Darbufelone
(Synonyms: (5Z)-2-氨基-5-[(4-羟基-3,5-二叔丁基苯基)亚甲基]-1,3-噻唑-4-酮,CI-1004) 目录号 : GC38764
Darbufelone 是细胞 PGF2α 和 LTB4 产生的双重抑制剂。Darbufelone 有效抑制 PGHS-2 (IC50=0.19 μM)，但对 PGHS-1 效力要低得多 (IC50=20 μM)。
Cas No.:139226-28-1
Sample solution is provided at 25 µL, 10mM.
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Quality Control & SDS
- View current batch:
- Purity: >98.00%
- COA (Certificate Of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
Kinase experiment: | The effect of Darbufelone on the cyclooxygenase activity of PGHS-2 is determined in assays with no enzyme-inhibitor preincubation. HoloPGHS-2 (30 nM final concentration) is added to reaction mixtures that contain 20 mM Tris-HCl buffer (pH 7.4), 100 μM TMPD, and varying levels of Arachidonic acid (0-60 μM) and Darbufelone (0-30 μM). The cyclooxygenase activity is measured by monitoring the oxidation of TMPD at 610 nm using a microplate reader[1]. |
Cell experiment: | A549 (CCL-185, lung adenocarcinoma cancer cell line), NCI-H520 (HTB-182, lung squamous cancer cell line) and NCI-H460 (HTB-177, lung large cell cancer cell line) cells are cultured in RPMI-1640, supplemented with 10% (v/v) heat-inactivated fetal bovine serum, 2 mM L-glutamine, 100 U/mL Penicillin G, and 100μg/mL Streptomycin. Cells are grown at 37°C in a humidified atmosphere of 95% air and 5% CO2 and routinely passaged using 0.25% trypsin–EDTA. The effect of Darbufelone on human lung carcinoma cell viability is determined by MTT reduction assay. In brief, tumor cells growing in log-phase are trypsinized and seeded at 5×103 cells per well into 96-well plates and allowed to attach overnight. Medium in each well is replaced with fresh medium or medium containing various concentrations of Darbufelone (5-60 μM) in at least triplicate wells. Cells are cultured to another 72 h. After treatment, 1/10 volume of MTT solution (5 mg/mL) is added to each well, and the plate is incubated at 37°C for another 4 h. Two hundred microliters of DMSO is added to each well to solubilize the MTT-formazan product after removal of the medium. Absorbance at 595 nm is measured with a multi-well spectrophotometer. Growth inhibition is calculated as a percentage of the untreated controls[2]. |
Animal experiment: | Mice[2]C57Bl/6 male mice at 4-5 weeks are used. These mice are housed in air-conditioned quarters and are provided food and water ad libitum. On the day 0, Lewis Lung Carcinoma cells (1×106) are implanted into the left armpit of C57Bl/6 mice. The mice are randomly divided into four treatment groups of ten animals each. The day after inoculation (day 1), control group is treated with CMC-Na, and other groups are administered Darbufelone by gavage at doses of 20, 40, and 80 mg/kg/day. The treatment is continued till the end of the study. On day 14, animals are killed, and tumors are excised, weighed, and fixed in formalin for the further histochemical analysis. |
References: [1]. Johnson AR, et al. Slow-binding inhibition of human prostaglandin endoperoxide synthase-2 with darbufelone, an isoform-selective antiinflammatory di-tert-butyl phenol. Biochemistry. 2001 Jun 26;40(25):7736-45. | |
Darbufelone is a dual inhibitor of cellular PGF2α and LTB4 production. Darbufelone potently inhibits PGHS-2 (IC50= 0.19 μM) but is much less potent with PGHS-1 (IC50=20 μM).
