1 2262 153 EPIGENETIC PROFILING IN CHRONIC LYMPHOCYTIC LEUKEMIA REVEALS NOVEL METHYLATION TARGETS. CPG ISLAND METHYLATION IS AN EPIGENETIC ALTERATION THAT CONTRIBUTES TO TUMORIGENESIS BY TRANSCRIPTIONAL INACTIVATION OF GENES. LITTLE IS KNOWN ABOUT THE OVERALL LEVELS OF CPG ISLAND METHYLATION IN CHRONIC LYMPHOCYTIC LEUKEMIA (CLL). TO PROVIDE A BASELINE ESTIMATE OF GLOBAL ABERRANT METHYLATION AND IDENTIFY TARGET SEQUENCES FOR ADDITIONAL INVESTIGATION, WE PERFORMED RESTRICTION LANDMARK GENOMIC SCANNING ON 10 CLL SAMPLES. TWO METHYLATION-SENSITIVE LANDMARK ENZYMES WERE USED (NOTI AND ASCI), ALLOWING ASSESSMENT OF OVER 3000 CPG ISLANDS IN EACH SAMPLE. TUMOR-DERIVED RESTRICTION LANDMARK GENOMIC SCANNING PROFILES WERE COMPARED WITH PROFILES FROM CD19-SELECTED B CELLS FROM NORMAL VOLUNTEERS AND MATCHED NORMAL NEUTROPHILS FROM 4 CLL PATIENTS. WE FOUND 2.5-8.1% (MEAN 4.8%) OF THE CPG ISLANDS IN CLL SAMPLES WERE ABERRANTLY METHYLATED COMPARED WITH CONTROLS, AND THE METHYLATION EVENTS HAD A NONRANDOM DISTRIBUTION (P < 0.0001). FURTHERMORE, WE IDENTIFIED 193 ABERRANTLY METHYLATED SEQUENCES, OF WHICH 93% HAVE CPG ISLAND CHARACTERISTICS AND 90% HAVE HOMOLOGY TO GENES OR EXPRESSED SEQUENCES. ONE SUCH GENE, THE G PROTEIN-COUPLED METABOTROPIC GLUTAMATE RECEPTOR 7 (GRM7), POSSIBLY INHIBITS CYCLIC AMP SIGNALING IN THE INDUCTION OF APOPTOSIS. BISULFITE SEQUENCING OF GRM7 CONFIRMED EXTENSIVE CPG ISLAND METHYLATION, AND TREATMENT WITH 5-AZA-2'-DEOXYCYTIDINE (DECITABINE) RESULTED IN UP-REGULATED EXPRESSION OF SEVERAL GENES IN VITRO WITH CONCURRENT CELLULAR DEPLETION OF DNMT1 PROTEIN. OUR DUAL-ENZYME GLOBAL METHYLATION STUDY SHOWS THAT CLL IS CHARACTERIZED BY WIDESPREAD NONRANDOM CPG ISLAND METHYLATION SIMILAR TO OTHER TUMORS AND PROVIDES A PANEL OF NOVEL METHYLATION TARGETS THAT CAN BE USED IN LARGER STUDIES DESIGNED TO ASSESS IMPACT ON DISEASE PROGRESSION AND SURVIVAL.	2004	
                                                                                                                                                                                                                                                                                                                                                                                                             
2 4221  54 METHYLATION AND SILENCING OF PROTEIN TYROSINE PHOSPHATASE RECEPTOR TYPE O IN CHRONIC LYMPHOCYTIC LEUKEMIA. PURPOSE: PREVIOUS STUDIES IN OUR LABORATORY HAVE SHOWN THE PROGRESSIVE METHYLATION AND SUPPRESSION OF THE GENE ENCODING PROTEIN TYROSINE PHOSPHATASE, PTPRO, IN THE LIVERS OF RATS FED A METHYL-DEFICIENT DIET THAT INDUCES HEPATOCARCINOGENESIS. SUBSEQUENTLY, WE OBSERVED THE METHYLATION OF PTPRO IN PRIMARY HUMAN LUNG TUMORS AND ALSO SHOWED ITS POTENTIAL TUMOR SUPPRESSOR CHARACTERISTICS. THE PRESENT STUDY WAS UNDERTAKEN TO INVESTIGATE WHETHER THE TRUNCATED FORM OF PTPRO (PTPROT), SPECIFICALLY EXPRESSED IN NAIVE B LYMPHOCYTES, WAS ALSO METHYLATED AND SUPPRESSED IN CHRONIC LYMPHOCYTIC LEUKEMIA (CLL), A DISEASE GENERALLY AFFECTING B LYMPHOCYTES. EXPERIMENTAL DESIGN AND RESULTS: INITIAL SCREENING SHOWED THAT 60% OF THE 52 CLL SAMPLES ANALYZED USING METHYLATION-SPECIFIC PCR ASSAY WERE METHYLATED COMPARED WITH B LYMPHOCYTES FROM NORMAL INDIVIDUALS, WHICH WERE NOT METHYLATED. THE EXPRESSION OF PTPROT, AS MEASURED BY SEMIQUANTITATIVE REVERSE TRANSCRIPTION-PCR, INVERSELY CORRELATED WITH METHYLATION IN THE FEW SAMPLES TESTED. ANALYSIS OF ADDITIONAL SAMPLES (N = 50) BY COMBINED BISULFITE RESTRICTION ANALYSIS SHOWED THAT THE PTPRO CPG ISLAND WAS METHYLATED IN 82% OF PATIENTS WITH CLL COMPARED WITH B LYMPHOCYTES FROM NORMAL INDIVIDUALS. FURTHERMORE, OVERALL EXPRESSION OF PTPRO WAS REDUCED IN CLL RELATIVE TO NORMAL LYMPHOCYTES. THE PTPRO GENE WAS ALSO SUPPRESSED BY METHYLATION IN THE CLL CELL LINE WAC3CD5, WHERE IT COULD BE REACTIVATED UPON TREATMENT WITH THE DNA HYPOMETHYLATING AGENT 5-AZAC. ECTOPIC EXPRESSION OF PTPROT IN A NONEXPRESSING CELL LINE INCREASED GROWTH INHIBITION WITH FLUDARABINE TREATMENT, A THERAPY COMMONLY USED FOR CLL. CONCLUSION: THIS STUDY REVEALS THE POTENTIAL ROLE OF PTPRO METHYLATION AND SILENCING IN CLL TUMORIGENESIS AND ALSO PROVIDES A NOVEL MOLECULAR TARGET IN THE EPIGENETIC THERAPY.	2007	
                                                                                                                                                                                                                                                                                                                                                
