Adavosertib

Manufactured by MedChemExpress

Sourced in United States

Adavosertib is a small molecule inhibitor that targets the WEE1 kinase. It is commonly used in cancer research to study cell cycle regulation and DNA damage response pathways.

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Lab products found in correlation

Oxaliplatin, by Selleck Chemicals (2 mentions) Ceralasertib, by MedChemExpress (2 mentions) 5 fluorouracil, by Selleck Chemicals (2 mentions) Sn 38, by Selleck Chemicals (2 mentions) Berzosertib, by MedChemExpress (2 mentions) Rabusertib, by MedChemExpress (2 mentions) Azd0156, by MedChemExpress (2 mentions) Nedisertib, by MedChemExpress (2 mentions) Olaparib, by Selleck Chemicals (2 mentions) Mg132, by Merck Group (2 mentions) Thz531, by Apexbio (1 mentions) Cgp57380, by Abcam (1 mentions) Simyc, by Horizon Discovery (1 mentions) Rnaimax, by Thermo Fisher Scientific (1 mentions) Puromycin, by Merck Group (1 mentions) Formaldehyde, by Merck Group (1 mentions) Basic fibroblast growth factor (bfgf), by Thermo Fisher Scientific (1 mentions) Epidermal growth factor (egf), by Thermo Fisher Scientific (1 mentions) Dmem f12 medium, by Welgene (1 mentions) Poly hema, by Merck Group (1 mentions)

8 protocols using adavosertib

Molecular Mechanisms of Cancer Regulation

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ZBC260 was provided by S.W., University of Michigan. dBET6 was provided by J. Bradner, Harvard Medical School. ETC-168 was shared from the Experimental Drug Development Centre, Agency for Science, Technology and Research, Singapore. Other key chemicals used in this study included THZ531 (ApexBio), CGP57380 (Abcam), Adavosertib (Medchem Express), TMZ (ApexBio), and formaldehyde (Calbiochem, Merck).
On-TARGETplus small interfering RNA (siRNA) pools including control (si-NT), si-ETV6, si-MYC, and si-NR2F2 were purchased from Dharmacon. RNAiMAX (Life Technologies) was used to transfect siRNAs. Stable knockdown cells were generated by lentiviral infection, followed by antibiotic (puromycin, Sigma-Aldrich) selection. SHC002 was used as a negative control (sh-Ctrl). All pLKO.1-based short hairpin RNA (shRNA) vectors are listed in table S1N. 293T cells were used for viral production.

Xu L., Chen Y., Huang Y., Sandanaraj E., Yu J.S., Lin R.Y., Dakle P., Ke X.Y., Chong Y.K., Koh L., Mayakonda A., Nacro K., Hill J., Huang M.L., Gery S., Lim S.W., Huang Z., Xu Y., Chen J., Bai L., Wang S., Wakimoto H., Yeo T.T., Ang B.T., Müschen M., Tang C., Tan T.Z, & Koeffler H.P. (2021). Topography of transcriptionally active chromatin in glioblastoma. Science Advances, 7(18), eabd4676.

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Culturing Primary Tumor Cells for Drug Screening

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Primary cultures of patient tumors were established as previously reported [13 (link)]. Tumor sample collection was approved by the local ethics committee (Kantonale Ethikkommission Zürich, BASEC 2017-00771) and written informed consent was obtained from all subjects prior to tumor sampling. Cells from benign pheochromocytoma and benign adrenal hyperplasia were plated at 250,000 cells /well in the spherical plate 5D (Kugelmeiers, Erlenbach, Switzerland). NCI-H295R, MUC-1 and ACC115m primary cells were cultured as previously described [13 (link), 18 (link)] and cultivated in Sphericalplates 5D for either 7 or 14 days at the densities of 75,000 cells/well, 150,000 cells/well and 75,000 cells/well respectively. The 7 day spheroids were treated with 650 nM Adavosertib (MedChemExpress) for 72 h. Additionally, ACC115m cells were cultivated in Sphericalplates 5D for 7 days and subsequently treated with 25 µM Gemcitabine and 40 µM Cisplatin (both Teva Pharma) for 72 h.

