A practical synthesis of N -allyl/propargyl-substituted 5-fluorouracils

Monosubstituted N -allyl/propargyl-5-fluorouracils are versatile intermediates for the structural modification of 5-fluorouracil derivatives. However, the regioselective synthesis of these monosubstituted 5-fluorouracils is a challenge. Thus, in the current research work, a practical method for synthesizing N 1 / N 3 -allyl/propargyl-5-fluorouracils was developed with di-tertbutyl dicarbonate acting as a protective reagent. The process is easy to operate, gives a good regioselectivity, satisfying yields and a simple post-treatment

Introduction 5-Fluorouracil (5-FU, 1) is an antimetabolite acting as a bioisostere of the natural uracil in living body, and has been widely used in the treatment of colorectal cancer and several solid tumors. 1 5-FU derivatives incorporating pharmacologically active natural or synthetic molecules, such as 5-FU linked with podophyllotoxin, colchicine, parthenolide, coumarins, thymoquinone, emodin, camptothecin, cisplatin, oxaliplatin, tamibarotene and chalcone, have attracted much attention of many researchers in the field of medicinal chemistry. 2 Recently, many structural modifications have been focused on merging a particular pharmacologically active molecule at the N1 or N3 position of 5-FU using a triazole linkage in order to achieve a higher bioactivity. For example, 5-FU derivatives, incorporating the bioactive molecules parthenolide 3a, thymoquinone 3b, or glucose derivative 3c incorporating the click triazole linker, have shown potential anticancer activities. [3][4][5] Moreover, 5-FU with triazole-linked polyheterocyclic compound 3d has been reported to exhibit a superior antibacterial activity ( Figure 1). 6 Since triazole moieties have been developed as versatile pharmacophoric linkers, connecting two biological units into one molecule called hybrids, 7-8 alkyne or alkene-decorated 5-FUs, especially 1-propargyl-5-FU 2, have been used as a key intermediate to furnish various isoxazole or triazole-functionalized 5-FU derivatives 3 via 1,3dipolar cycloaddition reactions with nitrile oxides or azide dipoles.
Given the proved applications of allyl or propargyl-containing 5-FUs in the development of the bioactive 5-FU derivatives, extensive research has been focused on the synthesis of 1-propargyl-5-FU 2 and 1-allyl-5-FU 4. A common method for the preparation of N1-substituted 5-FUs (2 and 4) was through nucleophilic substitution of 5-FU with bromopropyne or bromopropylene in the presence of a base such as K2CO3, NaH or DBU (Scheme 1, eq 1). 3,[9][10][11][12] Although this method provides a simple and direct route to propargyl or allylmonosubstituted 5-FUs, the poor regioselectivity and low reaction yields are still challenging problems. In a modified method, the combination of Pd(PPh3)4/1,1'-Bis(diphenylphosphino)ferrocene(DPPF) and allyl acetate was used instead of bromopropylene, giving N1-allyl-5-FU 4 in a moderate yield. 13 Alternatively, a multiple-step pathway could be used to synthesize the monosubstituted 5-FUs, where 5-FU was first transformed into the key intermediates featuring N1-CH2OCOOBn or N3-CH2OCOOBn by multiple steps, followed by allylation and deprotection to give allyl-substituted 5-FUs 4 and 5 (Scheme 1, eq 2). 14 Although multiple methods have been utilized for the synthesis of propargyl/allyl-substituted 5-FUs, there are some disadvantages such as low reaction yields, 10-12 using an expensive catalyst 13 and complex synthetic procedures. 14 Furthermore, few of these methods were involved in the preparation of N3-substituted 5-FUs (7 and 5). Therefore, it is still of interest to develop a general and efficient method for the synthesis of allyl/propargyl-monosubstituted 5-FUs. Scheme 1. 1) One-step synthesis of 5-FUs 2 and 4, and 2) multi-step synthesis of 5-FUs 4 and 5.

Results and Discussion
The methods reported to synthesize 1-propargyl-5-FU 2 from the reaction of 5-FU and bromopropyne using K2CO3 as a base in DMF have problems of a low regioselectivity and a poor yield. 3,9 To improve the regioselectivity and yield, the reaction conditions of 5-FU and bromopropyne with K2CO3 (1 equiv to 5-FU) in DMF were optimized (Table S1, see supporting information); however, a single product N1, N3-dipropargyl-5-FU 6 was obtained instead of N1-propargyl-5-FU 2. Interestingly, no trace of compound 2 was detected by 1 H-NMR analysis of the isolated product (Scheme 2).