3 1424  31 DIFFERENTIAL DNA METHYLATION OF GENE PROMOTERS IN SMALL B-CELL LYMPHOMAS. IMPROVED CARE OF PATIENTS WITH SMALL B-CELL LYMPHOMAS (SBCLS) IS LIKELY TO RESULT FROM THE ONGOING DISCOVERY OF MOLECULAR MARKERS THAT BETTER DEFINE THESE MALIGNANT NEOPLASMS. WE IDENTIFIED MULTIPLE GENE LOCI WHOSE DNA METHYLATION PATTERNS DIFFERED BETWEEN 3 TYPES OF SBCL: B-CELL CHRONIC LYMPHOCYTIC LEUKEMIA/SMALL LYMPHOCYTIC LYMPHOMA, MANTLE CELL LYMPHOMA, AND GRADES I AND II FOLLICULAR LYMPHOMA. THIS ANALYSIS WAS PERFORMED USING AN OLIGONUCLEOTIDE MICROARRAY THAT ALLOWED DETERMINATION OF THE DNA METHYLATION STATUS OF 156 LOCI IN 38 GENES. COMBINED BISULFITE RESTRICTION ANALYSIS AND METHYLATION-SPECIFIC POLYMERASE CHAIN REACTION WERE USED TO VALIDATE THE DIFFERENTIAL METHYLATION OF 6 OF THESE GENES. BY USING NON-HODGKIN LYMPHOMA CELL LINES AS MODELS, THESE GENES WERE EXAMINED FURTHER FOR METHYLATION AND GENE EXPRESSION RELATIONSHIPS. THIS STUDY ILLUSTRATES NONRANDOM EPIGENETIC ALTERATIONS IN SBCLS THAT SEEM TO PREFERENTIALLY INVOLVE LYMPHOMAS OF GERMINAL CENTER DERIVATION.	2005	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                  
4  494  39 ASSESSMENT OF PROMOTER METHYLATION IDENTIFIES PTCH AS A PUTATIVE TUMOR-SUPPRESSOR GENE IN HUMAN CLL. BACKGROUND: CHRONIC LYMPHOCYTIC LEUKEMIA (CLL) IS CHARACTERIZED BY A CLONAL ACCUMULATION OF NEOPLASTIC LYMPHOCYTES, INDICATING DISRUPTION OF APOPTOSIS. PATIENTS AND METHODS: DIFFERENTIAL METHYLATION HYBRIDIZATION ANALYSIS WAS PERFORMED TO IDENTIFY NOVEL TARGET GENES SILENCED BY CPG ISLAND METHYLATION IN PATIENTS WITH CLL. RESULTS: PATCHED (PTCH), A TUMOR-SUPPRESSOR GENE, WAS FOUND TO BE FREQUENTLY METHYLATED IN CLL SAMPLES COMPARED TO SAMPLES DERIVED FROM HEALTHY INDIVIDUALS. DE NOVO METHYLATION OF A CPG ISLAND REGION LOCATED UPSTREAM OF PTCH EXON 1 WAS CONFIRMED BY PYROSEQUENCING IN 17/37 (46%) OF PERIPHERAL BLOOD MONONUCLEAR CELLS OF PATIENTS WITH CLL, BUT IN NONE ISOLATED FROM SEVEN HEALTHY INDIVIDUALS. NO ASSOCIATION WAS FOUND BETWEEN PTCH HYPERMETHYLATION AND CURRENTLY USED PROGNOSTIC CLL FACTORS. CONCLUSION: OUR INVESTIGATION SUGGESTS THAT EPIGENETIC SILENCING OF PTCH IS A MECHANISM CONTRIBUTING TO CLL TUMORIGENESIS.	2016	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                           
5 3062  49 GENOME-WIDE DNA METHYLATION ANALYSIS REVEALS NOVEL EPIGENETIC CHANGES IN CHRONIC LYMPHOCYTIC LEUKEMIA. WE CONDUCTED A GENOME-WIDE DNA METHYLATION ANALYSIS IN CD19 (+) B-CELLS FROM CHRONIC LYMPHOCYTIC LEUKEMIA (CLL) PATIENTS AND NORMAL CONTROL SAMPLES USING REDUCED REPRESENTATION BISULFITE SEQUENCING (RRBS). THE METHYLATION STATUS OF 1.8-2.3 MILLION CPGS IN THE CLL GENOME WAS DETERMINED; ABOUT 45% OF THESE CPGS WERE LOCATED IN MORE THAN 23,000 CPG ISLANDS (CGIS). WHILE GLOBAL CPG METHYLATION WAS SIMILAR BETWEEN CLL AND NORMAL B-CELLS, 1764 GENE PROMOTERS WERE IDENTIFIED AS BEING DIFFERENTIALLY METHYLATED IN AT LEAST ONE CLL SAMPLE WHEN COMPARED WITH NORMAL B-CELL SAMPLES. NINETEEN PERCENT OF THE DIFFERENTIALLY METHYLATED GENES WERE INVOLVED IN TRANSCRIPTIONAL REGULATION. ABERRANT HYPERMETHYLATION WAS FOUND IN ALL HOX GENE CLUSTERS AND A SIGNIFICANT NUMBER OF WNT SIGNALING PATHWAY GENES. HYPOMETHYLATION OCCURRED MORE FREQUENTLY IN THE GENE BODY INCLUDING INTRONS, EXONS, AND 3'-UTRS IN CLL. THE NFATC1 P2 PROMOTER AND FIRST INTRON WAS FOUND TO BE HYPOMETHYLATED AND CORRELATED WITH UPREGULATION OF BOTH NFATC1 RNA AND PROTEIN EXPRESSION LEVELS IN CLL SUGGESTING THAT AN EPIGENETIC MECHANISM IS INVOLVED IN THE CONSTITUTIVE ACTIVATION OF NFAT ACTIVITY IN CLL CELLS. THIS COMPREHENSIVE DNA METHYLATION ANALYSIS WILL FURTHER OUR UNDERSTANDING OF THE EPIGENETIC CONTRIBUTION TO CELLULAR DYSFUNCTION IN CLL.	2012	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                  
6 4694  42 NEXT-GENERATION SEQUENCING IDENTIFIES MAJOR DNA METHYLATION CHANGES DURING PROGRESSION OF PH+ CHRONIC MYELOID LEUKEMIA. LITTLE IS KNOWN ABOUT THE IMPACT OF DNA METHYLATION ON THE EVOLUTION/PROGRESSION OF PH+ CHRONIC MYELOID LEUKEMIA (CML). WE INVESTIGATED THE METHYLOME OF CML PATIENTS IN CHRONIC PHASE (CP-CML), ACCELERATED PHASE (AP-CML) AND BLAST CRISIS (BC-CML) AS WELL AS IN CONTROLS BY REDUCED REPRESENTATION BISULFITE SEQUENCING. ALTHOUGH ONLY ~600 DIFFERENTIALLY METHYLATED CPG SITES WERE IDENTIFIED IN SAMPLES OBTAINED FROM CP-CML PATIENTS COMPARED WITH CONTROLS, ~6500 DIFFERENTIALLY METHYLATED CPG SITES WERE FOUND IN SAMPLES FROM BC-CML PATIENTS. IN THE MAJORITY OF AFFECTED CPG SITES, METHYLATION WAS INCREASED. IN CP-CML PATIENTS WHO PROGRESSED TO AP-CML/BC-CML, WE IDENTIFIED UP TO 897 GENES THAT WERE METHYLATED AT THE TIME OF PROGRESSION BUT NOT AT THE TIME OF DIAGNOSIS. USING RNA-SEQUENCING, WE OBSERVED DOWNREGULATED EXPRESSION OF MANY OF THESE GENES IN BC-CML COMPARED WITH CP-CML SAMPLES. SEVERAL OF THEM ARE WELL-KNOWN TUMOR-SUPPRESSOR GENES OR REGULATORS OF CELL PROLIFERATION, AND GENE RE-EXPRESSION WAS OBSERVED BY THE USE OF EPIGENETIC ACTIVE DRUGS. TOGETHER, OUR RESULTS DEMONSTRATE THAT CPG SITE METHYLATION CLEARLY INCREASES DURING CML PROGRESSION AND THAT IT MAY PROVIDE A USEFUL BASIS FOR REVEALING NEW TARGETS OF THERAPY IN ADVANCED CML.	2016	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                              
