| Size | Price | Stock | Qty |
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| 10mg |
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| 50mg |
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| 100mg |
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| 500mg | |||
| Other Sizes |
| Targets |
Methotrexate metabolite
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|---|---|
| ln Vitro |
Researchers have tested the hypothesis that 2,4-diamino-6-hydroxymethyl-pteridine (DAP), 2,4-diaminopteroic acid (DAPA), and 2,4 diamino-N10-methyl-pteroic acid (DAMPA) could be converted into aminopterin (from DAP and DAPA) and methotrexate (from DAMPA), both of which are potent inhibitors of dihydrofolate reductase, a proven drug target for Plasmodium falciparum. DAP, DAPA, and DAMPA inhibited parasite growth in the micromolar range; DAMPA was the most active, with 50% inhibitory concentrations in vitro of 446 ng/ml against the antifolate-sensitive strain and 812 ng/ml against the highly resistant strain under physiological folate conditions. DAMPA potentiates the activity of the sulfone dapsone, an inhibitor of dihydropteroate synthase, but not that of chlorcycloguanil, a known inhibitor of dihydrofolate reductase (DHFR). Experiments with a Saccharomyces cerevisiae strain dependent upon the P. falciparum DHFR enzyme showed that DHFR is a target of DAMPA in that system. We hypothesize that DAMPA is converted to methotrexate by the parasite dihydrofolate synthase, which explains the synergy of DAMPA with dapsone but not with chlorcycloguanil. This de novo synthesis will not occur in the host, since it lacks the complete folate pathway. If this hypothesis holds true, the de novo synthesis of the toxic compounds could be used as a framework for the search for novel potent antimalarial antifolates[2].
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| Enzyme Assay |
A rapid and simple turbulent flow liquid chromatography (TFC-LC) method implementing positive heated electrospray ionization (HESI) for the accurate and precise determination of methotrexate (MTX), 7-hydroxy methotrexate (7-OH MTX), and 4-amino-4-deoxy-N(10)-methylpteroic acid (DAMPA) concentrations in serum was developed. MTX was isolated from serum samples (100μL) after protein precipitation with methanol containing formic acid and internal standard (MTX-D3) followed by centrifugation. The supernatant was injected into the turbulent flow liquid chromatography which is followed by electrospray positive ionization tandem mass spectrometry (TFC-LC-MS/MS) and quantified using a six-point calibration curve. For MTX and DAMPA the assays were linear from 10 to 1000nmol/L and for 7-OH MTX from 20 to 2000nmol/L. Dilutions of 10, 100 and 1000-fold were validated giving a clinically reportable range of 10nmol/L to 5×10(5)nmol/L. Within-day and between-day precisions at concentrations spanning the analytical measurement ranges were less than 10% for all three analytes. MTX, DAMPA and 7-OH MTX were sufficiently stable under all relevant analytical conditions. No significant matrix effect was observed during the method validation. The TFC-LC-MS/MS MTX method was also compared with three other clinically validated MTX assays: a dihydrofolate reductase (DHFR) inhibition assay, an immunoassay based on fluorescence polarization and a previously developed LC-MS/MS assay[1].
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| ADME/Pharmacokinetics |
novel methotrexate (MTX) rescuer, carboxypeptidase-G(2) (CPDG(2)), converts over 98% of MTX in plasma into 2,4-diamino-N(10)-methylpteroic acid (DAMPA) and glutamate, for the treatment of MTX-induced renal failure and delayed MTX excretion. In these patients, DAMPA was cleared faster than MTX, suggesting possible non-renal clearance. The pharmacokinetics and metabolism of DAMPA were investigated in four non-human primates using reversed-phase high-performance liquid chromatography-ultraviolet detection, photodiode array detection, and mass spectrometry. The mean peak plasma concentration of DAMPA was 51 μM, and its plasma distribution conformed to a three-compartment open model of first-order elimination. The mean clearance of DAMPA was 1.9 L/kg/h, with a mean terminal half-life of 51 minutes. 46% of the dose was excreted unchanged in the urine. Three DAMPA metabolites were identified in plasma and urine: hydroxy-DAMPA, DAMPA-glucuronide, and hydroxy-DAMPA-glucuronide. These metabolites were also identified in the plasma of patients treated with CPDG(2) as an MTX rescue agent. The cytotoxicity of DAMPA and its effect on MTX cytotoxicity were evaluated in the Molt-4 human leukemia cell line. DAMPA was non-cytotoxic and did not significantly alter the cytotoxicity of MTX. In non-human primates, DAMPA metabolism is the main pathway of clearance, and metabolism is the reason why DAMPA is cleared faster than MTX after CPDG(2) treatment in patients with MTX-induced renal impairment. [3]
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| References |
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| Additional Infomation |
Deoxyaminopteroic acid is a metabolite produced by the carboxypeptidase-mediated cleavage of methotrexate. Deoxyaminopteroic acid can serve as a biomarker for methotrexate exposure.
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| Molecular Formula |
C15H15N7O2
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|---|---|
| Molecular Weight |
325.33
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| Exact Mass |
325.128
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| Elemental Analysis |
C, 55.38; H, 4.65; N, 30.14; O, 9.84
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| CAS # |
19741-14-1
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| Related CAS # |
Methotrexate metabolite-d3;1794780-00-9
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| PubChem CID |
72441
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| Appearance |
Yellow to brown solid powder
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| Density |
1.532
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| Boiling Point |
689.3±65.0 °C at 760 mmHg
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| Melting Point |
242 ºC
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| Flash Point |
370.7±34.3 °C
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| Vapour Pressure |
0.0±2.3 mmHg at 25°C
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| Index of Refraction |
1.799
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| LogP |
1.75
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| Hydrogen Bond Donor Count |
3
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| Hydrogen Bond Acceptor Count |
9
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| Rotatable Bond Count |
4
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| Heavy Atom Count |
24
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| Complexity |
444
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| Defined Atom Stereocenter Count |
0
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| InChi Key |
LWCXZSDKANNOAR-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C15H15N7O2/c1-22(10-4-2-8(3-5-10)14(23)24)7-9-6-18-13-11(19-9)12(16)20-15(17)21-13/h2-6H,7H2,1H3,(H,23,24)(H4,16,17,18,20,21)
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| Chemical Name |
4-[(2,4-diaminopteridin-6-yl)methyl-methylamino]benzoic acid
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| Synonyms |
NSC131463; Deoxyaminopteroic acid; Deoxyaminopteroic acid; 4-(((2,4-Diaminopteridin-6-yl)methyl)(methyl)amino)benzoic acid; Methotrexate metabolite; 4-[[(2,4-diamino-6-pteridinyl)methyl](methyl)amino]benzenecarboxylic acid; 4-[n-(2,4-diamino-6-pteridinylmethyl)-n-methylamino]benzoic acid; NSC131463; DAMPA
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| HS Tariff Code |
2934.99.9001
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| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
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| Solubility (In Vitro) |
DMSO : ~5 mg/mL (~15.37 mM)
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|---|---|
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (7.68 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL. Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution. Solubility in Formulation 2: ≥ 2.5 mg/mL (7.68 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly. Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution. (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 3.0738 mL | 15.3690 mL | 30.7380 mL | |
| 5 mM | 0.6148 mL | 3.0738 mL | 6.1476 mL | |
| 10 mM | 0.3074 mL | 1.5369 mL | 3.0738 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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