Bornstein S., Shapiro I., Malyukov M., Züllig R., Luca E., Gelfgat E., Beuschlein F., Nölting S., Berruti A., Sigala S., Peitzsch M., Steenblock C., Ludwig B., Kugelmeier P, & Hantel C. (2022). Innovative multidimensional models in a high-throughput-format for different cell types of endocrine origin. Cell Death & Disease, 13(7), 648.

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Adavosertib Induces SKOV3 Spheroid Formation

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SKOV3 cells were seeded at 1 × 10⁵ cells/mL in six-well plates coated with poly 2-hydroxylethyl methacrylate (poly-HEMA; Sigma-Aldrich) and cultured in serum-free DMEM/F12 medium (Welgene) supplemented with 20 ng/mL epidermal growth factor (Invitrogen), 10 ng/mL basic fibroblast growth factor (Invitrogen), 0.4% bovine serum albumin (GenDEPOT), and 5 mg/mL insulin (Sigma-Aldrich). Adavosertib (HY-10993; MedChemExpress, NJ, USA) was added at 200 nM. Spheroid formation was assessed 5 days after seeding. The cells were observed under a microscope at 100× magnification. The sizes of the spheroids were measured using ImageJ software.

Cho J.G., Kim S.W., Lee A., Jeong H.N., Yun E., Choi J., Jeong S.J., Chang W., Oh S., Yoo K.H., Lee J.B., Yoon S., Lee M.S., Park J.H., Jung M.H., Kim S.W., Kim K.H., Suh D.S., Choi K.U., Choi J., Kim J, & Kwon B.S. (2022). MicroRNA-dependent inhibition of WEE1 controls cancer stem-like characteristics and malignant behavior in ovarian cancer. Molecular Therapy. Nucleic Acids, 29, 803-822.

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HNSCC Cell Line Culturing and Inhibitor Treatments

Cited 1 time

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All cell lines were grown in RPMI (Sigma-Aldrich, St. Louis, MO, USA) supplemented with 10% fetal bovine serum (FBS) (Biochrom AG, Berlin, Germany) at 37°C, 5% CO₂ and 100% humidification. HPV-positive HNSCC cells UD-SCC-2, UM-SCC-47 and UPCI-SCC-154, UPCI-SCC-90, 93VU-147T, UT-SCC-45, and normal human fibroblasts F184 were described previously (21 (link), 33 (link), 39 (link)). Tumor cell lines were identified by a short tandem repeat multiplex assay (Applied Biosystems, Waltham, MD, USA). PARP inhibition was performed using 1 µM olaparib (MyBiosource, San Diego, CA, USA). Wee1 inhibition was performed using 240 nM adavosertib (Selleckchem, Houston, TX, USA) and combined Wee1/Chk1 inhibition was performed at a dose of 60 nM adavosertib and 1 nM prexasertib (MedChemExpress, Monmouth Junction, NJ, USA) unless stated otherwise. Supplementation with nucleosides (EmbryoMax 100×, Sigma-Aldrich, St. Louis, MO, USA) was performed at a final dilution of 1/12.5.

Hintelmann K., Berenz T., Kriegs M., Christiansen S., Gatzemeier F., Struve N., Petersen C., Betz C., Rothkamm K., Oetting A, & Rieckmann T. (2021). Dual Inhibition of PARP and the Intra-S/G2 Cell Cycle Checkpoints Results in Highly Effective Radiosensitization of HPV-Positive HNSCC Cells. Frontiers in Oncology, 11, 683688.

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Compound Preparation for Molecular Studies

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SNDX-5613, SNDX-50469, ziftomenib (KO-539), ATRA, SY-1425, Cytarabine, Daunorubicin, Selinexor (KPT-330), Entinostat, Panobinostat, and Adavosertib were obtained from MedChem Express (Monmouth Junction, NJ). Cycloheximide was obtained from Santa Cruz Biotechnology, Inc. (Dallas, TX). All compounds were prepared as 10 mM stocks in 100% DMSO and frozen at −80 °C in 5–10 µL aliquots to allow for single use, thus avoiding multiple freeze-thaw cycles that could result in compound decomposition and loss of activity. For in vivo studies, SNDX-5613 was obtained from Syndax Pharmaceuticals under an MTA and reconstituted per the manufacturer’s instructions.