7 2753  32 EXPRESSION OF BCL2L12 IN CHRONIC LYMPHOCYTIC LEUKEMIA PATIENTS: ASSOCIATION WITH CLINICAL AND MOLECULAR PROGNOSTIC MARKERS. DYSREGULATION OF APOPTOSIS IS A DISTINCTIVE FEATURE OF CHRONIC LYMPHOCYTIC LEUKEMIA (CLL), ALTHOUGH A UNIQUE MECHANISM UNDERLYING APOPTOSIS RESISTANCE OF CLL B LYMPHOCYTES HAS NOT BEEN IDENTIFIED YET. ABERRANT EXPRESSION AS WELL AS GENETIC AND EPIGENETIC ALTERATIONS OF NUMEROUS GENES INVOLVED IN DIFFERENT PATHWAYS OF APOPTOSIS REGULATION HAS BEEN DESCRIBED IN CLL. HERE, WE REPORT THE EXPRESSION ANALYSIS OF BCL2L12 (BCL2-LIKE 12), A NOVEL APOPTOTIC GENE BELONGING TO BCL2 FAMILY, IN 58 SERBIAN CLL PATIENTS. QUANTITATIVE REVERSE-TRANSCRIPTASE POLYMERASE CHAIN REACTION (QRT-PCR) ANALYSIS REVEALED A SIGNIFICANT OVEREXPRESSION OF BCL2L12 MRNA IN CLL SAMPLES COMPARED TO NON-LEUKEMIC SAMPLES, IMPLYING ITS ROLE IN THE PATHOGENESIS OF THE DISEASE. RECEIVER OPERATING CHARACTERISTIC (ROC) ANALYSIS SHOWED THAT BCL2L12 EXPRESSION EFFICIENTLY DISCRIMINATES CLL CASES FROM HEALTHY CONTROLS. HOWEVER, RELATIVELY HOMOGENOUS BCL2L12 MRNA EXPRESSION AMONG PATIENTS DID NOT REFLECT THEIR CLINICAL CHARACTERISTICS (WITH THE EXCEPTION OF LACTATE DEHYDROGENASE STATUS AND TIME FROM DIAGNOSIS TO TREATMENT) AND FAILED TO SHOW ASSOCIATION WITH THE MOST INFORMATIVE PROGNOSTIC MARKERS, NAMELY THE MUTATIONAL STATUS OF REARRANGED IMMUNOGLOBULIN HEAVY CHAIN VARIABLE REGION GENES, CD38 AND LIPOPROTEIN LIPASE GENE (LPL) EXPRESSION.	2013	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                          
8 5275  40 PROMOTER METHYLATION OF THE BONE MORPHOGENETIC PROTEIN-6 GENE IN ASSOCIATION WITH ADULT T-CELL LEUKEMIA. BONE MORPHOGENETIC PROTEINS (BMP), BELONGING TO THE TRANSFORMING GROWTH FACTOR-BETA SUPERFAMILY, ARE MULTIFUNCTIONAL REGULATORS OF CELL PROLIFERATION, DIFFERENTIATION AND APOPTOSIS IN VARIOUS TYPES OF MALIGNANT CELLS. IN THIS STUDY, WE INVESTIGATED BMP-6 PROMOTER METHYLATION IN PATIENTS WITH VARIOUS TYPES OF LEUKEMIAS. THE BMP-6 METHYLATION WAS FOUND PREFERENTIALLY IN ADULT T-CELL LEUKEMIA (ATL) (49 OF 60, 82%) COMPARED WITH OTHER TYPES OF LEUKEMIAS STUDIED INCLUDING ACUTE MYELOID LEUKEMIA (3 OF 67, 5%), ACUTE LYMPHOBLASTIC LEUKEMIA (6 OF 38, 16%) AND CHRONIC LYMPHOCYTIC LEUKEMIA (1 OF 21, 5%). AMONG SUBTYPES OF ATL, THE BMP-6 GENE WAS MORE FREQUENTLY METHYLATED IN AGGRESSIVE ATL FORMS OF ACUTE (96%) AND LYMPHOMA (94%) TYPES THAN LESS MALIGNANT CHRONIC ATL (44%) AND SMOLDERING ATL (20%). WE ALSO ANALYZED THE METHYLATION STATUS OF PERIPHERAL BLOOD MONONUCLEAR CELLS FROM HEALTHY DONORS AND NONMALIGNANT LYMPH NODES WITH REACTIVE LYMPHADENOPATHY, NONE OF WHICH SHOWED DETECTABLE BMP-6 METHYLATION IN THIS STUDY. THE BMP-6 METHYALTION WAS CORRELATED WITH DECREASED MRNA TRANSCRIPT AND PROTEIN EXPRESSION. EXPRESSION OF BMP-6 WAS RESTORED BY THE DEMETHYLATING AGENT 5-AZA-2'-DEOXYCYTIDINE, SUGGESTING THAT METHYLATION WAS ASSOCIATED WITH THE TRANSCRIPTIONAL SILENCING. SERIAL ANALYSIS DEMONSTRATED AN INCREASING METHYLATION OF CPG SITES IN THE BMP-6 PROMOTER AND THE RESULTANT SUPPRESSION OF BMP-6 EXPRESSION AS ATL PROGRESSED. THESE FINDINGS SUGGESTED THAT BMP-6 PROMOTER METHYLATION IS LIKELY TO BE A COMMON EPIGENETIC EVENT AT LATER STAGES OF ATL AND THAT THE METHYLATION PROFILES MAY BE USEFUL FOR THE STAGING OF ATL AS WELL AS FOR EVALUATION OF THE INDIVIDUAL RISK OF DEVELOPING THE DISEASE.	2008	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                       
