1 2678 136 EVALUATION OF A PROGNOSTIC EPIGENETIC CLASSIFICATION SYSTEM IN CHRONIC LYMPHOCYTIC LEUKEMIA PATIENTS. BACKGROUND: METHYLATION AT 5 CPG SITES WAS PREVIOUSLY SHOWN TO CLASSIFY CHRONIC LYMPHOCYTIC LEUKEMIA (CLL) INTO 3 PROGNOSTIC SUBGROUPS. HERE, WE AIMED TO VALIDATE THE MARKER SET IN AN ADDITIONAL COHORT AND TO EVALUATE ITS CLINICAL UTILITY FOR CLL PATIENT STRATIFICATION. METHODS: WE EVALUATED THIS EPIGENETIC MARKER SET IN 79 GERMAN PATIENTS USING BISULFITE TREATMENT FOLLOWED BY PYROSEQUENCING AND CLASSIFICATION USING A SUPPORT VECTOR MACHINE-LEARNING TOOL. RESULTS: THE N-CLL, I-CLL, AND M-CLL CLASSIFICATION WAS DETECTED IN 28 (35%), 10 (13%), AND 41 (51%) PATIENTS, RESPECTIVELY. EPIGENETIC GROUPING WAS ASSOCIATED WITH IGHV MUTATIONAL STATUS (P = 2 X 10(-12)), ISOLATED DEL13Q (P = 9 X 10(-6)), DEL17P (P = .015), COMPLEX KARYOTYPE (P = .005), VH-USAGE, AND CLINICAL OUTCOME AS TIME TO FIRST TREATMENT (P = 1.4 X 10(-12)) AND OVERALL SURVIVAL (P = .003). MULTIVARIATE COX REGRESSION ANALYSIS IDENTIFIED N-CLL AS A FACTOR FOR EARLIER TREATMENT HAZARD RATIO (HR), 6.3 (95% CONFIDENCE INTERVAL [CI] 2.4-16.4; P = .0002) COMPARED TO IGHV MUTATIONAL STATUS (HR 4.6, 95% CI 1.9-11.3, P = .0008). IN ADDITION, WHEN COMPARING THE PROGNOSTIC VALUE OF THE EPIGENETIC CLASSIFICATION SYSTEM WITH THE IGHV CLASSIFICATION, EPIGENETIC GROUPING PERFORMED BETTER COMPARED TO IGHV MUTATIONAL STATUS USING KAPLAN-MEIER ESTIMATION AND ALLOWED THE IDENTIFICATION OF A THIRD, INTERMEDIATE (I-CLL) GROUP. THUS, OUR STUDY CONFIRMED THE PROGNOSTIC VALUE OF THE EPIGENETIC MARKER SET FOR PATIENT STRATIFICATION IN ROUTINE CLINICAL DIAGNOSTICS.	2022	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                  
2 5243  51 PROGNOSTIC IMPACT OF EPIGENETIC CLASSIFICATION IN CHRONIC LYMPHOCYTIC LEUKEMIA: THE CASE OF SUBSET #2. BASED ON THE METHYLATION STATUS OF 5 SINGLE CPG SITES, A NOVEL EPIGENETIC CLASSIFICATION OF CHRONIC LYMPHOCYTIC LEUKEMIA (CLL) WAS RECENTLY PROPOSED, CLASSIFYING CLL PATIENTS INTO 3 CLINICO-BIOLOGICAL SUBGROUPS WITH DIFFERENT OUTCOME, TERMED MEMORY LIKE CLL (M-CLL), NAIVE LIKE CLL (N-CLL), AND A THIRD INTERMEDIATE CLL SUBGROUP (I-CLL). WHILE M-CLL AND N-CLL PATIENTS AT LARGE CORRESPONDED TO PATIENTS CARRYING MUTATED AND UNMUTATED IGHV GENES, RESPECTIVELY, LIMITED INFORMATION EXISTS REGARDING THE LESS DEFINED I-CLL GROUP. USING PYROSEQUENCING, WE INVESTIGATED THE PROGNOSTIC IMPACT OF THE PROPOSED 5 CPG SIGNATURE IN A WELL-CHARACTERIZED CLL COHORT (135 CASES), INCLUDING IGHV-MUTATED AND UNMUTATED PATIENTS AS WELL AS CLINICALLY AGGRESSIVE STEREOTYPED SUBSET #2 PATIENTS. OVERALL, WE CONFIRMED THE SIGNATURE'S ASSOCIATION WITH ESTABLISHED PROGNOSTIC MARKERS. MOREOVER, IN THE PRESENCE OF THE IGHV MUTATIONAL STATUS, THE EPIGENETIC SIGNATURE REMAINED INDEPENDENTLY ASSOCIATED WITH BOTH TIME-TO-FIRST-TREATMENT AND OVERALL SURVIVAL IN MULTIVARIATE ANALYSES. AS A PRIME FINDING, WE OBSERVED THAT SUBSET #2 PATIENTS WERE PREDOMINANTLY CLASSIFIED AS I-CLL, PROBABLY REFLECTING THEIR BORDERLINE IGHV MUTATIONAL STATUS (97-99% GERMLINE IDENTITY), THOUGH HAVING A SIMILARLY POOR PROGNOSIS AS N-CLL PATIENTS. IN SUMMARY, WE VALIDATED THE EPIGENETIC CLASSIFIER AS AN INDEPENDENT FACTOR IN CLL PROGNOSTICATION AND PROVIDE FURTHER EVIDENCE THAT SUBSET #2 IS A MEMBER OF THE I-CLL GROUP, HENCE SUPPORTING THE EXISTENCE OF A THIRD, INTERMEDIATE EPIGENETIC SUBGROUP.	2016	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                               
3   27  45 A B-CELL EPIGENETIC SIGNATURE DEFINES THREE BIOLOGIC SUBGROUPS OF CHRONIC LYMPHOCYTIC LEUKEMIA WITH CLINICAL IMPACT. PROSPECTIVE IDENTIFICATION OF PATIENTS WITH CHRONIC LYMPHOCYTIC LEUKEMIA (CLL) DESTINED TO PROGRESS WOULD GREATLY FACILITATE THEIR CLINICAL MANAGEMENT. RECENTLY, WHOLE-GENOME DNA METHYLATION ANALYSES IDENTIFIED THREE CLINICOBIOLOGIC CLL SUBGROUPS WITH AN EPIGENETIC SIGNATURE RELATED TO DIFFERENT NORMAL B-CELL COUNTERPARTS. HERE, WE DEVELOPED A CLINICALLY APPLICABLE METHOD TO IDENTIFY THESE SUBGROUPS AND TO STUDY THEIR CLINICAL RELEVANCE. USING A SUPPORT VECTOR MACHINE APPROACH, WE BUILT A PREDICTION MODEL USING FIVE EPIGENETIC BIOMARKERS THAT WAS ABLE TO CLASSIFY CLL PATIENTS ACCURATELY INTO THE THREE SUBGROUPS, NAMELY NAIVE B-CELL-LIKE, INTERMEDIATE AND MEMORY B-CELL-LIKE CLL. DNA METHYLATION WAS QUANTIFIED BY HIGHLY REPRODUCIBLE BISULFITE PYROSEQUENCING ASSAYS IN TWO INDEPENDENT CLL SERIES. IN THE INITIAL SERIES (N=211), THE THREE SUBGROUPS SHOWED DIFFERENTIAL LEVELS OF IGHV (IMMUNOGLOBULIN HEAVY-CHAIN LOCUS) MUTATION (P<0.001) AND VH USAGE (P<0.03), AS WELL AS DIFFERENT CLINICAL FEATURES AND OUTCOME IN TERMS OF TIME TO FIRST TREATMENT (TTT) AND OVERALL SURVIVAL (P<0.001). A MULTIVARIATE COX MODEL SHOWED THAT EPIGENETIC CLASSIFICATION WAS THE STRONGEST PREDICTOR OF TTT (P<0.001) ALONG WITH BINET STAGE (P<0.001). THESE FINDINGS WERE CORROBORATED IN A VALIDATION SERIES (N=97). IN THIS STUDY, WE DEVELOPED A SIMPLE AND ROBUST METHOD USING EPIGENETIC BIOMARKERS TO CATEGORIZE CLLS INTO THREE SUBGROUPS WITH DIFFERENT CLINICOBIOLOGIC FEATURES AND OUTCOME.	2015	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                     
