Detection of c-Ki-ras Mutation by PCR/RFLP Analysis and Diagnosis of Pancreatic Adenocarcinomas

K- ras activation by point mutation in codon 12 has been reported in lung adenocarcinomas in various models of experimental lung tumours induced by chemical carcinogens. The hypothesis of the presence of cells containing K- ras mutation in non neoplastic bronchial carina, the main site of impaction of airborne contaminants, was investigated by evaluating concurrent lung tumour and non-neoplastic proximal bronchial carinae from 19 patients with lung adenocarcinomas. The restriction fragment length polymorphism enriched PCR method used can detect one mutant allele among 10 3 normal alleles. A mutation was detected in 42% of lung adenocarcinoma samples. No mutation was detected in either tumour or bronchial carinae in nine patients (47%). K- ras mutation was detected in the lung tumour but not in bronchial carinae in four patients (21%), in both the lung tumour and bronchial carinae in four other patients (21%). In two patients (11%), K- ras mutation was detected in at least one bronchial carina, but not in the lung tumour. Mutations of codon 12, confirmed by sequencing analysis of ten samples, were G to T transversion, mostly TGT and GTT in bronchial carinae and lung tumours. Our data show that activated K- ras by point mutation can be present in non-neoplastic bronchial carina mucosa even when no mutation is detected in tumour samples. © 2000 Cancer Research Campaign

Section: Discussionmentioning

confidence: 99%

Detection of codon 12 K- ras mutations in non-neoplastic mucosa from bronchial carina in patients with lung adenocarcinomas

Urban

Ricci²,

Danel

et al. 2000

“…The sensitivity of the assay was therefore tested by a series of titration experiments. The threshold of detection was one cell with a homozygous mutation in the midst of 102 cells with a wild-type Ki-ras gene (Figure 2) (Urban et al, 1993). Tables I and II. When using the one step PCR-RFLP method no mutation involving the codon 12 of Ki-ras gene was detected in the 20 squamous cell and four adenosquamous cell carcinomas.…”

Section: Methodsmentioning

confidence: 99%

Codon 12 Ki-ras mutation in non-small-cell lung cancer: comparative evaluation in tumoural and non-tumoural lung

Urban

Ricci²,

Lacave³

et al. 1996

Summary Ki-ras activation by point mutation on codon 12 has been reported in non-small-cell lung carcinomas and in various models of experimental lung tumours induced by chemical carcinogens. The cellular targets for carcinogenic compounds of tobacco smoke are usually considered to be the cells of the bronchial mucosa or alveolar epithelium. However, little is known about preneoplastic events in bronchopulmonary carcinogenesis. The hypothesis of the presence of widespread target cells containing Ki-ras mutation was investigated by evaluating concurrent neoplastic and non-neoplastic bronchial and alveolar samples from 51 patients with non-small-cell lung carcinomas. The polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method used can detect one cell with a mutation on codon 12 among 102 normal cells. In tumour samples, a mutation was detected in 20% of adenocarcinomas, but in none of the adenosquamous or squamous cell carcinomas. No mutation was detected in the non-neoplastic bronchial or parenchymal samples. When using an enriched PCR-RFLP method detecting one mutated allele among 103 normal alleles a mutation was detected in 23% of adenocarcinomas. In conclusion, Ki-ras activation by mutation on codon 12 was not observed in non-neoplastic bronchial or parenchymal tissues in patients with bronchopulmonary cancers and does not appear to be a genetic event present in non-malignant epithelial target cells exposed to tobacco smoke.

“…In pancreatic cancer, point mutations of the k-ras oncogene have been shown with a prevalence of up to 90% (Almoguera et al, 1988;Hruban et al, 1993;Urban et al, 1993). Several studies have demonstrated the possibility to detect these mutations in pancreatic juice specimens or duodenal fluid (Tada et al, 1993;Trümper et al, 1994;Berthelemy et al, 1995;Van Laethem et al, 1995).…”

mentioning

confidence: 99%

Methylation status of p14ARF and p16INK4a as detected in pancreatic secretions

Klump¹,

Hsieh²,

Nehls³

et al. 2003

The clinical management of pancreatic disease is often hampered by a lack of tissue diagnosis. Endoscopic pancreatography offers the opportunity to investigate exfoliated cells. However, the significance of mere cytological investigation is compromised by an insufficient sensitivity. The evaluation of the molecular background of carcinogenesis hopefully is capable of providing more sensitive diagnostic markers. The p16INK4a-/retinoblastoma tumour-suppressive pathway has been shown to be involved in the development of near to all pancreatic neoplasms. p14ARF is another tumour suppressor located in the immediate neighbourhood of p16INK4a. Promoter methylation has been demonstrated to be a major inactivating mechanism of both genes. We sought to further evaluate the role of the gene locus INK4a methylation status in the endoscopic differentiation of chronic inflammatory and neoplastic pancreatic disease. Pancreatic fluid specimens of 61 patients with either pancreatic carcinoma (PCA: 39), chronic pancreatitis (CP: 16) or a normal pancreatogram (NAD: 6) were retrieved. In order to detect methylation of either the p14ARF or the p16INK4a promoter a methylation-specific PCR protocol was applied. While 19 out of 39 patients with PCA showed p16 promoter methylation (49%), none of the 16 patients with CP revealed p16 promoter methylation. p14ARF methylation was found in a lower percentage of PCA specimens and in none of the samples of patients with CP. These results suggest a specific significance of INK4a for the development of malignant pancreatic disease. Our data further indicate a potential role for INK4a methylation as a diagnostic marker in the endoscopic differentiation of benign and malignant pancreatic disease.