRISPR/CAS9 RESTORES GENE FUNCTION IN LEUKEMIA CELLS AND INCREASES SURVIVAL IN MOUSE XENOGRAFTS. RECURRENT SOMATIC MUTATIONS OF THE EPIGENETIC MODIFIER AND TUMOR SUPPRESSOR ASXL1 ARE COMMON IN MYELOID MALIGNANCIES, INCLUDING CHRONIC MYELOID LEUKEMIA (CML), AND ARE ASSOCIATED WITH POOR CLINICAL OUTCOME. CRISPR/CAS9 HAS RECENTLY EMERGED AS A POWERFUL AND VERSATILE GENOME EDITING TOOL FOR GENOME ENGINEERING IN VARIOUS SPECIES. WE HAVE USED THE CRISPR/CAS9 SYSTEM TO CORRECT THE ASXL1 HOMOZYGOUS NONSENSE MUTATION PRESENT IN THE CML CELL LINE KBM5, WHICH LACKS ASXL1 PROTEIN EXPRESSION. CRISPR/CAS9-MEDIATED ASXL1 HOMOZYGOUS CORRECTION RESULTED IN PROTEIN RE-EXPRESSION WITH RESTORED NORMAL FUNCTION, INCLUDING DOWN-REGULATION OF POLYCOMB REPRESSIVE COMPLEX 2 TARGET GENES. SIGNIFICANTLY REDUCED CELL GROWTH AND INCREASED MYELOID DIFFERENTIATION WERE OBSERVED IN ASXL1 MUTATION-CORRECTED CELLS, PROVIDING NEW INSIGHTS INTO THE ROLE OF ASXL1 IN HUMAN MYELOID CELL DIFFERENTIATION. MICE XENOGRAFTED WITH MUTATION-CORRECTED KBM5 CELLS SHOWED SIGNIFICANTLY LONGER SURVIVAL THAN UNCORRECTED XENOGRAFTS. THESE RESULTS SHOW THAT THE SOLE CORRECTION OF A DRIVER MUTATION IN LEUKEMIA CELLS INCREASES SURVIVAL IN VIVO IN MICE. THIS STUDY PROVIDES PROOF-OF-CONCEPT FOR DRIVER GENE MUTATION CORRECTION VIA CRISPR/CAS9 TECHNOLOGY IN HUMAN LEUKEMIA CELLS AND PRESENTS A STRATEGY TO ILLUMINATE THE IMPACT OF ONCOGENIC MUTATIONS ON CELLULAR FUNCTION AND SURVIVAL. 2015 2 2781 37 EZH2 IN MYELOID MALIGNANCIES. OUR UNDERSTANDING OF THE SIGNIFICANCE OF EPIGENETIC DYSREGULATION IN THE PATHOGENESIS OF MYELOID MALIGNANCIES HAS GREATLY ADVANCED IN THE PAST DECADE. ENHANCER OF ZESTE HOMOLOG 2 (EZH2) IS THE CATALYTIC CORE COMPONENT OF THE POLYCOMB REPRESSIVE COMPLEX 2 (PRC2), WHICH IS RESPONSIBLE FOR GENE SILENCING THROUGH TRIMETHYLATION OF H3K27. EZH2 DYSREGULATION IS HIGHLY TUMORIGENIC AND HAS BEEN OBSERVED IN VARIOUS CANCERS, WITH EZH2 ACTING AS AN ONCOGENE OR A TUMOR-SUPPRESSOR DEPENDING ON CELLULAR CONTEXT. WHILE LOSS-OF-FUNCTION MUTATIONS OF EZH2 FREQUENTLY AFFECT PATIENTS WITH MYELODYSPLASTIC/MYELOPROLIFERATIVE NEOPLASMS, MYELODYSPLASTIC SYNDROME AND MYELOFIBROSIS, CASES OF CHRONIC MYELOID LEUKEMIA (CML) SEEM TO BE LARGELY CHARACTERIZED BY EZH2 OVEREXPRESSION. A VARIETY OF OTHER FACTORS FREQUENTLY ABERRANT IN MYELOID LEUKEMIA CAN AFFECT PRC2 FUNCTION AND DISEASE PATHOGENESIS, INCLUDING ADDITIONAL SEX COMBS LIKE 1 (ASXL1) AND SPLICING GENE MUTATIONS. AS THE GENETIC BACKGROUND OF MYELOID MALIGNANCIES IS LARGELY HETEROGENEOUS, IT IS NOT SURPRISING THAT EZH2 MUTATIONS ACT IN CONJUNCTION WITH OTHER ABERRATIONS. SINCE EZH2 MUTATIONS ARE CONSIDERED TO BE EARLY EVENTS IN DISEASE PATHOGENESIS, THEY ARE OF THERAPEUTIC INTEREST TO RESEARCHERS, THOUGH TARGETING OF EZH2 LOSS-OF-FUNCTION DOES PRESENT UNIQUE CHALLENGES. PRELIMINARY RESEARCH INDICATES THAT COMBINED TYROSINE KINASE INHIBITOR (TKI) AND EZH2 INHIBITOR THERAPY MAY PROVIDE A STRATEGY TO ELIMINATE THE RESIDUAL DISEASE BURDEN IN CML TO ALLOW PATIENTS TO REMAIN IN TREATMENT-FREE REMISSION. 