Background
CORM-401 contains manganese (Mn) as a metal center, CORM-401 can release CO[1].
When 20 μM CORM-401 was added to a PBS solution containing 100 μM Mb, the formation of MbCO can be detected. By calculating the concentration of MbCO over time and by performing curve fitting, it was established that the half-lives of CO release were 14.1, 13.4 and 11.9 min for 5, 10 and 20 μM CORM-401, respectively. The amount of CO released from the compound diminishes as the ratio of CORM-401/Mb increases[2].
After exposure to 20 μM CORM-401 loaded EA.hy926 endothelial cells displayed increased levels of intracellular CO[2]. Upon CORM-401 exposure EROD activity of recombinant Cytochrome P450-dependent monooxygenases (CYP) decreased concentration dependently. Treatment with CORM-401 decreased EROD activity in HepG2 cells at concentrations higher than 50 μM CORM-401 [4]. CORM-401 partially reduced TNF-α/CHX-induced total cellular ROS,and reduced the effect of antimycin-A[5]. CORM-401 (10-100μM) induced a persistent increase in the oxygen consumption rate (OCR) that was accompanied by inhibition of glycolysis (extracellular acidification rate, ECAR) and a decrease in ATP-turnover. Furthermore, CORM-401 increased proton leak, diminished mitochondrial reserve capacity and enhanced non-mitochondrial respiration[6]. CORM-401 caused an apparent suppression of NO production through inhibition of iNOS at both the mRNA and protein levels in RAW264.7 cells stimulated with P. intermedia LPS. CORM-401 upregulated the expression of both the HO-1 gene and its protein in LPS-activated cells, and treatment with the HO-1 inhibitor significantly reversed the attenuating influence of CORM-401 against LPS-induced generation of NO[7].
Oral administration of CORM-401 reduces body weight gain and improves insulin resistance in HFD-induced obesity in mice.The transient uncoupling activity of CO elicited by repetitive administration of CORM-401 is associated with lower weight gain and increased insulin sensitivity during HFD[3].
参考文献:
[1]. Crook SH, Mann BE, et,al. [Mn(CO)4{S2CNMe(CH2CO2H)}], a new water-soluble CO-releasing molecule. Dalton Trans. 2011 Apr 28;40(16):4230-5. doi: 10.1039/c1dt10125k. Epub 2011 Mar 14. PMID: 21403944.
[2]. Fayad-Kobeissi S, Ratovonantenaina J, et,al.Vascular and angiogenic activities of CORM-401, an oxidant-sensitive CO-releasing molecule. Biochem Pharmacol. 2016 Feb 15;102:64-77. doi: 10.1016/j.bcp.2015.12.014. Epub 2015 Dec 22. PMID: 26721585.
[3]. Braud L, Pini M, et,al.Carbon monoxide-induced metabolic switch in adipocytes improves insulin resistance in obese mice. JCI Insight. 2018 Nov 15;3(22):e123485. doi: 10.1172/jci.insight.123485. PMID: 30429365; PMCID: PMC6302946.
[4]. Walter M, Stahl W, et,al. Carbon monoxide releasing molecule 401 (CORM-401) modulates phase I metabolism of xenobiotics. Toxicol In Vitro. 2019 Sep;59:215-220. doi: 10.1016/j.tiv.2019.04.018. Epub 2019 Apr 17. PMID: 31004742.
[5]. Babu D, Leclercq G, et,al.Differential Effects of CORM-2 and CORM-401 in Murine Intestinal Epithelial MODE-K Cells under Oxidative Stress. Front Pharmacol. 2017 Feb 8;8:31. doi: 10.3389/fphar.2017.00031. PMID: 28228725; PMCID: PMC5296622.
[6]. Kaczara P, Motterlini R, et,al.Carbon monoxide released by CORM-401 uncouples mitochondrial respiration and inhibits glycolysis in endothelial cells: A role for mitoBKCa channels. Biochim Biophys Acta. 2015 Oct;1847(10):1297-309. doi: 10.1016/j.bbabio.2015.07.004. Epub 2015 Jul 14. PMID: 26185029.
[7]. Choi EY, Lee JE, et,al. Carbon monoxide-releasing molecule-401, a water-soluble manganese-based metal carbonyl, suppresses Prevotella intermedia lipopolysaccharide-induced production of nitric oxide in murine macrophages. Immunopharmacol Immunotoxicol. 2022 Sep 8:1-8. doi: 10.1080/08923973.2022.2119998. Epub ahead of print. PMID: 36053007.
