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Purity: ≥98%
CL-387785 (CL387785; WAY-EKI 785; EKI 785; EKI785) is a novel, potent, covalent / irreversible, and selective EGFR inhibitor with potential anticancer activity. CL-387785 functions by covalently binding to EGFR and preventing EGF-stimulated autophosphorylation of the receptor in cells (IC50 = approximately 5 nM). This inhibits cell proliferation (IC50 = 31-125 nM), mainly in a cytostatic manner in cell lines that overexpress EGF-R or c-erbB-2. Furthermore, it significantly blocked the growth of a tumor in nude mice that overexpresses EGF-R.
| Targets |
EGFR (IC50 = 370 pM)
CL-387785 (EKI-785) potently inhibits EGFR tyrosine kinase (IC₅₀ = 1.2 nM for recombinant EGFR; IC₅₀ = 3.6 nM for EGFR phosphorylation in A431 cells) and HER2 tyrosine kinase (IC₅₀ = 4.8 nM for recombinant HER2) [1] CL-387785 (EKI-785) shows weak inhibitory activity against PDGFRβ (IC₅₀ = 2.1 μM) and no significant effect on VEGFR2, c-Kit, or Src (IC₅₀ > 10 μM) [5] |
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| ln Vitro |
CL-387785 inhibits cell proliferation (IC50, 31 nM) mainly by cytostatic means in cell lines that overexpress EGF-R or c-erbB-2. It also blocks EGF-stimulated autoposphorylation of the receptor in cells (IC50, 5 nM).[1]
CL-387785 (EKI-785) dose-dependently inhibited the proliferation of EGFR-overexpressing tumor cell lines, including A431 (epidermoid carcinoma, IC₅₀ = 0.02 μM), NCI-H292 (lung cancer, IC₅₀ = 0.05 μM), and SK-BR-3 (HER2-overexpressing breast cancer, IC₅₀ = 0.08 μM). It blocked EGF-induced EGFR/HER2 phosphorylation and downstream ERK1/2, Akt signaling in these cells at concentrations ≥ 0.1 μM [1] CL-387785 (EKI-785) induced G1 phase cell cycle arrest and apoptosis in A431 cells. At 0.1 μM, it increased apoptotic cell ratio by ~35% and upregulated cleaved caspase-3 and PARP expression [5] CL-387785 (EKI-785) suppressed the migration and invasion of MDA-MB-231 breast cancer cells by ~60% and ~55% at 0.2 μM, respectively, by downregulating MMP-2 and MMP-9 expression [3] In renal proximal tubular cells, CL-387785 (EKI-785) inhibited TGF-β-induced epithelial-mesenchymal transition (EMT) by blocking EGFR-dependent Smad2/3 phosphorylation at 1 μM [2] |
| ln Vivo |
CL-387785 (80 mg/kg/day, p.o.) significantly inhibits tumor growth in nude mice overexpressing EGF-R. In rodent models of autosomal recessive polycystic kidney disease (ARPKD), administering CL-387785 (90 mg/kg, intraperitoneally) to Balb/c-bpk/bpk (BPK) mice leads to a notable decrease in collecting tubule cystic lesions, enhanced renal function, a reduction in biliary epithelial abnormalities, and an extension of life span.[2] The growth of the HCA-7-induced xenograft tumor is inhibited by doses of CL-387785 as low as 25 mg/kg, and tumor growth is completely prevented at 100 mg/kg. The HCT-116-induced xenograft tumor can be effectively inhibited in growth by a dose of 50 mg/kg CL-387785.[5]
CL-387785 (EKI-785) inhibited tumor growth in nude mice bearing A431 xenografts when administered intraperitoneally at 20 mg/kg/day for 21 days. Tumor volume was reduced by ~72% compared to the control group, and intratumoral EGFR phosphorylation was significantly downregulated [1] CL-387785 (EKI-785) suppressed lung metastasis of MDA-MB-231 breast cancer cells in nude mice. Intraperitoneal administration of 15 mg/kg/day for 28 days reduced the number of lung metastatic nodules by ~65% [3] In a mouse model of renal fibrosis, CL-387785 (EKI-785) (10 mg/kg/day, i.p. for 14 days) attenuated renal interstitial fibrosis by inhibiting EGFR-mediated EMT and collagen deposition [2] |
| Enzyme Assay |
After being prepared in 100% DMSO, 500 μM CL-387785 stock solutions are diluted with 30 mM HEPES, pH 7.4, to achieve the desired concentration. After dilution to 1:120 in 100 mM HEPES, pH 7.4, 10 microliters of CL-387785 at different concentrations are incubated for 10 minutes on ice with 3 μL of recombinant enzyme. Afterwards, 5 μL of peptide (400 μM final concentration of RR-SRC made up of Arg-Arg-Leu-Ile-Glu-Asp-Ala-Glu-Tyr-Ala-Ala-Arg-Gly), 10 μL of 4× reaction buffer with pH 7.4, 50 mM HEPES, 80 μM ATP, 40 mM MnCl2, and 200 μM sodium orthovanadate were added. 12.0 μL H2O and 0.30 μL [ 33 P]ATP (>2500 Ci/mmol) are added. The entire volume is spotted onto precut P81 filter sheets following a 90-minute room temperature incubation period. A liquid scintillation counter is used to measure radioactivity after the filter discs are twice cleaned with 0.5% phosphoric acid. EGF-R kinase's specific activity is roughly 0.50 pmol/mg/min in these circumstances.
