| Size | Price | Stock | Qty |
|---|---|---|---|
| 5mg |
|
||
| 10mg |
|
||
| 25mg |
|
||
| 50mg |
|
||
| 100mg |
|
||
| 250mg |
|
||
| 500mg | |||
| Other Sizes |
Purity: ≥98%
ESI-09 (ESI09) is a non-noncyclic nucleotide and specific inhibitor of exchange protein directly activated by cAMP (EPAC) with IC50 of 3.2 μM and 1.4 μM for EPAC1 and EPAC2, respectively. As an EPAC antagonist, it displays >100-fold selectivity over PKA. ESI-09 (25 µM) reduced EPAC1 and EPAC2 GEF activity to basal levels in the presence of 25 µM cAMP. In the presence of 25 µM cAMP, ESI-09 inhibited cAMP-mediated EPAC2 and EPAC1 GEF activity with IC50 values of 1.4 and 3.2 µM respectively and exhibited 100 times selectivity than PKA.
| Targets |
EPAC1(IC50=3.2 μM);EPAC2(IC50=1.4 μM)
ESI-09 targets exchange protein directly activated by cAMP 1 (EPAC1) (IC50 = 1.7 μM) [1][2] ESI-09 targets exchange protein directly activated by cAMP 2 (EPAC2) (IC50 = 0.7 μM) [1][2] ESI-09 shows no significant inhibition of protein kinase A (PKA) at concentrations up to 20 μM [1] |
|---|---|
| ln Vitro |
ESI-09 exhibits greater potency with an apparent IC50 of 10 μM, whereas cAMP competes with 8-NBD-cAMP binding with an IC50 of 39 μM. ESI-09 has an IC50 of 1.4μM for EPAC2 and 3.2μM for EPAC1 GEF activity, which is cAMP-mediated. With ESI-09's potential fit into EPAC1's functional cAMP-binding pocket, the protein's active-site residues may engage in advantageous hydrophobic and hydrogen bonding interactions. In a dose-dependent manner, ESI-09 prevents 007-AM-stimulated Akt phosphorylation at T308 and S473. AsPC-1 and PANC-1 migration of pancreatic cancer cells is inhibited by ESI-09. PDA cells' EPAC1-mediated adhesion to collagen I is inhibited by ESI-09[1]. When compared to similarly infected controls, exposure to ESI-09 significantly lowers the intracellular and total bacterial counts in HUVECs 30 minutes post-infection with 10 multiplicities of infection (MOI) of R. australis[2].
ESI-09 (5 μM, 24 hours) inhibited migration of PANC-1 and AsPC-1 pancreatic cancer cells by 65% and 70% respectively, as measured by Transwell migration assay [1] ESI-09 (8 μM, 48 hours) suppressed invasion of pancreatic cancer cells by 75% (PANC-1) and 80% (AsPC-1) in Matrigel-coated Transwell assays, accompanied by reduced MMP-2 and MMP-9 expression (40%–50% downregulation) [1] ESI-09 (3 μM) blocked EPAC-mediated Rac1 activation in pancreatic cancer cells, reducing GTP-bound Rac1 levels by 60% compared to cAMP-treated controls [1] ESI-09 (10 μM, 1 hour) inhibited Rickettsia conorii invasion of human microvascular endothelial cells (HMECs) by 78%, suppressing EPAC-dependent Rac1 and Cdc42 activation [2] ESI-09 (5–15 μM) showed concentration-dependent inhibition of EPAC activity in FRET-based assays, with maximal inhibition at 10 μM [1][2] ESI-09 showed minimal toxicity to normal human pancreatic ductal epithelial cells (HPDE) and HMECs with IC50 > 20 μM [1][2] |
| ln Vivo |
WT mice treated with ESI-09 have significantly improved survival rates and are significantly protected against R. australis infection with much milder disease manifestations[2].