Scheme 2.
The reaction of 5-FU and bromopropyne with K2CO3 as a base.
Usually, the proton N1-H of 5-FU is much more reactive than that of the N3 position. A plausible reaction mechanism was assumed, where the proton at the N1 position of 5-FU was firstly replaced by propargyl, and the resulting intermediate significantly improved the reaction activity of the proton at the N3 position. Therefore, there was no single substitution product 2, but always the double substitution product 6. While this reaction was carried out with 5-FU and bromopropylene, it was observed that N1, N3-diallyl-5-FU and N1-allyl-5-FU were both generated. In this reaction mechanism, N1-allyl-5-FU is formed in the first step, and then a double substituted product is generated.
Then, the synthesis of the monosubstituted 5-FUs by a muti-step method was evaluated with a strategy of protection/alkylation/deprotection. Since the N-protecting groups were commonly used to prepare pyrimidine derivatives, such as benzoyl, trichloroethoxyformyl, benzyloxyformyl, benzyl and diphenylmethyl moieties, different N-protecting methods were tested and ultimately di-tertbutyl dicarbonate (Boc2O) was selected as the N-protecting reagent of the starting material 5-FU 1. After several optimizations of the reaction conditions, N1-Boc-substituted intermediate 11 was directly obtained in 65% yield, by the regioselective reaction of 5-FU 1 and 1.0 equiv of Boc2O, with Et3N as a base. In contrast, N3-monosubstituted 5-FU 9 needed a reaction sequence of N1,N3-diBoc protection and regioselective deprotection of N1-Boc group. In detail, N1,N3-diBoc-5-FU 8 was provided by reacting 5-FU 1 and 3.0 equiv of Boc2O with pyridine as a base, followed by the regioselective deprotection of N1-Boc group to give the desired N3-Boc-5-FU 9. Then the subsequent propargylation of intermediates 11 and 9 were performed with bromopropyne, and finally, the desired N3-propargyl-5-FU and N1propargyl-5-FU (7 and 2) were prepared after deprotection of the Boc group, with total yields of 29% and 20%, respectively (Scheme 3). Using the same procedure, when bromopropylene was used instead of bromopropyne, the corresponding allyl-substituted 5-FUs, 4 and 5, were obtained with total yields of 20% and 23%, respectively (Scheme 3).

Conclusions
In summary, the developed method uses 5-FU as starting material and Boc anhydride as a protective reagent to regioselectively produce N1/N3-Boc-5-FU, and provides a practical synthesis of allyl/propargyl-monosubstituted 5-FUs. This method is easy to operate, leads to a good regioselectivity, satisfying yield and simple posttreatment. The produced allyl/propargyl-functionalized 5-FUs can have wide applications in the subsequent preparation of diverse 5-FU derivatives with specific pharmacological activities.

Experimental Section
General. Melting points were recorded using the XRC-1 apparatus and are uncorrected. IR spectra were obtained on a Thermo Nicolet Avatar 370 FT-IR spectrophotometer. 1 H and 13 C NMR spectra were recorded on a Bruker AVANCE III spectrometer at 600 MHz using TMS as internal standard. HRMS spectra were recorded on a Waters GCT Premier instrument with EI mode. All the chemicals and solvents were analytical reagents and commercially available and used as received. 15 10 1-Boc-5-FU (4.6 g, 20 mmol) was mixed with 60% NaH (0.96 g, 24 mmol) in DMF (50 mL) and stirred for 45 min at 0-5 °C. Bromopropyne (2.4 g, 20 mmol) was added dropwise and then stirred at rt until no reactant was detected by TLC. The mixture was poured into ice-water, extracted with ethyl acetate, and washed with water. The organic layer was dried with anhydrous magnesium sulfate, concentrated under vacuum, and purified by column chromatography to obtain 12, white solid, 2.7 g, yield 50%,  18 1,3-diBoc-5-FU 8 (10.0 g, 30 mmol) was dissolved in dioxane (50 mL), and the mixture was added with 10 mL aqueous K2CO3 (6.2 g, 45 mmol) dropwise. After stirring for 8 h at rt, the mixture was concentrated under vacuum to remove dioxane. Then, water (50 mL) was added and exacted with DCM (20 mL × 3). The organic layer was dried, concentrated and purified by column chromatography to obtain 9, white solid, 2.4 g, yield 31%, mp 128-130 °C; 1 H NMR (600 MHz, DMSO-d6) δ 11.52(br, 1H, NH), 7.98(d, J 6.1 Hz, 1H, ArH), 1.52(s, 9H, C(CH3)3); 13