Concise synthesis of 2-methoXyestradiol from 17β-estradiol through the C(sp2)-H hydroXylation


A four-step route for the synthesis of 2-methoXyestradiol (5) starting from 17β-estradiol (1) has been achieved with a 51% overall yield. The key step was the ruthenium-catalyzed ortho-C(sp2)-H bond hydroXylation of aryl carbamates. Using dimethyl carbamate as the directing group, [RuCl2(p-cymene)]2 as the catalyst, PhI(OAc)2 as the oXidant and trifluoroacetate/trifluoroacetic anhydride (1:1) as the co-solvent, the hydroXyl group could be singly installed at the 2-position of 3-dimethylcarbamoyloXyestradiol (2) with 65% yield. Subsequent methy- lation of hydroXy and removal of dimethyl carbamate afforded 2-methoXyestradiol (5).

1. Introduction

2-MethoXyestradiol (2-ME, 5), a major estrogen metabolite found in human blood and urine, is the product of the sequential biochemical hydroXylation and methylation of 17β-estradiol (1) by enzymes cyto- chrome 450 and catechol-O-methyl transferase(COMT) mainly in the liver and erythrocytes [1]. Several biological and pharmacological studies have shown that 2-ME turned out to effectively inhibit cancer cell proliferation both in vitro and in vivo [2]. In addition, 2-ME has been demonstrated to act as an anti-angiogenic agent that prevents the growth of new blood vessels around cancer tissues and is currently being investigated in advanced phases of clinical trials for treatment of a variety of cancers [3]. Unlike other biologically active estrogens, 2- ME binds poorly to known estrogen receptors and consequently is well tolerated with some typical hormone-related side effects, which is a remarkable advantage for a potential chemotherapeutic agent [4]. It has also been reported that 2-ME has low oral bioavailability [5]. As of now, the nanocrystal dispersion system (NCD) [6] has significantly improved the oral bioavailability of 2-ME and some of its analogues like 3,17-bis sulfamate estradiol (STX140) [7], the promising anticancer agent (see Fig. 1.).

Some clinical trials have been conducted to evaluate the combina- tion therapy study of 2-ME. For example, combined with TamoXifen to chemotherapy for breast cancer [8] and combined with Epirubicin,Docetaxel, Fluorouracil, Cyclophosphamide, Carboplatin, Bortezomib, etc. can significantly inhibit the proliferation of tumor cells, such as breast cancer, multiple myeloma, urothelial carcinoma, colorectal me- tastases and many others [9]. At the same time, it has made good progress in union radiotherapy [10] and union targeted therapy [11] (Scheme 1).

Over the past decades, different strategies were adopted to in- troduce a suitable group at the 2-position of 17β-estradiol, which can be classified mainly into four types. The first type (Scheme 2. Type I) in- volved formylation or acylation at 2-position via various system or reaction such as MgCl2/Et3N/paraformaldehyde [12], sec-butyllithium/ DMF [13], DMF-POCl3 [14], Fries rearrangement [15] or hexamethy- lenetetramine-acid [16], then Dakin oXidation and subsequent hydro- lysis produced 2-position hydroXy. Structure-activity relationship stu- dies in our laboratories and elsewhere require regular supply of 2-ME and accordingly this strategy is unattractive for us due to its siX or more steps procedure. The second one (Scheme 2. Type II) is to halogenate of C-2, followed by a catalyzed substitution of the halide with methoXide [17]. Xiang Hua [17b] have reported an 84% yield for 2-bromo inter- mediate, however, our attempt affords a much lower yield (less than 30%) and the 2- and 4-bromo isomers are difficult to separate in chromatography. Other halogenation routes also suffer poor selectivity between C-2 and C-4 [17a,c,d]. The third approach (Scheme 2. Type III) focus on the insertion of the 2-hydroXy group via one-pot reaction using superbase sec-butyllithium, along with trimethyl borate and hydrogen peroXide [18]. These type of substitution proceeds with certain diffi- culty and high cost, stringent conditions being required. The last one (Scheme 2. Type IV) is the preferable one that attracts us in particular, a key step being the direct introduction the methoXy group onto the li- thiated C-2 by using cumyl methyl peroXide in the presence of tert- butyllithium [19]. Disappointedly, when the dimethyl sulfate was added into the suspension of cumyl hydroperoXide in DMF, a large powerful tool for catechol synthesis, therein the choice of a readily installable and removable directing group is the key to catalysis. Carbamate was identified as a matched directing group pair for phe- nols. Accordingly, our studies were initiated by investigating the hy- droXylation of phenol dimethyl carbamate 2, which was easily accessed from 17β-estradiol (1) and dimethyl carbamoyl chloride in the presence of K2CO3 in sharp refluXing CH3CN for three hours.

