Tozasertib (VX-680)

Catalog No.S1048 Batch:S104805

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Technical Data

Formula

C23H28N8OS
Molecular Weight 464.59 CAS No. 639089-54-6
Solubility (25°C)* In vitro DMSO 93 mg/mL (200.17 mM)
Water Insoluble
Ethanol Insoluble
In vivo (Add solvents to the product individually and in order)
Homogeneous suspension
CMC-NA
≥5mg/ml Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.
* <1 mg/ml means slightly soluble or insoluble.
* Please note that Selleck tests the solubility of all compounds in-house, and the actual solubility may differ slightly from published values. This is normal and is due to slight batch-to-batch variations.
* Room temperature shipping (Stability testing shows this product can be shipped without any cooling measures.)

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Biological Activity

Description Tozasertib (VX-680) is a pan-Aurora inhibitor, mostly against Aurora A with Kiapp of 0.6 nM in a cell-free assay, less potent towards Aurora B/Aurora C and 100-fold more selective for Aurora A than 55 other kinases. The only exceptions are Fms-related tyrosine kinase-3 (FLT-3) and BCR-ABL tyrosine kinase, which are inhibited by the Tozasertib with both Ki of 30 nM. Tozasertib induces apoptosis and autophagy. Phase 2.
Targets
Aurora A [1]
(Cell-free assay)		 Aurora C [1]
(Cell-free assay)		 Aurora B [1]
(Cell-free assay)		 FLT3 [4]
(Cell-free assay)		 Bcr-Abl [4]
(Cell-free assay)
0.6 nM(Ki app) 4.6 nM(Ki app) 18 nM(Ki app) 30 nM(Ki) 30 nM(Ki)
In vitro Although its multi-kinase profile, VX-680 induces similar cytotoxicity with IC50 of approximately 300 nM and exhibits an AUR B-like inhibitory phenotype of G2/M arrest, endoreduplication and apoptosis in BaF3 cells transfected with ABL or FLT-3 (mutant and wild type) kinases. VX-680 prevents the CAL-62 proliferation in a time-dependent manner. VX-680 treatment for 14 days significantly decreases the number and size of colonies by approximately 70% in the 8305C and 90% in the CAL-62, 8505C and BHT-101. Treatment of the different ATC cells with VX-680 inhibits proliferation with the IC50 between 25 and 150  nM. The VX-680 significantly impairs the ability of the different cell lines to form colonies in soft agar. Analysis of caspase-3 activity indicates that VX-680 induces apoptosis in the different cell lines. CAL-62 cells exposed for 12  hours to VX-680 showed an accumulation of cells with ≥4N DNA content. Time-lapse analysis demonstrates that VX-680-treated CAL-62 cells exit metaphase without dividing. Moreover, histone H3 phosphorylation is abrogated following VX-680 treatment. [2] VX-680 has significant inhibitory activity against BCR-Abl bearing the T315I mutation in patient-derived samples. [3]
In vivo VX-680 gives rise to a marked decrease in tumor size in a human AML (HL-60) xenograft model. In mude mice treateed with VX-680 at 75 mg/kg, twice a day intraperitoneally (b.i.d. i.p.) for 13 days, mean tumor volumes are reduced by 98%. Tumor growth decrease is dose dependent and significant at a dose of 12.5 mg/kg b.i.d. VX-680 is well tolerated, with a small decrease in body weight observed only at the highest dose. VX-680 also triggers tumor regresson in pancreatic and colon xenograft models. VX-680 also displays potent antitumor activity when infused i.v. in mude rats bearing established HCT116 tumors. A higher dose of VX-680 (2 mg/kg/h) improves efficacy with a 56% decrease in mean tumor volume. [1]

Protocol (from reference)

Kinase Assay:

[3]
  • Kinase inhibition assays

    The consumption of ATP is coupled via the pyruvate kinase/lactic dehydrogenase enzyme pair to the oxidation of NADH, which can be monitored through the decrease in absorption at 340 nm. Reactions contains 100 mM Tris (pH 8), 10 mM MgCl2, 2.2 mM ATP, 1 mM phosphoenolpyruvate, 0.6 mg/mL NADH, 75 units/mL pyruvate kinase, 105 units/mL lactate dehydrogenase, and 0.5 mM substrate peptide (sequence: EAIYAAPFAKKK). Reactions (75 μL) are started by adding sufficient kinase to bring the reactions to 30 nM kinase concentration and the decrease in absorbance is monitored over 30 minutes at 30°C in a microtiter plate spectrophotometer. Inhibitory constants are obtained through addition of 3.75 μL VX-680 in 100% DMSO or DMSO alone. Ki values are calculated as follows, K i = IC50 / (1 + [S]/Kd), where [S] = [ATP] = 2.2 mM, and Kd (of ATP to Abl) = 70 μM. These values are calculated assuming a Kd (ATP) of 70 μM for wild type and H396P Abl kinase domain.
Cell Assay:

[2]
  • Cell lines

    CAL-62 cells

  • Concentrations

    5-500 nM

  • Incubation Time

    4 days

  • Method

    The CAL-62 cells are cultured in the absence (dimethyl sulfoxide, DMSO) or the presence of 500  nM VX-680 for different periods of time (1-5 days). The dose-dependent effects of VX-680 on cell proliferation are evaluated by treating the different ATC cells for 4 days with different concentrations of the Aurora inhibitor (5–500  nM). The cells are pulse labeled with 30  mM BrdU for 2  hours before the end of the incubation time. The BrdU incorporation is analyzed by means of a colorimetric immunoassay using the cell proliferation ELISA kit. The results from VX-680-treated cells are compared with those observed in control cells and expressed as a fold of variation versus control.
Animal Study:

[1]
  • Animal Models

    Female athymic NCr-nu mice bearing HL-60 leukemia cells

  • Dosages

    50 mg/kg, 75 mg/kg

  • Administration

    Administered via i.p.