2020 3 2237 36 EPIGENETIC MODIFIERS IN MYELOID MALIGNANCIES: THE ROLE OF HISTONE DEACETYLASE INHIBITORS. MYELOID HEMATOLOGICAL MALIGNANCIES ARE CLONAL BONE MARROW NEOPLASMS, COMPRISING OF ACUTE MYELOID LEUKEMIA (AML), THE MYELODYSPLASTIC SYNDROMES (MDS), CHRONIC MYELOMONOCYTIC LEUKEMIA (CMML), THE MYELOPROLIFERATIVE NEOPLASMS (MPN) AND SYSTEMIC MASTOCYTOSIS (SM). THE FIELD OF EPIGENETIC REGULATION OF NORMAL AND MALIGNANT HEMATOPOIESIS IS RAPIDLY GROWING. IN RECENT YEARS, HETEROZYGOUS SOMATIC MUTATIONS IN GENES ENCODING EPIGENETIC REGULATORS HAVE BEEN FOUND IN ALL SUBTYPES OF MYELOID MALIGNANCIES, SUPPORTING THE RATIONALE FOR TREATMENT WITH EPIGENETIC MODIFIERS. HISTONE DEACETYLASE INHIBITORS (HDACI) ARE EPIGENETIC MODIFIERS THAT, IN VITRO, HAVE BEEN SHOWN TO INDUCE GROWTH ARREST, APOPTOTIC OR AUTOPHAGIC CELL DEATH, AND TERMINAL DIFFERENTIATION OF MYELOID TUMOR CELLS. THESE EFFECTS WERE OBSERVED BOTH AT THE BULK TUMOR LEVEL AND IN THE MOST IMMATURE CD34(+)38(-) CELL COMPARTMENTS CONTAINING THE LEUKEMIC STEM CELLS. THUS, THERE IS A STRONG RATIONALE SUPPORTING HDACI THERAPY IN MYELOID MALIGNANCIES. HOWEVER, DESPITE INITIAL PROMISING RESULTS IN PHASE I TRIALS, HDACI IN MONOTHERAPY AS WELL AS IN COMBINATION WITH OTHER DRUGS, HAVE FAILED TO IMPROVE RESPONSES OR SURVIVAL. THIS REVIEW PROVIDES AN OVERVIEW OF THE RATIONALE FOR HDACI IN MYELOID MALIGNANCIES, CLINICAL RESULTS AND SPECULATIONS ON WHY CLINICAL TRIALS HAVE THUS FAR NOT MET THE EXPECTATIONS, AND HOW THIS MAY BE IMPROVED IN THE FUTURE. 2018 4 1674 34 DRIVER MUTATIONS IN LEUKEMIA PROMOTE DISEASE PATHOGENESIS THROUGH A COMBINATION OF CELL-AUTONOMOUS AND NICHE MODULATION. STUDIES OF PATIENTS WITH ACUTE MYELOID LEUKEMIA (AML) HAVE LED TO THE IDENTIFICATION OF MUTATIONS THAT AFFECT DIFFERENT CELLULAR PATHWAYS. SOME OF THESE HAVE BEEN CLASSIFIED AS PRELEUKEMIC, AND A STEPWISE EVOLUTION PROGRAM WHEREBY CELLS ACQUIRE ADDITIONAL MUTATIONS HAS BEEN PROPOSED IN THE DEVELOPMENT OF AML. HOW THE TIMING OF ACQUISITION OF THESE MUTATIONS AND THEIR IMPACT ON TRANSFORMATION AND THE BONE MARROW (BM) MICROENVIRONMENT OCCURS HAS ONLY RECENTLY BEGUN TO BE INVESTIGATED. WE SHOW THAT CONSTITUTIVE AND EARLY LOSS OF THE EPIGENETIC REGULATOR, TET2, WHEN COMBINED WITH CONSTITUTIVE ACTIVATION OF FLT3, RESULTS IN TRANSFORMATION OF CHRONIC MYELOMONOCYTIC LEUKEMIA-LIKE OR MYELOPROLIFERATIVE NEOPLASM-LIKE PHENOTYPE TO AML, WHICH IS MORE PRONOUNCED IN DOUBLE-MUTANT MICE RELATIVE TO MICE CARRYING MUTATIONS IN SINGLE GENES. FURTHERMORE, WE SHOW THAT IN PRELEUKEMIC AND LEUKEMIC MICE THERE ARE ALTERATIONS IN THE BM NICHE AND SECRETED CYTOKINES, WHICH CREATES A PERMISSIVE ENVIRONMENT FOR THE GROWTH OF MUTATION-BEARING CELLS RELATIVE TO NORMAL CELLS. 2020 5 5687 37 SIGNIFICANCE OF INACTIVATED GENES IN LEUKEMIA: PATHOGENESIS AND PROGNOSIS. EPIGENETIC AND GENETIC ALTERATIONS ARE TWO MECHANISMS PARTICIPATING IN LEUKEMIA, WHICH CAN INACTIVATE GENES INVOLVED IN LEUKEMIA PATHOGENESIS OR PROGRESSION. THE PURPOSE OF THIS REVIEW WAS TO INTRODUCE VARIOUS INACTIVATED GENES AND EVALUATE THEIR POSSIBLE ROLE IN LEUKEMIA PATHOGENESIS AND PROGNOSIS. BY SEARCHING THE MESH WORDS "GENE, SILENCING AND LEUKEMIA" IN PUBMED WEBSITE, RELEVANT ENGLISH ARTICLES DEALT WITH HUMAN SUBJECTS AS OF 2000 WERE INCLUDED IN THIS STUDY. GENE INACTIVATION IN LEUKEMIA IS LARGELY MEDIATED BY PROMOTER'S HYPERMETHYLATION OF GENE INVOLVING IN CELLULAR FUNCTIONS SUCH AS CELL CYCLE, APOPTOSIS, AND GENE TRANSCRIPTION. INACTIVATED GENES, SUCH AS ASPP1, TP53, IKZF1 AND P15, MAY CORRELATE WITH POOR PROGNOSIS IN ACUTE LYMPHOID LEUKEMIA (ALL), CHRONIC LYMPHOID LEUKEMIA (CLL), CHRONIC MYELOGENOUS LEUKEMIA (CML) AND ACUTE MYELOID LEUKEMIA (AML), RESPECTIVELY. GENE INACTIVATION MAY PLAY A CONSIDERABLE ROLE IN LEUKEMIA PATHOGENESIS AND PROGNOSIS, WHICH CAN BE CONSIDERED AS COMPLEMENTARY DIAGNOSTIC TESTS TO DIFFERENTIATE DIFFERENT LEUKEMIA TYPES, DETERMINE LEUKEMIA PROGNOSIS, AND ALSO DETECT RESPONSE TO THERAPY. IN GENERAL, THIS REVIEW SHOWED SOME GENES INACTIVATED ONLY IN LEUKEMIA (WITH DIFFERENCES BETWEEN B-ALL, T-ALL, CLL, AML AND CML). THESE DIFFERENCES COULD BE OF INTEREST AS AN ADDITIONAL TOOL TO BETTER CATEGORIZE LEUKEMIA TYPES. FURTHERMORE; BASED ON INACTIVATED GENES, A DIVERSE CLASSIFICATION OF LEUKEMIAS COULD REPRESENT A POWERFUL METHOD TO ADDRESS A TARGETED THERAPY OF THE PATIENTS, IN ORDER TO MINIMIZE SIDE EFFECTS OF CONVENTIONAL THERAPIES AND TO ENHANCE NEW DRUG STRATEGIES. 