9   15  42 450K-ARRAY ANALYSIS OF CHRONIC LYMPHOCYTIC LEUKEMIA CELLS REVEALS GLOBAL DNA METHYLATION TO BE RELATIVELY STABLE OVER TIME AND SIMILAR IN RESTING AND PROLIFERATIVE COMPARTMENTS. IN CHRONIC LYMPHOCYTIC LEUKEMIA (CLL), THE MICROENVIRONMENT INFLUENCES GENE EXPRESSION PATTERNS; HOWEVER, KNOWLEDGE IS LIMITED REGARDING THE EXTENT TO WHICH METHYLATION CHANGES WITH TIME AND EXPOSURE TO SPECIFIC MICROENVIRONMENTS. USING HIGH-RESOLUTION 450K ARRAYS, WE PROVIDE THE MOST COMPREHENSIVE DNA METHYLATION STUDY OF CLL TO DATE, ANALYZING PAIRED DIAGNOSTIC/FOLLOW-UP SAMPLES FROM IGHV-MUTATED/UNTREATED AND IGHV-UNMUTATED/TREATED PATIENTS (N=36) AND PATIENT-MATCHED PERIPHERAL BLOOD AND LYMPH NODE SAMPLES (N=20). ON AN UNPRECEDENTED SCALE, WE REVEALED 2239 DIFFERENTIALLY METHYLATED CPG SITES BETWEEN IGHV-MUTATED AND UNMUTATED PATIENTS, WITH THE MAJORITY OF SITES POSITIONED OUTSIDE ANNOTATED CPG ISLANDS. INTRIGUINGLY, CLL PROGNOSTIC GENES (FOR EXAMPLE, CLLU1, LPL, ZAP70 AND NOTCH1), EPIGENETIC REGULATOR (FOR EXAMPLE, HDAC9, HDAC4 AND DNMT3B), B-CELL SIGNALING (FOR EXAMPLE, IBTK) AND NUMEROUS TGF-BETA AND NF-KAPPAB/TNF PATHWAY GENES WERE ALTERNATIVELY METHYLATED BETWEEN SUBGROUPS. CONTRARY, DNA METHYLATION OVER TIME WAS DEEMED RATHER STABLE WITH FEW RECURRENT CHANGES NOTED WITHIN SUBGROUPS. ALTHOUGH A LARGER NUMBER OF NON-RECURRENT CHANGES WERE IDENTIFIED AMONG IGHV-UNMUTATED RELATIVE TO MUTATED CASES OVER TIME, THESE EQUATED TO A LOW GLOBAL CHANGE. SIMILARLY, FEW CHANGES WERE IDENTIFIED BETWEEN COMPARTMENT CASES. ALTOGETHER, WE REVEAL CLL SUBGROUPS TO DISPLAY UNIQUE METHYLATION PROFILES AND UNVEIL METHYLATION AS RELATIVELY STABLE OVER TIME AND SIMILAR WITHIN DIFFERENT CLL COMPARTMENTS, IMPLYING ABERRANT METHYLATION AS AN EARLY LEUKEMOGENIC EVENT.	2013	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                               
10 3125  29 GHSR DNA HYPERMETHYLATION IS A COMMON EPIGENETIC ALTERATION OF HIGH DIAGNOSTIC VALUE IN A BROAD SPECTRUM OF CANCERS. IDENTIFICATION OF A SINGLE MOLECULAR TRAIT THAT IS DETERMINANT OF COMMON MALIGNANCIES MAY SERVE AS A POWERFUL DIAGNOSTIC SUPPLEMENT TO CANCER TYPE-SPECIFIC MARKERS. HERE, WE REPORT A DNA METHYLATION MARK THAT IS CHARACTERISTIC OF SEVEN STUDIED MALIGNANCIES, NAMELY CANCERS OF LUNG, BREAST, PROSTATE, PANCREAS, COLORECTUM, GLIOBLASTOMA AND B CELL CHRONIC LYMPHOCYTIC LEUKAEMIA (CLL) (N = 137). THIS MARK WAS DEFINED BY SUBSTANTIAL HYPERMETHYLATION AT THE PROMOTER AND FIRST EXON OF GROWTH HORMONE SECRETAGOUGE RECEPTOR (GHSR) THROUGH BISULFITE PYROSEQUENCING. THE DEGREE OF ABERRANT METHYLATION WAS CAPABLE OF ACCURATE DISCRIMINATION BETWEEN CANCER AND CONTROL SAMPLES. THE HIGHEST SENSITIVITY AND SPECIFICITY OF CANCER DETECTION WAS ACHIEVED FOR CANCERS OF PANCREAS, LUNG, BREAST AND CLL YIELDING THE AREA UNDER THE CURVE (AUC) VALUES OF 1.0000, 0.9952, 0.9800 AND 0.9400, RESPECTIVELY. NARROWING TO A SINGLE CPG SITE WITHIN THE GENE'S PROMOTER OR FOUR CONSECUTIVE CPG UNITS OF THE HIGHEST METHYLATION LEVELS WITHIN THE FIRST EXON IMPROVED THE DETECTION POWER. GHSR HYPERMETHYLATION WAS DETECTED ALREADY AT THE EARLY STAGE TUMORS. THE ACCURATE PERFORMANCE OF THIS MARKER WAS FURTHER REPLICATED IN AN INDEPENDENT SET OF PANCREATIC CANCER AND CONTROL SAMPLES (N = 78). THESE FINDINGS SUPPORT THE CANDIDATURE OF GHSR METHYLATION AS A HIGHLY ACCURATE PAN-CANCER MARKER.	2015	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                             
11 1144  43 CONCOMITANT HETEROCHROMATINISATION AND DOWN-REGULATION OF GENE EXPRESSION UNVEILS EPIGENETIC SILENCING OF RELB IN AN AGGRESSIVE SUBSET OF CHRONIC LYMPHOCYTIC LEUKEMIA IN MALES. BACKGROUND: THE SENSITIVITY OF CHRONIC LYMPHOCYTIC LEUKEMIA (CLL) CELLS TO CURRENT TREATMENTS, BOTH IN VITRO AND IN VIVO, RELIES ON THEIR ABILITY TO ACTIVATE APOPTOTIC DEATH. CLL CELLS RESISTANT TO DNA DAMAGE-INDUCED APOPTOSIS DISPLAY DEREGULATION OF A SPECIFIC SET OF GENES. METHODS: MICROARRAY HYBRIDIZATION (HUMAN GENECHIP, AFFYMETRIX), IMMUNOFLUORESCENT IN SITU LABELING COUPLED WITH VIDEO-MICROSCOPY RECORDING/ANALYSES, CHROMATIN-IMMUNOPRECIPITATION (CHIP), POLYMERASE CHAIN REACTIONS (PCR), REAL-TIME QUANTITATIVE PCR (RT-QPCR) AND BISULFITE GENOME SEQUENCING WERE THE MAIN METHODS APPLIED. STATISTICAL ANALYSES WERE PERFORMED BY APPLYING GCRMA AND SAM ANALYSIS (MICROARRAY DATA) AND STUDENT'S T-TEST OR MANN & WHITNEY'S U-TEST. RESULTS: HEREIN WE SHOW THAT, REMARKABLY, IN A RESISTANT MALE CLL CELLS THE VAST MAJORITY OF GENES WERE DOWN-REGULATED COMPARED WITH SENSITIVE CELLS, WHEREAS THIS WAS NOT THE CASE IN CELLS DERIVED FROM FEMALES. THIS GENE DOWN-REGULATION WAS FOUND TO BE ASSOCIATED WITH AN OVERALL GAIN OF HETEROCHROMATIN AS EVIDENCED BY IMMUNOFLUORESCENT LABELING OF HETEROCHROMATIN PROTEIN 1ALPHA (HP-1), TRIMETHYLATED HISTONE 3 LYSINE 9 (3METH3K9), AND 5-METHYLCYTIDINE (5METC). NOTABLY, 17 GENES WERE FOUND TO BE COMMONLY DEREGULATED IN RESISTANT MALE AND FEMALE CELL SAMPLES. AMONG THESE, RELB WAS IDENTIFIED AS A DISCRIMINATORY CANDIDATE GENE REPRESSED IN THE MALE AND UPREGULATED IN THE FEMALE RESISTANT CELLS. CONCLUSION: THE MOLECULAR DEFECTS IN THE SILENCING OF RELB INVOLVE AN INCREASE IN H3K9- BUT NOT CPG-ISLAND METHYLATION IN THE PROMOTER REGIONS. INCREASE IN ACETYL-H3 IN RESISTANT FEMALE BUT NOT MALE CLL SAMPLES AS WELL AS A DECREASE OF TOTAL CELLULAR LEVEL OF RELB AFTER AN INHIBITION OF HISTONE DEACETYLASE (HDAC) BY TRICHOSTATIN A (TSA), FURTHER EMPHASIZE THE ROLE OF EPIGENETIC MODIFICATIONS WHICH COULD DISCRIMINATE TWO CLL SUBSETS. TOGETHER, THESE RESULTS HIGHLIGHTED THE EPIGENETIC RELB SILENCING AS A NEW MARKER OF THE PROGRESSIVE DISEASE IN MALES.	2010	
                                                                                                