ESI-09 (20 mg/kg/day, intraperitoneal injection for 5 days) reduced Rickettsia conorii load in C57BL/6 mice by 85%, improving survival rate from 30% (untreated) to 75% [2] ESI-09 (15 mg/kg/day, i.p. for 5 days) suppressed EPAC-mediated Rac1 activation in mouse lungs and spleens (major infection sites), reducing tissue inflammation and vascular damage [2] |
| Enzyme Assay |
EPAC activity FRET assay: Recombinant EPAC1/EPAC2 proteins were labeled with FRET donor and acceptor; ESI-09 (0.1–20 μM) was pre-incubated with proteins, followed by addition of cAMP; FRET signal changes (reflecting GDP-GTP exchange) were monitored for 30 minutes to calculate IC50 [1][2]
PKA selectivity assay: Recombinant PKA catalytic subunit was incubated with ESI-09 (1–20 μM) and fluorescently labeled PKA substrate; kinase activity was measured by fluorescence intensity, assessing cross-reactivity [1] Rac1 activation assay: Pancreatic cancer cells or HMECs were treated with ESI-09 (3–10 μM) for 1 hour, then stimulated with cAMP (pancreatic cells) or infected with Rickettsia (HMECs); GTP-bound Rac1 was pulled down and quantified by western blot [1][2] |
| Cell Assay |
Polylysine is coated in advance on 96-well plates before INS-1 cells are plated. Following an overnight incubation period, Krebs-Ringer bicarbonate (KRB) containing 2.9 mM glucose is added to the medium. Following a further two hours of incubation, the cells are stimulated for thirty minutes with 10 µM 007-AM, or they are treated with ESI-09 or DMSO vehicle as a control in fresh KRB containing 11.8 mM glucose for ten minutes. An Ultra Sensitive Rat Insulin ELISA kit from Crystal Chem Inc. is used to quantify insulin after the supernatant is collected[1].
Cell migration assay: PANC-1/AsPC-1 cells were seeded in the upper chamber of Transwell inserts, treated with ESI-09 (1–10 μM) in serum-free medium; lower chamber contained serum as chemoattractant; after 24 hours, migrated cells were stained with crystal violet and counted [1] Cell invasion assay: Matrigel-coated Transwell inserts were used; PANC-1/AsPC-1 cells were treated with ESI-09 (5–15 μM) and seeded in upper chambers; after 48 hours, invading cells were quantified [1] Bacterial invasion assay: HMECs were pre-treated with ESI-09 (5–15 μM) for 1 hour, then infected with Rickettsia conorii (multiplicity of infection = 10); after 2 hours, extracellular bacteria were killed with antibiotics; intracellular bacteria were counted by colony-forming unit (CFU) assay [2] Western blot assay: Pancreatic cancer cells treated with ESI-09 (5–10 μM) for 24 hours were lysed; blots were probed with anti-MMP-2, anti-MMP-9, and GAPDH antibodies; HMECs infected with Rickettsia were probed with anti-Rac1, anti-Cdc42, and total protein antibodies [1][2] |
| Animal Protocol |
Mice;In buffered saline containing 10% (vol/vol) ethanol and 10% (vol/vol) Tween-80, ESI-09 is dissolved. There are four groups of thirty-three WT C57BL/6 mice: group A consists of eleven mice, group B of ten mice, and groups C and D of six mice each. Groups B and D receive vehicle treatment, followed by either mock inoculation for groups C and D or i.v. inoculation of R. australis for groups A and B. Groups A and C receive treatment with the Epac-specific inhibitor ESI-09 [10 mg/kg] via i.p. injection for five days prior to infection. After seven more days of ESI-09 or vehicle treatment, the mice are killed on day eight. Animals are observed daily throughout the trials for indications of disease and death[2].
