Tubulin polymerization-IN-59
Based on 1 Customer Validation
Tubulin polymerization-IN-59 is a tubulin polymerization inhibitor and colchicine binding site inhibitor (CBSI) (IC50 = 6.1 μM). Tubulin polymerization-IN-59 exerts potent antiproliferative activity against cancer cells, while showing lower cytotoxicity to normal cells. Tubulin polymerization-IN-59 arrests colorectal cancer HCT 116 cells in G2/M phase, induces cell apoptosis, and suppresses tumor cell colony formation and migration. Tubulin polymerization-IN-59 can be used for the study of colorectal cancer (CRC).
For research use only. We do not sell to patients.
- Purity : 99.85%
- Formula: C20H21FO5
- Molecular Weight:360.38
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Storage:Powder -20°C, 3 years , 4°C, 2 years ; In solvent -80°C, 6 months , -20°C, 1 month
To place orders, for customer services and technical support, please contact: MedChemExpress USA
Tel: 609-228-6898 E-mail: [email protected] [email protected]
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Biological Activity
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Chemical Information
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Solvent & Solubility
- Protocols
- Purity & Documentation
- References
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Help & FAQs
Biological Activity
Description
Cellular Effect
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Cell Line
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Type | Value | Description | References |
|---|---|---|---|---|
| A2780 | IC50 |
0.59 μM
Compound: (R)-9k
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Growth inhibition of human A2780 cells incubated for 1.5 hrs by MTT assay
Growth inhibition of human A2780 cells incubated for 1.5 hrs by MTT assay
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[PMID: 38107178] |
| CT26 | IC50 |
4.11 μM
Compound: (R)-9k
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Growth inhibition of CT26 cells incubated for 1.5 hrs by MTT assay
Growth inhibition of CT26 cells incubated for 1.5 hrs by MTT assay
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[PMID: 38107178] |
| HCT-116 | IC50 |
0.26 μM
Compound: (R)-9k
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Growth inhibition of human HCT-116 cells incubated for 1.5 hrs by MTT assay
Growth inhibition of human HCT-116 cells incubated for 1.5 hrs by MTT assay
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[PMID: 38107178] |
| HEK293 | IC50 |
7.45 μM
Compound: (R)-9k
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Cytotoxicity against HEK293 cells assessed as inhibition of cell growth incubated 1.5 hrs by MTT assay
Cytotoxicity against HEK293 cells assessed as inhibition of cell growth incubated 1.5 hrs by MTT assay
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[PMID: 38107178] |
| HeLa | IC50 |
0.38 μM
Compound: (R)-9k
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Growth inhibition of human HeLa cells incubated for 1.5 hrs by MTT assay
Growth inhibition of human HeLa cells incubated for 1.5 hrs by MTT assay
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[PMID: 38107178] |
| HUVEC | IC50 |
15.01 μM
Compound: (R)-9k
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Cytotoxicity against human HUVEC cells assessed as inhibition of cell growth incubated 1.5 hrs by MTT assay
Cytotoxicity against human HUVEC cells assessed as inhibition of cell growth incubated 1.5 hrs by MTT assay
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[PMID: 38107178] |
| SW480 | IC50 |
1.31 μM
Compound: (R)-9k
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Growth inhibition of human SW480 cells incubated for 1.5 hrs by MTT assay
Growth inhibition of human SW480 cells incubated for 1.5 hrs by MTT assay
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[PMID: 38107178] |
In Vitro
Tubulin polymerization-IN-59 ((R)-9k) (48-72 h) exerts potent antiproliferative activity against HeLa, A2780, HCT 116, SW480, and CT-26 cells, with IC50 values of 0.38 μM, 0.59 μM, 0.26 μM, 1.31 μM, and 4.11 μM, respectively[1].
Tubulin polymerization-IN-59 (3-12 μM) inhibits tubulin polymerization in a dose-dependent manner, with an IC50 value of 6.1 μM[1].
Tubulin polymerization-IN-59 (1-25 μM, 3.5 h) reduces the formation of EBI-β-tubulin adduct in HCT 116 cells by competing for the colchicine binding site on tubulin[1].
Tubulin polymerization-IN-59 (1.5-6.0 μM, 24 h) arrests HCT 116 cells in the G2/M phase in a dose-dependent manner and induces apoptosis of HCT 116 cells[1].
