Methotrexate |
Catalog No.GC10405
One-Click Copy Product Info
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Methotrexate is an antifolate antimetabolite that effectively inhibits the activity of dihydrofolate reductase (DHFR), with an IC50 of approximately 7nM.
Products are for research use only. Not for human use. We do not sell to patients.
Cas No.: 59-05-2
Sample solution is provided at 25 µL, 10mM.
Methotrexate is an antifolate antimetabolite that effectively inhibits the activity of dihydrofolate reductase (DHFR), with an IC50 of approximately 7nM [1]. Folic acid is essential for the biosynthesis of purine and pyrimidine bases, so methotrexate interferes with the synthesis of purines and pyrimidines required for DNA replication and cell proliferation [2]. Methotrexate also inhibits adenosine deaminase (ADA), resulting in increased extracellular adenine nucleotide levels [3]. Methotrexate has anticancer, antirheumatic, anti-inflammatory and immunomodulatory activities [4].
In vitro, methotrexate (5 μg/ml) has a wide range of IC50s against various cancer cell lines, from 6.05±0.81 nM to >1,000 nM. Methotrexate resistance in Saos-2 and MCF-7 cells The most potent, AGS and HCT-116 cells are highly sensitive to methotrexate, with IC50 of 6.05±0.81nM and 13.56±3.76nM respectively. NCI-H23 and A549 cells show similar sensitivity to methotrexate [5]. Methotrexate (3ng/ml-1μg/ml) measured the degree of drug resistance of the U-2OS cell line and found that the cell resistance was related to the increase in intracellular DHFR levels [6].
In vivo, treatment of arthritic DBA/1J mice with methotrexate (2-50 mg/kg; s.c.) dose-dependently reduced disease activity and mean paw volume and increased mean body weight percentage [7]. Methotrexate (2 mg/kg; i.p.) treats arthritis in rats and has significant anti-arthritic effects and prevents the occurrence of hematological toxicity [8].
References:
[1]Gurdag S, Khandare J, Stapels S, et al. Activity of dendrimer−methotrexate conjugates on methotrexate-sensitive and-resistant cell lines[J]. Bioconjugate chemistry, 2006, 17(2): 275-283.
[2] Rajagopalan P T R , Zhang Z , Mccourt L ,et al.Interaction of dihydrofolate reductase with methotrexate: Ensemble and single-molecule kinetics[J].Proceedings of the National Academy of Sciences, 2002, 99(21):13481-13486.
[3] Sramek M, Neradil J, Veselska R. Much more than you expected: the non-DHFR-mediated effects of methotrexate[J]. Biochimica et Biophysica Acta (BBA)-General Subjects, 2017, 1861(3): 499-503.
[4] Bedoui Y, Guillot X, Sélambarom J, et al. Methotrexate an old drug with new tricks[J]. International journal of molecular sciences, 2019, 20(20): 5023.
[5] Yoon S A , Choi J R , Kim J O ,et al. Influence of Reduced Folate Carrier and Dihydrofolate Reductase Genes on Methotrexate-Induced Cytotoxicity[J].Cancer Research & Treatment, 2010, 42(3):163-171.
[6]Serra M , Reverter B G , Maurici D ,et al.Analysis of dihydrofolate reductase and reduced folate carrier gene status in relation to methotrexate resistance in osteosarcoma cells.[J].Annals of oncology, 2004.
[7] Singh, Rakesh K.van Haandel, LeonKiptoo, et al . Methotrexate disposition, anti-folate activity and efficacy in the collagen-induced arthritis mouse model[J].European Journal of Pharmacology: An International Journal, 2019, 853.
[8] Banji D , Banji O F , Reddy K , et al. Evaluation of the concomitant use of methotrexate and curcumin on Freund's complete adjuvant-induced arthritis and hematological indices in rats[J]. Indian Journal of Pharmacology, 2011, 43(5):546-550.
