Detection of a novel point mutation in the p53 gene in grade II astrocytomas by PCR-SSCP analysis with additional Klenow treatment

Abstract

Using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) with additional Klenow treatment and direct sequencing, mutations in the p53 tumor suppressor gene were analyzed from 21 cases of human astrocytomas. Three cases had p53 gene mutations: two of them were glioblastomas showing a point mutation, one in exon 5 and the other in 6. The last one was a gemistocytic astrocytoma showing a point mutation in exon 5. The frequency of p53 gene mutations in the astrocytomas examined was 14.3% (3 out of 21). No SSCP alterations were observed in any of the p53 fragments amplified from WHO grade I pilocytic astrocytomas and WHO grade III anaplastic astrocytomas. Further examination by direct sequencing showed that two mutations of glioblastomas had single-base substitutions resulting in silent and missense mutations, whereas one of the gemistocytic astrocytomas had a double-base substitution resulting in a missense mutation. The present studies revealed that all mutations were located outside the hot spots and, interestingly, one of them disclosed a missense mutation in exon 5 at codon 166, which was first detected in a grade H astrocytoma (gemistocytic type). It is possible that the missense mutation at this codon may be associated with special risk factors for the development of astrocytic tumors in Thai patients.