Darbufelone is a noncompetitive inhibitor of PGHS-2 (Ki=10±5 μM). Darbufelone quenches the fluorescence of PGHS-2 at 325 nm (lambda(ex)=280 nm) with Kd=0.98±0.03 μM[1].To test the putative anti-proliferative effect of Darbufelone, A549, H520 and H460 cell lines are used, which are established from three distinct pathological subtypes of NSCLC (adenocarcinoma, squamous and large cell lung cancer respectively). Increasing concentrations of Darbufelone, ranging from 5 to 60 μM, are tested for 72 h. The cell growth inhibition of these three cell lines gradually increases with higher drug concentration. The IC50 of A549 and H520 are 20±3.6 and 21±1.8 μM, respectively, while the H460 has much lower IC50 (15±2.7 μM)[2].
Darbufelone is a dual inhibitor of cellular PGF2R and LTB4 production. Darbufelone is orally active and nonulcerogenic in animal models of inflammation and arthritis[1]. When mice are treated with Darbufelone at dosage of 80 mg/kg/day, the tumor volumes decrease in a time-dependent manner. In contrast, lower dose of Darbufelone (20 or 40 mg/kg/day) dos not show any significant inhibition of tumor weight. At necropsy, the tumor weight in mice treated with Darbufelone (80 mg/kg/day) is reduced by 30.2% in comparison with control group[2].
[1]. Johnson AR, et al. Slow-binding inhibition of human prostaglandin endoperoxide synthase-2 with darbufelone, an isoform-selective antiinflammatory di-tert-butyl phenol. Biochemistry. 2001 Jun 26;40(25):7736-45. [2]. Ye X, et al. Darbufelone, a novel anti-inflammatory drug, induces growth inhibition of lung cancer cells both in vitro and in vivo. Cancer Chemother Pharmacol. 2010 Jul;66(2):277-85.
| Cas No. | 139226-28-1 | SDF | |
| 别名 | (5Z)-2-氨基-5-[(4-羟基-3,5-二叔丁基苯基)亚甲基]-1,3-噻唑-4-酮,CI-1004 | ||
| Canonical SMILES | O=C1N=C(N)S/C1=C\C2=CC(C(C)(C)C)=C(O)C(C(C)(C)C)=C2 | ||
| 分子式 | C18H24N2O2S | 分子量 | 332.46 |
| 溶解度 | Soluble in DMSO | 储存条件 | Store at -20°C |
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1 mg | 5 mg | 10 mg |
| 1 mM | 3.0079 mL | 15.0394 mL | 30.0788 mL |
| 5 mM | 0.6016 mL | 3.0079 mL | 6.0158 mL |
| 10 mM | 0.3008 mL | 1.5039 mL | 3.0079 mL |
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Darbufelone, a novel anti-inflammatory drug, induces growth inhibition of lung cancer cells both in vitro and in vivo
Cancer Chemother Pharmacol 2010 Jul;66(2):277-85.PMID:20352217DOI:10.1007/s00280-009-1161-z.
Purpose: Inflammation plays a crucial role in the development of lung cancer. Accumulated studies have proved that non-steroidal anti-inflammatory drugs (NSAIDs) which block inflammation by their actions on arachidonic acid (AA) metabolism have a potential role in cancer chemotherapy and chemoprevention. The aim of our study was to investigate whether Darbufelone, a novel anti-inflammatory drug, has anticancer effects in lung cancer. Methods: Human non-small cell lung cancer cell lines were treated with Darbufelone at various doses and time points for analysis of cell viability, cell cycle, and apoptosis in vitro. The in vivo effect of Darbufelone was assessed in Lewis lung carcinoma mice model. Results: Darbufelone inhibited the proliferation of non-small cell lung cancer cell lines in a dose-dependent manner, and induced cell cycle arrest at G0/G1 phase through up-regulation of p27 expression. Treatment with Darbufelone also induced apoptosis by activating caspase-3 and caspase-8. Lewis lung carcinoma growth was also significantly inhibited by Darbufelone treatment at daily dose of 80 mg/kg. Conclusions: Taken together, these studies suggested that Darbufelone, an anti-inflammation drug, might represent a novel therapeutic approach for lung cancer treatment.
