Highly sensitive determination of the methylated p16 gene in cancer patients by microchip electrophoresis.

The p16 tumor suppressor gene is inactivated by promoter region hypermethylation in many types of tumor. Recent studies showed that aberrant methylation of the p16 gene is an early event in many tumors, especially in lung cancer, and may constitute a new biomarker for early detection and monitoring of prevention trials. We detected tumor-associated aberrant hypermethylation of the p16 gene in plasma and tissue DNA from 153 specimens using a modified semi-nested methylation-specific PCR (MSP) combining plastic microchip electrophoresis or slab gel electrophoresis, respectively. Specimens were from 79 lung cancer patients, 15 abdominal tumor patients, 30 positive controls and 30 negative controls. The results showed that the positive rate obtained by microchip electrophoresis was more than 26.6% higher and the same specificity was kept when compared with slab gel electrophoresis. The microchip electrophoresis can rapidly and accurately analyze the PCR products of methylated DNA and obviously improve the positive rate of diagnosis of cancer patients when compared with gel electrophoresis. This method with the high assay sensitivity might be used for detection of methylation of p16 gene and even to facilitate early diagnosis of cancer patients.

[Detection of hypermethylation of p16 gene in plasma DNA from lung cancer patients].

OBJECTIVE: To detect hyper methylation of p16 gene in plasma DNA from patients with lung cancer, and to assess its potential as a malignant marker. METHODS: Using a modified semi-nested methylation-specific PCR (MSP), the status of methylation of the p16 was investigated in plasma DNA from 137 lung cancer patients and 112 matched tumor tissues. RESULTS: Hypermethylation of the p16 was present in 75.2% (103/137) of the plasma samples and 80.4% (90/112) of the tumor tissues. Hypermethylation of the p16 in the plasma was detected in 77.9% squamous-cell carcinoma, 65.1% adenocarcionma, 75.1% adeno-squamous-cell carcinoma, and 91.7% small-cell lung cancer. Only in those patients whose tumor tissues had hypermethylation of p16 gene, similar changes could be detected in their plasma samples. Hypermethylation of the p16 in plasma and the corresponding tumor tissues was not significantly correlated with the clinical stage and pathological type of the tumor. CONCLUSION: The result indicates that hypermethylation of the p16 may be a useful marker in the auxiliary diagnosis of lung cancer.

[Hypermethylation of p16 gene in clinical specimens of patients with lung cancer].

OBJECTIVE: To evaluate aberrant methylation of the p16 promoter as a useful biomarker of lung cancer. METHODS: A modified methylation-specific semi-nested PCR was performed to detect p16 hypermethylation in the matched samples of tumor tissue, blood plasma and sputum derived from 51 cases of lung cancer patients. RESULTS: Hypermethylation of p16 promoter was demonstrated in 84.3% of the tumor tissues, 70.6% of the blood plasma and 76.5% of the sputum specimens, respectively. Only the patients whose tumor tissues had p16 hypermethylation exhibited aberrant methylation in their plasma and/or sputum specimens. Combining with cytological examination, 92.2% of the patients with lung cancer could be detected by p16 hypermethylation assay in both sputum and plasma samples. CONCLUSION: The results indicate that p16 hypermethylation in plasma and sputum identified by semi-nested PCR is a biomarker of lung cancer which can be useful as an auxillary diagnostic parameter.