Detection of K-ras mutations in pancreatic and hepatic neoplasms by non-isotopic mismatched polymerase chain reaction.
Mutations within codon 12 leading to activation of Kirsten-ras (K-ras) genes occur in a wide variety of human tumors, but have been reported most frequently in pancreatic carcinomas. We studied twenty-four paraffin-embedded pancreatic and hepatic tumors and two colon carcinoma cell lines with a rapid and simple approach that exploits allele-specific amplification of genomic DNA in a polymerase chain reaction (PCR). We extend the utility of this technique, which is dependent on an exact match at the 3' nucleotide between synthetic oligonucleotides and template DNA, to analyse paraffin-embedded tumor samples for the presence of point mutations at the first and second base of codon 12 of the K-ras gene. The PCR mismatch amplification technique demonstrated a 66% incidence of K-ras mutations at codon 12 in the group of pancreatic neoplasms as a whole. The percentage of mutations varied only slightly in the pancreatic cancer subcategories: 75% in ampullary, 66% in bile duct and 57% in the ductal adenocarcinomas. One islet cell carcinoma and normal tissues adjacent to the tumors revealed wild-type alleles only. One hepatoblastoma and one of six hepatocellular carcinomas also had codon 12 mutations. The PCR mismatch is a sensitive and rapid method that may be useful in screening neoplasms for K-ras point mutation and can be applied to archival material. This application allows a retrospective analyses of a wide range of pathological specimens to determine the role of K-ras mutations in human tumorigenesis.
Stork
P
P
New England Medical Center Hospital, Department of Pathology, Boston, Massachusetts 02111.
Loda
M
M
Bosari
S
S
Wiley
B
B
Poppenhusen
K
K
Wolfe
H
H
D000595:Amino Acid Sequence; D001483:Base Sequence; D006528:Carcinoma, Hepatocellular / Q000235:genetics; D002460:Cell Line; D003062:Codon / Q000235:genetics; D003110:Colonic Neoplasms; D004273:DNA, Neoplasm / Q000235:genetics / Q000302:isolation & purification; D005091:Exons; D011905:Genes, ras; D006801:Humans; D008113:Liver Neoplasms / Q000235:genetics* / Q000473:pathology; D008969:Molecular Sequence Data; D009154:Mutation; D015345:Oligonucleotide Probes; D010190:Pancreatic Neoplasms / Q000235:genetics* / Q000473:pathology; D016133:Polymerase Chain Reaction / Q000379:methods*
D003062:Codon; D004273:DNA, Neoplasm; D015345:Oligonucleotide Probes