2017 6 913 30 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 7 825 33 CHARACTERIZATION OF FUNCTIONAL TRANSPOSABLE ELEMENT ENHANCERS IN ACUTE MYELOID LEUKEMIA. TRANSPOSABLE ELEMENTS (TES) HAVE BEEN SHOWN TO HAVE IMPORTANT GENE REGULATORY FUNCTIONS AND THEIR ALTERATION COULD LEAD TO DISEASE PHENOTYPES. ACUTE MYELOID LEUKEMIA (AML) DEVELOPS AS A CONSEQUENCE OF A SERIES OF GENETIC CHANGES IN HEMATOPOIETIC PRECURSOR CELLS, INCLUDING MUTATIONS IN EPIGENETIC FACTORS. HERE, WE SET OUT TO STUDY THE GENE REGULATORY ROLE OF TES IN AML. WE FIRST EXPLORED THE EPIGENETIC LANDSCAPE OF TES IN AML PATIENTS USING ATAC-SEQ DATA. WE SHOW THAT A LARGE NUMBER OF TES IN GENERAL, AND MORE SPECIFICALLY MAMMALIAN-WIDE INTERSPERSED REPEATS (MIRS), ARE MORE ENRICHED IN AML CELLS THAN IN NORMAL BLOOD CELLS. WE OBTAINED A SIMILAR FINDING WHEN ANALYZING HISTONE MODIFICATION DATA IN AML PATIENTS. GENE ONTOLOGY ENRICHMENT ANALYSIS SHOWED THAT GENES NEAR MIRS IN OPEN CHROMATIN REGIONS ARE INVOLVED IN LEUKEMOGENESIS. TO FUNCTIONALLY VALIDATE THEIR REGULATORY ROLE, WE SELECTED 19 MIR REGIONS IN AML CELLS, AND TESTED THEM FOR ENHANCER ACTIVITY IN AN AML CELL LINE (KASUMI-1) AND A CHRONIC MYELOID LEUKEMIA (CML) CELL LINE (K562); THE RESULTS REVEALED SEVERAL MIRS TO BE FUNCTIONAL ENHANCERS. TAKEN TOGETHER, OUR RESULTS SUGGEST THAT TES ARE POTENTIALLY INVOLVED IN MYELOID LEUKEMOGENESIS AND HIGHLIGHT THESE SEQUENCES AS POTENTIAL CANDIDATES HARBORING AML-ASSOCIATED VARIATION. 2020 8 2911 39 GENE EXPRESSION PROFILING OF LOSS OF TET2 AND/OR JAK2V617F MUTANT HEMATOPOIETIC STEM CELLS FROM MOUSE MODELS OF MYELOPROLIFERATIVE NEOPLASMS. MYELOPROLIFERATIVE NEOPLASMS (MPNS) ARE CLINICALLY CHARACTERIZED BY THE CHRONIC OVERPRODUCTION OF DIFFERENTIATED PERIPHERAL BLOOD CELLS AND THE GRADUAL EXPANSION OF MALIGNANT INTRAMEDULLARY/EXTRAMEDULLARY HEMATOPOIESIS. IN MPNS MUTATIONS IN JAK2 MPL OR CALR ARE DETECTED MUTUALLY EXCLUSIVE IN MORE THAN 90% OF CASES [1,2]. MUTATIONS IN THEM LEAD TO THE ABNORMAL ACTIVATION OF JAK/STAT SIGNALING AND THE AUTONOMOUS GROWTH OF DIFFERENTIATED CELLS THEREFORE THEY ARE CONSIDERED AS "DRIVER" GENE MUTATIONS. IN ADDITION TO THE ABOVE DRIVER GENE MUTATIONS MUTATIONS IN EPIGENETIC REGULATORS SUCH AS TET2 DNMT3A ASXL1 EZH2 OR IDH1/2 ARE DETECTED IN ABOUT 5%-30% OF CASES RESPECTIVELY [3]. MUTATIONS IN TET2 DNMT3A EZH2 OR IDH1/2 COMMONLY CONFER THE INCREASED SELF-RENEWAL CAPACITY ON NORMAL HEMATOPOIETIC STEM CELLS (HSCS) BUT THEY DO NOT LEAD TO THE AUTONOMOUS GROWTH OF DIFFERENTIATED CELLS AND ONLY EXHIBIT SUBTLE CLINICAL PHENOTYPES [4,6-8,5]. IT WAS UNCLEAR HOW MUTATIONS IN SUCH EPIGENETIC REGULATORS INFLUENCED ABNORMAL HSCS WITH DRIVER GENE MUTATIONS HOW THEY INFLUENCED THE DISEASE PHENOTYPE OR WHETHER A SINGLE DRIVER GENE MUTATION WAS SUFFICIENT FOR THE INITIATION OF HUMAN MPNS. THEREFORE WE FOCUSED ON JAK2V617F AND LOSS OF TET2-THE FORMER AS A REPRESENTATIVE OF DRIVER GENE MUTATIONS AND THE LATTER AS A REPRESENTATIVE OF MUTATIONS IN EPIGENETIC REGULATORS-AND EXAMINED THE INFLUENCE OF SINGLE OR DOUBLE MUTATIONS ON HSCS (LINEAGE(-)SCA-1(+)C-KIT(+) CELLS (LSKS)) BY FUNCTIONAL ANALYSES AND MICROARRAY WHOLE-GENOME EXPRESSION ANALYSES [9]. GENE EXPRESSION PROFILING SHOWED THAT THE HSC FINGERPRINT GENES [10] WAS STATISTICALLY EQUALLY ENRICHED IN TET2-KNOCKDOWN-LSKS BUT NEGATIVELY ENRICHED IN JAK2V617F-LSKS COMPARED TO THAT IN WILD-TYPE-LSKS. DOUBLE-MUTANT-LSKS SHOWED THE SAME TENDENCY AS JAK2V617F-LSKS IN TERMS OF THEIR HSC FINGERPRINT GENES BUT THE EXPRESSION OF INDIVIDUAL GENES DIFFERED BETWEEN THE TWO GROUPS. AMONG 245 HSC FINGERPRINT GENES 100 WERE MORE HIGHLY EXPRESSED IN DOUBLE-MUTANT-LSKS THAN IN JAK2V617F-LSKS. THESE ALTERED GENE EXPRESSIONS MIGHT PARTLY EXPLAIN THE MECHANISMS OF INITIATION AND PROGRESSION OF MPNS WHICH WAS OBSERVED IN THE FUNCTIONAL ANALYSES [9]. HERE WE DESCRIBE GENE EXPRESSION PROFILES DEPOSITED AT THE GENE EXPRESSION OMNIBUS (GEO) UNDER THE ACCESSION NUMBER GSE62302 INCLUDING EXPERIMENTAL METHODS AND QUALITY CONTROL ANALYSES. 2015 9 390 38 AN INTEGRATIVE MODEL OF PATHWAY CONVERGENCE IN GENETICALLY HETEROGENEOUS BLAST CRISIS CHRONIC MYELOID LEUKEMIA. TARGETED THERAPIES AGAINST THE BCR-ABL1 KINASE HAVE REVOLUTIONIZED TREATMENT OF CHRONIC PHASE (CP) CHRONIC MYELOID LEUKEMIA (CML). IN CONTRAST, MANAGEMENT OF BLAST CRISIS (BC) CML REMAINS CHALLENGING BECAUSE BC CELLS ACQUIRE COMPLEX MOLECULAR ALTERATIONS THAT CONFER STEMNESS FEATURES TO PROGENITOR POPULATIONS AND RESISTANCE TO BCR-ABL1 TYROSINE KINASE INHIBITORS. COMPREHENSIVE MODELS OF BC TRANSFORMATION HAVE PROVED ELUSIVE BECAUSE OF THE RARITY AND GENETIC HETEROGENEITY OF BC, BUT ARE IMPORTANT FOR DEVELOPING BIOMARKERS PREDICTING BC PROGRESSION AND EFFECTIVE THERAPIES. TO BETTER UNDERSTAND BC, WE PERFORMED AN INTEGRATED MULTIOMICS ANALYSIS OF 74 CP AND BC SAMPLES USING WHOLE-GENOME AND EXOME SEQUENCING, TRANSCRIPTOME AND METHYLOME PROFILING, AND CHROMATIN IMMUNOPRECIPITATION FOLLOWED BY HIGH-THROUGHPUT SEQUENCING. EMPLOYING PATHWAY-BASED ANALYSIS, WE FOUND THE BC GENOME WAS SIGNIFICANTLY ENRICHED FOR MUTATIONS AFFECTING COMPONENTS OF THE POLYCOMB REPRESSIVE COMPLEX (PRC) PATHWAY. WHILE TRANSCRIPTOMICALLY, BC PROGENITORS WERE ENRICHED AND DEPLETED FOR PRC1- AND PRC2-RELATED GENE SETS RESPECTIVELY. BY INTEGRATING OUR DATA SETS, WE DETERMINED THAT BC PROGENITORS UNDERGO PRC-DRIVEN EPIGENETIC REPROGRAMMING TOWARD A CONVERGENT TRANSCRIPTOMIC STATE. SPECIFICALLY, PRC2 DIRECTS BC DNA HYPERMETHYLATION, WHICH IN TURN SILENCES KEY GENES INVOLVED IN MYELOID DIFFERENTIATION AND TUMOR SUPPRESSOR FUNCTION VIA SO-CALLED EPIGENETIC SWITCHING, WHEREAS PRC1 REPRESSES AN OVERLAPPING AND DISTINCT SET OF GENES, INCLUDING NOVEL BC TUMOR SUPPRESSORS. ON THE BASIS OF THESE OBSERVATIONS, WE DEVELOPED AN INTEGRATED MODEL OF BC THAT FACILITATED THE IDENTIFICATION OF COMBINATORIAL THERAPIES CAPABLE OF REVERSING BC REPROGRAMMING (DECITABINE+PRC1 INHIBITORS), NOVEL PRC-SILENCED TUMOR SUPPRESSOR GENES (NR4A2), AND GENE EXPRESSION SIGNATURES PREDICTIVE OF DISEASE PROGRESSION AND DRUG RESISTANCE IN CP. 2020 10 358 31 ALTERNATIVE SPLICING IN CHRONIC MYELOID LEUKEMIA (CML): A NOVEL THERAPEUTIC TARGET? ALTHOUGH THE IMATINIB BASED THERAPY OF CHRONIC MYELOID LEUKEMIA (CML) REPRESENTS A TRIUMPH OF MEDICINE, NOT ALL PATIENTS WITH CML BENEFIT FROM THIS DRUG DUE TO THE DEVELOPMENT OF RESISTANCE AND INTOLERANCE. THE INTERRUPTION OF IMATINIB TREATMENT IS OFTEN FOLLOWED BY CLINICAL RELAPSE, SUGGESTING A FAILURE IN THE KILLING OF RESIDUAL LEUKAEMIC STEM CELLS. THERE IS NEED TO IDENTIFY ALTERNATIVE SELECTIVE MOLECULAR TARGETS FOR THIS DISEASE AND DEVELOP MORE