PYR-41 is a ubiquitin-activating enzyme inhibitor (UBE1 inhibitor). PYR-41 blocks ubiquitination reactions but paradoxically leads to the accumulation of high MW ubiquitinated proteins. PYR-41 also mediated cross-linking of specific protein kinases (Bcr-Abl, Jak2) to inhibit their signaling activity. PYR-41 has demonstrated anti-tumor activity in animal studies, partially selective protein cross-linking may represent an alternate approach to affect signal transduction modules and ubiquitin cycle-regulatory proteins for cancer therapy. (Biochem Pharmacol. 2011 Aug 15;82(4):341-9.) ( PYR-41 in solution may exist as a mixture of E- and Z- isomers. HPLC may show two peaks depending on conditions).
Physicochemical Properties
| Molecular Formula | C17H13N3O7 |
| Molecular Weight | 371.30102 |
| Exact Mass | 371.075 |
| Elemental Analysis | C, 54.99; H, 3.53; N, 11.32; O, 30.16 |
| CAS # | 418805-02-4 |
| Related CAS # | 418805-02-4 |
| PubChem CID | 5335621 |
| Appearance | Brown solid powder |
| Density | 1.5±0.1 g/cm3 |
| Index of Refraction | 1.652 |
| LogP | 2.87 |
| Hydrogen Bond Donor Count | 1 |
| Hydrogen Bond Acceptor Count | 7 |
| Rotatable Bond Count | 5 |
| Heavy Atom Count | 27 |
| Complexity | 664 |
| Defined Atom Stereocenter Count | 0 |
| SMILES | O1C(=C([H])C([H])=C1/C(/[H])=C1/C(N([H])N(C/1=O)C1C([H])=C([H])C(C(=O)OC([H])([H])C([H])([H])[H])=C([H])C=1[H])=O)[N+](=O)[O-] |
| InChi Key | ARGIPZKQJGFSGQ-LCYFTJDESA-N |
| InChi Code | InChI=1S/C17H13N3O7/c1-2-26-17(23)10-3-5-11(6-4-10)19-16(22)13(15(21)18-19)9-12-7-8-14(27-12)20(24)25/h3-9H,2H2,1H3,(H,18,21)/b13-9- |
| Chemical Name | ethyl 4-(4-((5-nitrofuran-2-yl)methylene)-3,5-dioxopyrazolidin-1-yl)benzoate |
| Synonyms | PYR41; PYR 41; PYR-41. |
| HS Tariff Code | 2934.99.9001 |
| 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) |
Biological Activity
| Targets | E1(IC50: < 10 μM) |
| ln Vitro | PYR-41 not only prevents ubiquitylation but also raises total sumoylation in cells. PYR-41 reduces nuclear factor-κB activation mediated by cytokines. Additionally, PYR-41 stops IκBα from being ubiquitylated downstream and from being broken down by proteases. Moreover, PYR-41 stimulates this tumor suppressor's transcriptional activity and prevents p53 from being degraded[1]. The accumulation of ubiquitinated proteins is promoted by PYR-41 (50 μM).After 4 hours, Z138 cells exhibit a concentration-dependent (10-50 μM) decrease in DUB activity due to PYR-41. Potent inhibition of USP5 DUB activity is observed even at the lowest concentration (10 μM) of PYR-41. Potently (10-50 μM), PYR-41 inhibits the activity of multiple DUBs, which have been identified as USP9x, USP5, USP14, UCH37, and UCH-L3. When DTT and PYR-41 are administered together to Z138 cells, the accumulation of ubiquitinated proteins is totally eliminated[2]. |
| Enzyme Assay | For 15 minutes, 32P-ubiquitin is incubated at room temperature in 1× reaction buffer (50 mM Tris (pH 7.4), 0.2 mM ATP, and 0.5 mM MgCl2) with rabbit or mouse E1 (apper 250 ng). Before conducting incubations and reactions, the His-tagged mouse E1 is bound to TALON cobalt affinity resin in certain experiments. Beads in 1× reaction buffer are mixed with mouse E1 and 32P-ubiquitin, and they are then incubated similarly to E1 reactions. Samples are resolved by SDS-PAGE after being heated in nonreducing SDS-PAGE sample buffer. Storm PhosphoImager visualizes thioesters that have ubiquitin. |
| References |
[1]. Inhibitors of ubiquitin-activating enzyme (E1), a new class of potential cancer therapeutics. Cancer Res. 2007 Oct 1;67(19):9472-81. [2]. Protein cross-linking as a novel mechanism of action of a ubiquitin-activating enzyme inhibitor with anti-tumor activity. Biochem Pharmacol. 2011 Aug 15;82(4):341-9. |
| Additional Infomation | PYR-41 is an ethyl ester resulting from the formal condensation of the carboxy group of 4-{4-[(5-nitrofuran-2-yl)methylidene]-3,5-dioxopyrazolidin-1-yl}benzoic acid with ethanol. It is an irreversible and cell-permeable inhibitor of ubiquitin-activating enzyme E1. It has a role as an EC 6.2.1.45 (E1 ubiquitin-activating enzyme) inhibitor and an antineoplastic agent. It is an ethyl ester, a member of furans, a C-nitro compound, a member of pyrazolidines and a benzoate ester. |
Solubility Data
| Solubility (In Vitro) | DMSO : 46~74 mg/mL ( 123.89~199.29 mM ) |
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (6.73 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% DMSO + 30% PEG300 + 5% Tween80 + 63% ddH2O: 2mg/ml (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 2.6932 mL | 13.4662 mL | 26.9324 mL | |
| 5 mM | 0.5386 mL | 2.6932 mL | 5.3865 mL | |
| 10 mM | 0.2693 mL | 1.3466 mL | 2.6932 mL |