|A 115-bp MethyLight assay for detection of p16 (CDKN2A) methylation as a diagnostic biomarker in human tissues|
|Zhou, Jing1; Cao, Jie2; Lu, Zheming1; Liu, Hongwei2; Deng, Dajun1|
|刊名||BMC MEDICAL GENETICS|
|WOS标题词||Science & Technology|
|类目[WOS]||Genetics & Heredity|
|研究领域[WOS]||Genetics & Heredity|
|关键词[WOS]||ORAL EPITHELIAL DYSPLASIA ; DE-NOVO METHYLATION ; CPG ISLAND ; BARRETTS-ESOPHAGUS ; PROMOTER HYPERMETHYLATION ; MALIGNANT-TRANSFORMATION ; NEOPLASTIC PROGRESSION ; DNA ; ASSOCIATION ; VALIDATION|
Background: p16 Methylation is a potential biomarker for prediction of malignant transformation of epithelial dysplasia. A probe-based, quantitative, methylation-specific PCR (MSP) called MethyLight may become an eligible method for detecting this marker clinically. We studied oral mucosa biopsies with epithelial dysplasia from 78 patients enrolled in a published 4-years′ followup cohort, in which cancer risk for patients with p16 methylation positive dysplasia was significantly higher than those without p16 methylation (by 150-bp MSP and bisulfite sequencing; +133 similar to +283, transcription starting site, +1). The p16 methylation status in samples (N = 102) containing sufficient DNA was analyzed by the 70-bp classic (+238 similar to +307) and 115-bp novel (+157 similar to +272) MethyLight assays, respectively.
Results: p16 Methylation was detectable in 75 samples using the classic MethyLight assay. The methylated-p16 positive rate and proportion of methylated-p16 by the MethyLight in MSP-positive samples were higher than those in MSP-negative samples (positive rate: 37/44 vs. 38/58, P=0.035, two-sided; proportion [median]: 0.78 vs. 0.02, P <0.007). Using the published results of MSP as a golden standard, we found sensitivity, specificity, and accuracy for this MethyLight assay to be 70.5%, 84.5%, and 55.0%, respectively. Because amplicon of the classic MethyLight procedure only partially overlapped with the MSP amplicon, we further designed a 115-bp novel MethyLight assay in which the amplicon on the sense-strand fully overlapped with the MSP amplicon on the antisense-strand. Using the 115-bp MethyLight assay, we observed methylated-p16 in 26 of 44 MSP-positive samples and 2 of 58 MSP-negative ones (P = 0.000). These results were confirmed with clone sequencing. Sensitivity, specificity, and accuracy using the 115-bp MethyLight assay were 59.1%, 98.3%, and 57.4%, respectively. Significant differences in the oral cancer rate were observed during the followup between patients (>= 60 years) with and without methylated-p16 as detected by the 115-bp MethyLight assay (6/8 vs. 6/22, P = 0.034, two-sided).
Conclusions: The 115-bp MethyLight assay is a useful and practical assay with very high specificity for the detection of p16 methylation clinically.
|项目编号||434 ; Z090507017709016 ; 2006AA020902 ; 2006AA02A402|
|资助机构||Capital Program for Development of Health Science ; Beijing Science and Technology Commission ; Key Technologies Research & ; Development Programs (863)|
|作者单位||1.Peking Univ, Key Lab Carcinogenesis & Translat Res, Minist Educ, Dept Aetiol,Canc Hosp & Inst, Beijing 100142, Peoples R China|
2.Peking Univ, Dept Oral Med, Sch Stomatol, Beijing 100081, Peoples R China
|Zhou, Jing,Cao, Jie,Lu, Zheming,et al. A 115-bp MethyLight assay for detection of p16 (CDKN2A) methylation as a diagnostic biomarker in human tissues[J]. BMC MEDICAL GENETICS,2011,12.|
|APA||Zhou, Jing,Cao, Jie,Lu, Zheming,Liu, Hongwei,&Deng, Dajun.(2011).A 115-bp MethyLight assay for detection of p16 (CDKN2A) methylation as a diagnostic biomarker in human tissues.BMC MEDICAL GENETICS,12.|
|MLA||Zhou, Jing,et al."A 115-bp MethyLight assay for detection of p16 (CDKN2A) methylation as a diagnostic biomarker in human tissues".BMC MEDICAL GENETICS 12(2011).|