Selective Elimination of Osteosarcoma Cell Lines with Short Telomeres by ATR Inhibitors

The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acsptsci.0c00125. Origin and characteristics of the osteosarcoma cell lines, telomere length measurement from TRF, IC50 drug response in osteosarcoma cell lines, oligonucleotides used for this study, characterization of telomere status in osteosarcoma cell lines, APB assay, ATRi concentration–survival curves in osteosarcoma lines, relationship between sensitivity to methotrexate and telomere length, comparison of ATRi sensitivities between ALT-negative and positive lines, selective death of osteosarcoma with short telomeres exposed to ATR inhibitor BAY-1895344 (PDF). To avoid replicative senescence or telomere-induced apoptosis, cancers employ telomere maintenance mechanisms (TMMs) involving either the upregulation of telomerase or the acquisition of recombination-based alternative telomere lengthening (ALT). The choice of TMM may differentially influence cancer evolution and be exploitable in targeted therapies. Here, we examine TMMs in a panel of 17 osteosarcoma-derived cell lines, defining three separate groups according to TMM and the length of telomeres maintained. Eight were ALT-positive, including the previously uncharacterized lines, KPD and LM7. While ALT-positive lines all showed excessive telomere length, ALT-negative cell lines fell into two groups according to their telomere length: HOS-MNNG, OHSN, SJSA-1, HAL, 143b, and HOS displayed subnormally short telomere length, while MG-63, MHM, and HuO-3N1 displayed long telomeres. Hence, we further subcategorized ALT-negative TMM into long-telomere (LT) and short-telomere (ST) maintenance groups. Importantly, subnormally short telomeres were significantly associated with hypersensitivity to three different therapeutics targeting the protein kinase ataxia telangiectasia and Rad3-related (ATR) (AZD-6738/Ceralasertib, VE-822/Berzoserib, and BAY-1895344) compared to long telomeres maintained via ALT or telomerase. Within 24 h of ATR inhibition, cells with short but not long telomeres displayed chromosome bridges and underwent cell death, indicating a selective dependency on ATR for chromosome stability. Collectively, our work provides a resource to identify links between the mode of telomere maintenance and drug sensitivity in osteosarcoma and indicates that telomere length predicts ATR inhibitor sensitivity in cancer. KT and LCC were supported by Cancer Research UK (C36439/A12097) and European research council (281722-HRMCB). GZ and CAM were supported by Children with Cancer UK (17-244). CM was supported through a Cancer Research UK studentship (C416/A23233).