Fig. 1. Structure of 17β-estradiol (1), 2-ME (5) and two synthetic analogs.

2. Experimental
2.1. General procedures

Estradiol was purchased from Wuhan Dongkang Source Technology Co., Ltd., China. Melting points were determined on a WC-1 melting point apparatus and were uncorrected. High-resolution mass spectra were recorded with a Q-TOF micro mass spectrometer. 1H NMR and 13C NMR were respectively recorded on a Bruker AV-400 (400 MHz) nu- clear magnetic resonance spectrometer at 400 and 100 MHz as deut- erated chloroform (CDCl3) or deuterated dimethyl sulfoXide (DMSO-d6) solutions using tetramethyl silane (TMS) as an internal standard (δ = 0). Column chromatography (CC) was carried out on silica gel (200–300 mesh, Qingdao Ocean Chemical Company, China). Thin-layer chromatography (TLC) analyses were carried out on silica gel GF254 (Qingdao Ocean Chemical Company, China) glass plates (2.5 cm × 10 cm with 250 μm layer). TLC visualization was accom- plished by dipping in a solution of 5% phosphomolybdic acid in ethanol and charring at 120 °C. All chemicals were purchased from commercial sources and were used without further purification unless otherwise noted.

2.2. Chemical synthesis
2.2.1. 3-O-[17β-Hydroxy-estra-1,3,5(10)-trien]-yl-N,N- dimethylcarbamate (2) amount of purple smog diffuses immediately in hoods, albeit under ice-

A miXture of the 17β-estradiol (1) (3 g, 11.029 mmol) and bath condition. Thus, the development of a more effective reactions enabling to overcome the above limitations is in high demand.Herein, we report a novel, regioselective and operationally simple procedure for the preparation of 2-methoXyestradiol from 17β-estra- diol, which is outlined in Scheme 3. In this approach, the key step (Scheme 1) is the Ru-catalyzed CeH hydroXylation of aryl carbamates in the trifluoroacetic acid/trifluoroacetic anhydride (TFA/TFAA) system, which was recently published as a protocol for the synthesis of catechol and pyrogallol [20]. This reaction was regioselective and showed no indication of hydroXylation at the C-4 position. The syn- thetic route presented herein reduces the number of steps compared with most of those previously reported. Moreover, this approach is also amendable for large-scale preparations of 5.

With [RuCl2(p-cymene)]2 as the catalyst and PhI(OAc)2 as the oXi- dant, we attempted the hydroXylation of 2 in TFA/TFAA (1:1) at 80 °C for 24 h, in accordance to the reported condition. However, only 20% isolated yield of desired product 3 was observed, as shown in Table 1 (entry 2). When the temperature was increased to 90 °C, the yield was increased significantly to 65% (entry 3). Nevertheless, no improvement in the yield was observed on further raising the temperature (entry 4). Encouraged by promising preliminary results, we start an optimization of the reaction condition with phenol carbamate 2. A variety of catalyst was examined in the reaction, such as [RuCl2(p-cymene)]2, [RuCl2(PPh3)3], [Ru3(CO)12], [RuCl3(H2O)n], Pd2DBA3, Pd(OAc)2, XPhos PdG1 and PdCl2(dppf)CH2Cl2. Among those, [RuCl2(p-cymene)]2 had the highest catalytic activity and gave the best yield (65%, entry 3). In comparison to Pd-catalyst, the overall catalytic activity of Ru-catalyst is better in terms of efficiency. (entry 3, 7–13). Other oXidants including OXone and TBHP were next investigated and no desired product was detected when [RuCl2(p-cymene)]2 was used as the catalyst. (entry 5, 6).

In all attempts, we did not detect any of the 4-substituted isomer of 2 by TLC and 1H NMR-analyses. EXcept for the desired products, no double-hydroXylation products were observed in all case. As a method of C-2 functionalization, this method is superior to halogenation, which gives miXtures of the 2- and 4- monosubstituted halides as well as the 2,4-disubstituted dihalides as crude reaction miXture.
After determining the optimized reaction condition, compound 3 was prepared on a gram-scale reaction. Subsequently, methylation at hydroXyl group of compounds 3 with Me2SO4 produced compound 4, then the removal of directing groups gave compound 1 in high yield. In summary, a simple and straight forward process to access 2-ME (5) has been achieved starting from 17β-estradiol (1) with an overall yield of 51% in four steps. This method has the advantages of high efficiency and experimental simplicity. Notably, this protocol was conducted without the need for air- or moisture- free or drastic reaction conditions and all the steps can be up-scale to industrial process.