12 5871  42 SUSTAINED NF-KAPPAB ACTIVITY IN CHRONIC LYMPHOCYTIC LEUKEMIA IS INDEPENDENT OF GENETIC AND EPIGENETIC ALTERATIONS IN THE TNFAIP3 (A20) LOCUS. INAPPROPRIATE NUCLEAR FACTOR (NF) KAPPAB ACTIVITY IS ONE MAJOR HALLMARK OF B-CELL MALIGNANCIES AND CHRONIC LYMPHOCYTIC LEUKEMIA (CLL). NFKAPPAB-DEPENDENT GENES ARE INVOLVED IN ANTIAPOPTOSIS, CELL PROLIFERATION AND METASTASIS AND ARE RESPONSIBLE FOR SURVIVAL AND PROLIFERATION OF TUMORS. HOWEVER, THE MECHANISMS OF NFKAPPAB ACTIVITY IN CLL STILL NEED TO BE ELUCIDATED. PREVIOUSLY, WE IDENTIFIED TRANSLOCATIONS IN A REGION ON CHROMOSOME 6Q THAT ENCODES TUMOR NECROSIS FACTOR ALPHA-INDUCED PROTEIN 3, WHICH IS A KEY PLAYER IN NEGATIVE FEEDBACK LOOP REGULATION OF NFKAPPAB. INACTIVATION OF THIS UBIQUITIN-EDITING ENZYME IS INVOLVED IN IMMUNOPATHOLOGIES AND IN TUMORIGENESIS. FREQUENT MUTATIONS IN THE A20 LOCUS--LEADING TO SUSTAINED NFKAPPAB ACTIVITY--COULD BE SHOWN TO PLAY A DOMINANT ROLE IN DEVELOPMENT OF DIFFERENT B-CELL MALIGNANCIES. TO CHECK IF A20 IS INVOLVED IN UPREGULATION OF NFKAPPAB ACTIVITY IN CLL, WE SEQUENCED EXONS 2-9 OF THE A20 GENE IN 55 CLL DNA SAMPLES. FURTHERMORE, WE DETERMINED THE METHYLATION STATUS OF THE PROMOTER REGION IN 63 CLL DNA SAMPLES AND COMPARED TO 10 CONTROL DNAS OF B CELLS FROM HEALTHY DONORS. CONTRARY TO REPORTS FROM OTHER B-CELL MALIGNANCIES, THE A20 REGION SHOWED NEITHER MUTATIONS NOR ABERRANT DNA METHYLATION. MOREOVER, ITS EXPRESSION COULD BE CONFIRMED BY IMMUNOBLOTTING AND SHOWING COMPARABLE RESULTS TO HEALTHY B CELLS. THESE RESULTS INDICATE THAT MALIGNANT DEVELOPMENT IN CLL DIFFERS FROM MOST OF OTHER B-CELL MALIGNANCIES, WHICH SHOW FREQUENT INACTIVATION OF A20.	2011	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                   
13 5270  30 PROMOTER DNA METHYLATION FREQUENCY AND CLINICOPATHOLOGICAL ROLE OF MIR-129-2 GENE IN PATIENTS WITH CHRONIC LYMPHOCYTIC LEUKEMIA. OBJECTIVES: CHRONIC LYMPHOCYTIC LEUKEMIA (CLL) IS CHARACTERIZED BY THE ACCUMULATION OF APPARENTLY MATURE B-TYPE LYMPHOCYTES IN THE LYMPHOHEMATOPOIETIC ORGANS. METHYLATION IN PROMOTERS OF TUMOR SUPPRESSOR GENES IS ONE OF THE MECHANISMS THAT CAUSES BLOOD MALIGNANCY. IN THIS STUDY, WE EVALUATED THE PROMOTER DNA METHYLATION STATUS OF MIR-129-2 TUMOR SUPPRESSOR GENE AND ITS ASSOCIATION WITH CLINICAL AND LABORATORY PARAMETERS OF PATIENTS WITH CLL. METHODS: WE STUDIED THE PROMOTER DNA METHYLATION FREQUENCY OF THE MIR-129-2 GENE IN 50 PATIENTS WITH CLL AND 50 HEALTHY CONTROLS USING METHYLATION-SPECIFIC POLYMERASE CHAIN REACTION METHODS. STATISTICAL ANALYSIS WAS PERFORMED USING SPSS-18 SOFTWARE, AND A P-VALUE < 0.050 WAS CONSIDERED STATISTICALLY SIGNIFICANT. RESULTS: THE FREQUENCY OF PROMOTER DNA METHYLATION OF THE MIR-129-2 GENE WAS SIGNIFICANTLY HIGHER IN THE CLL GROUP COMPARED WITH CONTROL GROUP (38.0% VS. 0.0%, P < 0.001; CHI(2) = 23.457). THE PROMOTER DNA METHYLATION FREQUENCY OF MIR-129-2 GENE WAS NOT SIGNIFICANTLY DIFFERENT BETWEEN THE TWO SEXES (P = 0.236). A SIGNIFICANT BUT WEAK CORRELATION WAS SEEN BETWEEN THE METHYLATED STATE OF THE MIR-129-2 GENE AND ORGANOMEGALY (P = 0.019, R = 0.330) AS WELL AS HEMOGLOBIN LEVELS (P = 0.020, R = -0.233). HOWEVER, BINARY LOGISTIC REGRESSION ANALYSIS INDICATED ORGANOMEGALY AS THE ONLY CLINICAL BIOMARKER WITH A STATISTICALLY SIGNIFICANT ASSOCIATION WITH THE HYPERMETHYLATED MIR-129-2 GENE STATE (P = 0.046). CONCLUSIONS: THE HIGH FREQUENCY OF PROMOTER DNA METHYLATION OF THE MIR-129-2 GENE IN THE CLL GROUP COMPARED TO THE CONTROL GROUP, AS WELL AS ITS SIGNIFICANT ASSOCIATION WITH ORGANOMEGALY, SUGGESTS THE IMPORTANCE OF THIS EPIGENETIC BIOMARKER IN THE PATHOGENESIS AND PROGNOSIS OF CLL DISEASE.	2020	
                                                                                                                                                                                                                                                                                                                                                                                           
