Elevated frequency and functional activity of a specific germ-line p53 intron mutation in familial breast cancer

Previous studies have determined that the frequency of germ-line p53 mutati ons in familial breast cancer patients is 1 % or less, but these reports ha ve not investigated the importance of polymorphic intron base changes in th e p53 gene. Therefore, we investigated the frequency of both exon and intro n germ-line p53 base changes in 42 breast canter patients with a strong fam ily history of breast cancer. The mean age of presentation of these patient s was 44.0 years (range, 29-69), and 12 of 42 (29%) were of known Ashkenazi ancestry, Purified DNA obtained from the 42 index cases was screened for g erm-line p53 mutations in exons 2-11 and surrounding introns using a combin ation of intron based primers for PCR-single strand conformation polymorphi sm analysis, direct sequencing, and microarray sequencing using the Affymet rix p53 gene chip methodology, Morphological analysis of apoptosis and cell survival determination were performed on EBV-immortalized Lymphoblastoid c ell lines from patients with the p53 intron 6 mutation. A germ-line mutatio n in the p53 gene at nucleotide 13964 with a G to C base change (13964(GC)) was identified in 3 of 42 (7.1%) hereditary breast cancer patients, Two pa tients were heterozygous for this mutation, and one patient had a homozygou s mutation, In comparison, 0 of 171 (0%) of sporadic breast cancer patients had the p53 13964(GC) mutation (P = 0.0003), We found that 0 of 42 (0%) of these hereditary breast cancer patients had other germ-line p53 mutation i n exons 2-11, However, pedigree analysis demonstrated that all three patien ts had strong family histories of multiple types of cancers consistent with Li-Fraumeni syndrome but with late age of onset. Comprehensive BRCA1 and B RCA2 nucleotide analysis from patients with the p53 13964(GC) mutation reve aled no concomitant deleterious BRCA1 or BRCA2 mutations, although they wer e found in the other hereditary breast cancer patients, Functional analysis of tno immortalized lymphoblastoid cell lines derived from patients with t he p53 13964GC mutation demonstrated prolonged in vitro survival in respons e to cisplatinum treatment and showed decreased chemotherapy-induced apopto sis, Immunohistochemical analysis of breast tumors from these patients reve aled high levels of mutant p53 protein, suggesting a functional mutation in the p53 gene. In summary, we have identified a single p53 intron mutation in familial breast cancer patients that is present at elevated frequency an d has functional activity.