Slow-binding inhibition of human prostaglandin endoperoxide synthase-2 with Darbufelone, an isoform-selective antiinflammatory di-tert-butyl phenol
Biochemistry 2001 Jun 26;40(25):7736-45.PMID:11412128DOI:10.1021/bi002343f.
The antiinflammatory agent Darbufelone, ((Z)-5-[[3,5-bis(1,1-dimethylethyl)-4-hydroxyphenyl] methylene]-2-imino-4-thiazolidinone, methanesulfonate salt), was discovered as a dual inhibitor of cellular prostaglandin and leukotriene production. To study the mechanism of action of this drug, we expressed human prostaglandin endoperoxide synthase-1 (PGHS-1) and PGHS-2 and purified the recombinant enzymes using buffers that contain octylglucoside. In cyclooxygenase assays following a 15-min incubation of enzyme with inhibitor, Darbufelone potently inhibits PGHS-2 (IC(50) = 0.19 microM) but is much less potent with PGHS-1 (IC(50) = 20 microM). Interestingly, when the assay buffer contains traces of Tween 20 (0.0001%), Darbufelone appears inactive with PGHS-2 due to a detergent interaction that is detectable by absorption spectroscopy. We therefore used octylglucoside, which does not affect Darbufelone in this way, in place of Tween 20 in our PGHS buffers. Inhibition of PGHS-2 with Darbufelone is time dependent: with no preincubation, Darbufelone is a weak inhibitor (IC(50) = 14 microM), but after a 30-min incubation it is 20-fold more potent. Plots of PGHS-2 activity vs preincubation time at various Darbufelone concentrations reach a plateau. This finding is inconsistent with irreversible or one-step slow-binding inhibition. A two-step slow-binding inhibition model is proposed in which the E.I complex (K(i) = 6.2 +/- 1.9 to 14 +/- 1 microM) slowly transforms (k(5) = 0.015-0.030 s(-)(1)) to a tightly bound E.I form with K(i) = 0.63 +/- 0.07 microM and k(6) = 0.0034 s(-)(1). In steady-state kinetics inhibition experiments performed with no preincubation, we find that Darbufelone is a noncompetitive inhibitor of PGHS-2 (K(i) = 10 +/- 5 microM). Darbufelone quenches the fluorescence of PGHS-2 at 325 nm (lambda(ex) = 280 nm) with K(d) = 0.98 +/- 0.03 microM. The PGHS substrate, arachidonate, and various cyclooxygenase inhibitors do not alter this binding affinity of Darbufelone but a structural analogue of Darbufelone competes directly for binding to PGHS-2. Di-tert-butyl phenols such as Darbufelone may inhibit PGHS-2 by exploiting a previously unrecognized binding site on the enzyme.
Dual inhibition of COX-2/5-LOX blocks colon cancer proliferation, migration and invasion in vitro
Oncol Rep 2016 Mar;35(3):1680-8.PMID:26707712DOI:10.3892/or.2015.4506.
