Mismatch repair (MMR)-deficient cancers are characterized by microsatellite instability (MSI) and hypermutation. However, it remains unclear how MSI and hypermutation arise and contribute to cancer development. Here, we show that MSI and hypermutation are triggered by replication stress in an MMR-deficient background, enabling clonal expansion of cells harboring ARF/p53-module mutations and cells that are resistant to the anti-cancer drug camptothecin. While replication stress-associated DNA double-strand breaks (DSBs) caused chromosomal instability (CIN) in an MMR-proficient background, they induced MSI with concomitant suppression of CIN via a PARP-mediated repair pathway in an MMR-deficient background. This was associated with the induction of mutations, including cancer-driver mutations in the ARF/p53 module, via chromosomal deletions and base substitutions. Immortalization of MMR-deficient mouse embryonic fibroblasts (MEFs) in association with ARF/p53-module mutations was ~60-fold more efficient than that of wild-type MEFs. Thus, replication stress-triggered MSI and hypermutation efficiently lead to clonal expansion of cells with abrogated defense systems.

Genomic destabilisation is associated with the induction of mutations, including those in cancer-driver genes, and subsequent clonal evolution of cells with abrogated defence systems. Such mutations are not induced when genome stability is maintained; however, the mechanisms involved in genome stability maintenance remain elusive. Here, resveratrol (and related polyphenols) is shown to enhance genome stability in mouse embryonic fibroblasts, ultimately protecting the cells against the induction of mutations in the ARF/p53 pathway. Replication stress-associated DNA double-strand breaks (DSBs) that accumulated with genomic destabilisation were effectively reduced by resveratrol treatment. In addition, resveratrol transiently stabilised the expression of histone H2AX, which is involved in DSB repair. Similar effects on the maintenance of genome stability were observed for related polyphenols. Accordingly, we propose that polyphenol consumption can contribute to the suppression of cancers that develop with genomic instability, as well as lifespan extension. Most cancers are associated with genomic instability, which can be categorised as chromosomal instability or microsatellite instability (MSI) 1. Genomic destabilisation is a major cause of mutations, including those in cancer-driver genes, and can lead to clonal evolution of cells with abrogated defence systems, such as those containing mutations in the ARF/p53 pathway 2. Genome stability maintenance would likely prevent the formation of mutations and suppress cancer development; however, it is still unclear if genome stability can be maintained in vivo and whether this process can indeed suppress the occurrence of cancer. Genomic instability is caused by the erroneous repair of DNA double-strand breaks (DSBs); paradoxically, the DNA repair systems of most cancers that develop with genomic instability are genetically normal 3. The mechanisms by which normal cells accumulate DSBs remain unclear, but DSBs widely accumulate in pre-cancerous cells and are accompanied by genomic instability 2,4-6. In vitro, replication stress-associated DSBs and the associated genomic instability are observed in cells subjected to aberrant growth stimulation 2 or overexpression of oncogenes such as c-Myc and E2F1 6-8. Possibly reflecting the correlation between cancer development and age, DSBs accumulate with age in vivo and with cultivating passages in vitro 9 , suggesting that ageing cells are defective in DSB repair. DSB repair deficiency is probably related at least in part to a reduction in the level of H2AX. This histone mediates DSB repair and is required for genome stability maintenance, and H2AX expression levels are attenuated when the growth rate of normal cells slows down 10,11. In fact, such cells are defective in repairing replication stress-associated DSBs 10 ,

The development of cancer is driven by genomic instability and mutations. In general, cancer develops via multiple steps. Each step involves the clonal evolution of cells with abrogated defense systems, such as cells with mutations in cancer-suppressor genes. However, it remains unclear how cellular defense systems are abrogated and the associated clonal evolution is triggered and propagated. In this manuscript, we review current knowledge regarding mutagenesis associated with genomic destabilization and its relationship with the clonal evolution of cells over the course of cancer development, focusing especially on mechanistic aspects.

Most cancers develop with one of two types of genomic instability, namely, chromosomal instability (CIN) or microsatellite instability (MSI). Both are induced by replication stress-associated DNA double-strand breaks (DSBs). The type of genomic instability that arises is dependent on the choice of DNA repair pathway. Specifically, MSI is induced via a PolQ-dependent repair pathway called microhomology-mediated end joining (MMEJ) in a mismatch repair (MMR)-deficient background. However, it is unclear how the MMR status determines the choice of DSB repair pathway. Here, we show that replication stress-associated DSBs initially targeted by the homologous recombination (HR) system were subsequently hijacked by PolQ-dependent MMEJ in MMR-deficient cells, but persisted as HR intermediates in MMR-proficient cells. PolQ interacting with MMR factors was effectively loaded onto damaged chromatin in an MMR-deficient background, in which merged MRE11/γH2AX foci also effectively formed. Thus, the choice of DNA repair pathway according to the MMR status determines whether CIN or MSI is induced.