Citation

Peng Wang, Fei Liu, Qiu Zhong, Shi-Long Zheng, Yue Chen, Guang-Di Wang, Ling He. Chemoselective synthesis and cytotoxic activity of a series of novel benzo[1, 4]oxazin-3-one derivatives[J]. Chin. Chem. Lett., 2017, 28(6): 1243-1247  

Peng Wang^a, Fei Liu^a, Qiu Zhong^b,c, Shi-Long Zheng^b,c, Yue Chen^d, Guang-Di Wang^b,c, Ling He^a    

The construction of nitrogen-containing heterocycles catalyzed by transition metal complexes is a fascinating methodology and has been applied to the synthesis of many natural products and biologically active compounds [1-10]. Copper complexes are especially attractive catalysts due to their low cost, readily availability and versatile reactivity [11-34]. In our continuing effort to explore the application of transition metal as catalyst in synthetic methodology for discovering bioactive molecules [37-40]. We investigated the amidation reactions of arenes and found that [Cu(CH₃CN)₄]ClO₄ can synergistically catalyze the intramolecular functionalization of arenes to afford the corresponding 1, 4-benzoxazin-3-ones in moderate to high yield by C—H bond activation and concomitant amidation.

To the best of our knowledge, the construction of C—N bonds by oxidative amidation via insertion into C—H bonds with transition metal complexes is a significant approach [11-19]. Insertion requires activated C—N bonds (e.g., allylic or benzylic C—N bonds), and insertion and coupling with inactive benzene C—N bonds remain a formidable task. Although a few of studies have demonstrated that the intramolecular catalytic amidation reactions of C—H bonds are feasible [20-34], the intramolecular catalytic reactions of oxidatively constructing the C-N bonds between N—H and inactive C—N bonds have rarely been studied. Only a few studies have identified such a biphenyl compound with an amide, an aniline or a sulfonamide as potential nitrogen sources and substrates for this kind reactions to afford five-membered nitrogen-containing heterocyclic products or six-membered N-MeO, N-phenyl and N-alkyl substituted nitrogen-containing heterocyclic products, and the reaction conditions are harsh and the catalysts are expensive and not easily available [20-24].

2H-1, 4-Benzoxazin-3-(4H)-one derivatives are now known to possess useful biological and medicinal activities [36, 37]. The reported synthetic methods of 2H-1, 4-benzoxazin-3-(4H)-ones are all using 2-aminophenols, substituted 2-nitrophenols, or 2-halophenols as precursors [35-37], their disadvantage is that only a limited number of substrates can be employed or the reaction conditions are not mild. With respect to these considerations, we are thinking to develop a more convenient and novel method through arene oxidation and C-N bond formation to produce 2H-1, 4-benzoxazin-3-(4H)-ones. In this paper, we describe a mild and efficient synthesis of 2H-1, 4-benzoxazin-3-(4H)-one derivatives by the oxidative intramolecular amidation between inactive C—N and N—H functionalization from N-(1, 3-diphenyl-1H-1, 2, 4-triazol-5-yl)-2-phenoxyacetamides as a nitrogen source (Scheme 1).

Scheme1

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Scheme1. Synthetic procedure for benzo[1, 4]oxazin-3-one derivatives with a selective oxidative amidation.

N-(1, 3-Diphenyl-1H-1, 2, 4-triazol-5-yl)-2-phenoxyacetamide (1a) was chosen as a model substrate for optimizing the reaction conditions. Firstly, oxidants and catalysts were investigated to develop a mild and environmentally friendly methodology to accomplish the intramolecular cyclization reactions. The effects of the tested oxidants and catalysts are listed in Table 1.

