Inducing specific chromosome mis-segregation for the study of cancer aneuploidy patterns

Cancer genomes commonly display an abnormal number of chromosomes – aneuploidy – and persistent underlying chromosomal instability (CIN). Although these features are common to nearly all human cancer types, subsets of aneuploidies are represented more frequently in particular tumours than others. Our lab has recently shown that aneuploidies of chromosomes 1 and 2 are likely to arise in RPE1 cells when the mitotic spindle is compromised by the use of a small molecule inhibitor. Following these observations, I investigated the susceptibility of chromosomes to mis-segregate when their alignment at the metaphase plate is compromised by the inhibition of CENP-E, a motor protein crucial for chromosome congression. I observed a bias towards large chromosomes, supporting the hypothesis that chromosome identity can influence its ability in segregate correctly during mitosis, likely resulting in different aneuploidy patterns in daughter cells. In addition to CIN mechanisms generating distinct aneuploidy landscapes, there is likely preferential selection for some genomic alterations that can be tolerated under selective circumstances. Although models of chromosome aneuploidy have been developed to study their role in cancer evolution, the acute cellular responses to specific aneuploidy and how this is tolerated still remain to be addressed. The impact of specific chromosome changes are also mostly unknown. For this purpose, I investigated the possibility of interfering with the mitotic segregation of specific chromosomes in order to obtain desired chromosomal changes, by implementing a dCas9-targeting approach. The devised strategies are designed to either assemble an ectopic kinetochore (by recruiting dCas9-CENPT) on a target chromosome or to interfere with the chromosome position in mitosis (by recruiting an exogenous protein domain, PACT, fused to dCas9). With this approach, I was able to induce specific chromosome mis-segregation and this can be potentially applied to any human chromosome. This thesis work thus provides the technical basis for the creation of the recurrent patterns of aneuploidy seen in human cancer and the study of the downstream cellular response