4 1062  53 CLINICAL SIGNIFICANCE OF DNA METHYLATION IN CHRONIC LYMPHOCYTIC LEUKEMIA PATIENTS: RESULTS FROM 3 UK CLINICAL TRIALS. CHRONIC LYMPHOCYTIC LEUKEMIA PATIENTS WITH MUTATED IMMUNOGLOBULIN HEAVY-CHAIN GENES (IGHV-M), PARTICULARLY THOSE LACKING POOR-RISK GENOMIC LESIONS, OFTEN RESPOND WELL TO CHEMOIMMUNOTHERAPY (CIT). DNA METHYLATION PROFILING CAN SUBDIVIDE EARLY-STAGE PATIENTS INTO NAIVE B-CELL-LIKE CLL (N-CLL), MEMORY B-CELL-LIKE CLL (M-CLL), AND INTERMEDIATE CLL (I-CLL), WITH DIFFERING TIMES TO FIRST TREATMENT AND OVERALL SURVIVAL. HOWEVER, WHETHER DNA METHYLATION CAN IDENTIFY PATIENTS DESTINED TO RESPOND FAVORABLY TO CIT HAS NOT BEEN ASCERTAINED. WE CLASSIFIED TREATMENT-NAIVE PATIENTS (N = 605) FROM 3 UK CHEMO AND CIT CLINICAL TRIALS INTO THE 3 EPIGENETIC SUBGROUPS, USING PYROSEQUENCING AND MICROARRAY ANALYSIS, AND PERFORMED EXPANSIVE SURVIVAL ANALYSIS. THE N-CLL, I-CLL, AND M-CLL SIGNATURES WERE FOUND IN 80% (N = 245/305), 17% (53/305), AND 2% (7/305) OF IGHV-UNMUTATED (IGHV-U) CASES, RESPECTIVELY, AND IN 9%, (19/216), 50% (108/216), AND 41% (89/216) OF IGHV-M CASES, RESPECTIVELY. MULTIVARIATE COX PROPORTIONAL ANALYSIS IDENTIFIED M-CLL AS AN INDEPENDENT PROGNOSTIC FACTOR FOR OVERALL SURVIVAL (HAZARD RATIO [HR], 0.46; 95% CONFIDENCE INTERVAL [CI], 0.24-0.87; P = .018) IN CLL4, AND FOR PROGRESSION-FREE SURVIVAL (HR, 0.25; 95% CI, 0.10-0.57; P = .002) IN ARCTIC AND ADMIRE PATIENTS. THE ANALYSIS OF EPIGENETIC SUBGROUPS IN PATIENTS ENTERED INTO 3 FIRST-LINE UK CLL TRIALS IDENTIFIES M-CLL AS AN INDEPENDENT MARKER OF PROLONGED SURVIVAL AND MAY AID IN THE IDENTIFICATION OF PATIENTS DESTINED TO DEMONSTRATE PROLONGED SURVIVAL AFTER CIT.	2019	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                      
5 3560  32 IMPACT OF CLINICAL, CYTOGENETIC, AND MOLECULAR PROFILES ON LONG-TERM SURVIVAL AFTER TRANSPLANTATION IN PATIENTS WITH CHRONIC MYELOMONOCYTIC LEUKEMIA. CHRONIC MYELOMONOCYTIC LEUKEMIA (CMML) IS A HETEROGENEOUS GROUP OF CLONAL HEMATOPOIETIC MALIGNANCIES WITH VARIABLE CLINICAL AND MOLECULAR FEATURES. WE ANALYZED LONG-TERM RESULTS OF ALLOGENEIC HEMATOPOIETIC CELL TRANSPLANTATION IN PATIENTS WITH CMML AND DETERMINED CLINICAL AND MOLECULAR RISK FACTORS ASSOCIATED WITH OUTCOMES. DATA FROM 129 PATIENTS, AGED 7-74 (MEDIAN 55) YEARS, AT VARIOUS STAGES OF THE DISEASE AND TRANSPLANTED FROM RELATED OR UNRELATED DONORS WERE ANALYZED. USING A PANEL OF 75 GENES SOMATIC MUTATIONS PRESENT BEFORE HEMATOPOIETIC CELL TRANSPLANTATION WERE IDENTIFIED IN 52 PATIENTS. THE PROGRESSION-FREE SURVIVAL RATE AT 10 YEARS WAS 29%. THE MAJOR CAUSE OF DEATH WAS RELAPSE (32%), WHICH WAS SIGNIFICANTLY ASSOCIATED WITH ADVERSE CYTOGENETICS (HAZARD RATIO, 3.77; P=0.0002), CMML PROGNOSTIC SCORING SYSTEM (HAZARD RATIO, 14.3, P=0.01), AND MD ANDERSON PROGNOSTIC SCORES (HAZARD RATIO, 9.4; P=0.005). MORTALITY WAS ASSOCIATED WITH HIGH-RISK CYTOGENETICS (HAZARD RATIO, 1.88; P=0.01) AND HIGH HEMATOPOIETIC CELL TRANSPLANTATION COMORBIDITY INDEX (SCORE >/=4: HAZARD RATIO, 1.99; P=0.01). HIGH OVERALL MUTATION BURDEN (>/=10 MUTATIONS: HAZARD RATIO, 3.4; P=0.02), AND >/=4 MUTATED EPIGENETIC REGULATORY GENES (HAZARD RATIO 5.4; P=0.003) WERE LINKED TO RELAPSE. UNSUPERVISED CLUSTERING OF THE CORRELATION MATRIX REVEALED DISTINCT HIGH-RISK GROUPS WITH UNIQUE ASSOCIATIONS OF MUTATIONS AND CLINICAL FEATURES. CMML WITH A HIGH MUTATION BURDEN APPEARED TO BE DISTINCT FROM HIGH-RISK GROUPS DEFINED BY COMPLEX CYTOGENETICS. NEW TRANSPLANT STRATEGIES MUST BE DEVELOPED TO TARGET SPECIFIC DISEASE SUBGROUPS, STRATIFIED BY MOLECULAR PROFILING AND CLINICAL RISK FACTORS.	2020	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                      
