Ceralasertib

Manufactured by Selleck Chemicals

Sourced in United States

Ceralasertib is a laboratory equipment product manufactured by Selleck Chemicals. It is a small molecule that functions as a selective inhibitor of the Ataxia Telangiectasia and Rad3-related (ATR) kinase.

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Lab products found in correlation

7 protocols using ceralasertib

Cell culture and ATR inhibitor treatment

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Human H460 and A549 NSCLC and U2OS osteosarcoma cells were grown in DMEM with GlutaMAX-I, and SW900 and H1975 NSCLC cells in RPMI 1640 medium with GlutaMAX-I (both media from Gibco by Life Technologies), at 37°C with humidified 5% CO₂ atmosphere. The media were supplemented with 10% fetal bovine serum (Biowest) and 1% penicillin–streptomycin solution (50 IU/ml) (Gibco). Cells were tested for Mycoplasma infection, and their identity was confirmed by short tandem repeat analysis. ATR inhibitors VE822 (berzosertib/VX970, Selleckchem) and AZD6738 (ceralasertib, Selleckchem) were added 10–30 minutes before irradiation (160 kV X-rays, 1 Gy/min, Faxitron CP-160).

Eek Mariampillai A., Hauge S., Øynebråten I., Rødland G.E., Corthay A, & Syljuåsen R.G. (2022). Caspase activation counteracts interferon signaling after G2 checkpoint abrogation by ATR inhibition in irradiated human cancer cells. Frontiers in Oncology, 12, 981332.

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Studying Cell Line Responses to ATR and Caspase Inhibitors

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Human U2OS osteosarcoma and H460 NSCLC cells were grown in Dulbecco’s modified Eagle’s medium with GlutaMAX-I (Gibco by Life Technologies, ThermoFisher Scientific #61965059), and human H1975 and SW900 NSCLC cells were grown in Roswell Park Memorial Institute 1640 medium with GlutaMAX-I (Gibco by Life Technologies, ThermoFisher Scientific #61870044) in a humidified 5% CO₂ atmosphere at 37°C. Both media were supplemented with 10% foetal bovine serum (FBS, Biowest #S1810) and 1% penicillin–streptomycin solution (10,000 IU/ml; 10,000 µg/ml) (Pen Strep, Gibco by Life Technologies, ThermoFisher Scientific #15140122). Cell line identity was confirmed by short tandem repeat analysis, and the cultures were tested for Mycoplasma infection. The cells were treated with ATR inhibitors VE822 (berzosertib/VX970, Selleckchem #S7102) at 250 nM or 50 nM and AZD6738 (ceralasertib, Selleckchem #S7693) at 1250 nM, and the pan-caspase inhibitor Q-VD-OPh (quinoline-Val-Asp-difluorophenoxymethylketone, MedChemExpress #HY12305) at 10 µM, for 10-30 minutes before X-irradiation (160 kV Faxitron Corporation CP-160, dose rate 1 Gy/min). Caspase-1 inhibitors Ac-YVAD-cmk (acetyl-Tyr-Val-Ala-Asp-chloromethylketone) and VX-765 (belnacasan) (both from InvivoGen, #inh-yvad and #inh-vx765i-1, respectively) were employed at 60 and 120 µM.

Eek Mariampillai A., Hauge S., Kongsrud K, & Syljuåsen R.G. (2023). Immunogenic cell death after combined treatment with radiation and ATR inhibitors is dually regulated by apoptotic caspases. Frontiers in Immunology, 14, 1138920.

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Glioblastoma Cell Lines and Primary Culture

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Glioblastoma cell lines U373, LN229, and T98G were available from the American Tissue Type Culture Collection (USA). KUGBM8 primary cell line was established by Dr. Filiz Şenbabaoğlu from patient samples in collaboration with Koç University Hospital Neurosurgery Department; ethical approval for KUGBM8 cell line was obtained from the Koç University Institutional Review Board (2014.079.IRB2.022) [48 (link)]. Protocol of primary cell line generation was adapted from [69 (link)]. All parental and irradiated cell populations cells were cultured in DMEM (Gibco, Gaithersburg, MD, USA) supplemented with 10% FBS (Gibco, Gaithersburg, MD, USA) and 1% Penicillin-Streptomycin (Gibco, Gaithersburg, MD, USA). All cells were maintained at 37 °C in a humidified incubator with 5% CO₂. To achieve hypoxic conditions, cells were maintained at 37 °C in a humidified incubator with 5% CO₂ and 1% O₂. AZD7762 (Selleckchem, S1532), AZD6738 (Ceralasertib, Selleckchem, S7693), LY2603618 (Rabusertib, Selleckchem, S2626), KU55933 (Selleckchem, S1092), BML-277 (Selleckchem, S8632), Bleomycin (Selleckchem, S1214), and Temozolomide (Selleckchem, S1237) were used for drug treatment experiments.