Table 1

Table 1 Effect of catalysts and oxidants on the amidation reaction.a

Table 1 Effect of catalysts and oxidants on the amidation reaction.a

The results of entries 1-8 in Table 1 show that PhI(OAc)₂, PhI (CF₃CO₂)₂, and PhI = O (Table 1, entries 1, 2, and 3) are active oxidants for the oxidative amidation of 1a to form 2a with Cu (CF₃SO₃)₂ catalyst. Next, using PhI(CF₃CO₂)₂ as oxidant, both Cu-ophenanthroline and Cu[(CH₃CN)₄]ClO₄ (Table 1, entries 17 and 18) display as the best catalysts to transform 1a to 2a by C—N bond formation. Cu[(CH₃CN)₄]ClO₄ is much less expensive than Cu-ophenanthroline, so we chosed Cu[(CH₃CN)₄]ClO₄ as catalyst and PhI(CF₃CO₂)₂ as oxidant in further condition optimization for the intramolecular amidation reaction. From the entry 19, we can find that the yield reaches to 66% from 60% (entry 18) by increasing the Cu[(CH₃CN)₄] ClO₄ loading from 10 to 20% molar equivalent to 1a. However, the yield maintains 61% when the catalyst was loaded to 25% molar of 1a (entry 20). Thus, 4-(1, 3-diphenyl-1H-1, 2, 4-triazol-5-yl)-2H-benzo[b][1, 4]oxazin-3 (4H)-one (2a) was obtained in the best yield of 66% when using N-(1, 3-diphenyl-1H-1, 2, 4-triazol-5-yl)-2-phenoxyacetamide (1a) as the substrate at room temperature with a molar ratio of substrate/PhI(CF₃CO₂)₂/Cu [(CH₃CN)₄] ClO₄ of 1:2.5:0.2. The solvent effects of this reaction was also explored. Among CH₂Cl₂, ClCH₂CH₂Cl, CH₃CN, CHCl₃, THF, and DMF, halogenated hydrocarbon solvents provided good yields under identical experimental conditions, and the best one was CH₂Cl₂ which provided the highest yield (Table 1, entry 19). Finally, to test the effect of temperature on the yield, identical experimental conditions were utilized while varying the temperature from 0 ℃ to 60 ℃. The optimal yield and shortest reaction time was achieved at room temperature. Therefore, the optimized conditions for this intramolecular oxidative amidation were as follows: substrate, 1a (1.0 equiv.); oxidant, PhI(CF₃CO₂)₂ (2.5 equiv.); catalyst, Cu[(CH₃CN)₄]ClO₄ (20 mol%); solvent, CH₂Cl₂; and temperature, room temperature.

We further investigated the scope of this method and performed intramolecular oxidative amidation reactions using different arenes under the above optimized conditions. As showed in Fig. 1, various substrates were subjected to intramolecular oxidative amidation, affording corresponding N-1H-1, 2, 4-triazol-2H-1, 4-benzoxazin-3-(4H)-ones in moderate to good yields. It is worth mentioning that, our substrates have three aromatic rings (Scheme 1), which bear different substituent.

Figure 1

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Fig. 1. Intramolecular oxidative amidation catalyzed by Cu[(CH3CN)4] ClO4. All reactions were performed at room temperature, CH2Cl2 as the solvent for 4-10 h with a molar ratio of substrate oxidant/catalyst of 1:2.5:0.2. The reaction was conducted at 0 ℃ for 5 h. (2c and 2c').

Interestingly, only the cyclization products on ring C were obtained in the amidation reaction, no cyclizationproducts on ring B were found (Scheme 1). The reason of chemoselective formation of N-1H-1, 2, 4-triazol-2H-1, 4-benzoxazin-3-(4H)-ones (2a-2r) from N-(1, 3-diphenyl-1H-1, 2, 4-triazol-5-yl)-2-phenoxy-acetamide derivative-s may be that the amidation reaction favors the formation of more stable N-1H-1, 2, 4-triazol-2H-1, 4-benzoxazin-3-(4H)-ones which could be enolated to fully conjugated structure. In Fig. 1, the substrate with methoxyl in the meta-position of ring C (1c) gave two corresponding six-membered heterocyclic products 2c and 2c', the ratio of 2c and 2c' reflects the regioselectivity difference in the reaction which arises from steric and electronic effect in the formation of products. At the same time, we notice that the reaction of the methoxy group substituent in the para-position gave only corresponding five-membered triazole-spirodienone product in 94% yield (Scheme 2).

Scheme2

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Scheme2. The reaction results of p-methoxy substituted aromatic ether derivatives as substrates.