6 5665  41 SF3B1, RUNX1 AND TP53 MUTATIONS SIGNIFICANTLY IMPACT THE OUTCOME OF PATIENTS WITH LOWER-RISK MYELODYSPLASTIC SYNDROME. INTRODUCTION: PROGNOSIS OF PATIENTS WITH MYELODYSPLASTIC SYNDROME (MDS), PARTICULARLY THE GROUP WITH LOWER-RISK DISEASE (LR-MDS) IS VERY HETEROGENEOUS. SEVERAL STUDIES HAVE DESCRIBED THE PROGNOSTIC VALUE OF RECURRENT SOMATIC MUTATIONS IN MDS INCLUDING ALL RISK CATEGORIES. RECENTLY, THE INCORPORATION OF GENOMIC DATA TO CLINICAL PARAMETERS DEFINED THE NEW MOLECULAR INTERNATIONAL PROGNOSTIC SCORING SYSTEM (IPSS-M). MATERIALS AND METHODS: IN THIS STUDY, WE EVALUATED THE IMPACT OF MOLECULAR PROFILE IN A SERIES OF 181 PATIENTS WITH LR-MDS AND NON-PROLIFERATIVE CHRONIC MYELOMONOCYTIC LEUKEMIA. RESULTS: EPIGENETIC REGULATORS (TET2, ASXL1) AND SPLICING (SF3B1) WERE THE MOST RECURRENT MUTATED PATHWAYS. IN UNIVARIATE ANALYSIS, RUNX1 OR TP53 MUTATIONS CORRELATED WITH LOWER MEDIAN OVERALL SURVIVAL (OS). IN CONTRAST, SF3B1 MUTATION WAS ASSOCIATED WITH PROLONGED MEDIAN OS [95 MONTHS (95% IC, 32-157) VS. 33 MONTHS (95% CI, 19-46) IN UNMUTATED PATIENTS (P < 0.01)]. IN A MULTIVARIATE COX REGRESSION MODEL, RUNX1 MUTATIONS INDEPENDENTLY ASSOCIATED WITH SHORTER OS, WHILE SF3B1 MUTATION RETAINED ITS FAVORABLE IMPACT ON OUTCOME (HR: 0.24, 95% CI, 0.1-0.5; P = 0.001). IN ADDITION, TP53 OR RUNX1 MUTATIONS WERE IDENTIFIED AS PREDICTIVE COVARIATES FOR THE PROBABILITY OF LEUKEMIC PROGRESSION (P < 0.001). CONCLUSION: INCORPORATION OF MOLECULAR TESTING IN LR-MDS IDENTIFIED A SUBSET OF PATIENTS WITH EXPECTED POORER OUTCOME, EITHER DUE TO LOWER SURVIVAL OR PROBABILITY OF LEUKEMIC PROGRESSION.	2022	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                       
7 3071  43 GENOME-WIDE DNA METHYLATION PROFILING INTEGRATED WITH GENE EXPRESSION PROFILING IDENTIFIES PAX9 AS A NOVEL PROGNOSTIC MARKER IN CHRONIC LYMPHOCYTIC LEUKEMIA. BACKGROUND: IN CHRONIC LYMPHOCYTIC LEUKEMIA (CLL), EPIGENOMIC AND GENOMIC STUDIES HAVE EXPANDED THE EXISTING KNOWLEDGE ABOUT THE DISEASE BIOLOGY AND LED TO THE IDENTIFICATION OF POTENTIAL BIOMARKERS RELEVANT FOR IMPLEMENTATION OF PERSONALIZED MEDICINE. IN THIS STUDY, AN ATTEMPT HAS BEEN MADE TO EXAMINE AND INTEGRATE THE GLOBAL DNA METHYLATION CHANGES WITH GENE EXPRESSION PROFILE AND THEIR IMPACT ON CLINICAL OUTCOME IN EARLY STAGE CLL PATIENTS. RESULTS: THE INTEGRATION OF DNA METHYLATION PROFILE (N = 14) WITH THE GENE EXPRESSION PROFILE (N = 21) REVEALED 142 GENES AS HYPERMETHYLATED-DOWNREGULATED AND; 62 GENES AS HYPOMETHYLATED-UPREGULATED IN EARLY STAGE CLL PATIENTS COMPARED TO CD19+ B-CELLS FROM HEALTHY INDIVIDUALS. THE MRNA EXPRESSION LEVELS OF 17 GENES IDENTIFIED TO BE DIFFERENTIALLY METHYLATED AND/OR DIFFERENTIALLY EXPRESSED WAS FURTHER EXAMINED IN EARLY STAGE CLL PATIENTS (N = 93) BY QUANTITATIVE REAL TIME PCR (RQ-PCR). SIGNIFICANT DIFFERENCES WERE OBSERVED IN THE MRNA EXPRESSION OF MEIS1, PMEPA1, SOX7, SPRY1, CDK6, TBX2, AND SPRY2 GENES IN CLL CELLS AS COMPARED TO B-CELLS FROM HEALTHY INDIVIDUALS. THE ANALYSIS IN THE IGHV MUTATION BASED CATEGORIES (UNMUTATED = 39, MUTATED = 54) REVEALED SIGNIFICANTLY HIGHER MRNA EXPRESSION OF CRY1 AND PAX9 GENES IN THE IGHV UNMUTATED SUBGROUP (P < 0.001). THE RELATIVE RISK OF TREATMENT INITIATION WAS SIGNIFICANTLY HIGHER AMONG PATIENTS WITH HIGH EXPRESSION OF CRY1 (RR = 1.91, P = 0.005) OR PAX9 (RR = 1.87, P = 0.001). HIGH EXPRESSION OF CRY1 (HR: 3.53, P < 0.001) OR PAX9 (HR: 3.14, P < 0.001) GENE WAS SIGNIFICANTLY ASSOCIATED WITH SHORTER TIME TO FIRST TREATMENT. THE HIGH EXPRESSION OF PAX9 GENE (HR: 3.29, 95% CI 1.172-9.272, P = 0.016) WAS ALSO PREDICTIVE OF SHORTER OVERALL SURVIVAL IN CLL. CONCLUSIONS: THE DNA METHYLATION CHANGES ASSOCIATED WITH MRNA EXPRESSION OF CRY1 AND PAX9 GENES ALLOW RISK STRATIFICATION OF EARLY STAGE CLL PATIENTS. THIS COMPREHENSIVE ANALYSIS SUPPORTS THE CONCEPT THAT THE EPIGENETIC CHANGES ALONG WITH THE ALTERED EXPRESSION OF GENES HAVE THE POTENTIAL TO PREDICT CLINICAL OUTCOME IN EARLY STAGE CLL PATIENTS.	2017	
                                                                                                                                                                  
8 5270  35 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	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                               
9  817  37 CHARACTERISTIC PATTERNS OF ALTERED DNA METHYLATION PREDICT EMERGENCE OF HUMAN HEPATOCELLULAR CARCINOMA. WE AIMED TO IDENTIFY THE SPECIFIC SUBSET OF TUMOR SUPPRESSOR GENES (TSGS) THAT ARE METHYLATION-SILENCED DURING THE EARLIEST STEPS OF HEPATOCARCINOGENESIS, AND TO FURTHER EVALUATE WHETHER THESE GENES CAN SERVE AS PREDICTIVE BIOMARKERS OF HEPATOCELLULAR CARCINOMA (HCC) EMERGENCE. A TOTAL OF 482 LIVER TISSUES INCLUDING 177 PAIRS OF HCCS AND MATCHED NONTUMOR LIVERS AND 128 LIVER BIOPSIES FROM CHRONIC HEPATITIS C (CHC) PATIENTS WERE ANALYZED FOR QUANTITATIVE METHYLATION ANALYSIS IN 24 TSG PROMOTERS AND THREE MINT LOCI. THE TUMORS WERE CLASSIFIED AS EARLY, LESS-PROGRESSED, AND