Pinarbasi-Degirmenci N., Sur-Erdem I., Akcay V., Bolukbasi Y., Selek U., Solaroglu I, & Bagci-Onder T. (2022). Chronically Radiation-Exposed Survivor Glioblastoma Cells Display Poor Response to Chk1 Inhibition under Hypoxia. International Journal of Molecular Sciences, 23(13), 7051.

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Ceralasertib ATR Inhibitor Protocol

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Ceralasertib (ADZ6738)-Ataxia-telangiectasia-mutated (ATR) inhibitor (Selleck Chemicals, Houston, TX, Cat#S7693) was dissolved at 5 mM stock.

Grosshans D., Thomas R., Zhang D., Cronkite C., Thomas R., Singh S., Bronk L., Morales R, & Duman J. (2024). Subcellular functions of tau mediates repair response and synaptic homeostasis in injury. Research Square.

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ATR Inhibitor-Induced Micronuclei in DZ-Expressing Cell Lines

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HT and SUDHL-5 cell lines were selected based on the high expression of the DZ spatial signature according to the 23Q2 DepMap gene expression dataset (https://depmap.org/portal/download/all/). HT and SUDHL-5 cells were cultured in RPMI media supplemented with 1% glutamine, 10% fetal bovine serum (FBS) and penicillin-streptomycin. Suspension cultures were maintained in flasks in 5% CO₂, at 37°C. The cells were treated for 48 hours with increasing dosages of the clinical-grade ATR inhibitor Ceralasertib (AZD6738 S7693 Selleckchem, 1 or 2 μM) and DMSO was added at a similar concentration in the untreated control. Additional informations are provided in Supplementary Methods section. Lamin B1 staining for analysis of micronuclei formation, RNA extraction, qPCR, and RNAseq on ATRi and control (DMSO-treated) cells are detailed in the Supplementary Methods.

Cancila V., Morello G., Bertolazzi G., Chan A.S., Bastianello G., Paysan D., Jaynes P.W., Schiavoni G., Mattei F., Piconese S., Revuelta M.V., Noto F., De Ninno A., Cammarata I., Pagni F., Venkatachalapathy S., Sangaletti S., Di Napoli A., Vacca D., Lonardi S., Lorenzi L., Ferreri A.J., Belmonte B., Varano G., Colombo M.P., Bicciato S., Inghirami G., Cerchietti L., Ponzoni M., Zappasodi R., Facchetti F., Foiani M., Casola S., Jeyasekharan A.D, & Tripodo C. (2024). Germinal Center Dark Zone harbors ATR-dependent determinants of T-cell exclusion that are also identified in aggressive lymphoma. Research Square.

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Glioblastoma Stem Cell Line Establishment

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The GSC lines were established by isolating neurosphere-forming cells from fresh surgical specimens of human GBM tissue obtained from patients at The University of Texas MD Anderson Cancer Center from 2005 to 2008, as described previously.^{21 (link)} Eight GSC lines [4 with MGMT-unmethylated/MGMT expression (MGMT+)] and [4 with MGMT-methylated/no MGMT expression (MGMT-)] were cultured in DMEM/F12 medium containing B27 supplement (Invitrogen), basic fibroblast growth factor, and epidermal growth factor. Short tandem repeats using the Applied Biosystems AmpFISTR Identifier kit were used to authenticate cells. The last authentication test was performed in July 2017. All cell lines tested negative for mycoplasma contamination using the MycoAlert Detection Kit.
TMZ was from Sigma–Aldrich and lomeguatrib, elimusertib, ceralasertib, and berzosertib were from Selleckchem. For in vitro use, all inhibitors were dissolved in dimethyl sulfoxide. For irradiation experiments, we performed photon irradiation using an X-RAD 320-Precision X-Ray Biological Radiator to deliver a precise dosage of 2 Gy to 12 Gy to the cultured cells. After irradiation, cells were transferred to a cell culture–grade incubator for downstream applications. Sham-treated cultures were kept in close proximity to the X-RAD device for the same amount of time without exposure to X-rays.

Leong V.W., Khan S., Sharma P., Wu S., Thomas R.R., Li X., Singh S.K., Lang F.F., Yung A.W, & Koul D. (2023). MGMT function determines the differential response of ATR inhibitors with DNA-damaging agents in glioma stem cells for GBM therapy. Neuro-Oncology Advances, 6(1), vdad165.

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Inhibition of DNA-PK and ATR

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The DNA-PK inhibitor AZD7648 and the ATR inhibitor Ceralasertib (AZD6738) were purchased from Selleck Chemicals (Houston, TX, USA, product codes: S8843 and S7693).

Chughtai A.A., Pannhausen J., Dinger P., Wirtz J., Knüchel R., Gaisa N.T., Eble M.J, & Rose M. (2022). Effective Radiosensitization of Bladder Cancer Cells by Pharmacological Inhibition of DNA-PK and ATR. Biomedicines, 10(6), 1277.

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