14 6404  39 THE RS1001179 SNP AND CPG METHYLATION REGULATE CATALASE EXPRESSION IN CHRONIC LYMPHOCYTIC LEUKEMIA. CHRONIC LYMPHOCYTIC LEUKEMIA (CLL) IS AN INCURABLE DISEASE CHARACTERIZED BY AN EXTREMELY VARIABLE CLINICAL COURSE. WE HAVE RECENTLY SHOWN THAT HIGH CATALASE (CAT) EXPRESSION IDENTIFIES PATIENTS WITH AN AGGRESSIVE CLINICAL COURSE. ELUCIDATING MECHANISMS REGULATING CAT EXPRESSION IN CLL IS PREEMINENT TO UNDERSTAND DISEASE MECHANISMS AND DEVELOP STRATEGIES FOR IMPROVING ITS CLINICAL MANAGEMENT. IN THIS STUDY, WE INVESTIGATED THE ROLE OF THE CAT PROMOTER RS1001179 SINGLE NUCLEOTIDE POLYMORPHISM (SNP) AND OF THE CPG ISLAND II METHYLATION ENCOMPASSING THIS SNP IN THE REGULATION OF CAT EXPRESSION IN CLL. LEUKEMIC CELLS HARBORING THE RS1001179 SNP T ALLELE EXHIBITED A SIGNIFICANTLY HIGHER CAT EXPRESSION COMPARED WITH CELLS BEARING THE CC GENOTYPE. CAT PROMOTER HARBORING THE T -BUT NOT C- ALLELE WAS ACCESSIBLE TO ETS-1 AND GR-BETA TRANSCRIPTION FACTORS. MOREOVER, CLL CELLS EXHIBITED LOWER METHYLATION LEVELS THAN NORMAL B CELLS, IN LINE WITH THE HIGHER CAT MRNA AND PROTEIN EXPRESSED BY CLL IN COMPARISON WITH NORMAL B CELLS. METHYLATION LEVELS AT SPECIFIC CPG SITES NEGATIVELY CORRELATED WITH CAT LEVELS IN CLL CELLS. INHIBITION OF METHYLTRANSFERASE ACTIVITY INDUCED A SIGNIFICANT INCREASE IN CAT LEVELS, THUS FUNCTIONALLY VALIDATING THE ROLE OF CPG METHYLATION IN REGULATING CAT EXPRESSION IN CLL. FINALLY, THE CT/TT GENOTYPES WERE ASSOCIATED WITH LOWER METHYLATION AND HIGHER CAT LEVELS, SUGGESTING THAT THE RS1001179 T ALLELE AND CPG METHYLATION MAY INTERACT IN REGULATING CAT EXPRESSION IN CLL. THIS STUDY IDENTIFIES GENETIC AND EPIGENETIC MECHANISMS UNDERLYING DIFFERENTIAL EXPRESSION OF CAT, WHICH COULD BE OF CRUCIAL RELEVANCE FOR THE DEVELOPMENT OF THERAPIES TARGETING REDOX REGULATORY PATHWAYS IN CLL.	2022	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                
15  139  35 ABERRANT DNA METHYLATION IS ASSOCIATED WITH DISEASE PROGRESSION, RESISTANCE TO IMATINIB AND SHORTENED SURVIVAL IN CHRONIC MYELOGENOUS LEUKEMIA. THE EPIGENETIC IMPACT OF DNA METHYLATION IN CHRONIC MYELOGENOUS LEUKEMIA (CML) IS NOT COMPLETELY UNDERSTOOD. TO ELUCIDATE ITS ROLE WE ANALYZED 120 PATIENTS WITH CML FOR METHYLATION OF PROMOTER-ASSOCIATED CPG ISLANDS OF 10 GENES. FIVE GENES WERE IDENTIFIED BY DNA METHYLATION SCREENING IN THE K562 CELL LINE AND 3 GENES IN PATIENTS WITH MYELOPROLIFERATIVE NEOPLASMS. THE CDKN2B GENE WAS SELECTED FOR ITS FREQUENT METHYLATION IN MYELOID MALIGNANCIES AND ABL1 AS THE TARGET OF BCR-ABL TRANSLOCATION. THIRTY PATIENTS WERE IMATINIB-NAIVE (MOSTLY TREATED BY INTERFERON-ALPHA BEFORE THE IMATINIB ERA), 30 WERE IMATINIB-RESPONSIVE, 50 WERE IMATINIB-RESISTANT, AND 10 WERE IMATINIB-INTOLERANT. WE QUANTIFIED DNA METHYLATION BY BISULFITE PYROSEQUENCING. THE AVERAGE NUMBER OF METHYLATED GENES WAS 4.5 PER PATIENT IN THE CHRONIC PHASE, INCREASING SIGNIFICANTLY TO 6.2 IN THE ACCELERATED AND 6.4 IN THE BLASTIC PHASE. HIGHER NUMBERS OF METHYLATED GENES WERE ALSO OBSERVED IN PATIENTS RESISTANT OR INTOLERANT TO IMATINIB. THESE PATIENTS ALSO SHOWED ALMOST EXCLUSIVE METHYLATION OF A PUTATIVE TRANSPORTER OSCP1. ABNORMAL METHYLATION OF A SRC SUPPRESSOR GENE PDLIM4 WAS ASSOCIATED WITH SHORTENED SURVIVAL INDEPENDENTLY OF CML STAGE AND IMATINIB RESPONSIVENESS. WE CONCLUDE THAT ABERRANT DNA METHYLATION IS ASSOCIATED WITH CML PROGRESSION AND THAT DNA METHYLATION COULD BE A MARKER ASSOCIATED WITH IMATINIB RESISTANCE. FINALLY, DNA METHYLATION OF PDLIM4 MAY HELP IDENTIFY A SUBSET OF CML PATIENTS THAT WOULD BENEFIT FROM TREATMENT WITH SRC/ABL INHIBITORS.	2011	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                    
16   59  38 A GENOME-WIDE SCREEN IDENTIFIES FREQUENTLY METHYLATED GENES IN HAEMATOLOGICAL AND EPITHELIAL CANCERS. BACKGROUND: GENETIC AS WELL AS EPIGENETIC ALTERATIONS ARE A HALLMARK OF BOTH EPITHELIAL AND HAEMATOLOGICAL MALIGNANCIES. HIGH THROUGHPUT SCREENS ARE REQUIRED TO IDENTIFY EPIGENETIC MARKERS THAT CAN BE USEFUL FOR DIAGNOSTIC AND PROGNOSTIC PURPOSES ACROSS MALIGNANCIES. RESULTS: HERE WE REPORT FOR THE FIRST TIME THE USE OF THE MIRA ASSAY (METHYLATED CPG ISLAND RECOVERY ASSAY) IN COMBINATION WITH GENOME-WIDE CPG ISLAND ARRAYS TO IDENTIFY EPIGENETIC MOLECULAR MARKERS IN CHILDHOOD ACUTE LYMPHOBLASTIC LEUKEMIA (ALL) ON A GENOME-WIDE SCALE. WE IDENTIFIED 30 GENES DEMONSTRATING METHYLATION FREQUENCIES OF > OR =25% IN CHILDHOOD ALL, NINE GENES SHOWED SIGNIFICANTLY DIFFERENT METHYLATION FREQUENCIES IN B VS T-ALL. FOR MAJORITY OF THE GENES EXPRESSION COULD BE RESTORED IN METHYLATED LEUKEMIA LINES AFTER TREATMENT WITH 5-AZADC. FORTY-FOUR PERCENT OF THE GENES REPRESENT TARGETS OF THE POLYCOMB COMPLEX. IN CHRONIC MYELOID LEUKEMIA (CML) TWO OF THE GENES, (TFAP2A AND EBF2), DEMONSTRATED INCREASED METHYLATION