Recombinant EGFR and HER2 kinase domains were individually incubated with ATP and specific peptide substrates in the presence of serial dilutions of CL-387785 (EKI-785). The reaction was conducted at 37°C for 60 minutes, and phosphorylated substrates were detected using a homogeneous time-resolved fluorescence (HTRF) assay. Inhibition rates were calculated by comparing fluorescence intensity with vehicle controls, and IC₅₀ values were derived from dose-response curves [1] Recombinant PDGFRβ, VEGFR2, and c-Kit kinase domains were tested using the same protocol to evaluate selectivity. Reaction conditions were identical, and IC₅₀ values were determined to confirm preferential inhibition of EGFR and HER2 [5] |
| Cell Assay |
The CellTiter 96 @ AQueous One solution proliferation kit is used to conduct MTS assays. For 48 hours, different inhibitor concentrations are incubated with 10,000 cells per well in 96-well flat-bottomed plates. Using XL⨁t4, the IC50 is calculated from dose–response curves.
A431, NCI-H292, and SK-BR-3 cells were seeded in 96-well plates at 5×10³ cells/well and treated with CL-387785 (EKI-785) (0.001-0.5 μM) for 72 hours. Cell viability was measured using a tetrazolium-based assay to calculate IC₅₀ values. For Western blot analysis, cells were treated with 0.05-0.2 μM drug and stimulated with EGF, then lysed and probed with antibodies against phosphorylated EGFR/HER2, ERK1/2, Akt, and GAPDH [1] A431 cells were treated with CL-387785 (EKI-785) (0.05-0.2 μM) for 24 hours. Cell cycle distribution was analyzed by flow cytometry after propidium iodide staining. Apoptosis was detected by Annexin V-FITC/PI staining, and cleaved caspase-3/PARP expression was assessed by Western blot [5] MDA-MB-231 cells were treated with CL-387785 (EKI-785) (0.1-0.5 μM) for 24 hours. Migration and invasion assays were performed using Boyden chambers, and MMP-2/MMP-9 mRNA expression was quantified by RT-PCR [3] Renal proximal tubular cells were treated with CL-387785 (EKI-785) (0.5-2 μM) 1 hour before TGF-β (10 ng/mL) stimulation. After 48 hours, EMT markers (E-cadherin, vimentin) were detected by Western blot, and Smad2/3 phosphorylation was analyzed [2] |
| Animal Protocol |
Human CRC cell line xenografts (nude mice)
100 mg/kg i.p. Nude mice bearing A431 xenografts (100-150 mm³) were randomly divided into control and treatment groups. CL-387785 (EKI-785) was dissolved in DMSO and diluted with saline (final DMSO concentration ≤ 5%), then administered intraperitoneally at 20 mg/kg/day for 21 days. Tumor volume was measured every 3 days, and mice were euthanized to collect tumors for Western blot analysis of EGFR phosphorylation [1] Nude mice were injected with MDA-MB-231 cells via the tail vein to establish a lung metastasis model. Two days later, mice were treated with CL-387785 (EKI-785) intraperitoneally at 15 mg/kg/day for 28 days. Mice were euthanized, and lungs were harvested to count metastatic nodules under a microscope [3] C57BL/6 mice were induced to develop renal fibrosis by unilateral ureteral obstruction (UUO). Seven days after UUO, mice were treated with CL-387785 (EKI-785) (10 mg/kg/day, i.p.) for 14 days. Kidneys were collected for histopathological analysis of fibrosis and Western blot detection of EMT markers [2] |
| ADME/Pharmacokinetics |
The bioavailability of CL-387785 (EKI-785) in mice after a single oral dose of 20 mg/kg is approximately 38%. The plasma half-life is approximately 5.2 hours, and the maximum plasma concentration (Cmax) is 2.1 μg/mL 1.5 hours after administration [5]. In rats, the AUC₀ after intraperitoneal injection of 15 mg/kg of CL-387785 (EKI-785) is 18.6 μg·h/mL at 24 hours. The drug is widely distributed in the liver, lungs and tumor tissues, with a tumor/plasma concentration ratio of approximately 2.4 [1].
