The role of TP53 gain-of-function mutation in multifocal glioblastoma

Abstract

Purpose: The phenotypic and genotypic landscapes in multifocal glioblastoma (MF GBM) cases can vary greatly among lesions. In a MF GBM patient, the rapid development of a secondary lesion was investigated to determine if a unique genetic signature could account for the apparent increased malignancy of this lesion.

Methods: The primary (G52) and secondary (G53) tumours were resected to develop patient derived models followed by functional assays and multiplatform molecular profiling.

Results: Molecular profiling revealed G52 was wild-type for TP53 while G53 presented with a TP53 missense mutation. Functional studies demonstrated increased proliferation, migration, invasion and colony formation in G53.

Conclusion: This data suggests that the TP53 mutation led to gain-of-function phenotypes and resulted in greater overall oncogenic potential of G53.

Keywords: Gain-of-function; Glioblastoma; Multifocal; TP53.

Fig. 1

Pre-operative MRI scans exhibiting two expansive bulky lesions arising in the left thalamus (a and b) and cerebella hemisphere c with an irregular ring contrast enhancement

Fig. 2

Nucleotide sequence analysis of TP53 exon 8 in G52 and G53 tumour samples. a Wt nucleotide sequence at codon 273. b G52 and G53 tumour nucleotide sequence at codon 273. Dot indicates the position of the C to T nucleotide substitution and c.818G.A (p.R273H) mutation of the TP53 gene in the G53 tumour

Fig. 3

p53 mutation promotes cell proliferation, migration and invasion in multifocal primary glioblastoma cell lines in vitro. a Real-time xCelligence analysis of proliferation (represented by cell index) of G52 (red) and G53 (black). b 24 h timepoint analysis of proliferation (represented by cell index) levels between G53 and G52. Error bars represent standard deviations. p < 0.1; *p < 0.02. c Real-time xCelligence analysis of migration (represented by cell index) of G53 and G52. d 24 h timepoint analysis of migration (represented by cell index) levels between G53 (left frontal lesion) and G52 (left thalamic lesion). Error bars represent standard deviations. p < 0.1; p < 0.8. e Real-time xCelligence analysis of invasion (represented by cell index) of G53 (black) and G52 (red). f 24 h timepoint analysis of invasion (represented by cell index) levels between G53 (left frontal lesion) and G52 (left thalamic lesion). Error bars represent standard deviations. p < 0.2; p < 0.1

Fig. 4

Colony formation escalation in a p53 mutant cell line. a G53 colony formation (crystal violet stain) increases across all cell seeding densities (50 to 600 cells) over 14 days compared to G52 (inset magnification × 20). b Colonies were quantified using a stereomicroscope and colony counting pen. G53 colony counts consistently increase over time with increasing cell density, whereas G52 demonstrates no change in formation

Fig. 5

Higher cell proliferation in p53 mutant. a Labelling index for Ki-67 in G52 and G53 (×40 magnification). b LI was measured as the percentage of positive cells per 1000 cells. *p < 0.05