Tubulin polymerization-IN-59 (0.1-2 μM, 2 weeks) inhibits colony formation of HCT 116 cells[1].
Tubulin polymerization-IN-59 (0.5-5.0 μM, 24 h) inhibits migration of HCT 116 cells in a dose-dependent manner
Tubulin polymerization-IN-59 (0.16-40 μM) weakly inhibits the hERG potassium channel in CHO cells[1].
MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only. Further protocols information, click here.
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Cell Line:HCT 116 cells
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Concentration:1.5, 3, 6.0 μM
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Incubation Time:24 h
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Result:Arrested HCT 116 cells in the G2/M phase in a dose-dependent manner.
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Cell Line:HCT 116 cells
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Concentration:1.5, 3, 6.0 μM
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Incubation Time:24 h
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Result:Induced apoptosis of HCT 116 cells.
Chemical Information
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Appearance Solid
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Molecular Weight 360.38
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Formula C20H21FO5
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Color White to off-white
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SMILES
CCOC1=CC=C(C=C1F)[C@H]2CC(C3=CC(OC)=C(C(OC)=C32)OC)=O
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Shipping
Room temperature in continental US; may vary elsewhere.
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Storage
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month
Solvent & Solubility
In Vitro:
DMSO : ≥ 100 mg/mL (277.48 mM; Hygroscopic DMSO has a significant impact on the solubility of product, please use newly opened DMSO)
* "≥" means soluble, but saturation unknown.
Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 6 months; -20°C, 1 month. When stored at -80°C, please use it within 6 months. When stored at -20°C, please use it within 1 month.
Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 6 months; -20°C, 1 month. When stored at -80°C, please use it within 6 months. When stored at -20°C, please use it within 1 month.
Concentration (start) × Volume (start) = Concentration (final) × Volume (final)
In Vivo:
Select the appropriate dissolution method based on your experimental animal and administration route.
- For the following dissolution methods, please ensure to first prepare a clear stock solution using an In Vitro approach and then sequentially add co-solvents:
- To ensure reliable experimental results, the clarified stock solution can be appropriately stored based on storage conditions. As for the working solution for In Vivo experiments, it is recommended to prepare freshly and use it on the same day.
- The percentages shown for the solvents indicate their volumetric ratio in the final prepared solution. If precipitation or phase separation occurs during preparation, heat and/or sonication can be used to aid dissolution.
Add each solvent one by one: 10% DMSO 40% PEG300 5% Tween-80 45% Saline
Solubility: ≥ 2.5 mg/mL (6.94 mM); Clear solution
This protocol yields a clear solution of ≥ 2.5 mg/mL (saturation unknown).
Taking 1 mL working solution as an example, add 100 μL DMSO stock solution (25.0 mg/mL) to 400 μL PEG300, and mix evenly; then add 50 μL Tween-80 and mix evenly; then add 450 μL Saline to adjust the volume to 1 mL.
Preparation of Saline: Dissolve 0.9 g sodium chloride in ddH₂O and dilute to 100 mL to obtain a clear Saline solution.
In Vivo Dissolution Calculator
Please enter the basic information of animal experiments:
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Recommended: Prepare an additional quantity of animals to account for potential losses during experiments.
Please enter your animal formula composition:
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%DMSO +
Recommended: Keep the proportion of DMSO in working solution below 2% if your animal is weak.
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%+
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+%Tween-80 + +
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%Saline +
The co-solvents required include: DMSO, . All of co-solvents are available by MedChemExpress (MCE). , Tween 80. All of co-solvents are available by MedChemExpress (MCE).
Working solution concentration: 0.22 mg/mL
Method for preparing stock solution: mg drug dissolved in μL DMSO. Stock solution concentration: mg/mL.
1. Take μL DMSO stock solution;
2. Add μL .
μL , mix evenly;
3. Then add μL Tween 80, mix evenly;
4. Then add μL
Please ensure that the stock solution in the first step is dissolved to a clear state, and add co-solvents in sequence. You can use ultrasonic heating (ultrasonic cleaner, recommended frequency 20-40 kHz), vortexing, etc. to assist dissolution.
Protocols
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Apoptosis
Apoptosis, also called programmed cell death, is generally characterized by distinct morphological characteristics.
