Molecular approach to rapid assessment of p53 tumor suppressor mutations in esophageal tumors from stained histological slides.
The analysis of the tumor suppressor gene, p53, is of fundamental importance in prognosis and staging in many cancers; however, the molecular techniques required to analyze this gene have been expensive, time consuming, and unrelatable to the histological appearance of the samples. This research explored one model of clinically testing for specific mutations in the p53 gene by scraping selected areas of stained histological slides and analyzing for "hot-spot" p53 mutations. Selectively removing samples from the stained histological slide will be of special value in examining suspicious regions in adenomas, potential metastatic regions, and the margins of resected area. A polymerase chain reaction (PCR)-mediated restriction fragment length polymorphism (RFLP) analysis approach in which naturally occurring or primer-mediated mutagenesis-induced restriction enzyme sites were utilized to test seven hot-spot mutations. These assays were able to detect one mutated sequence in 100, and therefore, were sufficiently sensitive to be used with very heterogeneous tumors. Several of the assays could be multiplexed to reduce the number of PCRs necessary to screen for the seven mutational hot spots. Furthermore, an exact determination of the base change could be obtained by direct sequencing of the PCR products. Although this form of analysis may be applicable only to certain types of cancers (e.g., bladder, brain, colon, esophageal, gastric, thyroid, and ovarian tumors), this approach can obtain detailed mutational information from specific regions of a histological slide in a cost-effective and timely manner.
Whetsell
L H
LH
H.A. Chapman Research Institute for Medical Genetics, Children's Medical Center, Tulsa, OK 74135.
Ringer
D P
DP
Schaefer
F V
FV
D000236:Adenoma / Q000235:genetics*; D001483:Base Sequence; D015342:DNA Probes; D004938:Esophageal Neoplasms / Q000235:genetics*; D016158:Genes, p53 / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D016133:Polymerase Chain Reaction; D013194:Staining and Labeling; D014407:Tumor Cells, Cultured
D015342:DNA Probes