HIGHLY PROGRESSED HCCS USING HISTOLOGY AND RADIOLOGICAL APPROACHES. A SUBSET OF TSGS THAT HARBORED DISTINCTLY HIGH LEVELS OF METHYLATION IN EARLY HCCS WERE SELECTED. BASED ON THE METHYLATION PROFILES OF THESE GENES, KAPLAN-MEIER ANALYSES WERE PERFORMED TO DETERMINE TIME-TO-HCC OCCURRENCE IN CHC PATIENTS. SUBSEQUENTLY, MULTIVARIATE ANALYSIS WAS PERFORMED USING AGE, GENDER, FIBROSIS STAGE, AND NUMBER OF METHYLATED TSGS AS COVARIATES. AMONG TSGS ANALYZED, A SUBSET OF EIGHT TSGS (HIC1, GSTP1, SOCS1, RASSF1, CDKN2A, APC, RUNX3, AND PRDM2) DEMONSTRATED A DISTINCT CLUSTER BY HIERARCHICAL CLUSTERING AND RECEIVER OPERATING CHARACTERISTIC ANALYSES. THIS SUBSET OF TSGS SHOWED SIGNIFICANTLY HIGHER METHYLATION LEVELS IN THE EARLY HCCS (P < 0.0001). IN THE CHC PATIENTS, METHYLATION FREQUENCIES IN THESE TSGS WERE ASSOCIATED WITH SHORTER TIME-TO-HCC OCCURRENCE (P < 0.0001), AND NUMBER OF METHYLATED GENES WAS AN INDEPENDENT RISK FACTOR FOR HCC (HAZARD RATIO = 5.21, 95% CONFIDENCE INTERVAL = 2.25-11.76, P = 0.0002). CONCLUSION: EPIGENETIC INACTIVATION OF A SUBSET OF TSGS PLAYS A CRITICAL ROLE IN THE EARLIEST STEPS OF HEPATOCARCINOGENESIS. FURTHERMORE, EPIGENETIC INACTIVATION OF THESE GENES IN CHC PROVIDES A PROGNOSTIC VALUE FOR DETERMINING THE RISK FOR DEVELOPING HCC LATER IN LIFE.	2012	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                             
10 1473  33 DISTINCT PATTERNS OF GLOBAL PROMOTER METHYLATION IN EARLY STAGE CHRONIC LYMPHOCYTIC LEUKEMIA. GENOMIC AND EPIGENOMIC STUDIES OF CHRONIC LYMPHOCYTIC LEUKEMIA (CLL) ARE RESHAPING OUR UNDERSTANDING OF THE DISEASE AND HAVE PROVIDED NEW PERSPECTIVES FOR A MORE INDIVIDUALIZED DIAGNOSIS AND NEW POTENTIAL THERAPEUTIC TARGETS. IN THIS STUDY, THE GLOBAL PROMOTER METHYLATION PROFILE WAS DETERMINED IN HIGHLY PURIFIED B-CELLS FROM 37 (BINET STAGE A) CLL PATIENTS, USING HIGH-RESOLUTION METHYLATION MICROARRAYS (27,578 CPG). OVERALL, THE METHYLATION PATTERN CORRELATED WITH THE MAJOR BIOLOGICAL (ZAP-70 AND CD38), AND MOLECULAR (IGHV MUTATION) MARKERS, DISTINGUISHING CLL CASES ACCORDING TO IGHV MUTATIONAL STATUS. CELL ADHESION MOLECULES WERE ENRICHED IN THE SIGNATURE OF UNMUTATED (UM) VERSUS MUTATED (M-) CLL. MOREOVER, IN M-CLL CPG HYPER-METHYLATION IN THREE GENES, INCLUDING SPG20, WAS SIGNIFICANTLY ANTI-CORRELATED WITH THE CORRESPONDING GENE EXPRESSION LEVEL. FINALLY, THE CORRELATION BETWEEN THE METHYLATION PATTERN AND CLINICAL PARAMETERS WAS INVESTIGATED. NOTABLY, OUT OF 42 METHYL-PROBES THAT WERE SIGNIFICANTLY ASSOCIATED WITH PROGRESSION FREE SURVIVAL (PFS), HYPER-METHYLATION OF SPG20 WAS ALSO POSITIVELY ASSOCIATED WITH PFS. THESE DATA SUPPORT THE NOTION THAT EPIGENETIC CHANGES HAVE CLINICAL IMPACT IN CLL AND MAY CONTRIBUTE TO THE IDENTIFICATION OF NOVEL CANDIDATE DISEASE-ASSOCIATED GENES POTENTIALLY USEFUL TO PREDICT THE CLINICAL OUTCOME OF EARLY STAGE CLL PATIENTS.	2014	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                      
11 3995  38 LONGITUDINAL STUDY OF DNA METHYLATION OF INFLAMMATORY GENES AND CANCER RISK. BACKGROUND: CHRONIC INFLAMMATION PLAYS A KEY ROLE IN CANCER ETIOLOGY. DNA METHYLATION MODIFICATION, ONE OF THE EPIGENETIC MECHANISMS REGULATING GENE EXPRESSION, IS CONSIDERED A HALLMARK OF CANCER. HUMAN AND ANIMAL MODELS HAVE IDENTIFIED NUMEROUS LINKS BETWEEN DNA METHYLATION AND INFLAMMATORY BIOMARKERS. OUR OBJECTIVE WAS TO PROSPECTIVELY AND LONGITUDINALLY EXAMINE ASSOCIATIONS BETWEEN METHYLATION OF FOUR INFLAMMATORY GENES AND CANCER RISK. METHODS: WE INCLUDED 795 NORMATIVE AGING STUDY PARTICIPANTS WITH BLOOD DRAWN ONE TO FOUR TIMES FROM 1999 TO 2012 (MEDIAN FOLLOW-UP, 10.6 YEARS). PROMOTER DNA METHYLATION OF IL6, ICAM-1, IFN, AND TLR2 IN BLOOD LEUKOCYTES WAS MEASURED USING PYROSEQUENCING AT MULTIPLE CPG SITES AND AVERAGED BY GENE FOR DATA ANALYSIS. WE USED COX REGRESSION MODELS TO EXAMINE PROSPECTIVE ASSOCIATIONS OF BASELINE AND TIME-DEPENDENT METHYLATION WITH CANCER RISK AND COMPARED MEAN METHYLATION DIFFERENCES OVER TIME BETWEEN CANCER CASES AND CANCER-FREE PARTICIPANTS. RESULTS: BASELINE IFN HYPERMETHYLATION WAS ASSOCIATED WITH ALL-CANCER (HR, 1.49; P = 0.04) AND PROSTATE CANCER INCIDENCE (HR, 1.69; P = 0.02). BASELINE ICAM-1 AND IL6 HYPERMETHYLATION WERE ASSOCIATED WITH PROSTATE CANCER INCIDENCE (HR, 1.43; P = 0.02; HR, 0.70; P = 0.03, RESPECTIVELY). IN OUR TIME-DEPENDENT ANALYSES, IFN HYPERMETHYLATION WAS ASSOCIATED WITH ALL-CANCER (HR, 1.79; P = 0.007) AND PROSTATE CANCER (HR, 1.57; P = 0.03) INCIDENCE; AND ICAM-1 AND IL6 HYPERMETHYLATION WERE ASSOCIATED WITH PROSTATE CANCER INCIDENCE (HR, 1.39; P = 0.02; HR, 0.69; P = 0.03, RESPECTIVELY). WE DETECTED SIGNIFICANT ICAM-1 HYPERMETHYLATION IN CANCER CASES (P = 0.0003) 10 TO 13 YEARS PREDIAGNOSIS. CONCLUSION: HYPERMETHYLATION OF IFN AND ICAM-1 MAY PLAY IMPORTANT ROLES IN EARLY CARCINOGENESIS, PARTICULARLY THAT OF PROSTATE CANCER. IMPACT: THESE METHYLATION CHANGES COULD INFORM THE DEVELOPMENT OF EARLY DETECTION BIOMARKERS AND POTENTIAL TREATMENTS OF INFLAMMATION-RELATED CARCINOGENESIS.	