EFFECTIVE THERAPEUTIC APPROACHES. ALTERNATIVE PRE-MRNA SPLICING (AS) IS AN EPIGENETIC PROCESS THAT GREATLY DIVERSIFIES THE REPERTOIRE OF THE TRANSCRIPTOME. AS ORCHESTRATES INTERACTIONS BETWEEN VARIOUS TYPES OF PROTEINS AND BETWEEN PROTEINS AND NUCLEIC ACIDS. CHANGES CAUSED BY INDIVIDUAL SPLICING EVENTS IN THE CELLS ARE SMALL, HOWEVER, "SPLICING PROGRAMS" TYPICALLY REACT TO THESE INDIVIDUAL CHANGES WITH CONSIDERABLE EFFECTS IN CELL PROLIFERATION, CELL SURVIVAL, AND APOPTOSIS. CURRENT EVIDENCE SUGGESTS A PIVOTAL ROLE OF AS IN LEUKEMIAS, PARTICULARLY IN MYELODISPLASTIC SYNDROME (MDS) AND CHRONIC LYMPHOCYTE LEUKEMIA (CLL). FROM THESE STUDIES AND STUDIES IN OTHER MALIGNANCES, IT IS CLEAR THAT SPLICING ABNORMALITIES PLAY A SIGNIFICANT ROLE IN MALIGNANT TRANSFORMATION. EVALUATION OF AS EVENTS IN CML CAN BE USED TO IDENTIFY NOVEL DISEASE MARKERS AND DRUGSENSITIVE TARGETS TO OVERCOME THE LIMITS OF THE SMALL MOLECULE INHIBITORS CURRENTLY USED FOR TREATING PATIENTS WITH CML. THE USE OF ABERRANT SPLICE VARIANTS AS DISEASE MARKERS HAS BEEN REPORTED, HOWEVER, LITTLE IS KNOWN ABOUT THE USE OF SPLICING ABNORMALITIES AS DRUG TARGETS IN CML. HEREIN WE DISCUSS POTENTIAL THERAPEUTIC APPROACHES THAT CAN BE USED TO TARGET SPLICING ABNORMALITIES IN CML. 2013 11 3234 30 HEMATOPOIETIC AND CHRONIC MYELOID LEUKEMIA STEM CELLS: MULTI-STABILITY VERSUS LINEAGE RESTRICTION. THERE IS COMPELLING EVIDENCE TO SUPPORT THE VIEW THAT THE CELL-OF-ORIGIN FOR CHRONIC MYELOID LEUKEMIA IS A HEMATOPOIETIC STEM CELL. UNLIKE NORMAL HEMATOPOIETIC STEM CELLS, THE PROGENY OF THE LEUKEMIA STEM CELLS ARE PREDOMINANTLY NEUTROPHILS DURING THE DISEASE CHRONIC PHASE AND THERE IS A MILD ANEMIA. THE HALLMARK ONCOGENE FOR CHRONIC MYELOID LEUKEMIA IS THE BCR-ABLP210 FUSION GENE. VARIOUS STUDIES HAVE EXCLUDED A ROLE FOR BCR-ABLP210 EXPRESSION IN MAINTAINING THE POPULATION OF LEUKEMIA STEM CELLS. STUDIES OF BCR-ABLP210 EXPRESSION IN EMBRYONAL STEM CELLS THAT WERE DIFFERENTIATED INTO HEMATOPOIETIC STEM CELLS AND OF THE EXPRESSION IN TRANSGENIC MICE HAVE REVEALED THAT BCR-ABLP210 IS ABLE TO VEER HEMATOPOIETIC STEM AND PROGENITOR CELLS TOWARDS A MYELOID FATE. FOR THE TRANSGENIC MICE, GLOBAL CHANGES TO THE EPIGENETIC LANDSCAPE WERE OBSERVED. IN CHRONIC MYELOID LEUKEMIA, THE ABILITY OF THE LEUKEMIA STEM CELLS TO CHOOSE FROM THE MANY FATES THAT ARE AVAILABLE TO NORMAL HEMATOPOIETIC STEM CELLS APPEARS TO BE DEREGULATED BY BCR-ABLP210 AND CHANGES TO THE EPIGENOME ARE ALSO IMPORTANT. EVEN SO, WE STILL DO NOT HAVE A PRECISE PICTURE AS TO WHY NEUTROPHILS ARE ABUNDANTLY PRODUCED IN CHRONIC MYELOID LEUKEMIA. 2022 12 160 26 ABERRANT PROMOTER HYPOMETHYLATION IN CLL: DOES IT MATTER FOR DISEASE DEVELOPMENT? OVER THE LAST 30 YEARS, STUDIES OF ABERRANT DNA METHYLATION IN HEMATOLOGIC MALIGNANCIES HAVE BEEN DOMINATED BY THE PRIMARY FOCUS OF UNDERSTANDING PROMOTER HYPERMETHYLATION. THESE EFFORTS NOT ONLY RESULTED IN A BETTER UNDERSTANDING OF THE BASIS OF EPIGENETIC SILENCING OF TUMOR SUPPRESSOR GENES BUT ALSO RESULTED IN APPROVAL OF HYPOMETHYLATING AGENTS FOR THE TREATMENT OF SEVERAL MALIGNANCIES, SUCH AS MYELODYSPLASTIC SYNDROME AND ACUTE MYELOID LEUKEMIA. RECENT ADVANCES IN GLOBAL METHYLATION PROFILING COUPLED WITH THE USE OF MOUSE MODELS SUGGEST THAT ABERRANT PROMOTER HYPOMETHYLATION IS ALSO A FREQUENT EVENT IN HEMATOLOGIC MALIGNANCIES, PARTICULARLY IN CHRONIC LYMPHOCYTIC LEUKEMIA (CLL). PROMOTER HYPOMETHYLATION AFFECTS GENE EXPRESSION AND, THEREFORE, MAY PLAY AN IMPORTANT ROLE IN DISEASE PATHOGENESIS. HERE, WE REVIEW RECENT FINDINGS