Mill C.P., Fiskus W., Das K., Davis J.A., Birdwell C.E., Kadia T.M., DiNardo C.D., Daver N., Takahashi K., Sasaki K., McGeehan G.M., Ruan X., Su X., Loghavi S., Kantarjian H, & Bhalla K.N. (2023). Causal linkage of presence of mutant NPM1 to efficacy of novel therapeutic agents against AML cells with mutant NPM1. Leukemia, 37(6), 1336-1348.

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Drug Combination Treatment Protocol

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5-fluorouracil (5-FU; S1209), oxaliplatin (S1224), SN-38 (S4908), and olaparib (AZD2281, Ku-0059436, S1060) were purchased from Selleckchem. Berzosertib (ATRi; HY-13902), ceralasertib (ATRi; HY-19323), rabusertib [CHK1 inhibitor (CHKi); HY-14720], adavosertib [Wee1 inhibitor (Wee1i); HY-10993], AZD0156 [ATM inhibitor (ATMi); HY-100016], and nedisertib [DNA-PK inhibitor (DNA-PKi); HY-101570] were purchased from MedChemExpress, while MG-132 (474790) was purchased from Merck and hydroxyurea (HU; H8627) from Sigma.

Durinikova E., Reilly N.M., Buzo K., Mariella E., Chilà R., Lorenzato A., Dias J.M., Grasso G., Pisati F., Lamba S., Corti G., Degasperi A., Cancelliere C., Mauri G., Andrei P., Linnebacher M., Marsoni S., Siena S., Sartore-Bianchi A., Nik-Zainal S., Di Nicolantonio F., Bardelli A, & Arena S. (2022). Targeting the DNA Damage Response Pathways and Replication Stress in Colorectal Cancer. Clinical Cancer Research, 28(17), 3874-3889.

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Drug Combination Treatment Protocol

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Testis Extract Culture and Compound Screening

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Cultures were performed as described elsewhere (Brieño-Enríquez et al., 2016 (link); Wiltshire et al., 1995 (link)), with some modifications. Testis extracts were dissected into 1 x PBS and incubated in a shaking water bath at 34°C, 150rpm in 2 mg/ml Collagenase (Sigma C5138) in 1 x PBS for 15 minutes. Following multiple rounds of centrifugation and washing in 1 x PBS, the testis extract was re-suspended in 2.5 mg/ml Trypsin (Sigma T0303), 200 μg/ml DNase (Sigma DN25) in 1 x PBS and incubated in a shaking water bath at 34°C, 150rpm for 15 minutes. Extract was then centrifuged and washed five times in 4 mL spermatocyte culture medium (SCM) (GIBCO DMEM without red phenol (21063-029), fetal calf serum (GIBCO 10082139), penicillin-streptomycin 1003 (GIBCO 15140-122); lactic acid (Sigma L13750, NaHCO3 9s8761) and sodium pyruvate 1003 (Sigma 11360-070). Cells were re-suspended in 600μl of SCM and placed in 12-well treated culture dishes (Corning #3513). Adavosertib (MedChemExpress HY-10993) was diluted in 10% DMSO and added at 0.25, 0.5, 1, 2, 5 and 10 μg/ml. Nocodazole (Sigma SML1665-1ML) was diluted in 10% DMSO and added at 1, 5, 10, 20, 50 and 80 μg/ml. Cultures were incubated at 37°C for 6 hours followed by protein extraction.

Gray S., Santiago E.R., Chappie J.S, & Cohen P.E. (2020). Cyclin N-Terminal Domain-Containing-1 Coordinates Meiotic Crossover Formation with Cell-Cycle Progression in a Cyclin-Independent Manner. Cell Reports, 32(1), 107858.

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