Detection of loss of heterozygosity of p53 gene in paraffin‐embedded breast cancers by non‐isotopic PCR–SSCP

Abstract

A rapid non‐isotopic PCR‐SSCP (polymerase chain reaction‐single‐stranded conformation polymorphism) method was developed in this study to detect polymorphism and loss of heterozygosity (LOH) of p53 in formalin‐fixed and paraffin‐embedded samples of normal breast tissue and of breast cancer. p53 expression was also examined by immunohistochemistry. In 35 paired samples, heterozygosity in exon 4 of p53 was detected in 17 cases (49 per cent) and LOH of the p53 gene in breast cancer tissues was observed in 7 out of 15 informative cases (47 per cent). The correlation of LOH of p53 with positive p53 immunostaining did not reach statistical significance, but all immunostaining‐positive tumours among informative cases had LOH of p53. The results support the hypothesis that in most cases the allelic deletion of p53 may uncover the ‘recessive mutation’ in the remaining allele. However, LOH of p53 was more frequent than positive immunostaining and was significantly associated with poor differentiation of breast cancer (P<0·05). The results suggest that the allelic deletion of p53 may also contribute to the development and progression of breast cancer by reducing the amount of normal p53 protein. These results show that non‐isotopic PCR‐SSCP is a simple, fast, and effective method for detecting polymorphism and LOH of the p53 gene, which is especially useful for retrospective studies.