AND DISCUSS THE POTENTIAL INVOLVEMENT OF ABERRANT PROMOTER HYPOMETHYLATION IN CLL. 2016 13 3098 32 GENOMIC DISRUPTION OF THE HISTONE METHYLTRANSFERASE SETD2 IN CHRONIC LYMPHOCYTIC LEUKAEMIA. HISTONE METHYLTRANSFERASES (HMTS) ARE IMPORTANT EPIGENETIC REGULATORS OF GENE TRANSCRIPTION AND ARE DISRUPTED AT THE GENOMIC LEVEL IN A SPECTRUM OF HUMAN TUMOURS INCLUDING HAEMATOLOGICAL MALIGNANCIES. USING HIGH-RESOLUTION SINGLE NUCLEOTIDE POLYMORPHISM (SNP) ARRAYS, WE IDENTIFIED RECURRENT DELETIONS OF THE SETD2 LOCUS IN 3% (8/261) OF CHRONIC LYMPHOCYTIC LEUKAEMIA (CLL) PATIENTS. FURTHER VALIDATION IN TWO INDEPENDENT COHORTS SHOWED THAT SETD2 DELETIONS WERE ASSOCIATED WITH LOSS OF TP53, GENOMIC COMPLEXITY AND CHROMOTHRIPSIS. WITH NEXT-GENERATION SEQUENCING WE DETECTED MUTATIONS OF SETD2 IN AN ADDITIONAL 3.8% OF PATIENTS (23/602). IN MOST CASES, SETD2 DELETIONS OR MUTATIONS WERE OFTEN OBSERVED AS A CLONAL EVENT AND ALWAYS AS A MONO-ALLELIC LESION, LEADING TO REDUCED MRNA EXPRESSION IN SETD2-DISRUPTED CASES. PATIENTS WITH SETD2 ABNORMALITIES AND WILD-TYPE TP53 AND ATM FROM FIVE CLINICAL TRIALS EMPLOYING CHEMOTHERAPY OR CHEMO-IMMUNOTHERAPY HAD REDUCED PROGRESSION-FREE AND OVERALL SURVIVAL COMPARED WITH CASES WILD TYPE FOR ALL THREE GENES. CONSISTENT WITH ITS POSTULATED ROLE AS A TUMOUR SUPPRESSOR, OUR DATA HIGHLIGHT SETD2 ABERRATION AS A RECURRENT, EARLY LOSS-OF-FUNCTION EVENT IN CLL PATHOBIOLOGY LINKED TO AGGRESSIVE DISEASE. 2016 14 4547 24 MUTATION ALLELE BURDEN REMAINS UNCHANGED IN CHRONIC MYELOMONOCYTIC LEUKAEMIA RESPONDING TO HYPOMETHYLATING AGENTS. THE CYTIDINE ANALOGUES AZACYTIDINE AND 5-AZA-2'-DEOXYCYTIDINE (DECITABINE) ARE COMMONLY USED TO TREAT MYELODYSPLASTIC SYNDROMES, WITH OR WITHOUT A MYELOPROLIFERATIVE COMPONENT. IT REMAINS UNCLEAR WHETHER THE RESPONSE TO THESE HYPOMETHYLATING AGENTS RESULTS FROM A CYTOTOXIC OR AN EPIGENETIC EFFECT. IN THIS STUDY, WE ADDRESS THIS QUESTION IN CHRONIC MYELOMONOCYTIC LEUKAEMIA. WE DESCRIBE A COMPREHENSIVE ANALYSIS OF THE MUTATIONAL LANDSCAPE OF THESE TUMOURS, COMBINING WHOLE-EXOME AND WHOLE-GENOME SEQUENCING. WE IDENTIFY AN AVERAGE OF 14+/-5 SOMATIC MUTATIONS IN CODING SEQUENCES OF SORTED MONOCYTE DNA AND THE SIGNATURES OF THREE MUTATIONAL PROCESSES. SERIAL SEQUENCING DEMONSTRATES THAT THE RESPONSE TO HYPOMETHYLATING AGENTS IS ASSOCIATED WITH CHANGES IN DNA METHYLATION AND GENE EXPRESSION, WITHOUT ANY DECREASE IN THE MUTATION ALLELE BURDEN, NOR PREVENTION OF NEW GENETIC ALTERATION OCCURENCE. OUR FINDINGS INDICATE THAT CYTOSINE ANALOGUES RESTORE A BALANCED HAEMATOPOIESIS WITHOUT DECREASING THE SIZE OF THE MUTATED CLONE, ARGUING FOR A PREDOMINANTLY EPIGENETIC EFFECT. 2016 15 1465 23 DISSECTING THE ROLE OF ABERRANT DNA METHYLATION IN HUMAN LEUKAEMIA. CHRONIC MYELOID LEUKAEMIA (CML) IS A MYELOPROLIFERATIVE DISORDER CHARACTERIZED BY THE GENETIC TRANSLOCATION T(9;22)(Q34;Q11.2) ENCODING FOR THE BCR-ABL FUSION ONCOGENE. HOWEVER, MANY MOLECULAR MECHANISMS OF THE DISEASE PROGRESSION STILL REMAIN POORLY UNDERSTOOD. A GROWING BODY OF EVIDENCE SUGGESTS THAT THE EPIGENETIC ABNORMALITIES ARE INVOLVED IN TYROSINE KINASE RESISTANCE IN CML, LEADING TO LEUKAEMIC CLONE ESCAPE AND DISEASE PROPAGATION. HERE WE SHOW THAT, BY APPLYING CELLULAR REPROGRAMMING TO PRIMARY CML CELLS, ABERRANT DNA METHYLATION CONTRIBUTES TO THE DISEASE EVOLUTION. IMPORTANTLY, USING A BCR-ABL INDUCIBLE MURINE MODEL, WE DEMONSTRATE THAT A SINGLE ONCOGENIC LESION TRIGGERS DNA METHYLATION CHANGES, WHICH IN TURN ACT AS A PRECIPITATING EVENT IN LEUKAEMIA PROGRESSION. 