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Cell experiment [1]: |
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Cell lines |
MCF-7, AGS, A549, NCI-H23, HCT-116 and Saos-2 cell lines |
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Preparation method |
The cytotoxicity of Methotrexate was repeatedly tested at 8 different concentrations for control: 0.03, 0.1, 0.3, 1, 3, 10, 30 and 100µg/mL. The cells were incubated for 24h to allow sufficient cell adhesion, and all the microplates were incubated for a total of 72h after Methotrexate administration. |
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Reaction Conditions |
0.03, 0.1, 0.3, 1, 3, 10, 30 and 100µg/mL; 72h |
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Applications |
The IC50s of Methotrexate was in an extensively broad range from 6.05±0.81nM to>1,000nM in the cell lines. The Saos-2 and MCF-7 cells were most resistant to Methotrexate; in contrast, the AGS and HCT-116 cells were highly sensitive to Methotrexate with an IC50 of 6.05±0.81nM and 13.56±3.76nM, respectively. The NCI-H23 and A549 cells demonstrated similar sensitivity to Methotrexate. |
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Animal experiment [2]: |
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Animal models |
Male DBA/1J mice |
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Preparation method |
The Methotrexate treatment group were further stratified into 4 subgroups and administered subcutaneous Methotrexate at doses of 2, 10, 20, or 50mg/kg weekly starting on day 14 for a total of 6 doses. Disease activity scores, paw volume measurements, and animal weight were collected. |
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Dosage form |
2, 10, 20, or 50mg/kg; s.c. |
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Applications |
Methotrexate treatment resulted in a reduction in disease activity that was variable and dose-dependent. Mean paw volume measurements were significantly reduced in mice treated with Methotrexate. Mean percent body weight increased significantly in mice treated with Methotrexate. |
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References: [1] Yoon S A , Choi J R , Kim J O ,et al. Influence of Reduced Folate Carrier and Dihydrofolate Reductase Genes on Methotrexate-Induced Cytotoxicity[J].Cancer Research & Treatment, 2010, 42(3):163-171. [2] Singh, Rakesh K.van Haandel, LeonKiptoo, et al . Methotrexate disposition, anti-folate activity and efficacy in the collagen-induced arthritis mouse model[J].European Journal of Pharmacology: An International Journal, 2019, 853. |
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| Cas No. | 59-05-2 | SDF | |
| Chemical Name | (2S)-2-[[4-[(2,4-diaminopteridin-6-yl)methyl-methylamino]benzoyl]amino]pentanedioic acid | ||
| Canonical SMILES | CN(CC1=CN=C2C(=N1)C(=NC(=N2)N)N)C3=CC=C(C=C3)C(=O)NC(CCC(=O)O)C(=O)O | ||
| Formula | C20H22N8O5 | M.Wt | 454.44 |
| Solubility | ≥ 21.55 mg/mL in DMSO | Storage | 4°C, protect from light |
| General tips | Please select the appropriate solvent to prepare the stock solution according to the
solubility of the product in different solvents; once the solution is prepared, please store it in
separate packages to avoid product failure caused by repeated freezing and thawing.Storage method
and period of the stock solution: When stored at -80°C, please use it within 6 months; when stored
at -20°C, please use it within 1 month. To increase solubility, heat the tube to 37°C and then oscillate in an ultrasonic bath for some time. |
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| Shipping Condition | Evaluation sample solution: shipped with blue ice. All other sizes available: with RT, or with Blue Ice upon request. | ||
| Prepare stock solution | |||
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1 mg | 5 mg | 10 mg |
| 1 mM | 2.2005 mL | 11.0026 mL | 22.0051 mL |
| 5 mM | 440.1 μL | 2.2005 mL | 4.401 mL |
| 10 mM | 220.1 μL | 1.1003 mL | 2.2005 mL |
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Method for preparing in vivo formulation: Take μL DMSO master liquid, next addμL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL saline, mix and clarify.
Method for preparing in vivo formulation: Take μL DMSO master liquid, next add μL Corn oil, mix and clarify.
Note: 1. Please make sure the liquid is clear before adding the next solvent.
2. Be sure to add the solvent(s) in order. You must ensure that the solution obtained, in the previous addition, is a clear solution before proceeding to add the next solvent. Physical methods such as vortex, ultrasound or hot water bath can be used to aid dissolving.
3. All of the above co-solvents are available for purchase on the GlpBio website.
Quality Control & SDS
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- Purity: >98.00% Appearance: A solid
- COA (Certificate of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
Average Rating: 5 (Based on Reviews and 30 reference(s) in Google Scholar.)