Inflammation is emerging as a new hallmark of cancer. Arachidonic acid (AA) metabolism, the family of cyclooxygenases (COXs) and lipoxygenase (LOX) play important roles in AA-related inflammatory cascades. In 94 colorectal cancer samples collected from the Han population, the immunohistochemical results indicated that 68% of the patients with colorectal cancer had a co-expression of both COX-2 and 5-LOX, while both displayed low expression in the matched normal tissues. In cell lines, three colorectal cancer cell lines exhibited high expression of COX-2 and 5-LOX. During stable silencing of the expression of COX-2 or 5-LOX in LoVo cancer cells, we found that downregulation of either COX-2 or 5-LOX significantly diminished the growth, migration and invasion of the colon cancer cells and specifically, downregulation of COX-2 could elicit upregulation of 5-LOX protein and vice versa. The above results suggested that the simultaneous blocking of COX-2 and 5-LOX activity may bring more potential benefits in managing the progression of colon cancer. Therefore, we sought to explore the effectiveness of a dual COX-2/5-LOX inhibitor Darbufelone on the proliferation, migration, invasion and apoptosis of colon cancer cells, as well as the underlying mechanism of action. The results indicated that Darbufelone significantly decreased the proliferative and invasive abilities of the colon cancer cells, in a dose-dependent manner. During the study of the related mechanisms, we found an upregulation of p27 and downregulation of cyclin D1 as well as CDK4 after Darbufelone treatment, which indicated that Darbufelone could arrest the cell cycle of LoVo cells at the G0/G1 phase. Furthermore, the activation of caspase-3 and -9, upregulation of Bax and downregulation of Bcl-2 demonstrated the occurrence of apoptosis by Darbufelone. Finally, Darbufelone also prevented the migration and invasion of LoVo cells, which may be ascribed to the upregulation of E-cadherin and ZO-1. In summary, our data suggest that the inhibition of both COX-2/5-LOX may be an effective therapeutic approach for colon cancer management, particularly for those patients with high expression of COX-2/5-LOX.
Anti-inflammatory and antinociceptive activity profile of a new lead compound - LQFM219
Int Immunopharmacol 2020 Nov;88:106893.PMID:32892073DOI:10.1016/j.intimp.2020.106893.
LQFM219 is a molecule designed from celecoxibe (COX-2 inhibitor) and Darbufelone (inhibitor of COX-2 and 5-LOX) lead compounds through a molecular hybridisation strategy. Therefore, this work aimed to investigate the antinociceptive and anti-inflammatory activities of this new hybrid compound. The acute oral systemic toxicity of LQFM219 was evaluated via the neutral red uptake assay. Acetic acid-induced abdominal writhing and CFA-induced mechanical hyperalgesia were performed to evaluate the antinociceptive activity, and the anti-oedematogenic activity was studied by CFA-induced paw oedema and croton oil-induced ear oedema. Moreover, the acute anti-inflammatory activity was determined by carrageenan-induced pleurisy. In addition, cell migration, myeloperoxidase enzyme activity, and TNF-α and IL-1β levels were determined in pleural exudate. Moreover, a redox assay was conducted using electroanalytical and DPPH methods. The results demonstrated that LQFM219 was classified as GHS category 4, and it showed better free radical scavenger activity compared to BHT. Besides, LQFM219 decreased the number of writhings induced by acetic acid and the response to the mechanical stimulus in the CFA-induced mechanical hyperalgesia test. Furthermore, LQFM219 reduced oedema formation, cell migration, and IL-1β and TNF-α levels in the pleural cavity and inhibited myeloperoxidase enzyme activity. Thus, our study provides that the new pyrazole derivative, LQFM219, demonstrated low toxicity, antinociceptive and anti-inflammatory potential in vitro and in vivo.
Synthesis and biological evaluation of 3-[4-(amino/methylsulfonyl)phenyl]methylene-indolin-2-one derivatives as novel COX-1/2 and 5-LOX inhibitors
Bioorg Med Chem Lett 2010 Dec 15;20(24):7349-53.PMID:21055929DOI:10.1016/j.bmcl.2010.10.056.
Fourteen new 3-[4-(amino/methylsulfonyl)phenyl]methylene-indolin-2-one derivatives were synthesized. Six compounds displayed potent inhibitory activities against COX-1/2 and 5-LOX with IC(50) in the range of 0.10-9.87 μM. Particularly, 10f exhibited well balanced inhibitory action on these enzymes (IC(50)=0.10-0.56 μM). More importantly, 10f and several other compounds had comparable or stronger anti-inflammatory and analgesic activities, but better gastric tolerability in vivo, as compared with Darbufelone mesilate and tenidap sodium. Therefore, our findings may aid in the design of new and safe anti-inflammatory reagents for the intervention of painful inflammatory diseases, such as rheumatoid arthritis at clinic.