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| 5mg |
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| Targets |
Amg-1 directly targets IDO1 with an IC50 of 3.0 microM. It exhibits high selectivity over the related isoenzymes IDO2 (IC50 > 250 microM, >80-fold selectivity) and tryptophan 2,3-dioxygenase (TDO, IC50 > 62.5 microM, >20-fold selectivity). By inhibiting IDO1, Amg-1 blocks the conversion of tryptophan to kynurenine, thereby preventing T-cell suppression and restoring anti-tumor immunity.
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| ln Vitro |
Amg-1 is a reversible inhibitor of IDO1 enzymatic activity in vitro. In cell-free enzymatic assays using purified recombinant human IDO1, Amg-1 inhibits enzyme activity with an IC50 of 3.0 microM. This inhibition is reversible upon compound washout. The compound shows high selectivity for IDO1 compared to IDO2 and TDO, with no significant activity against a panel of other metabolic enzymes at 100 microM.
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| ln Vivo |
In vivo, Amg-1 has been used to study the effects of IDO1 inhibition in animal models of cancer and cardiovascular disease. As an IDO1 inhibitor, Amg-1 can restore T-cell proliferation and cytokine production in the tumor microenvironment, leading to reduced tumor growth. Preclinical studies have shown that Amg-1 has beneficial effects in hypotension models by influencing vascular function through NO pathway.
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| Enzyme Assay |
A fluorescence-based enzymatic assay is performed. Recombinant human IDO1 (50 nM) is incubated with varying concentrations of Amg-1 (0.01-300 microM) in 100 microL of 50 mM potassium phosphate buffer pH 6.5 containing 40 mM ascorbic acid, 20 microg/mL catalase, and 200 microM L-tryptophan. After 60 min at 37degC, the reaction is stopped by adding 50 microL of 30% trichloroacetic acid and heated at 50degC for 30 min to convert N-formylkynurenine to kynurenine. After centrifugation (4000 rpm, 10 min), 100 microL of supernatant is mixed with an equal volume of 2% p-dimethylaminobenzaldehyde in acetic acid. Absorbance is measured at 480 nm. IC50 is calculated by nonlinear regression.
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| Cell Assay |
For cell-based potency, human cancer cell lines expressing high levels of IDO1 (e.g., HeLa cells) are seeded in 96-well plates at 2×10⁴ cells/well and stimulated with IFN-gamma (100 ng/mL) for 24 hours to induce IDO1 expression. Cells are treated with Amg-1 (0.1-100 microM) for 48 hours. Kynurenine concentration in culture supernatant is measured by LC-MS/MS or colorimetric assay with Ehrlich‘s reagent. Cell viability is assessed by MTT assay to differentiate growth inhibition from direct cytotoxicity.
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| Animal Protocol |
For an in vivo model of IDO1-dependent immunosuppression, female BALB/c mice are subcutaneously implanted with IDO1-expressing tumor cells. When tumors reach 100-150 mm3, mice are treated orally or intraperitoneally with Amg-1 at 10-50 mg/kg daily for 14 days. Tumor volumes are measured every 2-3 days. Plasma kynurenine levels are measured by LC-MS/MS as a pharmacodynamic biomarker of target engagement. Splenocytes are collected for T-cell proliferation assay and flow cytometry analysis of immune cell subsets.
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| ADME/Pharmacokinetics |
Pharmacokinetic studies in rodents show that Amg-1 is orally bioavailable and distributes into tissues including the tumor microenvironment. Based on its structure as a small molecule (MW 424.50), it is expected to have moderate plasma clearance and a half-life suitable for once or twice daily dosing. Metabolites may be formed by oxidation and glucuronidation.
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| Toxicity/Toxicokinetics |
Acute toxicity studies in rodents indicate that Amg-1 has a favorable safety margin. No overt toxicity or body weight loss is observed at doses up to 100 mg/kg. The high selectivity for IDO1 over other tryptophan catabolizing enzymes reduces the risk of off-target effects. Standard safety assays show no significant hERG inhibition or genotoxicity potential at concentrations up to 10 microM. Long-term toxicity studies have not been reported.
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| References | |
| Additional Infomation |
This compound has CAS number 496023-55-3. Its molecular formula is C20H16N4O3S2 and molecular weight is 424.50. It is typically stored as powder at -20degC for up to 3 years or in solvent at -80degC for 6 months. The compound is commercially available for research purposes only.
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| Molecular Formula |
C20H16N4O3S2
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| Molecular Weight |
424.50
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| CAS # |
496023-55-3
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| Appearance |
Light brown to brown solid powder
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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) |
May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples
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| Solubility (In Vivo) |
Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.
Injection Formulations
Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)(e.g. IP/IV/IM/SC) *Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution. Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil) Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals). View More
Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)] Oral Formulations
Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium) Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals). View More
Oral Formulation 3: Dissolved in PEG400 (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 2.3557 mL | 11.7786 mL | 23.5571 mL | |
| 5 mM | 0.4711 mL | 2.3557 mL | 4.7114 mL | |
| 10 mM | 0.2356 mL | 1.1779 mL | 2.3557 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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