2015	
                                                                                                                                                                                                                                                                                                                                                                                          
12 2960  29 GENETIC AND EPIGENETIC MARKERS IN THE EVALUATION OF PANCREATIC MASSES. BACKGROUND: METHYLATION MARKERS HAVE SHOWN PROMISE IN THE EARLY DIAGNOSIS OF PANCREATIC CARCINOMA. THE AIM OF THIS STUDY WAS TO ASSESS THE DIAGNOSTIC UTILITY OF HYPERMETHYLATION STATUS OF CANDIDATE GENES IN COMBINATION WITH KRAS MUTATION DETECTION IN THE EVALUATION OF PANCREATIC MASSES. EXPERIMENTAL DESIGN: SIXTY-ONE FINE NEEDLE ASPIRATES OF PANCREATIC MASSES (43 PANCREATIC ADENOCARCINOMAS AND 18 CHRONIC PANCREATITIS) WERE STUDIED. METHYLATION STATUS OF HRH2, EN1, SPARC, CDH13 AND APC WERE ANALYSED USING MELTING CURVE ANALYSIS AFTER DNA BISULFITE TREATMENT. KRAS MUTATIONS WERE ALSO ANALYSED. RESULTS: THE METHYLATION PANEL HAD A SENSITIVITY OF 73% (27 OF 37, CI 95% 56 TO 86%) AND A SPECIFICITY OF 100% WHENEVER TWO OR MORE PROMOTERS WERE FOUND HYPERMETHYLATED. KRAS MUTATIONS SHOWED A SENSITIVITY OF 77% (33 OF 43, CI 95% 62 TO 88%) AND A SPECIFICITY OF 100%. BOTH MOLECULAR ANALYSES ADDED USEFUL INFORMATION TO CYTOLOGY BY INCREASING THE NUMBER OF INFORMATIVE CASES. WHEN GENETIC AND EPIGENETIC ANALYSES WERE COMBINED SENSITIVITY WAS 84% (36 OF 43 CI 95% 69 TO 93%) MAINTAINING A 100% SPECIFICITY. CONCLUSIONS: ANALYSIS OF HYPERMETHYLATION STATUS OF A PANEL OF GENES AND KRAS MUTATION DETECTION OFFER A SIMILAR DIAGNOSTIC YIELD IN THE EVALUATION OF PANCREATIC MASSES. THE COMBINED MOLECULAR ANALYSIS INCREASES THE NUMBER OF INFORMATIVE CASES WITHOUT DIMINISHING SPECIFICITY.	2013	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                            
13 1433  34 DIFFERENTIAL GENOME-WIDE ARRAY-BASED METHYLATION PROFILES IN PROGNOSTIC SUBSETS OF CHRONIC LYMPHOCYTIC LEUKEMIA. GLOBAL HYPOMETHYLATION AND REGIONAL HYPERMETHYLATION ARE WELL-KNOWN EPIGENETIC FEATURES OF CANCER; HOWEVER, IN CHRONIC LYMPHOCYTIC LEUKEMIA (CLL), STUDIES ON GENOME-WIDE EPIGENETIC MODIFICATIONS ARE LIMITED. HERE, WE ANALYZED THE GLOBAL METHYLATION PROFILES IN CLL, BY APPLYING HIGH-RESOLUTION METHYLATION MICROARRAYS (27,578 CPG SITES) TO 23 CLL SAMPLES, BELONGING TO THE IMMUNOGLOBULIN HEAVY-CHAIN VARIABLE (IGHV) MUTATED (FAVORABLE) AND IGHV UNMUTATED/IGHV3-21 (POOR-PROGNOSTIC) SUBSETS. OVERALL, RESULTS DEMONSTRATED SIGNIFICANT DIFFERENCES IN METHYLATION PATTERNS BETWEEN THESE SUBGROUPS. SPECIFICALLY, IN IGHV UNMUTATED CLL, WE IDENTIFIED METHYLATION OF 7 KNOWN OR CANDIDATE TUMOR SUPPRESSOR GENES (EG, VHL, ABI3, AND IGSF4) AS WELL AS 8 UNMETHYLATED GENES INVOLVED IN CELL PROLIFERATION AND TUMOR PROGRESSION (EG, ADORA3 AND PRF1 ENHANCING THE NUCLEAR FACTOR-KAPPAB AND MITOGEN-ACTIVATED PROTEIN KINASE PATHWAYS, RESPECTIVELY). IN CONTRAST, THESE LATTER GENES WERE SILENCED BY METHYLATION IN IGHV MUTATED PATIENTS. THE ARRAY DATA WERE VALIDATED FOR SELECTED GENES USING METHYLATION-SPECIFIC POLYMERASE CHAIN REACTION, QUANTITATIVE REVERSE TRANSCRIPTASE-POLYMERASE CHAIN REACTION, AND BISULFITE SEQUENCING. FINALLY, THE SIGNIFICANCE OF DNA METHYLATION IN REGULATING GENE PROMOTERS WAS SHOWN BY REINDUCING 4 METHYLATED TUMOR SUPPRESSOR GENES (EG, VHL AND ABI3) IN IGHV UNMUTATED SAMPLES USING THE METHYL-INHIBITOR 5-AZA-2'-DEOXYCYTIDINE. TAKEN TOGETHER, OUR DATA FOR THE FIRST TIME REVEAL DIFFERENCES IN GLOBAL METHYLATION PROFILES BETWEEN PROGNOSTIC SUBSETS OF CLL, WHICH MAY UNFOLD EPIGENETIC SILENCING MECHANISMS INVOLVED IN CLL PATHOGENESIS.	2010	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                       
