TP53 mutation analysis of low-grade astrocytomas and their recurrences

A33 Introduction: TP53 is a key tumor suppressor gene on chromosome 17p13.1 that encodes a transcription factor involved in several cellular mechanisms including growth arrest, DNA repair, and induction of apoptosis. TP53 mutations are the most frequent and earliest detectable genetic alterations in diffusely infiltrating astrocytomas and are already present in about 50-75% of low-grade astrocytomas. De novo glioblastomas typically lack TP53 mutations while secondary glioblastomas are characterized by the presence of TP53 mutations. These mutations are believed to be already present in the precursor low-grade astrocytic tumors. The data underlying this assumption are derived from transversal investigations of cohorts of gliomas of various malignancy grades. In the present study we investigated and compared the presence of TP53 mutations in primary low-grade astrocytomas and their recurrences in a longitudinal setting, in order to study the consistence of such mutations over time.
 Materials and Methods: We analysed a serie of 31 primary low-grade astrocytomas with 14 recurrences and 9 primary high-grade astrocytic tumors with 4 recurrences. DNA from routinely formalin-fixed paraffin-embedded tumor specimens was isolated and mutation analysis for exons 4 to 9 of the TP53 gene was performed by bi-directional DNA sequencing. All cases were investigated for p53 expression by immunohistochemistry.
 Results: P53 immunohistochemistry was positive in 80.6% (25/31) of the primary low-grade astrocytomas and in 87.5% (14/16) of their recurrences, although the number of positive cells varied between sections. All 9 primary high-grade astrocytic tumors and their 4 recurrences were p53 positive. We could effectively perform TP53 mutation analysis in DNA from 93.1% (54/58) of the glioma tissues. TP53 mutations were present in 64.5% (20/31) of primary low-grade astrocytomas and in 56.3% (9/16) of recurrent tumors. In the high-grade lesions, TP53 mutations were found in 88.9% (8/9) of the primary tumors and in 100% (4/4) of the recurrences. In two patients a not previously reported in-frame duplication of 18 nucleotides (codon 107-112) in exon 4 was found. Mutational hotspots were codon 175 of exon 5 (13.0%), codon 220 of exon 7 (13.0%) and codon 273 of exon 8 (14.8%). Identical TP53 mutations in the primary and the recurrent tumor were present in 12 of 14 patients, suggestive of clonal relationship between the tumor pairs.
 Discussion and Conclusion: In this study, a successful DNA sequencing procedure was established for detection of TP53 mutations on archival neural tumors. We showed in a longitudinal setting that mutations in the TP53 gene are early and frequent events during progression from low-grade astrocytomas to secondary glioblastomas, and demonstrated genetic homology between primary and recurrent astrocytomas.