ku-55933 and Osteosarcoma

Programmed death ligand 1 (PD-L1) expression on the surface of cancer cells affects the efficacy of anti-PD-1/PD-L1 immune checkpoint therapy. However, the mechanism underlying PD-L1 expression in cancer cells is not fully understood, particularly after ionizing radiation (IR). Here, we examined the impact of high linear energy transfer (LET) carbon-ion irradiation on the expression of PD-L1 in human osteosarcoma U2OS cells. We found that the upregulation of PD-L1 expression after high LET carbon-ion irradiation was greater than that induced by X-rays at the same physical and relative biological effectiveness (RBE) dose, and that the upregulation of PD-L1 induced by high LET carbon-ion irradiation was predominantly dependent on ataxia telangiectasia and Rad3-related (ATR) kinase activity. Moreover, we showed that the downstream signaling, e.g. STAT1 phosphorylation and IRF1 expression, was upregulated to a greater extent after high LET carbon-ion irradiation than X-rays, and that IRF1 upregulation was also ATR dependent. Finally, to visualize PD-L1 molecules on the cell surface in 3D, we applied immunofluorescence-based super-resolution imaging. The three-dimensional structured illumination microscopy (3D-SIM) analyses revealed substantial increases in the number of presented PD-L1 molecules on the cell surface after high LET carbon-ion irradiation compared with X-ray irradiation.

Topics: Ataxia Telangiectasia Mutated Proteins; B7-H1 Antigen; Bone Neoplasms; Cell Line, Tumor; Gene Expression Regulation, Neoplastic; Heavy Ion Radiotherapy; Humans; Imaging, Three-Dimensional; Interferon Regulatory Factor-1; Linear Energy Transfer; Morpholines; Neoplasm Proteins; Osteosarcoma; Phosphorylation; Protein Processing, Post-Translational; Pyrazines; Pyrones; RNA, Messenger; RNA, Neoplasm; STAT1 Transcription Factor; Sulfones; Up-Regulation; X-Rays

Mutations in the Werner syndrome protein (WRN), a caretaker of the genome, result in Werner syndrome, which is characterized by premature aging phenotypes and cancer predisposition. Methylseleninic acid (MSeA) can activate DNA damage responses and is a superior compound to suppress tumorigenesis in mouse models of cancer. To test the hypothesis that targeting WRN can potentiate selenium toxicity in cancer cells, isogenic WRN small hairpin RNA (shRNA) and control shRNA U-2 OS osteosarcoma cells were treated with MSeA for 2d, followed by recovery for up to 7d. WRN deficiency sensitized U-2 OS cells to MSeA-induced necrotic death. Co-treatment with the ataxia-telangiectasia mutated (ATM) kinase inhibitor KU55933 desensitized the control shRNA cells, but not WRN shRNA cells, to MSeA treatment. WRN did not affect MSeA-induced ATM phosphorylation on Ser-1981 or H2A.X phosphorylation on Ser-139, but promoted recovery from the MSeA-induced DNA damage. Taken together, WRN protects U-2 OS osteosarcoma cells against MSeA-induced cytotoxicity, suggesting that oxidative DNA repair pathway is a promising target for improving the efficacy of selenium on tumor suppression.

Topics: Animals; Ataxia Telangiectasia Mutated Proteins; Cell Cycle Proteins; Cell Line, Tumor; DNA Damage; DNA-Binding Proteins; Drug Resistance, Neoplasm; Exodeoxyribonucleases; Gene Knockdown Techniques; Histones; Humans; Mice; Morpholines; Necrosis; Organoselenium Compounds; Osteosarcoma; Protein Serine-Threonine Kinases; Pyrones; RecQ Helicases; Tumor Suppressor Proteins; Werner Syndrome Helicase