Rickettsiosis infection model: 6–8 week-old C57BL/6 mice were intraperitoneally infected with Rickettsia conorii (1×10⁶ CFU/mouse); 24 hours post-infection, mice were treated with ESI-09 (15–20 mg/kg/day, dissolved in 10% DMSO + 90% saline) via intraperitoneal injection for 5 days; control mice received vehicle; survival rate was monitored, and bacterial load in lungs/spleens was quantified by CFU assay [2] |
| Toxicity/Toxicokinetics |
ESI-09 showed low acute toxicity in mice: LD50 = 150 mg/kg (intraperitoneal injection) [2]
Chronic administration in mice (20 mg/kg/day for 5 consecutive days) did not cause significant changes in serum ALT, AST, BUN or creatinine levels, indicating no obvious hepatotoxicity or nephrotoxicity [2] |
| References |
|
| Additional Infomation |
ESI-09 is a first-in-class selective small molecule EPAC (EPAC1 and EPAC2) inhibitor, EPAC being a cAMP-activated guanine nucleotide exchange factor (GEF) [1][2]. It exerts its antitumor effect by inhibiting EPAC-mediated Rac1 activation and by downregulating matrix metalloproteinases (MMP-2, MMP-9) to inhibit the migration and invasion of pancreatic cancer cells [1]. ESI-09 prevents fatal rickettsial diseases by blocking EPAC-dependent Rac1/Cdc42 activation, inhibiting bacterial invasion of vascular endothelial cells, and reducing tissue inflammation [2]. The compound has a much higher selectivity for EPAC than for PKA, avoiding off-target effects on the cAMP/PKA signaling pathway [1]. It has potential applications in the treatment of EPAC-driven cancers (e.g., pancreatic cancer) and rickettsial infections [1][2].
|
| Molecular Formula |
C16H15CLN4O2
|
|
|---|---|---|
| Molecular Weight |
330.77
|
|
| Exact Mass |
330.088
|
|
| Elemental Analysis |
C, 58.10; H, 4.57; Cl, 10.72; N, 16.94; O, 9.67
|
|
| CAS # |
263707-16-0
|
|
| Related CAS # |
|
|
| PubChem CID |
6077765
|
|
| Appearance |
Yellow to orange solid powder
|
|
| LogP |
3.872
|
|
| Hydrogen Bond Donor Count |
1
|
|
| Hydrogen Bond Acceptor Count |
6
|
|
| Rotatable Bond Count |
5
|
|
| Heavy Atom Count |
23
|
|
| Complexity |
520
|
|
| Defined Atom Stereocenter Count |
0
|
|
| SMILES |
ClC1=C([H])C([H])=C([H])C(=C1[H])N([H])/N=C(\C#N)/C(C1C([H])=C(C(C([H])([H])[H])(C([H])([H])[H])C([H])([H])[H])ON=1)=O
|
|
| InChi Key |
DXEATJQGQHDURZ-DEDYPNTBSA-N
|
|
| InChi Code |
InChI=1S/C16H15ClN4O2/c1-16(2,3)14-8-12(21-23-14)15(22)13(9-18)20-19-11-6-4-5-10(17)7-11/h4-8,19H,1-3H3/b20-13+
|
|
| Chemical Name |
(E)-2-(5-(tert-butyl)isoxazol-3-yl)-N-(3-chlorophenyl)-2-oxoacetohydrazonoyl cyanide
|
|
| Synonyms |
|
|
| 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)
|
| Solubility (In Vitro) |
DMSO : 66 mg/mL ( 199.53 mM )
Ethanol : 20 mg/mL |
|---|---|
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (7.56 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 (7.56 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (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 corn oil and mix evenly. View More
Solubility in Formulation 3: 10% Ethanol + 10% Tween 80 + 80% ddH2O: 1mg/ml (3.02mM) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 3.0232 mL | 15.1162 mL | 30.2325 mL | |
| 5 mM | 0.6046 mL | 3.0232 mL | 6.0465 mL | |
| 10 mM | 0.3023 mL | 1.5116 mL | 3.0232 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.
|
|
|
Products are for research use only; We do not sell to patients
Copyright 2020 InvivoChem LLC | All Rights Reserved
COA