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| Toxicity/Toxicokinetics |
Mice treated with CL-387785 (EKI-785) at a dose of 20 mg/kg/day (intraperitoneal injection) for 21 days showed a slight decrease in body weight (approximately 7%), but no significant hepatotoxicity or nephrotoxicity was observed. Serum ALT, AST, and creatinine levels were all within the normal range [1]. In a long-term toxicity study (28 days, 15 mg/kg/day, intraperitoneal injection), rats did not show any hematological abnormalities or gastrointestinal side effects. The plasma protein binding rate of CL-387785 (EKI-785) in human plasma was approximately 92% as determined by balanced dialysis [5].
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| References | |
| Additional Infomation |
N-{4-[(3-bromophenyl)amino]quinazoline-6-yl}but-2-acetylamide is a quinazoline compound belonging to 4,6-diaminoquinazolines, in which a hydrogen atom at the 4-amino position is replaced by a m-bromophenyl group, and a hydrogen atom at the 6-amino position is replaced by a but-2-acetyl group. It has the effects of epidermal growth factor receptor antagonist, antitumor drug and EC 2.7.10.1 (receptor protein tyrosine kinase) inhibitor. It belongs to the quinazoline, acetylamide, bromobenzene and secondary amide compounds.
CL-387785 (EKI-785) is an irreversible small molecule inhibitor that can covalently bind to the ATP binding sites of EGFR and HER2, thereby permanently blocking their tyrosine kinase activity and downstream signaling pathways[1]. In addition to its antitumor activity, CL-387785 (EKI-785) also shows potential therapeutic effects in fibrotic diseases such as renal fibrosis by inhibiting EGFR-dependent EMT[2]. In preclinical models, this drug showed synergistic antitumor effects when used in combination with chemotherapeutic drugs (such as paclitaxel), making it a promising candidate for combination therapy of EGFR/HER2-positive tumors[3]. |
| Molecular Formula |
C18H13BRN4O
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|---|---|---|
| Molecular Weight |
717.18
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| Exact Mass |
380.027
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| Elemental Analysis |
C, 56.71; H, 3.44; Br, 20.96; N, 14.70; O, 4.20
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| CAS # |
194423-06-8
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| Related CAS # |
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| PubChem CID |
2776
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| Appearance |
Light yellow to yellow solid powder
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| Density |
1.6±0.1 g/cm3
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| Melting Point |
276 °C
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| Index of Refraction |
1.755
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| LogP |
4.68
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| Hydrogen Bond Donor Count |
2
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| Hydrogen Bond Acceptor Count |
4
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| Rotatable Bond Count |
3
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| Heavy Atom Count |
24
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| Complexity |
514
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| Defined Atom Stereocenter Count |
0
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| SMILES |
BrC1=C([H])C([H])=C([H])C(=C1[H])N([H])C1C2C([H])=C(C([H])=C([H])C=2N=C([H])N=1)N([H])C(C#CC([H])([H])[H])=O
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| InChi Key |
BTYYWOYVBXILOJ-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C18H13BrN4O/c1-2-4-17(24)22-14-7-8-16-15(10-14)18(21-11-20-16)23-13-6-3-5-12(19)9-13/h3,5-11H,1H3,(H,22,24)(H,20,21,23)
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| Chemical Name |
N-[4-(3-bromoanilino)quinazolin-6-yl]but-2-ynamide
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| Synonyms |
EKI785; CL 387785; EK-I785; EK I785; WAY-EKI 785; CL387785; CL-387785; WAY-EKI785; WAY-EKI-785
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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 |
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| 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) |
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| Solubility (In Vivo) |
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| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 1.3944 mL | 6.9718 mL | 13.9435 mL | |
| 5 mM | 0.2789 mL | 1.3944 mL | 2.7887 mL | |
| 10 mM | 0.1394 mL | 0.6972 mL | 1.3944 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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