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TUNEL staining for apoptotic DNA fragmentation
TUNEL staining detects DNA strand breaks by using terminal deoxynucleotidyl transferase to add labeled nucleotides to exposed 3′-OH DNA termini, generating either microscopic staining in fixed cells or tissue sections, or fluorescence/cytometric signal in cell suspensions. TUNEL positivity reflects DNA fragmentation but should not be interpreted alone as definitive apoptosis, because TUNEL can also label necrotic, autolytic, mechanically damaged, or DNA-repair-associated DNA breaks.
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Cell Cytotoxicity Assay
Cytotoxicity assays are usually based on the assessment of cell membrane damage, which can also be indirectly detected by measuring cell viability. Detection methods include MTT assay, CKK-8 assay, LDH assay and ATP assay, etc.
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Mammalian live/dead viability and cytotoxicity staining
Live/dead viability and cytotoxicity staining assays are based on the simultaneous detection of intracellular esterase activity in metabolically active (viable) cells and membrane integrity loss in non-viable cells. In commonly used dual-staining approaches, membrane-permeant fluorogenic substrates are converted by intracellular esterases into fluorescent products in live cells, while impermeant DNA-binding dyes selectively enter cells with compromised plasma membranes and label nucleic acids in dead or dying cells, enabling discrimination between viable and non-viable populations by fluorescence microscopy or flow cytometry.
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Flow cytometric DNA-content cell-cycle staining
Flow cytometric DNA-content cell-cycle staining measures the fluorescence intensity of DNA-bound fluorochromes in single cells or nuclei to estimate DNA content distributions, allowing assignment of populations to G0/G1, S, and G2/M phases by DNA histogram deconvolution. Propidium iodide (PI) intercalates into DNA, and PI fluorescence is proportional to cellular DNA content when staining is performed under conditions that make DNA accessible and minimize non-DNA signal. Cells with G2/M DNA content are expected to show approximately twice the fluorescence intensity of G0/G1 cells, while S-phase cells occupy intermediate fluorescence values. PI-based DNA-content analysis can also detect cells with fractional DNA content, often reported as sub-G1, when DNA fragmentation and extraction during staining reduce retained DNA signal in apoptotic cells. DAPI is an alternative DNA fluorochrome for univariate DNA-content analysis, while bivariate approaches combining DNA content with proliferation
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Annexin V plus membrane-impermeant dye apoptosis staining
Annexin V-based apoptosis assays rely on the detection of phosphatidylserine (PS) externalization from the inner leaflet of the plasma membrane to the outer leaflet, an early biochemical hallmark of apoptosis. Fluorescently labeled Annexin V binds PS in a calcium-dependent manner, enabling identification of early apoptotic cells by flow cytometry or fluorescence microscopy. When combined with a membrane-impermeant DNA-binding dye (e. g. , propidium iodide), this approach allows discrimination between viable (Annexin V−/dye−), early apoptotic (Annexin V+/dye−), and late apoptotic or necrotic (Annexin V+/dye+) cell populations by assessing membrane integrity and PS exposure.
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Cell migration
Cell migration is a method that plays an important role in wound healing, cell differentiation, embryonic development, etc.
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Apoptosis Solutions
Apoptosis is a regulated, generally non-lytic cell-death pathway that removes unwanted, damaged, infected, or abnormal cells through coordinated morphological changes, caspase activation, DNA fragmentation, and membrane remodeling. The intrinsic apoptosis pathway is controlled mainly by mitochondrial outer membrane permeabilization, BCL-2 family proteins, cytochrome c release, apoptosome formation, caspase-9 activation, and downstream executioner caspase-3/7 activation. The extrinsic apoptosis pathway is initiated by death receptors such as Fas, TNFR, and TRAIL receptors, which recruit adaptor proteins and activate caspase-8 before engaging executioner caspases or mitochondrial amplification through BID cleavage. Apoptosis is linked to many phenotypes, including cancer cell killing, tissue homeostasis, immune regulation, neurodegeneration, infection response, and treatment-induced cytotoxicity; unresolved questions include how apoptosis interacts with necroptosis, pyroptosis, ferroptos
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Protocol for Cell Cycle
Cell-cycle analysis by flow cytometry measures DNA content in single cells to estimate the fraction of cells in G0/G1, S, and G2/M phases. Propidium iodide intercalates into DNA, and after RNA removal with RNase, fluorescence intensity reflects cellular DNA content: 2N cells are assigned to G0/G1, cells between 2N and 4N to S phase, and 4N cells to G2/M. DNA-content analysis alone cannot reliably separate G0 from G1 or G2 from M. Ki-67 can distinguish quiescent G0 cells from cycling cells, EdU or BrdU incorporation marks active DNA synthesis in S phase, and phospho-histone H3 staining identifies mitotic cells within the 4N population.