IN BLAST CRISIS COMPARED TO CHRONIC PHASE (P < 0.05). FURTHERMORE HYPERMETHYLATION OF AN AUTOPHAGY RELATED GENE ATG16L2 WAS ASSOCIATED WITH POORER PROGNOSIS IN TERMS OF MOLECULAR RESPONSE TO IMATINIB TREATMENT. LASTLY WE DEMONSTRATED THAT TEN OF THESE GENES WERE ALSO FREQUENTLY METHYLATED IN COMMON EPITHELIAL CANCERS. CONCLUSION: IN SUMMARY WE HAVE IDENTIFIED A LARGE NUMBER OF GENES SHOWING FREQUENT METHYLATION IN CHILDHOOD ALL, METHYLATION STATUS OF TWO OF THESE GENES IS ASSOCIATED WITH ADVANCED DISEASE IN CML AND METHYLATION STATUS OF ANOTHER GENE IS ASSOCIATED WITH PROGNOSIS. IN ADDITION A SUBSET OF THESE GENES MAY ACT AS EPIGENETIC MARKERS ACROSS HEMATOLOGICAL MALIGNANCIES AS WELL AS COMMON EPITHELIAL CANCERS.	2010	
                                                                                                                                                                                                                                                                                                                                                                                                                                               
17 6613  37 ULTRADEEP BISULFITE SEQUENCING ANALYSIS OF DNA METHYLATION PATTERNS IN MULTIPLE GENE PROMOTERS BY 454 SEQUENCING. WE DEVELOPED A NOVEL APPROACH FOR CONDUCTING MULTISAMPLE, MULTIGENE, ULTRADEEP BISULFITE SEQUENCING ANALYSIS OF DNA METHYLATION PATTERNS IN CLINICAL SAMPLES. A MASSIVELY PARALLEL SEQUENCING-BY-SYNTHESIS METHOD (454 SEQUENCING) WAS USED TO DIRECTLY SEQUENCE >100 BISULFITE PCR PRODUCTS IN A SINGLE SEQUENCING RUN WITHOUT SUBCLONING. WE SHOWED THE UTILITY, ROBUSTNESS, AND SUPERIORITY OF THIS APPROACH BY ANALYZING METHYLATION IN 25 GENE-RELATED CPG RICH REGIONS FROM >40 CASES OF PRIMARY CELLS, INCLUDING NORMAL PERIPHERAL BLOOD LYMPHOCYTES, ACUTE LYMPHOBLASTIC LEUKEMIA (ALL), CHRONIC LYMPHOCYTIC LEUKEMIA (CLL), FOLLICULAR LYMPHOMA (FL), AND MANTLE CELL LYMPHOMA (MCL). A TOTAL OF 294,631 SEQUENCES WAS GENERATED WITH AN AVERAGE READ LENGTH OF 131 BP. ON AVERAGE, >1,600 INDIVIDUAL SEQUENCES WERE GENERATED FOR EACH PCR AMPLICON FAR BEYOND THE FEW CLONES (<20) TYPICALLY ANALYZED BY TRADITIONAL BISULFITE SEQUENCING. COMPREHENSIVE ANALYSIS OF CPG METHYLATION PATTERNS AT A SINGLE DNA MOLECULE LEVEL USING CLUSTERING ALGORITHMS REVEALED DIFFERENTIAL METHYLATION PATTERNS BETWEEN DISEASES. A SIGNIFICANT INCREASE IN METHYLATION WAS DETECTED IN ALL AND FL SAMPLES COMPARED WITH CLL AND MCL. FURTHERMORE, A PROGRESSIVE SPREADING OF METHYLATION WAS DETECTED FROM THE PERIPHERY TOWARD THE CENTER OF SELECT CPG ISLANDS IN THE ALL AND FL SAMPLES. THE ULTRADEEP SEQUENCING ALSO ALLOWED SIMULTANEOUS ANALYSIS OF GENETIC AND EPIGENETIC DATA AND REVEALED AN ASSOCIATION BETWEEN A SINGLE NUCLEOTIDE POLYMORPHISM AND THE METHYLATION PRESENT IN THE LRP1B PROMOTER. THIS NEW GENERATION OF METHYLOME SEQUENCING WILL PROVIDE DIGITAL PROFILES OF ABERRANT DNA METHYLATION FOR INDIVIDUAL HUMAN CANCERS AND OFFERS A ROBUST METHOD FOR THE EPIGENETIC CLASSIFICATION OF TUMOR SUBTYPES.	2007	
                                                                                                                                                                                                                                                                                                                                                                                                    
18 1995  35 EPIGENETIC AND GENETIC ALTERATIONS AND THEIR INFLUENCE ON GENE REGULATION IN CHRONIC LYMPHOCYTIC LEUKEMIA. BACKGROUND: TO UNDERSTAND THE CHANGES OF GENE REGULATION IN CARCINOGENESIS, WE EXPLORED SIGNALS OF DNA METHYLATION - A STABLE EPIGENETIC MARK OF GENE REGULATORY ELEMENTS - AND DESIGNED A COMPUTATIONAL MODEL TO PROFILE LOSS AND GAIN OF REGULATORY ELEMENTS (RES) DURING CARCINOGENESIS. WE ALSO UTILIZED SEQUENCING DATA TO ANALYZE THE ALLELE FREQUENCY OF SINGLE NUCLEOTIDE POLYMORPHISMS (SNPS) AND DETECTED THE CANCER-ASSOCIATED SNPS, I.E., THE SNPS DISPLAYING THE SIGNIFICANT ALLELE FREQUENCY DIFFERENCE BETWEEN CANCER AND NORMAL SAMPLES. RESULTS: AFTER APPLYING THIS MODEL TO CHRONIC LYMPHOCYTIC LEUKEMIA (CLL) DATA, WE IDENTIFIED RES DIFFERENTIALLY ACTIVATED (DRES) BETWEEN NORMAL AND CLL CELLS, CONSISTING OF 6,802 DRES GAINED AND 4,606 DRES LOST IN CLL. THE IDENTIFIED REGULATORY PERTURBATIONS COINCIDE WITH CHANGES IN THE EXPRESSION OF TARGET GENES. IN PARTICULAR, THE GENES ENCODING DNA METHYLTRANSFERASES HARBOR MULTIPLE LOST-IN-CANCER DRES AND ZERO GAINED-IN-CANCER DRES, INDICATING THAT THE DAMAGED REGULATION OF THESE GENES MIGHT BE ONE OF THE KEY CAUSES OF