CORM-401含有锰(Mn)作为金属中心，CORM-401可以释放CO[1]。
当将 20 μM CORM-401 添加到含有 100 μM Mb 的 PBS 溶液中时，可以检测到 MbCO 的形成。通过计算随时间变化的 MbCO 浓度并进行曲线拟合，确定 5、10 和 20 μM CORM-401 的 CO 释放半衰期分别为 14.1、13.4 和 11.9 分钟。随着 CORM-401/Mb 比例的增加，化合物释放的 CO 量减少[2]。
暴露于 20 μM CORM-401 负载的 EA.hy926 内皮细胞后，细胞内 CO[2] 水平升高。在 CORM-401 暴露后，重组细胞色素 P450 依赖性单加氧酶 (CYP) 的 EROD 活性浓度依赖性降低。在浓度高于 50 μM CORM-401 [4] 时，用 CORM-401 处理可降低 HepG2 细胞中的 EROD 活性。 CORM-401部分降低了TNF-α/CHX诱导的总细胞ROS，降低了antimycin-A[5]的作用。 CORM-401 (10-100μM) 诱导耗氧率 (OCR) 持续增加，同时抑制糖酵解（细胞外酸化率，ECAR）和 ATP 周转率降低。此外，CORM-401 增加了质子泄漏，降低了线粒体储备能力并增强了非线粒体呼吸[6]。在用 P. intermedia LPS 刺激的 RAW264.7 细胞中，CORM-401 通过抑制 iNOS 在 mRNA 和蛋白质水平上明显抑制 NO 的产生。 CORM-401 上调 LPS 激活细胞中 HO-1 基因及其蛋白的表达，用 HO-1 抑制剂处理可显着逆转 CORM-401 对 LPS 诱导的 NO 生成的减弱作用[ 7].
口服 CORM-401 可减少体重增加并改善 HFD 诱导的肥胖小鼠的胰岛素抵抗。重复给予 CORM-401 引起的 CO 瞬时解偶联活性与体重增加减少和胰岛素敏感性增加有关高频头[3].
Protocol
| Quantification of CO release using a myoglobin assay [1]: |
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Preparation method
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1 mL horse heart myoglobin solution prepared in phosphate buffered saline at the desired final concentration was transferred to a cuvette and converted to deoxymyoglobin (deoxyMb) by addition of granular sodium dithionite. A deoxyMb spectrum (500 600 nm) was recorded and then a 2-fold excess of CORM-3 was added to the cuvette to obtain a maximum carbonmonoxy-myoglobin (MbCO) spectrum (saturated MbCO). In a separate cuvette, the release of CO from CORM-401 was then measured by adding the compound to the deoxyMb solution overlaid with olive oil to prevent loss of CO or diffusion of atmospheric oxygen back into the solution. Samples were scanned immediately after addition of the compounds and at consecutive time-points to monitor the formation of MbCO. Experiments were conducted either by: (1) using CORM-401 at a final concentration of 10 μM and varying the concentration of myoglobin (25, 50 and 100 μM) or (2) testing the release of CORM-401 at 5, 10 or 20 μM after addition to a fixed concentration of myoglobin (100 μM).
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Applications
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CO released by CORM-401 increases with the amount of Mb used in the assay.
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| Cell experiment [2]: |
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Cell lines
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EA.hy926 endothelial cells
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Preparation Method
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To assess the amount of CO delivered by CO-RMs intracellularly, EA.hy926 endothelial cells in DPBS (+Ca2+, +Mg2+) were treated with 20 μM CORM-401 or iCORM-401, CORM-3 or CORM-A1 for 15 min at 37 °C followed by 30 min incubation with 1 μM COP-1. The intracellular fluorescence, indicative of CO accumulation in cells, was quantified using a CyAn ADP LX7 Analyzer.
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Reaction Conditions
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20 μM CORM-401 for 15 min at 37 °C
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Applications
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COP-1 loaded EA.hy926 endothelial cells displayed increased levels of intracellular CO after exposure to 20 μM CORM-401.
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| Animal experiment [3]: |
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Animal models
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Eight-week-old male C57BL6 mice weighing approximately 25 g
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Preparation Method
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Determined the levels of blood carbonmonoxy hemoglobin (COHb) in mice receiving CORM-401 at 2 different doses after oral gavage.
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Dosage form
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CORM-401 15 mg/kg and 30 mg/kg, given orally 3 times a week.
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Applications
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Oral administration of CORM-401 reduces body weight gain and improves insulin resistance in HFD-induced obesity in mice.
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参考文献:
[1]. Fayad-Kobeissi S, Ratovonantenaina J, et,al. Vascular and angiogenic activities of CORM-401, an oxidant-sensitive CO-releasing molecule. Biochem Pharmacol. 2016 Feb 15;102:64-77. doi: 10.1016/j.bcp.2015.12.014. Epub 2015 Dec 22. PMID: 26721585.
[2]. Braud L, Pini M, et,al. Carbon monoxide-induced metabolic switch in adipocytes improves insulin resistance in obese mice. JCI Insight. 2018 Nov 15;3(22):e123485. doi: 10.1172/jci.insight.123485. PMID: 30429365; PMCID: PMC6302946.
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