Whole genome duplication is an early event leading to aneuploidy in IDH-wild type glioblastoma

Abstract

Glioblastoma, the most frequent and lethal form of glioma, displays chromosome instability and recurrent somatic copy number alterations (SCNA). Chromothripsis and whole genome duplication (WGD) have been recently identified in cancer. In the present study, we analyzed SCNA and determine the ploidy pattern in 123 IDH-wild-type glioblastomas, using SNP array data. WGD and chromothripsis events were validated using, respectively, FISH and CTLPScanner. WGD was detected in 11.4% glioblastomas (14/123) and was associated with TP53 mutation (p = 0.0068). It was an early event occurring after the recurrent SCNA observed in diffuse high-grade gliomas. Glioblastomas with WGD were more aneuploid compared to glioblastomas without WGD (p < 0.0001). Chromothripsis occurred in 29.3% glioblastomas (36/123) and mostly affected chromosomes 7, 9 and 12, with amplification of oncogenes (EGFR, MDM2/CDK4), and homozygous deletion of tumor suppressor genes (CDKN2A). There was a significant association between chromothripsis and gene rearrangement at a given locus. WGD is an early genetic event significantly associated to TP53 mutation and leading to chromosome instability and aneuploidy in IDH-wild-type glioblastoma. Chromothripsis recurrently targets oncogenes and tumor suppressor genes that are key players in gliomagenesis and tumor progression. The occurrence of chromothripsis points to underlying gene rearrangements (including gene fusions), potential therapeutic targets in glioblastoma.

Keywords: SNP arrays; aneuploidy; chromothripsis; glioblastoma; whole genome duplication.

Figure 1. Landscape of SCNA in the 123 GBM

Total chr losses are in dark green, partial losses in light green, chr gains in red and copy neutral LOH in light blue. Amplifications and homozygous deletions of key genes in GBM are shown in orange (top rows). WGD is in pink and CT in dark blue (bottom rows). Note that WGD cases present marked aneuploidy with many chr losses. Abbreviations: SCNA, somatic copy number alterations; chr, chromosome; ampl, amplification; HD, homozygous deletion; LOH, loss of heterozygosity; WGD, whole genome duplication; CT, chromothripsis.

Figure 2. WGD in an IDH-wild-type GBM

(A) SNP array profile (GAP method) showing a near tetraploid genome characterized by marked allelic imbalance. Chr 2 displayed a LogR ratio below 0 (2 DNA copies), with a heterozygous allelotype; chr 17 had a LogR ratio above 0 (4 DNA copies), with a heterozygous allelotype; chrs 16 and 19 displayed a LogR ratio near 0 (3 DNA copies). (B) FISH picture showing the presence of two copies of chr 2 (red signals). (C) FISH picture showing the presence of four copies of chr 17 (red signals) and three copies of chr 16 (green signals). (D) FISH picture showing the presence of three copies of chr 19q13 (green signals). Abbreviations: BAF, B-allele frequency; LRR, LogR Ratio; CEN, centromere.

Figure 3. Chronology of genetic events in WGD GBM

SNP array profile (GAP method) of a near-tetraploid GBM showing two types of chr losses: chr losses with a homozygous allelotype (2 DNA copies instead of 4; e.g., chrs 10, 13q, 14q) (solid boxes) and chr losses with a heterozygous allelotype (three DNA copies; e.g., chrs 2 and 4) (dashed boxes). According to these observations, chrs 10, 13q and 14q were lost before WGD occurrence, and chrs 2 and 4 were lost after genome doubling. Abbreviations: BAF, B-allele frequency; LRR, LogR Ratio; CEN, centromere.

Figure 4. Occurrence of CT

(A) SNP array profile (GAP method) showing a CT pattern on chr 12q with multiple amplifications involving, among others, MDM2 and CDK4 genes (dotted and solid arrows, respectively). Note interspersed loss and retention of heterozygosity with no more than three different copy number states except for focal amplifications. (B) SNP array profile (GAP method) showing a CT pattern on chr 17p with TP53 homozygous deletion (arrow). (C) Number of chrs affected by CT per GBM sample. In most cases, CT involved only one or two chrs. (D) CT occurrence across the whole genome in the 123 GBM. High-level event indicates gene amplification (³ 4 copies) or homozygous deletion (no copy); oscillating pattern indicates alternating copy number states (1, 2 or 3) without amplification or homozygous deletion of genomic regions. High-level events were more frequent than oscillating patterns. CT mostly involved chrs 7, 9, and 12, which harbor key oncogenes or tumor supressor genes in GBM. Abbreviations: CT, chromothripsis.

Figure 5. EGFR amplification and CDKN2A deletion with or without CT

SNP array profiles (GAP method) showing EGFR amplification (arrow) and chr 7 gain with (A) or without (B) CT. CDKN2A homozygous deletion (arrow) and chr 9p loss with (C) or without (D) CT. Note interspersed loss and retention of heterozygosity with no more than three different copy number states except for focal amplification or homozygous deletion.