A cancer genome carries the historic mutagenic activity that has occurred throughout the development of a tumour. While driver mutations were the main focus of cancer research for a long time, passenger mutational signatures – the imprints of DNA damage and DNA repair processes that have been operative during tumourigenesis – are also biologically informative. Since the inception of the field, many mutational signatures have been uncovered from the analysis of primary tumours. However, the causes and the mechanisms underlying many of these signatures are not fully understood nor have they been established with experimental evidence. Several pioneering experimental studies have shown that mutational signatures are not just mathematical abstraction; they can be recreated in vitro using isogenic experimental model systems.

In this dissertation, I build on these pioneering efforts to demonstrate the concept of mutational signatures by engineering gene knockouts in various isogenic human cell line models (i.e. HAP-1 cells and human-induced pluripotent stem cells) and using whole-genome sequencing as readouts to study their associated mutational patterns. In doing so, I have successfully validated some of the most widely-applied signatures, including those that are associated with deficiencies in homologous recombination-based repair and mismatch repair.

During this process, I have also identified key issues in designing and performing such mutational signature studies, and hence devised a set of guidelines for future experiments and analyses. The work described in this dissertation paves the way for future investigations aiming to understand the relationships between DNA repair and the causes of somatic mutations in human cancers. The experimental evidence established for some of the mutational signatures here may hopefully support their clinical applications henceforth.