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Detection of c-Ki-ras mutation by PCR/RFLP analysis and diagnosis of pancreatic adenocarcinomas.
J Natl Cancer Inst. 1993 Dec 15; 85(24):2008-12.JNCI

Abstract

BACKGROUND

The c-Ki-ras oncogene (also known as KRAS2) is activated by point mutations involving codon 12 in 72%-100% of primary pancreatic adenocarcinomas, but the gene is not activated in nonneoplastic tissues. Therefore, the detection of c-Ki-ras mutations can facilitate the diagnosis of pancreatic adenocarcinomas, which are not always identified with current tests. Detection is usually performed by oligonucleotide hybridization combined with polymerase chain reaction (PCR), RNAse mismatch cleavage assay, or non-isotopic mismatched PCR, methods that are not feasible for routine screening of large numbers of samples because they are time consuming and/or expensive.

PURPOSE

Our purpose was to evaluate a rapid, non-radioactive method of detection of a mutation in codon 12 of the c-Ki-ras gene in pancreatic tumor samples obtained by fine-needle aspiration for diagnostic screening.

METHODS

Twenty consecutive patients (15 with pancreatic adenocarcinoma, one with pancreatic cystadenocarcinoma, one with endocrine islet cell tumor, and three with chronic pancreatitis) were selected for this study. A sample of pancreatic tissue from each patient with a tumor or pancreatitis was obtained and evaluated by fine-needle aspiration biopsy under computerized tomography scan or ultrasound guidance using a two-needle coaxial technique. Pancreatic DNA from each of these samples was evaluated by PCR amplification and restriction fragment length polymorphism (RFLP) analysis with nucleotide substitution in PCR primers, creating BstNI restriction patterns that distinguished mutated from normal alleles. The accuracy of the PCR/RFLP assay was validated with DNA from SW480 and HT29 colonic carcinoma cell lines with known mutated and wild-type c-Ki-ras gene sequences. Sensitivity was tested with a series of titration experiments.

RESULTS

PCR/RFLP analysis can detect a mutation present in 1% of cells. No amplification could be performed in four (20%) samples because of the absence of cells in the aspirated sample. In the 16 samples adequate for PCR/RFLP analysis, a c-Ki-ras gene mutation was detected in 11 (92%) of 12 adenocarcinomas. Overall, diagnosis was obtained by pathologic (cytomorphologic) examination alone in 13 samples (81%). The presence of malignant cells and/or mutated c-Ki-ras gene was detected in 12 of 12 adenocarcinomas but not in chronic pancreatitis or islet cell tumor.

CONCLUSION

Screening of pancreatic tissue samples obtained by fine-needle aspiration for c-Ki-ras mutation using PCR/RFLP analysis combined with pathologic examination could facilitate diagnosis of pancreatic tumors.

Authors+Show Affiliations

Laboratory of Histology and Tumor Biology, Hôpital Tenon, Paris, France.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

7902444

Citation

Urban, T, et al. "Detection of c-Ki-ras Mutation By PCR/RFLP Analysis and Diagnosis of Pancreatic Adenocarcinomas." Journal of the National Cancer Institute, vol. 85, no. 24, 1993, pp. 2008-12.
Urban T, Ricci S, Grange JD, et al. Detection of c-Ki-ras mutation by PCR/RFLP analysis and diagnosis of pancreatic adenocarcinomas. J Natl Cancer Inst. 1993;85(24):2008-12.
Urban, T., Ricci, S., Grange, J. D., Lacave, R., Boudghene, F., Breittmayer, F., Languille, O., Roland, J., & Bernaudin, J. F. (1993). Detection of c-Ki-ras mutation by PCR/RFLP analysis and diagnosis of pancreatic adenocarcinomas. Journal of the National Cancer Institute, 85(24), 2008-12.
Urban T, et al. Detection of c-Ki-ras Mutation By PCR/RFLP Analysis and Diagnosis of Pancreatic Adenocarcinomas. J Natl Cancer Inst. 1993 Dec 15;85(24):2008-12. PubMed PMID: 7902444.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Detection of c-Ki-ras mutation by PCR/RFLP analysis and diagnosis of pancreatic adenocarcinomas. AU - Urban,T, AU - Ricci,S, AU - Grange,J D, AU - Lacave,R, AU - Boudghene,F, AU - Breittmayer,F, AU - Languille,O, AU - Roland,J, AU - Bernaudin,J F, PY - 1993/12/15/pubmed PY - 1993/12/15/medline PY - 1993/12/15/entrez SP - 2008 EP - 12 JF - Journal of the National Cancer Institute JO - J Natl Cancer Inst VL - 85 IS - 24 N2 - BACKGROUND: The c-Ki-ras oncogene (also known as KRAS2) is activated by point mutations involving codon 12 in 72%-100% of primary pancreatic adenocarcinomas, but the gene is not activated in nonneoplastic tissues. Therefore, the detection of c-Ki-ras mutations can facilitate the diagnosis of pancreatic adenocarcinomas, which are not always identified with current tests. Detection is usually performed by oligonucleotide hybridization combined with polymerase chain reaction (PCR), RNAse mismatch cleavage assay, or non-isotopic mismatched PCR, methods that are not feasible for routine screening of large numbers of samples because they are time consuming and/or expensive. PURPOSE: Our purpose was to evaluate a rapid, non-radioactive method of detection of a mutation in codon 12 of the c-Ki-ras gene in pancreatic tumor samples obtained by fine-needle aspiration for diagnostic screening. METHODS: Twenty consecutive patients (15 with pancreatic adenocarcinoma, one with pancreatic cystadenocarcinoma, one with endocrine islet cell tumor, and three with chronic pancreatitis) were selected for this study. A sample of pancreatic tissue from each patient with a tumor or pancreatitis was obtained and evaluated by fine-needle aspiration biopsy under computerized tomography scan or ultrasound guidance using a two-needle coaxial technique. Pancreatic DNA from each of these samples was evaluated by PCR amplification and restriction fragment length polymorphism (RFLP) analysis with nucleotide substitution in PCR primers, creating BstNI restriction patterns that distinguished mutated from normal alleles. The accuracy of the PCR/RFLP assay was validated with DNA from SW480 and HT29 colonic carcinoma cell lines with known mutated and wild-type c-Ki-ras gene sequences. Sensitivity was tested with a series of titration experiments. RESULTS: PCR/RFLP analysis can detect a mutation present in 1% of cells. No amplification could be performed in four (20%) samples because of the absence of cells in the aspirated sample. In the 16 samples adequate for PCR/RFLP analysis, a c-Ki-ras gene mutation was detected in 11 (92%) of 12 adenocarcinomas. Overall, diagnosis was obtained by pathologic (cytomorphologic) examination alone in 13 samples (81%). The presence of malignant cells and/or mutated c-Ki-ras gene was detected in 12 of 12 adenocarcinomas but not in chronic pancreatitis or islet cell tumor. CONCLUSION: Screening of pancreatic tissue samples obtained by fine-needle aspiration for c-Ki-ras mutation using PCR/RFLP analysis combined with pathologic examination could facilitate diagnosis of pancreatic tumors. SN - 0027-8874 UR - https://www.unboundmedicine.com/medline/citation/7902444/Detection_of_c_Ki_ras_mutation_by_PCR/RFLP_analysis_and_diagnosis_of_pancreatic_adenocarcinomas_ L2 - https://academic.oup.com/jnci/article-lookup/doi/10.1093/jnci/85.24.2008 DB - PRIME DP - Unbound Medicine ER -