The current preclinical and clinical findings demonstrate that, in addition to the conventional clinical and pathological indicators that have a prognostic value in radiation oncology, the number of cancer stem cells (CSCs) and their inherent radioresistance are important parameters for local control after radiotherapy. In this review, we discuss the molecular mechanisms of CSC radioresistance attributable to DNA repair mechanisms and the development of CSC-targeted therapies for tumor radiosensitization. We also discuss the current challenges in preclinical and translational CSC research including the high inter- and intratumoral heterogeneity, plasticity of CSCs, and microenvironment-stimulated tumor cell reprogramming.

Cancer stem cells (CSCs) are pluripotent and highly tumorigenic cells that can re-populate a tumor and cause relapses even after initially successful therapy. As with tissue stem cells, CSCs possess enhanced DNA repair mechanisms. An active DNA damage response alleviates the increased oxidative and replicative stress and leads to therapy resistance. On the other hand, mutations in DNA repair genes cause genomic instability, therefore driving tumor evolution and developing highly aggressive CSC phenotypes. However, the role of DNA repair proteins in CSCs extends beyond the level of DNA damage. In recent years, more and more studies have reported the unexpected role of DNA repair proteins in the regulation of transcription, CSC signaling pathways, intracellular levels of reactive oxygen species (ROS), and epithelial–mesenchymal transition (EMT). Moreover, DNA damage signaling plays an essential role in the immune response towards tumor cells. Due to its high importance for the CSC phenotype and treatment resistance, the DNA damage response is a promising target for individualized therapies. Furthermore, understanding the dependence of CSC on DNA repair pathways can be therapeutically exploited to induce synthetic lethality and sensitize CSCs to anti-cancer therapies. This review discusses the different roles of DNA repair proteins in CSC maintenance and their potential as therapeutic targets.

Chromosomal instability not only has a negative effect on survival in triple-negative breast cancer, but also on the well treatable subgroup of luminal A tumors. This suggests a general mechanism independent of subtypes. Increased chromosomal instability (CIN) in triple-negative breast cancer (TNBC) is attributed to a defect in the DNA repair pathway homologous recombination. Homologous recombination (HR) prevents genomic instability by repair and protection of replication. It is unclear whether genetic alterations actually lead to a repair defect or whether superior signaling pathways are of greater importance. Previous studies focused exclusively on the repair function of HR. Here, we show that the regulation of HR by the intra-S-phase damage response at the replication is of overriding importance. A damage response activated by Ataxia telangiectasia and Rad3 related-checkpoint kinase 1 (ATR-CHK1) can prevent replication stress and leads to resistance formation. CHK1 thus has a preferred role over HR in preventing replication stress in TNBC. The signaling cascade ATR-CHK1 can compensate for a double-strand break repair error and lead to resistance of HR-deficient tumors. Established methods for the identification of HR-deficient tumors for Poly(ADP-Ribose)-Polymerase 1 (PARP1) inhibitor therapies should be extended to include analysis of candidates for intra-S phase damage response.

Chromosomal instability (CIN) is an emerging hallmark of cancer and its role in therapeutic responses has been increasingly attracting the attention of the research community. To target the vulnerability of tumors with high CIN, it is important to identify the genes and mechanisms involved in the maintenance of CIN. In our work, we recognize the tumor suppressor gene Phosphatase and Tensin homolog (PTEN) as a potential gene causing CIN in triple-negative breast cancer (TNBC) and show that TNBC with low expression levels of PTEN can be sensitized for the treatment with poly-(ADP-ribose)-polymerase 1 (PARP1) inhibitors, independent of Breast Cancer (BRCA) mutations or a BRCA-like phenotype. In silico analysis of mRNA expression data from 200 TNBC patients revealed low expression of PTEN in tumors with a high CIN70 score. Western blot analysis of TNBC cell lines confirm lower protein expression of PTEN compared to non TNBC cell lines. Further, PTEN-deficient cell lines showed cellular sensitivity towards PARP1 inhibition treatment. DNA fiber assays and examination of chromatin bound protein fractions indicate a protective role of PTEN at stalled replication forks. In this study, we recognize PTEN as a potential CIN-causing gene in TNBC and identify its important role in the replication processes.

Aneuploidy is a consequence of chromosomal instability (CIN) that affects prognosis. Gene expression levels associated with aneuploidy provide insight into the molecular mechanisms underlying CIN. Based on the gene signature whose expression was consistent with functional aneuploidy, the CIN70 score was established. We observed an association of CIN70 score and survival in 519 HNSCC patients in the TCGA dataset; the 15% patients with the lowest CIN70 score showed better survival (p = 0.11), but association was statistically non-significant. This correlated with the expression of 39 proteins of the major repair complexes. A positive association with survival was observed for MSH2, XRCC1, MRE11A, BRCA1, BRCA2, LIG1, DNA2, POLD1, MCM2, RAD54B, claspin, a negative for ERCC1, all related with replication. We hypothesized that expression of these factors leads to protection of replication through efficient repair and determines survival and resistance to therapy. Protein expression differences in HNSCC cell lines did not correlate with cellular sensitivity after treatment. Rather, it was observed that the stability of the DNA replication fork determined resistance, which was dependent on the ATR/CHK1-mediated S-phase signaling cascade. This suggests that it is not the expression of individual DNA repair proteins that causes therapy resistance, but rather a balanced expression and coordinated activation of corresponding signaling cascades.