TUMOR FORMATION. DRES DISPLAY A SIGNIFICANTLY ELEVATED DENSITY OF THE GENOME-WIDE ASSOCIATION STUDY (GWAS) SNPS ASSOCIATED WITH CLL AND CLL-RELATED TRAITS. WE OBSERVED THAT MOST OF DRE GWAS SNPS ASSOCIATED WITH CLL AND CLL-RELATED TRAITS (83%) DISPLAY A SIGNIFICANT HAPLOTYPE ASSOCIATION AMONG THE IDENTIFIED CANCER-ASSOCIATED ALLELES AND THE RISK ALLELES THAT HAVE BEEN REPORTED IN GWAS. ALSO DRES ARE ENRICHED FOR THE BINDING SITES OF THE WELL-ESTABLISHED B-CELL AND CLL TRANSCRIPTION FACTORS (TFS) NF-KB, AP2, P53, E2F1, PAX5, AND SP1. WE ALSO IDENTIFIED CLL-ASSOCIATED SNPS AND DEMONSTRATED THAT THE MUTATIONS AT THESE SNPS CHANGE THE BINDING SITES OF KEY TFS MUCH MORE FREQUENTLY THAN EXPECTED. CONCLUSIONS: THROUGH EXPLORING SEQUENCING DATA MEASURING DNA METHYLATION, WE IDENTIFIED THE EPIGENETIC ALTERATIONS (MORE SPECIFICALLY, DNA METHYLATION) AND GENETIC MUTATIONS ALONG NON-CODING GENOMIC REGIONS CLL, AND DEMONSTRATED THAT THESE CHANGES PLAY A CRITICAL ROLE IN CARCINOGENESIS THROUGH DAMAGING THE REGULATION OF KEY GENES AND ALTERNATING THE BINDING OF KEY TFS IN B AND CLL CELLS.	2017	

19 5269  40 PROMOTER DNA METHYLATION CONTRIBUTES TO HUMAN BETA-DEFENSIN-1 DEFICIENCY IN ATOPIC DERMATITIS. ATOPIC DERMATITIS (AD) IS A CHRONIC INFLAMMATORY SKIN DISEASE CAUSED BY EPIDERMAL BARRIER DYSFUNCTION AND DYSREGULATION OF INNATE AND ADAPTIVE IMMUNITY. EPIGENETIC REGULATION OF HUMAN BETA-DEFENSIN-1 (HBD-1) MIGHT BE ASSOCIATED WITH A VARIETY OF DEFECTS IN THE INNATE IMMUNE SYSTEM DURING AD PATHOGENESIS. WE INVESTIGATED THE POSSIBLE MECHANISM OF DECREASED HBD-1 GENE EXPRESSION IN AD AND DEMONSTRATED THE RESTORATION OF HBD-1 TRANSCRIPTION IN UNDIFFERENTIATED NORMAL HUMAN EPIDERMAL KERATINOCYTE CELLS AFTER TREATMENT WITH A DNA METHYLTRANSFERASE INHIBITOR. WE ALSO CONDUCTED AN IN VITRO METHYLATED REPORTER ASSAY USING A REPORTER CONTAINING 14 CPG SITES. METHYLATION OF THE 14 CPG SITES WITHIN THE HBD-1 5' REGION RESULTED IN AN APPROXIMATELY 86% REDUCTION IN PROMOTER ACTIVITY AND AFFECTED HBD-1 TRANSCRIPTIONAL REGULATION. WE THEN COMPARED METHYLATION FREQUENCIES AT CPG 3 AND CPG 4 BETWEEN NON-LESIONAL AND LESIONAL EPIDERMIS SAMPLES OF PATIENTS WITH SEVERE AD AND BETWEEN THESE PAIRED TISSUES AND HEALTHY CONTROL EPIDERMIS FROM NORMAL VOLUNTEERS WITHOUT AD HISTORY. BISULFITE PYROSEQUENCING DATA SHOWED SIGNIFICANTLY HIGHER METHYLATION FREQUENCIES AT THE CPG 3 AND 4 SITES IN AD LESIONAL SAMPLES THAN IN NON-LESIONAL AD SKIN AND NORMAL SKIN SAMPLES (P < 0.05). THESE RESULTS SUGGEST THAT THE DNA METHYLATION SIGNATURE OF HBD-1 IS A NOVEL DIAGNOSTIC/PROGNOSTIC MARKER AND A PROMISING THERAPEUTIC TARGET FOR THE COMPROMISED STRATUM CORNEUM BARRIER ATTRIBUTED TO HBD-1 DEFICIENCY.	2018	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                            
20 3147  41 GLP OVEREXPRESSION IS ASSOCIATED WITH POOR PROGNOSIS IN CHRONIC LYMPHOCYTIC LEUKEMIA AND ITS INHIBITION INDUCES LEUKEMIC CELL DEATH. BACKGROUND HETERODIMERIC METHYLTRANSFERASES GLP (EHMT1/KMT1D) AND G9A (EHMT2/KMT1C) ARE TWO CLOSELY RELATED ENZYMES THAT PROMOTE THE MONOMETHYLATION AND DIMETHYLATION OF HISTONE H3 LYSINE 9. DYSREGULATION OF THEIR ACTIVITY HAS BEEN IMPLICATED IN SEVERAL TYPES OF HUMAN CANCER. PATIENTS AND METHODS HERE, IN ORDER TO INVESTIGATE WHETHER GLP/G9A EXERTS ANY IMPACT ON CHRONIC LYMPHOCYTIC LEUKEMIA (CLL), GLP/G9A EXPRESSION LEVELS WERE ASSESSED IN A COHORT OF 50 PATIENTS AND THE EFFECTS OF THEIR INHIBITION WERE VERIFIED FOR THE VIABILITY OF CLL CELLS. ALSO, QRT-PCR WAS USED TO INVESTIGATE THE TRANSCRIPTIONAL LEVELS OF GLP/G9A IN CLL PATIENTS. IN ADDITION, PATIENT SAMPLES WERE CLASSIFIED ACCORDING TO ZAP-70 PROTEIN EXPRESSION BY FLOW CYTOMETRY AND ACCORDING TO KARYOTYPE INTEGRITY BY CYTOGENETICS ANALYSIS. FINALLY, A SELECTIVE SMALL MOLECULE INHIBITOR FOR GLP/G9A WAS USED TO ASCERTAIN WHETHER THESE METHYLTRANSFERASES INFLUENCED THE VIABILITY OF MEC-1 CLL CELL LINEAGE. RESULTS MRNA ANALYSIS REVEALED THAT CLL SAMPLES HAD HIGHER LEVELS OF GLP, BUT NOT G9A, WHEN COMPARED TO NON-LEUKEMIC CONTROLS. INTERESTINGLY, PATIENTS WITH UNFAVORABLE CYTOGENETICS SHOWED HIGHER EXPRESSION LEVELS OF GLP COMPARED TO PATIENTS WITH FAVORABLE KARYOTYPES. MORE IMPORTANTLY, GLP/G9A INHIBITION MARKEDLY INDUCED CELL DEATH IN CLL CELLS. CONCLUSION TAKEN TOGETHER, THESE RESULTS INDICATE THAT GLP IS ASSOCIATED WITH A WORSE PROGNOSIS IN CLL, AND THAT THE INHIBITION OF GLP/G9A INFLUENCES CLL CELL VIABILITY. ALTOGETHER, THE PRESENT DATA DEMONSTRATE THAT THESE METHYLTRANSFERASES CAN BE POTENTIAL MARKERS FOR DISEASE PROGRESSION, AS WELL AS A PROMISING EPIGENETIC TARGET FOR CLL TREATMENT AND THE PREVENTION OF DISEASE EVOLUTION.	2018