References

  • https://pubmed.ncbi.nlm.nih.gov/14981513/
  • https://pubmed.ncbi.nlm.nih.gov/18430894/
  • https://pubmed.ncbi.nlm.nih.gov/16424036/
  • https://pubmed.ncbi.nlm.nih.gov/17240048/

Customer Product Validation

<p>Senescence induction upon PKCι depletion combined with aurora kinase inhibition. ( a) MCF7 cells were transfected as above to deplete PKCι . Two days after transfection, cells were treated for the indicated time period with 400 n M VX-680. Medium with VX-680 was then removed and fresh medium was added. Cells were stained for SA-b -gal activity 5 days after the start of transfection.* indicates a P value <0.05. ( b) MCF7 cells were treated as above. Five days after transfection, cells were fixed and assessed for the presence of gH2AX foci by immunofluorescence microscopy. (c, d) MCF7 cells were treated with dimethyl sulfoxide (DMSO) control or 400 n M VX-680 for the indicated time periods. Total cell lysates were then analyzed by western blotting for levels of p21 and GAPDH (as loading control). A representative blot is shown in panel c. Quantitation of changes in p21 levels (normalized to vehicle-treated controls) is shown in panel d. The data shown are the means ±s.e. of three independent experiments.</p>

Data from [ Oncogene , 2012 , 31, 3584-96 ]

<p>Senescence induction upon PKCι depletion combined with aurora kinase inhibition in glioblastoma cells. (a, b) U87MG cells were transfected as above to deplete PKCι. Two days after transfection, cells were treated for 72 ( a)or24h (b) with 400 nM VX-680. Medium with VX-680 was then removed and fresh medium was added. Cells were stained for SA-b-gal activity 5 days after the start of transfection. * indicates a P value <0.05. (c) U87MG cells were treated as described in panel a above. Five days after transfection, cells were fixed and assessed for the presence of gH2AX foci by immunofluorescence microscopy. (d) U87MG cells were treated with the dimethyl sulfoxide (DMSO) control or 400 n M VX-680 for the indicated time periods. Total cell lysates were then analyzed by western blotting for levels of p21. The bar graph shows quantitation of p21 levels (normalized to vehicle-treated controls) from three independent experiments. A representative blot is also shown, with lanes aligned to correspond to the labels on the graph.</p>

Data from [ Oncogene , 2012 , 31, 3584-96 ]

<p>MTT assay reveals a dose-dependent decrease in cell viability in mouse derived brainstem glioma cells treated with VX-680 ( P < 0.001) after 72 h of treatment. The error bars represent the standard deviation. Propidium iodide based cell sorting of mouse derived brainstem glioma cells after 72 h treatment with 5 μM reversine or 100 nM VX-680 respectively reveals increased cell populations with 4N and 8N DNA content as compared to vehicle control.</p>

Data from [ Brain Pathol , 2012 , 23, 244-53 ]

<p>Treatment of mouse derived brainstem glioma cells for 72 h with 5 μM reversine or 100 nM VX-680 increases cell size compared with vehicle-treated control and leads to irregular-shaped nuclei and micronuclei (F–H). Images F–H represent immunofluorescent staining for GFAP (green) with DAPI counter-stain (blue) and were taken at 400 ×magnification.</p>

Data from [ Brain Pathol , 2012 , 23, 244-53 ]

Selleck's Tozasertib (VX-680) Has Been Cited by 143 Publications

A patient-derived T cell lymphoma biorepository uncovers pathogenetic mechanisms and host-related therapeutic vulnerabilities [ Cell Rep Med, 2025, S2666-3791(25)00102-8] PubMed: 40147445
Overcoming MET-targeted drug resistance in MET-amplified lung cancer by aurora kinase B inhibition [ Biochim Biophys Acta Mol Cell Res, 2025, 1872(7):120001] PubMed: 40499687
Hedgehog signalling is involved in acquired resistance to KRASG12C inhibitors in lung cancer cells [ Cell Death Dis, 2024, 15(1):56] PubMed: 38225225
Dynamic phosphorylation of FOXA1 by Aurora B guides post-mitotic gene reactivation [ Cell Rep, 2024, 43(9):114739] PubMed: 39276350
The molecular basis of Abelson kinase regulation by its αI-helix [ Elife, 2024, 12RP92324] PubMed: 38588001
Tozasertib activates anti-tumor immunity through decreasing regulatory T cells in melanoma [ Neoplasia, 2024, 48:100966] PubMed: 38237304
Machine learning based androgen receptor regulatory gene-related random forest survival model for precise treatment decision in prostate cancer [ Heliyon, 2024, 10(17):e37256] PubMed: 39296076
Visualization strategies to aid interpretation of high-dimensional genotoxicity data [ Environ Mol Mutagen, 2024, 10.1002/em.22604] PubMed: 38757760
Molecular landscape and functional characterization of centrosome amplification in ovarian cancer [ Nat Commun, 2023, 14(1):6505] PubMed: 37845213
Molecular landscape and functional characterization of centrosome amplification in ovarian cancer [ Nat Commun, 2023, 14(1):6505] PubMed: 37845213

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