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Soft Agar Colony Formation Assay
Soft agar colony formation assay measures anchorage-independent growth, in which transformed or tumorigenic cells proliferate as colonies in a semisolid agar matrix while many non-transformed adherent cells fail to proliferate without attachment; classic studies showed that growth in semisolid medium correlates with tumorigenicity in nude mice, and later protocol papers describe the method as a stringent in vitro assay for malignant transformation. The readout is the number, size, morphology, or signal intensity of colonies formed within agar after incubation; published formats include manual colony counting after staining, 96-well or 384-well quantitative formats, DNA-binding dye detection, MTT/tetrazolium-based detection, digital image analysis, and PCR-based marker detection from soft agar cultures.
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Colony Formation (Clonogenic) Assay
The clonogenic (colony formation) assay measures the ability of a single cell to retain reproductive viability and form a macroscopic colony, typically defined as a cluster derived from one progenitor cell after a defined growth period. This assay is widely used to evaluate cell survival after exposure to ionizing radiation or cytotoxic treatments and is considered a standard method in radiation biology for generating dose-response relationships of reproductive cell death. Colony formation reflects long-term proliferative capacity rather than short-term metabolic activity, and survival is quantified by comparing treated versus untreated conditions based on colony number and derived survival fractions.
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Cell Viability Determination by MTT Colorimetric Assay
The following protocol uses the MTT colorimetric assay as a classic literature-established method for assessing cell viability/metabolic activity in cultured mammalian cells. MTT[3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] is reduced by metabolically active cells to a colored formazan product; the amount of formazan is quantified spectrophotometrically and provides an indirect measure of metabolically active viable cells. Importantly, MTT reduction reflects cellular oxidoreductase/metabolic activity rather than an absolute direct count of living cells, so changes in cellular metabolism can alter the signal independently of cell number.
Purity & Documentation
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Data Sheet (280 KB)
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SDS (251 KB)
- English - EN (251 KB)
- Français - FR (251 KB)
- Deutsch - DE (251 KB)
- Norwegian - NO (251 KB)
- Español - ES (251 KB)
- Swedish - SV (251 KB)
- Italian - IT (251 KB)
- Korean - KR (251 KB)
- Portuguese - PT (251 KB)
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Handling Instructions (2659 KB)
References
Complete Stock Solution Preparation Table
Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 6 months; -20°C, 1 month. When stored at -80°C, please use it within 6 months. When stored at -20°C, please use it within 1 month.
| Optional Solvent | Concentration Solvent Mass | 1 mg | 5 mg | 10 mg | 25 mg |
|---|---|---|---|---|---|
| DMSO | 1 mM | 2.7748 mL | 13.8742 mL | 27.7485 mL | 69.3712 mL |
| 5 mM | 0.5550 mL | 2.7748 mL | 5.5497 mL | 13.8742 mL | |
| 10 mM | 0.2775 mL | 1.3874 mL | 2.7748 mL | 6.9371 mL | |
| 15 mM | 0.1850 mL | 0.9249 mL | 1.8499 mL | 4.6247 mL | |
| 20 mM | 0.1387 mL | 0.6937 mL | 1.3874 mL | 3.4686 mL | |
| 25 mM | 0.1110 mL | 0.5550 mL | 1.1099 mL | 2.7748 mL | |
| 30 mM | 0.0925 mL | 0.4625 mL | 0.9249 mL | 2.3124 mL | |
| 40 mM | 0.0694 mL | 0.3469 mL | 0.6937 mL | 1.7343 mL | |
| 50 mM | 0.0555 mL | 0.2775 mL | 0.5550 mL | 1.3874 mL | |
| 60 mM | 0.0462 mL | 0.2312 mL | 0.4625 mL | 1.1562 mL | |
| 80 mM | 0.0347 mL | 0.1734 mL | 0.3469 mL | 0.8671 mL | |
| 100 mM | 0.0277 mL | 0.1387 mL | 0.2775 mL | 0.6937 mL |
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