Journal: Signal Transduction and Targeted Therapy

Article Title: A ribosomal gene panel predicting a novel synthetic lethality in non-BRCAness tumors

doi: 10.1038/s41392-023-01401-y

Figure Lengend Snippet: ATM signaling balances the cell fate by simultaneously controlling both HR repair and ribosomal stress during PARP inhibitor/cisplatin treatment. a OV56 cells were pretreated with KU55933 (ATM inhibitor), VX970 (ATR inhibitor), or AZD7648 (DNA-PK inhibitor) for 1 h (scale bar: 5 μm). The localization of NPM1 was detected by immunofluorescence to indicate the ribosomal stress induced by cisplatin or olaparib. b , c OV56 cells were pretreated with indicated concentrations of KU55933 for 1 h (scale bar: 5 μm). The localization of NPM1 was detected to indicate the ribosomal stress level caused by cisplatin or olaparib. d , e OV90 cells ( d ) and OV56 cells ( e ) were pretreated with indicated concentrations of KU55933 for 1 h followed by ionizing radiation (2 Gy) to trigger DNA double-strand breaks. RAD51 foci were stained and quantified to determine the HR status. f , g OV90 cells ( f ) and OV56 cells ( g ) were pretreated with indicated concentrations of KU55933, colony-formation assay was performed to determine the drug sensitivity to cisplatin or olaparib. h Proposed model of cell fate in response to PARP inhibitor/cisplatin treatment. Red labels: cell lines with low expression of the gene panel; Blue labels: cell lines with high expression of the gene panel. * P < 0.05, ** P < 0.01, (com p ared with CTRL)

Article Snippet: Indicated cells were cultured on coverslips and treated with 50 µM cisplatin (Sigma, P4394), 100 µM olaparib (LC Laboratories, O-9201), 10 µM DNA-PK inhibitor AZD7648 (ChemScene, CS-0091859), 10 µM ATM inhibitor KU55933 (Abcam, ab120637) or 10 µM ATR inhibitor VX970 (Selleckchem, S7102) for 6 h. After washing with PBS, cells were fixed in 3% paraformaldehyde for 15 min and permeabilized in 0.5% triton X-100 solution for 5 min at room temperature.

Techniques: Immunofluorescence, Staining, Colony Assay, Expressing

Journal: EMBO Molecular Medicine

Article Title: The evolving genetic landscape of telomere biology disorder dyskeratosis congenita

doi: 10.1038/s44321-024-00118-x

Figure Lengend Snippet: Reagents and tools table

Article Snippet: DNA-PK inhibitor NU7026 , STEMCELL Technologies , 74172.

Techniques: Recombinant, FLAG-tag, Sequencing, In Situ, Software

Journal: Biochemical and biophysical research communications

Article Title: Inhibiting DNA-PK CS Radiosensitizes Human Osteosarcoma Cells

doi: 10.1016/j.bbrc.2017.03.033

Figure Lengend Snippet: A) RNASeq analysis of DNA-PKCS mRNA in primary bone tumor specimens and cell lines. Specimens included four chondrosarcomas, eight chondroblastomas, five chordoma, five Ewing’s sarcoma (one tissue and four cell lines) and four OS (one tissue and three cell lines). B) Total DNA-PKCS protein levels in OS cell lines compared with HOB cells. C) Levels of DNA-PKCS autophosphorylation at Ser2056 induced with IR (10Gy) and with graded concentration of KU60648, in 143B cells. Results are representative of three independent experiments.

Article Snippet: The DNA-PK CS inhibitor, KU60648, was kindly provided by KuDOS Pharmaceuticals and was dissolved in DMSO as a 1 mM stock solution.

Techniques: Concentration Assay

Journal: Biochemical and biophysical research communications

Article Title: Inhibiting DNA-PK CS Radiosensitizes Human Osteosarcoma Cells

doi: 10.1016/j.bbrc.2017.03.033

Figure Lengend Snippet: Clonogenic survival for 143B cells (A) and U2OS cells (B) treated with vehicle control or 300 nM KU60648 for one hour pre-IR and 24 hours post-IR, rinsed with PBS, media changed and incubated for colony formation. Results are the mean ± SD of three independent experiments fitted to the LQ model.