2015 16 2971 27 GENETIC AND EPIGENETIC SILENCING OF MICRORNA-203 ENHANCES ABL1 AND BCR-ABL1 ONCOGENE EXPRESSION. THE MAMMALIAN GENOME CONTAINS SEVERAL HUNDRED MICRORNAS THAT REGULATE GENE EXPRESSION THROUGH MODULATION OF TARGET MRNAS. HERE, WE REPORT A FRAGILE CHROMOSOMAL REGION LOST IN SPECIFIC HEMATOPOIETIC MALIGNANCIES. THIS 7 MB REGION ENCODES ABOUT 12% OF ALL GENOMIC MICRORNAS, INCLUDING MIR-203. THIS MICRORNA IS ADDITIONALLY HYPERMETHYLATED IN SEVERAL HEMATOPOIETIC TUMORS, INCLUDING CHRONIC MYELOGENOUS LEUKEMIAS AND SOME ACUTE LYMPHOBLASTIC LEUKEMIAS. A PUTATIVE MIR-203 TARGET, ABL1, IS SPECIFICALLY ACTIVATED IN THESE HEMATOPOIETIC MALIGNANCIES IN SOME CASES AS A BCR-ABL1 FUSION PROTEIN (PHILADELPHIA CHROMOSOME). RE-EXPRESSION OF MIR-203 REDUCES ABL1 AND BCR-ABL1 FUSION PROTEIN LEVELS AND INHIBITS TUMOR CELL PROLIFERATION IN AN ABL1-DEPENDENT MANNER. THUS, MIR-203 FUNCTIONS AS A TUMOR SUPPRESSOR, AND RE-EXPRESSION OF THIS MICRORNA MIGHT HAVE THERAPEUTIC BENEFITS IN SPECIFIC HEMATOPOIETIC MALIGNANCIES. 2008 17 1542 34 DNA METHYLATION IN HAEMATOLOGICAL MALIGNANCIES: THE ROLE OF DECITABINE. NORMAL CELL DEVELOPMENT AND FUNCTION IS DEPENDENT UPON CONTROLLED GENE EXPRESSION. DNA METHYLATION IS AN EPIGENETIC MODIFICATION THAT CAN PLAY AN IMPORTANT ROLE IN THE CONTROL OF GENE EXPRESSION. DNA METHYLATION AT CYTOSINE RESIDUES IN GENE PROMOTER CPG SEQUENCES IS KNOWN TO INHIBIT GENE TRANSCRIPTION. INAPPROPRIATE INHIBITION OF THE TRANSCRIPTION OF TUMOUR SUPPRESSOR GENES, GENES THAT INHIBIT ANGIOGENESIS AND METASTASIS AND GENES INVOLVED IN DNA REPAIR BY UNCONTROLLED METHYLATION, CAN LEAD TO UNREGULATED GROWTH AND PROLIFERATION OF A CELL AND CARCINOGENESIS. PROMOTER HYPERMETHYLATION AFFECTING THE P16 GENE, RESULTING IN GENE SILENCING, HAS BEEN SHOWN TO OCCUR IN MANY HUMAN SOLID TUMOURS AND A 'HYPERMETHYLATION PROFILE' IN SOME LEUKAEMIAS HAS BEEN DEFINED. THE MOLECULAR MECHANISMS BY WHICH ABERRANT DNA METHYLATION TAKES PLACE DURING CARCINOGENESIS ARE STILL NOT CLEAR. HOWEVER, THE LARGE NUMBER OF TARGET GENES (INVOLVED IN TUMORIGENESIS) THAT ARE SILENCED BY ABERRANT METHYLATION SUGGESTS THAT INHIBITION OF THIS PROCESS MAY HAVE POTENTIAL AS CANCER THERAPY. DECITABINE (NSC-127716, DACOGEN; SUPERGEN) IS A POTENT AND SPECIFIC HYPOMETHYLATING AGENT AND AN INHIBITOR OF THE DNA METHYLTRANSFERASE ACTIVITY THAT MEDIATES DNA METHYLATION. DECITABINE HAS BEEN SHOWN TO HAVE A BROAD RANGE OF ANTINEOPLASTIC ACTIVITY IN PRECLINICAL STUDIES. THIS AGENT HAS EXHIBITED SIGNIFICANT ACTIVITY IN THE TREATMENT OF PATIENTS WITH MYELODYSPLASTIC SYNDROME, CHRONIC MYELOID LEUKAEMIA AND ACUTE MYELOID LEUKAEMIA, ALTHOUGH CLINICAL PHASE I AND II STUDIES WITH SOLID TUMOURS HAVE NOT BEEN VERY PROMISING. PHASE II AND III STUDIES ARE CURRENTLY ONGOING TO EVALUATE DECITABINE, BOTH ALONE AND IN COMBINATION, IN VARIOUS STAGES OF THESE HAEMATOLOGICAL MALIGNANCIES. 2003 18 6773 25 [ADVANCES OF RESEARCH ON DEMETHYLATION THERAPY FOR HEMATOLOGIC MALIGNANCIES]. DNA METHYLATION IS AN IMPORTANT AND REVERSIBLE EPIGENETIC MODIFICATION WHICH REGULATES GENOMIC STABILITY. METHYLATION IS ESSENTIAL FOR MAMMALIAN DEVELOPMENT. GENERALLY, GENE EXPRESSION LEVEL AND DNA METHYLATION ARE NEGATIVE CORRELATION. TRANSCRIPTIONAL SILENCING VIA METHYLATION OF CPG ISLANDS IN THE PROMOTER IS IMPORTANT FOR CELL GROWTH AND DIFFERENTIATION AND PLAYS A KEY ROLE IN TUMORIGENESIS. DEMETHYLATION DRUG CAN MODIFY CHROMATIN AND RESTORE THE ABILITY OF ANTI-ONCOGENE. DEMETHYLATION THERAPY AS A NEW THERAPY MAY TREAT EFFICIENTLY HEMATOLOGICAL MALIGNANCIES WITH RESISTANCE AND RELAPSE. IN THIS REVIEW, DNA METHYLATION MECHANISM, RELATIONSHIP BETWEEN ABERRANT METHYLATION AND HEMATOLOGIC MALIGNANCIES, MECHANISM OF DEMETHYLATION THERAPY, THE ADVANCE OF RESEARCH ON THE DEMETHYLATION THERAPY OF HEMATOLOGICAL MALIGNANCIES, SUCH AS ACUTE AND CHRONIC LEUKEMIA, LYMPHOMA, MYELODYSPLASTIC SYNDROME WERE SUMMARIZED. 