Whole-genome and whole-exome sequencing of bladder cancer identifies frequent alterations in genes involved in sister chromatid cohesion and segregation

Abstract

Bladder cancer is one of the most common cancers worldwide, with transitional cell carcinoma (TCC) being the predominant form. Here we report a genomic analysis of TCC by both whole-genome and whole-exome sequencing of 99 individuals with TCC. Beyond confirming recurrent mutations in genes previously identified as being mutated in TCC, we identified additional altered genes and pathways that were implicated in TCC. Notably, we discovered frequent alterations in STAG2 and ESPL1, two genes involved in the sister chromatid cohesion and segregation (SCCS) process. Furthermore, we also detected a recurrent fusion involving FGFR3 and TACC3, another component of SCCS, by transcriptome sequencing of 42 DNA-sequenced tumors. Overall, 32 of the 99 tumors (32%) harbored genetic alterations in the SCCS process. Our analysis provides evidence that genetic alterations affecting the SCCS process may be involved in bladder tumorigenesis and identifies a new therapeutic possibility for bladder cancer.

Figure 1

Significantly mutated genes in TCC as determined by exome sequencing. Significantly mutated genes are listed on the right. The percentage of bladder tumors with mutations detected by automated calling is noted on the left. The upper histogram shows the somatic mutation rate in each of the 99 tumors. The central heatmap shows the distribution of mutations across the sequenced samples. All the mutations shown were confirmed by Sanger sequencing. Genes with asterisks had mutations newly observed in TCC in this study.

Figure 2

STAG2 somatic mutations and copy number changes in TCC. (a) Somatic alterations overlaid on the STAG2 protein with the conserved protein domains highlighted. STAG, STAG domain; SCD, stromalin conservative domain. (b) Five tumors harboring genomic deletions of STAG2 on the X chromosome (ideogram shown on the left). (c) Kaplan-Meier survival analysis of individuals with TCC (n = 99) shows that the survival rate for individuals with somatic STAG2 alterations (n = 16) is significantly lower (log-rank test P < 0.001) than for individuals with wild-type STAG2 (n = 83).

Figure 3

FGFR3-TACC3 fusion was identified in TCC. (a) Genomic fusion of intron 17 of FGFR3 with intron 10 of TACC3 resulting in exon 17 of FGFR3 being spliced 5’ to exon 11 of TACC3 in the fused mRNA. Triangles indicate the genomic positions of the breakpoints. Detailed information on the positions and sequences of primers P1 and P3 is provided in Supplementary Table 14. (b) Outlier high expression of TACC3 in TCCs harboring FGFR3-TACC3 gene fusions. RPKM, reads per kilobase of exon region in a gene per million mapped reads, (c) RNA-seq coverage analysis of FGFR3 (top) and TACC3 (bottom) in the tumor and matched normal bladder tissue from B59-3. Three transcripts of FGFR3 and one transcript of TACC3 are shown. Black dotted lines indicate breakpoints. E, exon.

Figure 4

Frequent genetic alterations in genes from the cell cycle pathway in TCC. Alterations are defined as somatic mutations, focal amplifications and deletions, and, in some cases, as gene fusion events. Alteration frequencies are expressed as a percentage of all cases.