On the basis of the mechanisms proposed originally for the amidation by Antonchick [30], Zhao [41] and Himo [42], we proposed a mechanism for oxidative amidation reactions of arenes with N-triazoleamides (Scheme 3). Initially ditrifluoroacetoxyiodobenzene reacts with the amide to give intermediate A, which is then transformed into intermediate B through an oxidative insertion of N—I bond by copper(Ⅱ). The reductive elimination of intermediate C releases copper (Ⅰ) salt, trifluoroacetic acid and iodobenzene, leading to final product 4-(2, 5-diphenyl-2H-1, 2, 4-triazol-3-yl)-2H benzo[b] [1, 4] oxazin-3(4H)-one.

Scheme 3

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Scheme 3. Proposed mechanism for the oxidative amidation of arenes.

Some of benzo[1, 4]oxazin-3-one derivatives were examined for in vitro cytotoxic activities against human breast cancer cell line MDA-MB-231, and cervical cancer cell line HeLa. Table 2 lists the IC₅₀ values obtained with these compounds. Most compounds showed moderate antiproliferative effects. They displayed IC₅₀ values in the micromolar concentration range against MDA-MB-231 and HeLa cells. These biological data indicate that the benzo [1, 4]oxazin-3-ones can be useful templates upon which to develop potential small molecule anti-cancer agents. Further investigation of the scope, mechanism and synthetic application of the intramolecular oxidative amidation is currently underway.

Table 2

Table 2 Antiproliferation activity against cancer cell lines of some compounds.

Table 2 Antiproliferation activity against cancer cell lines of some compounds.

In summary, we developed a novel protocol for the synthesis of 2H-1, 4-benzoxazin-3-(4H)-one derivatives. This method efficiently performs the intramolecular oxidative amidation of arenes to install C—N bonds catalyzed by Cu[(CH₃CN)₄]ClO₄ under mild conditions. The approach enables the synthesis of various products from a wide range of aromatic ethers in moderate to good yields utilizing simple and inexpensive reagents and catalysts under mild conditions and shows good chemo-and regioselectivity. Preliminary cytotoxicity assays in two metastatic cancer cell lines suggests that benzo[1, 4]oxazin-3-ones can be useful templates upon which to develop potential small molecule anticancer agents.

All reactions were performed by using standard Schlenk techniques under nitrogen and argon atmosphere. Flash column chromatography was performed on silica gel. ¹H NMR and ¹³C NMR spectra were recorded on Varian Mercury 400 MHz spectrometers. High-resolution mass spectral (HRMS) analyses were carried out using a MALDI-TOF MS instrument.

A flame-dried round-bottom flask was charged, under nitrogen atmosphere, with the substrate (0.25 mmol, 1.0 equiv.), PhI (CF₃CO₂)₂ (0.625 mmol, 2.5 equiv.), Cu[(CH₃CN)₄]ClO₄ (0.05 mmol, 20 mol%), and distilled dichloromethane (10 mL). The resulting mixture was stirred at room temperature for 5 h. Then the mixture was washed with NaOH (5%), brine and dried over MgSO₄. The filtrate was purified by flash column chromatography (PE/EtOAc, 3:1) on silica gel.

Human breast adenocarcinoma (MDA-MB-231) and human cervix carcinoma (HeLa) cells were grown in DMEM medium supplemented with 115 units/mL of penicillin G, 115 mg/mL of streptomycin, and 10% fetal bovine serum (all from Life Technologies, Grand Island, NY). Cells were seeded in 96-well plates (5 ×10³ cells/well) containing 50 mL growth medium for 24 h. After medium removal, 100 mL fresh medium containing individual analog compounds at different concentrations was added to each well and incubated at 37 ℃ for 72 h. After 24 h of culture, the cells were supplemented with 50 mL of analog compounds dissolved in DMSO (less than 0.25% in each preparation). After 72 h of incubation, 20 mL of resazurin was added for 2 h before recording fluorescence at 560 nm (excitation) and 590 nm (emission) using a Victor microtiter plate fluorimeter (Perkin-Elmer, USA). The IC₅₀ was defined as the compound concentration required to inhibit cell proliferation by 50%, in comparison with cells treated with the maximum amount of DMSO (0.25%) and considered as 100% viability.

This work was financially supported by the National Natural Science FoundationofChina (No.21072131), and SichuanUniversity--Lu Zhou Strategic Cooperation Projects (No. 2013CDLZ-S18) (Ling He) and the NIH RCMI program at Xavier University of Louisiana (No. 2G12MD007595-07) (G. Wang).