2009 19 4728 35 NOTCH SIGNALING PROMOTES DISEASE INITIATION AND PROGRESSION IN MURINE CHRONIC LYMPHOCYTIC LEUKEMIA. NOTCH1 GAIN-OF-FUNCTION MUTATIONS ARE RECURRENT IN B-CELL CHRONIC LYMPHOCYTIC LEUKEMIA (B-CLL), WHERE THEY ARE ASSOCIATED WITH ACCELERATED DISEASE PROGRESSION AND REFRACTORINESS TO CHEMOTHERAPY. THE SPECIFIC ROLE OF NOTCH1 IN THE DEVELOPMENT AND PROGRESSION OF THIS MALIGNANCY IS UNCLEAR. HERE, WE ASSESS THE IMPACT OF LOSS OF NOTCH SIGNALING AND PATHWAY HYPERACTIVATION IN AN IN VIVO MOUSE MODEL OF CLL (IGH.TEMU) THAT FAITHFULLY REPLICATES MANY FEATURES OF THE HUMAN PATHOLOGY. ABLATION OF CANONICAL NOTCH SIGNALING USING CONDITIONAL GENE INACTIVATION OF RBP-J IN IMMATURE HEMATOPOIETIC OR B-CELL PROGENITORS DELAYED CLL INDUCTION AND REDUCED INCIDENCE OF MICE DEVELOPING DISEASE. IN CONTRAST, FORCED EXPRESSION OF A DOMINANT ACTIVE FORM OF NOTCH RESULTED IN MORE ANIMALS DEVELOPING CLL WITH EARLY DISEASE ONSET. COMPARATIVE ANALYSIS OF GENE EXPRESSION AND EPIGENETIC FEATURES OF NOTCH GAIN-OF-FUNCTION AND CONTROL CLL CELLS REVEALED DIRECT AND INDIRECT REGULATION OF CELL CYCLE-ASSOCIATED GENES, WHICH LED TO INCREASED PROLIFERATION OF NOTCH GAIN-OF-FUNCTION CLL CELLS IN VIVO. THESE RESULTS DEMONSTRATE THAT NOTCH SIGNALING FACILITATES DISEASE INITIATION AND PROMOTES CLL CELL PROLIFERATION AND DISEASE PROGRESSION. 2021 20 1304 34 DEFECTS IN SPLICEOSOMAL MACHINERY: A NEW PATHWAY OF LEUKAEMOGENESIS. PROPER SPLICING OF PRE-MRNA IS REQUIRED FOR PROTEIN SYNTHESIS AND THEREFORE IS A FUNDAMENTAL CELLULAR FUNCTION. THE DISCOVERY OF A VARIETY OF SOMATIC SPLICEOSOMAL MUTATIONS IN HAEMATOLOGICAL MALIGNANCIES, INCLUDING MYELOID NEOPLASMS AND CHRONIC LYMPHOCYTIC LEUKAEMIA HAS POINTED TO A NEW LEUKAEMOGENIC PATHWAY INVOLVING SPLICEOSOMAL DYSFUNCTION. THEORETICALLY, SPLICEOSOMAL MUTATIONS CAN LEAD TO ACTIVATION OF INCORRECT SPLICE SITES, INTRON RETENTION OR ABERRANT ALTERNATIVE SPLICING OCCURRING IN PATTERNS GENERATED BY MUTATIONS OF INDIVIDUAL SPLICEOSOMAL PROTEINS. SUCH EVENTS CAN PRODUCE A DEFECTIVE BALANCE BETWEEN PROTEIN ISOFORMS LEADING TO FUNCTIONAL CONSEQUENCES INCLUDING DEFECTIVE REGULATION OF PROLIFERATION AND DIFFERENTIATION. THE OBSERVED PATTERN OF OCCURRENCE OF HIGHLY SPECIFIC MISSENSE MUTATIONS, COUPLED WITH THE LACK OF NONSENSE MUTATIONS AND DELETIONS, IMPLIES A GAIN-OF-FUNCTION OR BETTER GAIN-OF-DYSFUNCTION MECHANISM. INCORRECT SPLICING OF DOWNSTREAM GENES, SUCH AS TUMOUR SUPPRESSOR GENES, MAY RESULT IN HAPLOINSUFFICIENT EXPRESSION THROUGH NONSENSE-MEDIATED MRNA DECAY. THUS, SPLICEOSOMAL MUTATIONS MAY, DEPENDING ON THE PATTERN OF AFFECTED PROTEINS, LEAD TO SIMILAR FUNCTIONAL EFFECTS ON TUMOUR SUPPRESSOR GENES AS CHROMOSOMAL DELETIONS, EPIGENETIC SILENCING OR INACTIVATING/HYPOMORPHIC MUTATIONS. THE PROGNOSTIC VALUE OF THE MOST COMMON MUTATIONS AND THEIR PHENOTYPIC ASSOCIATION IN THE CLINICAL SETTING IS CURRENTLY UNDER INVESTIGATION. IT IS LIKELY THAT SPLICEOSOMAL MUTATIONS MAY INDICATE SENSITIVITY TO SPLICEOSOME INHIBITORS APPLIED IN THE FORM OF A SYNTHETIC LETHAL APPROACH. THIS REVIEW DISCUSSES THE MOST CURRENT ASPECTS OF SPLICEOSOMAL RESEARCH IN THE CONTEXT OF HAEMATOLOGICAL MALIGNANCIES. 2012