Article Snippet: The DNA-PK CS inhibitor, KU60648, was kindly provided by KuDOS Pharmaceuticals and was dissolved in DMSO as a 1 mM stock solution.

Techniques: Control, Incubation

Journal: Biochemical and biophysical research communications

Article Title: Inhibiting DNA-PK CS Radiosensitizes Human Osteosarcoma Cells

doi: 10.1016/j.bbrc.2017.03.033

Figure Lengend Snippet: A–C) FACS histograms for U2OS cells treated with vehicle control (A), 5 Gy (B), and 5 Gy plus 100 nM KU60648 (C). Results are representative of three independent experiments. D–F) The summary of the cell cycle analyses for 143B cells (D), U2OS cells (E) and HOB cells (F). Results are mean ± SD of three or more independent experiments. (* P < 0.05)

Article Snippet: The DNA-PK CS inhibitor, KU60648, was kindly provided by KuDOS Pharmaceuticals and was dissolved in DMSO as a 1 mM stock solution.

Techniques: Control

Journal: Biochemical and biophysical research communications

Article Title: Inhibiting DNA-PK CS Radiosensitizes Human Osteosarcoma Cells

doi: 10.1016/j.bbrc.2017.03.033

Figure Lengend Snippet: Representative confocal microscopy images of γH2AX foci 24 hours post-IR in cells treated with vehicle control, 5 Gy or 5 Gy plus 100 nM KU60648 (added 1 hour before IR) in 143B cells (A) and U2OS cells (B). Quantification of percentage of cells with > 20 γH2AX foci in cells treated with vehicle control, 100 nM KU60648 alone, 5 Gy or 5 Gy plus 100 nM KU60648 in 143B cells (C) and U2OS cells (D). Results are mean ± SD from 2 independent experiments. For each treatment group, >150 and >90 nuclei were counted for 143B and U2OS cells, respectively.

Article Snippet: The DNA-PK CS inhibitor, KU60648, was kindly provided by KuDOS Pharmaceuticals and was dissolved in DMSO as a 1 mM stock solution.

Techniques: Confocal Microscopy, Control

Journal: Scientific Reports

Article Title: HERC2 regulates RPA2 by mediating ATR-induced Ser33 phosphorylation and ubiquitin-dependent degradation

doi: 10.1038/s41598-019-50812-x

Figure Lengend Snippet: Effect of suppression of HERC2 E3 ligase activity on RPA2 Ser33 phosphorylation and ubiquitination. ( a ) Wild type and HERC2 ΔE3/ΔE3 HCT116 cells were treated with 0.2 mM HU for the indicated time and subjected to immunoblotting with the indicated antibodies. HERC2 (N) : the antibody to an epitope 1781–1974 of HERC2. ( b ) HCT116-HERC2 ΔE3/ΔE3 cells were incubated with or without 10 μM ATR inhibitor VE821, 10 μM ATM inhibitor Ku55933, or 10 μM DNA-PK inhibitor Nu7026 for 2 h as indicated and subjected to immunoblotting with the indicated antibodies. ( c ) Wild type and HERC2 ΔE3/ΔE3 HCT116 cells were treated or not with MG132 and 5 μM ATR inhibitor for 12 h as indicated and subjected to immunoprecipitation in denature condition with control IgG or anti-RPA2 antibody followed by immunoblotting (left panels) or to direct immunoblotting (right panels).

Article Snippet: Chemical agents used in the present study were hydroxyurea (HU) (Sigma-Aldrich), mitomycin C (MMC) (Sigma-Aldrich), irinotecan hydrochloride (CPT) (Sigma-Aldrich), aphidicolin (APH) (Sigma-Aldrich), MG132 (Calbiochem), the ATR inhibitor VE821 (Toronto Research Chemicals), the ATM inhibitor Ku55933 (TOCRIS Bioscience), and the DNA-PK inhibitor Nu7026 (ChemScene).

Techniques: Activity Assay, Phospho-proteomics, Ubiquitin Proteomics, Western Blot, Incubation, Immunoprecipitation, Control