14 3588  29 IMPACT OF TP53 GENE PROMOTER METHYLATION ON CHRONIC LYMPHOCYTIC LEUKEMIA PATHOGENESIS AND PROGRESSION. BACKGROUND: CHRONIC LYMPHOCYTIC LEUKEMIA (CLL) IS A MALIGNANT LYMPHOID DISORDER THAT RESULTS FROM THE OVERGROWTH OF MATURE-LOOKING LYMPHOID CELLS IN THE BLOOD AND LYMPHATIC TISSUE. VARIOUS CLINICAL PRESENTATIONS HAVE BEEN ATTRIBUTED TO THE DISEASE AS A RESULT OF THE DIFFERENT UNDERLYING GENETIC AND EPIGENETIC ALTERATIONS. THE CURRENT STUDY HAS BEEN INITIATED TO STUDY THE ROLE OF AN EPIGENETIC ALTERATION AFFECTING THE PROMOTER OF THE TP53GENE ON CLL PATHOGENESIS AND PROGRESSION. METHODS: THE CURRENT STUDY INVOLVED 54 NEWLY DIAGNOSED PATIENTS PRESENTING WITH CLL AS WELL AS 30 NORMAL INDIVIDUALS AS CONTROLS. AFTER OBTAINING VERBAL CONSENT, DATA COLLECTION WAS DONE AND THE BLOOD COLLECTED FROM ALL ENROLLED INDIVIDUALS FOR HEMATOLOGICAL INVESTIGATIONS AS WELL AS FOR MOLECULAR CATEGORIZATION OF TP53 METHYLATION STATUS. METHYLATION-SPECIFIC POLYMERASE CHAIN REACTION (MS-PCR) TECHNIQUE WAS USED TO DEFINE THE METHYLATION STATUS OF THE TP53 GENE PROMOTER THAT ENCOMPASSES DNA EXTRACTION, BISULFITE CONVERSION, CONVENTIONAL PCR AMPLIFICATION, RUNNING ON AGAROSE GEL AND DOCUMENTATION. FINALLY, STATISTICAL ANALYSIS WAS DONE TO ASSESS ANY CORRELATION OF THE TP53 EPIGENETIC ALTERATION TO THE DISEASE ETIOLOGY AND THE PROGRESSION. RESULTS: IN THE CURRENT STUDY, ALL CONTROLS AND 42 OF 54 PATIENTS SHOW UNMETHYLATED TP53 GENE PROMOTER; ON THE OTHER HAND, THE METHYLATED PROMOTER WAS DETECTED AMONG 12 PATIENTS WITH A P-VALUE OF 0.001. TP53 GENE PROMOTER METHYLATION SIGNIFICANTLY LINKED TO REDUCED PLATELET COUNT (P-VALUE OF 0.047) AND ADVANCED STAGE AT PRESENTATION (P-VALUE OF 0.076). NO SIGNIFICANT DIFFERENCES WERE SEEN AMONG BOTH METHYLATED AND UNMETHYLATED TP53 PROMOTERS IN RELATION TO THE AGE OF THE AFFECTED INDIVIDUALS, TOTAL WHITE BLOOD CELL COUNTS AND HEMOGLOBIN LEVEL OF THE AFFECTED INDIVIDUALS. CONCLUSION: THE CURRENT STUDY REVEALED A SIGNIFICANT CORRELATION OF TP53 GENE PROMOTER METHYLATION TO CHRONIC LYMPHOCYTIC LEUKEMIA PATHOGENESIS AND LOWER PLATELET COUNTS.	2019	
                                                                                                                                                                                                                                                                                                                                                       
15 1581  25 DNA METHYLATION PROFILES IN CHRONIC LYMPHOCYTIC LEUKEMIA PATIENTS TREATED WITH CHEMOIMMUNOTHERAPY. BACKGROUND: IN ORDER TO GAIN INSIGHT INTO THE CONTRIBUTION OF DNA METHYLATION TO DISEASE PROGRESSION OF CHRONIC LYMPHOCYTIC LEUKEMIA (CLL), USING 450K ILLUMINA ARRAYS, WE DETERMINED THE DNA METHYLATION PROFILES IN PAIRED PRE-TREATMENT/RELAPSE SAMPLES FROM 34 CLL PATIENTS TREATED WITH CHEMOIMMUNOTHERAPY, MOSTLY (N = 31) WITH THE FLUDARABINE-CYCLOPHOSPHAMIDE-RITUXIMAB (FCR) REGIMEN. RESULTS: THE EXTENT OF IDENTIFIED CHANGES IN CLL CELLS VERSUS MEMORY B CELLS FROM HEALTHY DONORS WAS TERMED "EPIGENETIC BURDEN" (EB) WHEREAS THE NUMBER OF CHANGES BETWEEN THE PRE-TREATMENT VERSUS THE RELAPSE SAMPLE WAS TERMED "RELAPSE CHANGES" (RC). SIGNIFICANT (P < 0.05) ASSOCIATIONS WERE IDENTIFIED BETWEEN (I) HIGH EB AND SHORT TIME-TO-FIRST-TREATMENT (TTFT); AND, (II) FEW RCS AND SHORT TIME-TO-RELAPSE. BOTH THE EB AND THE RC CLUSTERED IN SPECIFIC GENOMIC REGIONS AND CHROMATIN STATES, INCLUDING REGULATORY REGIONS CONTAINING BINDING SITES OF TRANSCRIPTION FACTORS IMPLICATED IN B CELL AND CLL BIOLOGY. CONCLUSIONS: OVERALL, WE SHOW THAT DNA METHYLATION IN CLL FOLLOWS DIFFERENT DYNAMICS IN RESPONSE TO CHEMOIMMUNOTHERAPY. THESE EPIGENETIC ALTERATIONS WERE LINKED WITH SPECIFIC CLINICAL AND BIOLOGICAL FEATURES.	2019	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                      
16  780  40 CELL-FREE DNA PROMOTER HYPERMETHYLATION IN PLASMA AS A DIAGNOSTIC MARKER FOR PANCREATIC ADENOCARCINOMA. BACKGROUND: PANCREATIC CANCER HAS A 5-YEAR SURVIVAL RATE OF ONLY 5-7%. DIFFICULTIES IN DETECTING PANCREATIC CANCER AT EARLY STAGES RESULTS IN THE HIGH MORTALITY AND SUBSTANTIATES THE NEED FOR ADDITIONAL DIAGNOSTIC TOOLS. SURGERY IS THE ONLY CURATIVE TREATMENT AND UNFORTUNATELY ONLY POSSIBLE IN LOCALIZED TUMOURS. A DIAGNOSTIC BIOMARKER FOR PANCREATIC CANCER WILL HAVE A MAJOR IMPACT ON PATIENT SURVIVAL BY FACILITATING EARLY DETECTION AND THE POSSIBILITY FOR CURATIVE TREATMENT. DNA PROMOTER HYPERMETHYLATION IS A MECHANISM OF EARLY CARCINOGENESIS, WHICH CAN CAUSE INACTIVATION OF TUMOUR SUPPRESSOR GENES. THE AIM OF THIS STUDY WAS TO EXAMINE PROMOTER HYPERMETHYLATION IN A PANEL OF SELECTED GENES FROM CELL-FREE DNA, AS A DIAGNOSTIC MARKER FOR PANCREATIC ADENOCARCINOMA. METHODS: PATIENTS WITH SUSPECTED OR BIOPSY-VERIFIED PANCREATIC CANCER WERE INCLUDED PROSPECTIVELY AND CONSECUTIVELY. PATIENTS WITH CHRONIC/ACUTE PANCREATITIS WERE INCLUDED AS ADDITIONAL BENIGN CONTROL GROUPS. BASED ON AN OPTIMIZED ACCELERATED BISULFITE TREATMENT PROTOCOL, METHYLATION-SPECIFIC