Cancer chromosomal instability (CIN) results in an elevated rate of change of chromosome number and structure and generates intratumour heterogeneity1,2. CIN is observed in the majority of solid tumours and is associated with both poor prognosis and drug resistance3,4. Therefore, understanding a mechanistic basis for CIN is paramount. Here we find evidence for impaired replication fork progression and elevated DNA replication stress in CIN+ colorectal cancer (CRC) cells relative to CIN− CRC cells, with structural chromosome abnormalities precipitating chromosome missegregation in mitosis. We identify three novel CIN-suppressor genes (PIGN (MCD4), RKHD2 (MEX3C) and ZNF516 (KIAA0222)) encoded on chromosome 18q, which is subject to frequent copy number loss in CIN+ CRC. 18q loss was temporally associated with aneuploidy onset at the adenoma-carcinoma transition. CIN-suppressor gene silencing leads to DNA replication stress, structural chromosome abnormalities and chromosome missegregation. Supplementing cells with nucleosides, to alleviate replication-associated damage5, reduces the frequency of chromosome segregation errors following CIN-suppressor gene silencing and attenuates segregation errors and DNA damage in CIN+ cells. These data implicate a central role for replication stress in the generation of structural and numerical CIN, which may inform new therapeutic approaches to limit intratumour heterogeneity.

The contribution of whole-genome doubling to chromosomal instability (CIN) and tumor evolution is unclear. We use long-term culture of isogenic tetraploid cells from a stable diploid colon cancer progenitor to investigate how a genome-doubling event affects genome stability over time. Rare cells that survive genome doubling demonstrate increased tolerance to chromosome aberrations. Tetraploid cells do not exhibit increased frequencies of structural or numerical CIN per chromosome. However, the tolerant phenotype in tetraploid cells, coupled with a doubling of chromosome aberrations per cell, allows chromosome abnormalities to evolve specifi cally in tetraploids, recapitulating chromosomal changes in genomically complex colorectal tumors. Finally, a genome-doubling event is independently predictive of poor relapse-free survival in early-stage disease in two independent cohorts in multivariate analyses [discovery data: hazard ratio (HR), 4.70, 95% confi dence interval (CI), 1.04-21.37; validation data: HR, 1.59, 95% CI, 1.05-2.42]. These data highlight an important role for the tolerance of genome doubling in driving cancer genome evolution. SIGNIFICANCE:Our work sheds light on the importance of whole-genome-doubling events in colorectal cancer evolution. We show that tetraploid cells undergo rapid genomic changes and recapitulate the genetic alterations seen in chromosomally unstable tumors. Furthermore, we demonstrate that a genome-doubling event is prognostic of poor relapse-free survival in this disease type. Cancer Discov; 4(2);