生物活性

2-Methoxyestradiol(2-MeOE2; 2ME2; 2ME; NSC-659853), a naturalmetabolite of estradiol, possess anti-angiogenic and anti-cancer effectswithout any undesirable estrogen activity. 2-Methoxyestradiol is an endogenous estrogen metabolite with low affinity for the estrogen receptor but which disrupts microtubule function. Apoptotic, antiproliferative and antiangiogenic agent, in vitro and in vivo; acts via an estrogen receptor-independent mechanism. Induces p53-induced apoptosis via two pathways: activation of p38 and NF-κB; and activation of JNK and AP-1 leading to Bcl-2 phosphorylation. Also upregulates death receptor 5 and binds to tubulin, inhibiting its assembly. 2-Methoxyestradiol is formed by CYP450-mediated hydroxylation followed by catechol-O-methyltransferase (COMT) methylation of estradiol. 2-Methoxyestradiol is a derivative of an oestrogenic steroidal hormone and has demonstrated: potent inhibition of endothelial cell proliferation and migration and inhibition of bFGF and VEGF-induced corneal neovascularization in mice. In MDA-MB-231 cells, 2-Methoxyestradiol inhibits HIF mediated transcriptional activation of target genes without affecting the transcription of HIF-1α itself. 2-Methoxyestradiol has been shown to induce apoptosis through activation of the p53 pathway through either activation of p38 and NF-κB or JNK and AP-1 both leading to Bcl-2 phsophorylation.

2MEinhibits tubulin assembly with an IC50 of 2.2μM.[1]

2MEinduces microtubule depolymerization with an EC50 of7.5μM in A-10 cells.[2]

2MEarrests mitosis with an IC50 of 1.2μM in MCF7 cells.[3]

2MEinduces bovine pulmonary artery endothelial cells (BPAEC) apoptosis with an EC50of 0.45±0.09μM.[4]

2MEinhibits BPAEC migration with an IC50 of 0.71 ±0.11μM.[4]

2MEinhibits angiogenesis with an IC50 of 3.2μM.[5]

Cdc25phosphatases inhibition of 2-methoxyestradiol[6]

Anti-proliferationof 2-methoxyestradiol

Cytotoxic effects of 2-methoxyestradiol

体外研究

体内研究

激酶实验

Inhibition of tubulin assembly[1]

Tubulin was isolated from porcine brain andstored at –78 ℃. Samples were prepared directly in a 96-wellmicrotitre testplate that was preincubated at 4^oC in the fridge for30 min and contained Mes buffer [128μl (0.1M Mes, 1mM EGTA, 0.5mMMgCl₂, distilled water, pH 6.6)], GTP (20μl, 5mM in Mesbuffer), tubulin (50μl, 11mg/ml in Mes buffer) and the candidate drug (20μl,Csample in DMSO). The tubulin/drug samples were immediately placed in a 96-wellplate reader, alongside blank samples containing Mes buffer (198μl)and the analogs (10μl, same concentration). The absorbance (λ=350 nm) wasrecorded at 25 ℃temperature for a period of 60min, and the resultswere compared to untreated controls to evaluate the relative degree of changein optical density.

细胞实验

Cancer cell growth inhibition (cytotoxicity)[5]

K562 human chronic myelogeneous leukemiacells were cultivated in RMPI medium, free of antibiotics and containing2-mercaptoethanol (2μM) and L-glutamine (2mM), supplemented with fetal calf serum (FCS)(10% v/v). The cells were adjusted to a concentration depending on their observeddoubling time (ca. 40,000 cells/ml), in RPMI medium supplemented with FCS (10%v/v). The test compound was dissolved in DMSO. A solution of 100 ml in mediumwas added to 100 ml of cell solution (40,000cells/ml) in a 96-well microtitretestplate (4 ml of the compound solution diluted in medium in order to reachdecreasing concentrations). This series of dilutions was continued to affordsamples at different concentrations leaving one cell solution free of drugacting as a control. The plates were incubated at 37^oC (5% CO₂ in air) for 5 days. The plate was then removed from the incubator and 50 ml ofa solution of MTT (3mg/ml in PBS) was added to each well. After incubation (37^oC,5% CO₂ in air, 3h) the medium was carefully removed from each wellby suction and the resulting formazan precipitate re-dissolved in 200 ml DMSO.The optical density of each well was read at two wavelengths (λ540and 690 nm) using a Titretek Multiscan MCC/340 plate reader. After processingand analysis through the application of an ‘in-house’ software package, theresults enabled the calculation of the drug dose required to inhibit cellgrowth by 50% (IC50 value), determined by graphical means as percentage of the controlgrowth.

动物实验

In vivo treatment of transplantable liver tumors[6]

Fischer (F344 male) rat HCC-derived JM-1cell line was grown in MEM + 10%FBS culture medium up tosemi-conXuency. Cellswere harvested by trypsinization (0.2%) for 5 min at 37 °C, washed with ice-coldPBS, centrifuged (500g, 10 min), and resuspended in ice-cold PBS at aconcentration of 10⁶ cells/ml. A quantity of 1 ml of the cellsuspension containing 10⁶ cells was injected into a Fischer ratliver through its mesenteric vein. Tumor cell transplanted rat were dividedinto two groups (three rats per group). One group was treated intraperitoneallywith 10 mg/kg body weight of 2-ME dissolved in DMSO, every other day, for atotal of Wve injections. Treatment was started the day after cell transplantation.The other group was treated similarly with the solvent DMSO as control. Animalswere killed 2 weeks after cell transplantation and the numbers of tumor focivisible were counter per liver. The diameter of the tumors was also measured bya slide calipers.

不同实验动物依据体表面积的等效剂量转换表（数据来源于FDA指南）

例如，已知某工具药用于小鼠的剂量为88 mg/kg , 则用于大鼠的剂量换算方法：将88 mg/kg 乘以小鼠的Km系数（3），再除以大鼠的Km系数（6），得到该药物用于大鼠的等效剂量44 mg/kg。

参考文献

[1] Solum EJ, Vik A, Hansen TV. Synthesis, cytotoxic effects and tubulin polymerization inhibition of 1,4-disubstituted 1,2,3-triazole analogs of 2-methoxyestradiol. Steroids. 2014;87:46-53.
[2] Rao PN CJ, Tinley TL, Mooberry SL. Synthesis and antimitotic activity of novel 2-methoxyestradiol analogs. Steroids. 2002;67(13-14):1079-1089.
[3] Kamath K, Okouneva T, Larson G, Panda D, Wilson L, Jordan MA. 2-Methoxyestradiol suppresses microtubule dynamics and arrests mitosis without depolymerizing microtubules. Mol Cancer Ther. 2006;5(9):2225-2233.
[more]

体内溶解度
5 mg/mL

Clinical Trial Information ( data from http://clinicaltrials.gov )

注：以上所有数据均来自公开文献，并不保证对所有实验均有效，数据仅供参考。

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