PCR OF A 28 GENE PANEL WAS PERFORMED ON PLASMA SAMPLES. A DIAGNOSTIC PREDICTION MODEL WAS DEVELOPED BY MULTIVARIABLE LOGISTIC REGRESSION ANALYSIS USING BACKWARD STEPWISE ELIMINATION. RESULTS: PATIENTS WITH PANCREATIC ADENOCARCINOMA (N = 95), CHRONIC PANCREATITIS (N = 97) AND ACUTE PANCREATITIS (N = 59) AND PATIENTS SCREENED, BUT NEGATIVE FOR PANCREATIC ADENOCARCINOMA (N = 27), WERE INCLUDED. THE DIFFERENCE IN MEAN NUMBER OF METHYLATED GENES IN THE CANCER GROUP (8.41 (95% CI 7.62-9.20)) VS THE TOTAL CONTROL GROUP (4.74 (95% CI 4.40-5.08)) WAS HIGHLY SIGNIFICANT (P < 0.001). A DIAGNOSTIC PREDICTION MODEL (AGE >65, BMP3, RASSF1A, BNC1, MESTV2, TFPI2, APC, SFRP1 AND SFRP2) HAD AN AREA UNDER THE CURVE OF 0.86 (SENSITIVITY 76%, SPECIFICITY 83%). THE MODEL PERFORMANCE WAS INDEPENDENT OF CANCER STAGE. CONCLUSIONS: CELL-FREE DNA PROMOTER HYPERMETHYLATION HAS THE POTENTIAL TO BE A DIAGNOSTIC MARKER FOR PANCREATIC ADENOCARCINOMA AND DIFFERENTIATE BETWEEN MALIGNANT AND BENIGN PANCREATIC DISEASE. THIS STUDY BRINGS US CLOSER TO A CLINICAL USEFUL DIAGNOSTIC MARKER FOR PANCREATIC CANCER, WHICH IS URGENTLY NEEDED. EXTERNAL VALIDATION IS, HOWEVER, REQUIRED BEFORE THE TEST CAN BE APPLIED IN THE CLINIC. TRIAL REGISTRATION: CLINICALTRIALS.GOV, NCT02079363.	2016	

17 2135  34 EPIGENETIC INACTIVATION OF TUMOR SUPPRESSOR GENES IN SERUM OF PATIENTS WITH CUTANEOUS MELANOMA. SMALL AMOUNTS OF CELL-FREE DNA CIRCULATE IN BOTH HEALTHY AND DISEASED HUMAN BLOOD, WHILE INCREASED CONCENTRATIONS OF DNA ARE PRESENT IN THE SERUM OF CANCER PATIENTS. TUMOR-SPECIFIC MUTATIONS OR EPIGENETIC MODIFICATIONS HAVE PREDOMINANTLY BEEN DETECTED IN TISSUE SPECIMENS. THE PURPOSE OF THIS STUDY WAS TO INVESTIGATE METHYLATION OF FIVE DIFFERENT GENES INVOLVED IN TUMOR SUPPRESSION AND DNA REPAIR (SUPPRESSORS OF CYTOKINE SIGNALING 1 AND 2 (SOCS1, SOCS2)), RAS-ASSOCIATION DOMAIN FAMILY PROTEIN 1A (RASSF1A), D-TYPE P16(INK4A) CYCLIN-DEPENDENT KINASE INHIBITOR (CDKN), AND O6-METHYLGUANINE DNA-METHYLTRANSFERASE (MGMT)) IN THE SERUM OF 100 PATIENTS USING METHYLATION-SPECIFIC PCR. IN ALL, 41 MELANOMA PATIENTS (STAGE I = 18; STAGE II = 10; STAGE III/IV = 13), 13 HEALTHY CONTROLS WITHOUT NEVI, AND 10 INDIVIDUALS WITH MORE THAN 15 NEVI OF >5 MM IN SIZE WERE INVESTIGATED. FOR COMPARISON, SERA FROM PATIENTS WITH OTHER SKIN TUMORS (NINE BASAL CELL CANCERS, FIVE KAPOSI'S SARCOMA), DIFFERENT METASTASIZED CANCERS (FIVE BREAST CANCERS, FIVE COLON CANCERS), AND SEVERAL CHRONIC INFLAMMATORY DISEASES (N = 12) WERE ALSO ANALYZED. IN ADDITION, WE EXAMINED IF METHYLATION WAS INVOLVED IN SILENCING TRANSCRIPTION OF THESE GENES IN 12 MELANOMA SPECIMENS. SOCS1, SOCS2, RASSF1A, CDKN2A, AND MGMT WERE METHYLATED IN 75, 43, 64, 75, AND 64% OF MELANOMA SAMPLES, RESPECTIVELY. OF THE 41 MELANOMA PATIENTS, 83% HAD ONE HYPERMETHYLATED GENE, WHILE 66, 51, AND 41% HAD TWO, THREE, OR FOUR HYPERMETHYLATED GENES, RESPECTIVELY. ALSO, 20% OF THESE PATIENTS SHOWED HYPERMETHYLATION FOR ALL GENES, WHILE ONLY 17% SHOWED NO METHYLATION. IMPORTANTLY, THE METHYLATION PROFILE OF THE SELECTED GENES FROM MELANOMA PATIENTS WAS DISTINCT FROM THE OTHER ANALYZED TUMORS. TRANSCRIPTION OF SOCS1, SOCS2, CDKN2A, AND RASSF1A GENES WAS SIGNIFICANTLY REDUCED IN FRESH MELANOMA SAMPLES, WHILE MGMT SHOWED A 12-FOLD UPREGULATION AT THE MESSENGER RIBONUCLEIC ACID LEVEL (P < 0.001). OUR FINDINGS SUGGEST THAT EPIGENETIC SILENCING OF THE STUDIED TUMOR SUPPRESSOR GENES IS A COMMON AND PROBABLY IMPORTANT MECHANISM FOR MELANOMA FORMATION. THIS CONVENIENT METHOD USING A SIMPLE BLOOD SAMPLE MAY CONTRIBUTE TO CLASSIFICATION OF MELANOMA AND AWAITS CLINICAL VALIDATION.	2006	
                                                                                                     
18  831  34 CHARACTERIZATION OF TET AND IDH GENE EXPRESSION IN CHRONIC LYMPHOCYTIC LEUKEMIA: COMPARISON WITH NORMAL B CELLS AND PROGNOSTIC SIGNIFICANCE. BACKGROUND: CHRONIC LYMPHOCYTIC LEUKEMIA (CLL) IS THE MOST COMMON HEMATOLOGICAL MALIGNANCY IN WESTERN COUNTRIES, CHARACTERIZED BY A HETEROGENEOUS CLINICAL COURSE. ALTHOUGH GENETIC STUDIES HAVE IDENTIFIED CHROMOSOMAL ABERRATIONS OR SPECIFIC MUTATIONS, EPIGENETIC CHANGES HAVE BEEN POORLY CHARACTERIZED IN CLL. METHODS: WE ASSESSED TEN-ELEVEN TRANSLOCATIONS (TET) 1, 2, AND 3, ISOCITRATE DEHYDROGENASE (IDH) 1, AND 2 MESSENGER RNA (MRNA) EXPRESSION USING REAL-TIME PCR ON PURIFIED LEUKEMIC B CELLS FROM 214 CLL PATIENTS (MEDIAN FOLLOW-UP = 75 MONTHS, RANGE 1-380), NORMAL PERIPHERAL BLOOD B CELLS (N = 20), AND UMBILICAL CORD BLOOD B CELLS (N = 21). THE MICROENVIRONMENT INFLUENCE WAS ASSESSED AFTER 24 H CO-CULTURE OF CLL CELLS WITH BONE