ATR-CHEK1 signalling is critical for genomic stability. ATR-CHEK1 signalling may be deregulated in breast cancer and have prognostic, predictive and therapeutic significance. We investigated ATR, CHEK1 and phosphorylated CHEK1 (Ser345) protein (pCHEK1) levels in 1712 breast cancers. ATR and CHEK1 mRNA expression was evaluated in 1950 breast cancers. Pre-clinically, biological consequences of ATR gene knock down or ATR inhibition by the small molecule inhibitor (VE-821) were investigated in MCF7 and MDA-MB-231 breast cancer cell lines and in non-tumorigenic breast epithelial cells (MCF10A). High ATR and high cytoplasmic pCHEK1 levels were significantly associated with higher tumour stage, higher mitotic index, pleomorphism and lymphovascular invasion. In univariate analyses, high ATR and high cytoplasmic pCHEK1 levels were associated with poor breast cancer specific survival (BCSS). In multivariate analysis, high ATR level remains an independent predictor of adverse outcome. At the mRNA level, high CHEK1 remains associated with aggressive phenotypes including lymph node positivity, high grade, Her-2 overexpression, triple negative, aggressive molecular phenotypes and adverse BCSS. Pre-clinically, CHEK1 phosphorylation at serine(345) following replication stress was impaired in ATR knock down and in VE-821 treated breast cancer cells. Doxycycline inducible knockdown of ATR suppressed growth, which was restored when ATR was re-expressed. Similarly, VE-821 treatment resulted in a dose dependent suppression of cancer cell growth and survival (MCF7 and MDA-MB-231) but was less toxic in non-tumorigenic breast epithelial cells (MCF10A). We provide evidence that ATR and CHEK1 are promising biomarkers and rational drug targets for personalized therapy in breast cancer.

Photodynamic therapy that uses photosensitizers which only become toxic upon light-irradiation provides a strong alternative to conventional cancer treatment due to its ability to selectively target tumour material without affecting healthy tissue. Transition metal complexes are highly promising PDT agents due to intense visible light absorption, yet the majority are toxic even without light. This study introduces a small, photostable, charge-neutral platinum-based compound, Pt(II) 2,6-dipyrido-4-methyl-benzenechloride, complex 1, as a photosensitizer, which works under visible light. Activation of the new photosensitizer at low concentrations (0.1–1 μM) by comparatively low dose of 405 nm light (3.6 J cm−2) causes significant cell death of cervical, colorectal and bladder cancer cell lines, and, importantly, a cisplatin resistant cell line EJ-R. The photo-index of the complex is 8. We demonstrate that complex 1 induces irreversible DNA single strand breaks following irradiation, and that oxygen is essential for the photoinduced action. Neither light, nor compound alone led to cell death. The key advantages of the new drug include a remarkably fast accumulation time (diffusion-controlled, minutes), and photostability. This study demonstrates a highly promising new agent for photodynamic therapy, and attracts attention to photostable metal complexes as viable alternatives to conventional chemotherapeutics, such as cisplatin.

BRCA1, a key factor in homologous recombination (HR) repair may also regulate base excision repair (BER). Targeting BRCA1-BER deficient cells by blockade of ATM and DNA-PKcs could be a promising strategy in breast cancer. We investigated BRCA1, XRCC1 and pol β protein expression in two cohorts (n = 1602 sporadic and n = 50 germ-line BRCA1 mutated) and mRNA expression in two cohorts (n = 1952 and n = 249). Artificial neural network analysis for BRCA1-DNA repair interacting genes was conducted in 249 tumours. Pre-clinically, BRCA1 proficient and deficient cells were DNA repair expression profiled and evaluated for synthetic lethality using ATM and DNA-PKcs inhibitors either alone or in combination with cisplatin. In human tumours, BRCA1 negativity was strongly associated with low XRCC1, and low pol β at mRNA and protein levels (p < 0.0001). In patients with BRCA1 negative tumours, low XRCC1 or low pol β expression was significantly associated with poor survival in univariate and multivariate analysis compared to high XRCC1 or high pol β expressing BRCA1 negative tumours (ps < 0.05). Pre-clinically, BRCA1 negative cancer cells exhibit low mRNA and low protein expression of XRCC1 and pol β. BRCA1-BER deficient cells were sensitive to ATM and DNA-PKcs inhibitor treatment either alone or in combination with cisplatin and synthetic lethality was evidenced by DNA double strand breaks accumulation, cell cycle arrest and apoptosis. We conclude that XRCC1 and pol β expression status in BRCA1 negative tumours may have prognostic significance. BRCA1-BER deficient cells could be targeted by ATM or DNA-PKcs inhibitors for personalized therapy.

Bloom syndrome helicase (BLM) has key roles in homologous recombination repair, telomere maintenance, and DNA replication. Germ-line mutations in the BLM gene causes Bloom syndrome, a rare disorder characterized by premature aging and predisposition to multiple cancers, including breast cancer. The clinicopathologic significance of BLM in sporadic breast cancers is unknown. We investigated BLM mRNA expression in the Molecular Taxonomy of Breast Cancer International Consortium cohort (n ¼ 1,950) and validated in an external dataset of 2,413 tumors. BLM protein level was evaluated in the Nottingham Tenovus series comprising 1,650 breast tumors. BLM mRNA overexpression was significantly associated with high histologic grade, larger tumor size, estrogen receptor-negative (ER À ), progesterone receptor-negative (PR À ), and triple-negative phenotypes (ps < 0.0001). BLM mRNA overexpression was also linked to aggressive molecular phenotypes, including PAM50.Her2 (P < 0.0001), PAM50. Basal (P < 0.0001), and PAM50.LumB (P < 0.0001) and Genufu subtype (ER þ /Her2 À /high proliferation; P < 0.0001). PAM50. LumA tumors and Genufu subtype (ER þ /Her2 À /low proliferation) were more likely to express low levels of BLM mRNA (ps < 0.0001). Integrative molecular clusters (intClust) intClust.1 (P < 0.0001), intClust.5 (P < 0.0001), intClust.9 (P < 0.0001), and intClust.10 (P < 0.0001) were also more likely in tumors with high BLM mRNA expression. BLM mRNA overexpression was associated with poor breast cancer-specific survival (BCSS; ps < 0.000001). At the protein level, altered subcellular localization with high cytoplasmic BLM and low nuclear BLM was linked to aggressive phenotypes. In multivariate analysis, BLM mRNA and BLM protein levels independently influenced BCSS. This is the first and the largest study to provide evidence that BLM is a promising biomarker in breast cancer.