The Significance Of Promoter Hypermethylation Of P16 Gene In Circulating Dna On Early Diagnosis Of Non-small Cell Lung Cancer

ObjectiveTo evaluate the diagnostic value of detecting promoter hypermethylation of p16 gene in circulating DNA from non-small cell lung cancer (NSCLC) patients and high-risk groups of NSCLC.MethodsCirculating free DNA was extracted from plasma which derived from peripheral venous blood collecting from 21 cases of diagnosed NSCLC patients, 20 cases of healthy heavy smokers and 15 cases of healthy non-smokers. After the circulating DNA was treated by bisulfate, Methylation-specific PCR (MSP) was performed for the detection of promoter hypermethylation of p16 gene in circulating DNA from NSCLC patients and that from high-risk groups of NSCLC (i.d. healthy heavy smokers). Whole blood DNA from healthy non-smokers treated by SSI methyltransferase was used as a positive control of methylation, while placental blood served as a negative control of methylation and positive control of non-methylation, and pure water was used as a negative control of non-methylation.Results1.The concentration and purity of whole blood DNA extract and placental blood DNA extract was better enough to be treated by bisulfate and SSI methyltransferase.2.The content of circulating DNA in extract was enough to be treated by bisulfate.3.The detectable rate from NSCLC patients compared with that from the heavy smoking group and that from the non-smoking healthy group: The detectable rates of promoter hypermethylation of p16 gene in circulating DNA from NSCLC patients and the heavy smoking group were 28.6% (6/21) and 10%(2/20), respectively. However, there was no case of the non-smoking healthy group exhibited aberrant methylation in promoter of p16 gene in circulating DNA. The detectable rate from NSCLC patients higher than that from the non-smoking healthy group (P=0.030), while there was no significant difference between the detectable rate from NSCLC patients and that from the heavy smoking group (P=0.238), as well as that from the heavy smoking group and that from the non-smoking healthy group (P=0.496).4.The detectable rates from each tumor type and stage: The detectable rates of promoter hypermethylation of p16 gene in circulating DNA from squalors cell carcinoma patients and adenous cancer patients were 37.6% (3/8) and 23.1%(3/13), respectively. There was no significant difference between the detectable rate from squamous patients and that from Adenocarcinoma patients (P=0.631), as well as that from early NSCLC patients (16.7%, 1/6) and that from advanced NSCLC patients (33.3%, 5/15) (P=1.000).Conclusion1.The frequency of promoter hypermethylation of p16 gene in circulating DNA from NSCLC patients was significantly higher.2.There was no significant difference between the detectable rate from squamous patients and that from Adenocarcinoma patients, as well as that from early NSCLC patients and that from advanced NSCLC patients.3.The frequency of p16 aberrant methylation was promoted, may be related to the lung damage caused by smoking.4. Detecting promoter hypermethylation of p16 gene in circulating DNA may be helpful screening for NSCLC in high-risk population, and for the early diagnosis of NSCLC.