MARROW MESENCHYMAL STROMAL CELLS (BMSC). FINALLY, 5-HYDROXYMETHYLCYTOSINE LEVEL (%5-HMC) WAS ASSESSED BY ELISA IN CLL CELLS ALONE OR WITH MICROENVIRONMENT STIMULI. RESULTS: TET 1 AND 3 AND IDH2 WERE DECREASED IN CLL CELLS COMPARED WITH HEALTHY B CELLS (P = 0.0221, 0.0013, <0.0001, RESPECTIVELY), WHILE IDH1 WAS OVEREXPRESSED (P = 0.0037). TET2 AND IDH1 WERE SIGNIFICANTLY CORRELATED WITH TREATMENT-FREE SURVIVAL (TFS); PATIENTS WITH HIGH TET2/IDH1 EXPRESSION HAD A HIGHER MEDIAN TFS (111 MONTHS) THAN PATIENTS WITH LOW EXPRESSION (78 MONTHS, P = 0.0071/0.0123). MOREOVER, TET1 EXPRESSION DECREASED (P = 0.0371), WHILE TET3 AND IDH2 EXPRESSION INCREASED (P = 0.0273/0.0039) IN CO-CULTURES. HOWEVER, %5-HMC WAS NOT CORRELATED WITH CLINICAL DATA AND WAS UNCHANGED FOLLOWING MICROENVIRONMENT STIMULI. CONCLUSIONS: DESPITE A SLIGHT DEREGULATION IN CLL CELLS COMPARED WITH NORMAL B CELLS, WE IDENTIFIED A SIGNIFICANT ASSOCIATION BETWEEN TET/IDH GENE EXPRESSION AND PROGNOSIS, SUGGESTING THAT EPIGENETIC CHANGES COULD POTENTIALLY BE ASSOCIATED WITH DISEASE PROGRESSION. MOREOVER, DESPITE AN IDENTICAL %5-HMC, TET GENE EXPRESSION WAS INFLUENCED BY CONTACT WITH BMSC CONFIRMING THE CRUCIAL ROLE OF THE MICROENVIRONMENT IN CLL PATHOGENESIS.	2016	
                                                                                                                                                                                                                                                                                                                             
19 2847  21 FREQUENT P15 PROMOTER METHYLATION IN TUMOR AND PERIPHERAL BLOOD FROM HEPATOCELLULAR CARCINOMA PATIENTS. WE PROSPECTIVELY ANALYZED P15 METHYLATION PATTERNS IN 25 SURGICALLY RESECTED TUMORS AND 130 PLASMA, SERUM, AND BUFFY COAT SAMPLES FROM HEPATOCELLULAR CARCINOMA (HCC) PATIENTS, CONTROLS WITH CHRONIC HEPATITIS/CIRRHOSIS, AND HEALTHY SUBJECTS. USING METHYLATION-SPECIFIC PCR, WE DEMONSTRATED FOR THE FIRST TIME P15 PROMOTER METHYLATION IN 64% OF TUMORS AND 25% (4 OF 16) OF PATIENTS' PLASMA AND SERUM SAMPLES. CONCURRENT P15 AND P16 METHYLATION WAS SHOWN IN 48% OF TUMORS, AND P15/P16 METHYLATION WAS DETECTED IN THE PLASMA/SERUM OF 92% (11 OF 12) OF PATIENTS. OF NOTE, 75% OF 12 PATIENTS WITH CONCURRENT TUMOR METHYLATION DEVELOPED CLINICAL METASTASIS/RECURRENCE (P = 0.027). IN BUFFY COAT SAMPLES, P15 METHYLATION WAS DETECTED IN ALL EIGHT PATIENTS WITH TUMOR P15 METHYLATION, SUGGESTING THE PRESENCE OF CIRCULATING TUMOR CELLS. NONE OF THE CONTROL SAMPLES WERE METHYLATION POSITIVE. OUR DATA UNDERSCORE THE IMPORTANT ROLE(S) OF P15 AND P16 METHYLATION IN HEPATOCARCINOGENESIS AND TUMOR PROGRESSION. AMONG 92% (23 OF 25) OF PATIENTS WITH TUMOR P15/P16 METHYLATION, CIRCULATING TUMOR DNA AND HCC CELLS WERE DETECTED IN THE PERIPHERAL BLOOD OF 87% (20 OF 23) OF PATIENTS. THE COMBINATION OF THESE EPIGENETIC MARKERS MAY PROVE VALUABLE FOR NONINVASIVE HCC DIAGNOSIS AND DISEASE MONITORING.	2000	
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                             
20 1849  28 EIGHT WEEKS OF PHYSICAL TRAINING DECREASES 2 YEARS OF DNA METHYLATION AGE OF SEDENTARY WOMEN. PURPOSE: THE ACCELERATION OF EPIGENETIC AGE IS A PREDICTOR OF MORTALITY AND CONTRIBUTES TO THE INCREASE IN CHRONIC DISEASES. ADHERENCE TO A HEALTHY LIFESTYLE IS A STRATEGY TO REDUCE EPIGENETIC AGE. THE PRESENT STUDY AIMED TO DETERMINE WHETHER EIGHT WEEKS OF COMBINED (AEROBIC AND STRENGTH) TRAINING (CT) CAN INFLUENCE THE EPIGENETIC AGE OF WOMEN BETWEEN 50 AND 70 YEARS OLD AND THE DIFFERENCES IN SITES AND METHYLATED REGIONS. METHODS: EIGHTEEN WOMEN (AAR(LOW): LOWER AGE ACCELERATION RESIDUAL, N = 10; AAR(HIGH): HIGHER AGE ACCELERATION RESIDUAL, N = 8) PARTICIPATED IN A COMBINED EXERCISE TRAINING PROGRAM (60 MINUTES, 3X A WEEK) FOR EIGHT WEEKS. DNA WAS EXTRACTED FROM WHOLE BLOOD USING THE SALTING OUT TECHNIQUE. DNA METHYLATION WAS PERFORMED USING THE ARRAY TECHNIQUE (ILLUMINA'S INFINIUM METHYLATION BEADCHIP 850K). WE USED THE DNA METHYLATION AGE CALCULATOR PLATFORM TO CALCULATE THE BIOLOGICAL EPIGENETIC AGE. TWO-WAY ANOVA FOLLOWED BY FISHER LSD POSTHOC WAS APPLIED, ADOPTING P < .05. RESULTS: AFTER EIGHT WEEKS OF CT, THERE WERE NO CHANGES TO THE EPIGENETIC AGE ACCELERATION FOR THE AAR(LOW) GROUP (PRE: -2.3 +/- 3.2 TO POST: -2.3 +/- 3.6). HOWEVER, THE AAR(HIGH) GROUP SIGNIFICANTLY DECREASED THE AGE ACCELERATION (PRE: 3.6 +/- 2.6 TO POST: 2.2 +/- 2.7) (GROUP EFFECT, P = .01; TIME EFFECT, P = .31; GROUP VS. TIME EFFECT, P = .005). CONCLUSION: CT FOR EIGHT WEEKS BENEFITS THE EPIGENETIC CLOCK OF WOMEN WITH THE MOST ACCELERATED AGE.	2023