High-Yield Synthesis of N⁴-(3-Chloro-4-fluorophenyl)-7-((4-methoxybenzyl)thio)quinazoline-4,6-diamine via N-(3-Chloro-4-fluorophenyl)-7-((4-methoxybenzyl)thio)-6-nitroquinazolin-4-amine

Abstract

The target compound, N⁴-(3-chloro-4-fluorophenyl)-7-((4-methoxybenzyl)thio)quinazoline-4,6-diamine, was synthesized in two steps from N-(3-chloro-4-fluorophenyl)-7-fluoro-6-nitroquinazolin-4-amine by a nucleophilic aromatic substitution, yielding N-(3-chloro-4-fluorophenyl)-7-((4-methoxybenzyl)thio)-6-nitroquinazolin-4-amine which was characterized by ¹H NMR, ¹³C NMR, ¹⁹F NMR, and HRMS. Subsequent reduction yielded the target molecule, N⁴-(3-chloro-4-fluorophenyl)-7-((4-methoxybenzyl)thio)quinazoline-4,6-diamine. The structure of the target compound was confirmed by ¹H NMR, ¹³C NMR, ¹⁹F NMR, HSQC, HMBC, HRMS, and FTIR.

1. Introduction

The quinazoline scaffold is found in many biologically active molecules, such as compounds that have bronchodilatory activity [1,2], antimycobacterial activity [3], or antihypertensive activity [4]. Furthermore, quinazoline derivatives are attractive anticancer agents due to their ability to act as protein kinase inhibitors [5,6]. For example, the N-(3-chloro-4-fluorophenyl)quinazolin-4-amine derivatives gefitinib [7,8,9], afatinib [10,11,12], and dacomitinib [12,13,14] are FDA-approved drugs used in the treatment of non-small cell lung cancer (Figure 1).

While sulfur-substituted quinazolines have also been investigated as inhibitors of tyrosine kinases or epidermal growth factor receptors, their oxygen analogs have proven to be superior anticancer drug candidates [15,16,17]. We were, however, more interested in potential antibacterial or antimicrobial drug candidates for which sulfur-substituted quinazolines have shown some promise [18].

2. Results and Discussion

The target compound N⁴-(3-chloro-4-fluorophenyl)-7-((4-methoxybenzyl)thio)quinazoline-4,6-diamine (3) was prepared as an extension of the previous work in our group [19,20] to include biologically relevant scaffolds such as quinazolines. As illustrated in Scheme 1, commercially available N-(3-chloro-4-fluorophenyl)-7-fluoro-6-nitroquinazolin-4-amine (1) was treated with (4-methoxyphenyl)methanethiol which, upon the addition of aqueous sodium hydroxide, selectively afforded N-(3-chloro-4-fluorophenyl)-7-((4-methoxybenzyl)thio)-6-nitroquinazolin-4-amine (2) by a nucleophilic aromatic substitution reaction. Compound 2 was isolated in 99% yield as a bright yellow powder and fully characterized by ¹H NMR (Supplementary Information, Figure S1),¹³C NMR (Supplementary Information, Figure S2), ¹⁹F NMR (Supplementary Information, Figure S3), and HRMS (Supplementary Information, Figure S4). The subsequent reduction of 2 in the presence of iron powder and ammonium chloride yielded the target compound N⁴-(3-chloro-4-fluorophenyl)-7-((4-methoxybenzyl)thio)quinazoline-4,6-diamine (3) quantitatively in form of a pale yellow powder. Compounds 2 and 3 are stable in air at room temperature for more than a year (as determined by ¹H NMR). Molecules 2 and 3 are also stable in the presence of water such as humid air at room temperature for over a year (as determined by ¹H NMR) and in boiling water for over 24 h (as determined by ¹H NMR).

The molecular structure of the target compound N⁴-(3-chloro-4-fluorophenyl)-7-((4-methoxybenzyl)thio)quinazoline-4,6-diamine (3) was in agreement with its ¹H NMR (Figure 2 and Figure S5). The NH proton at 9.54 ppm and the NH₂ protons at 5.47 ppm were confirmed by the absence of a cross peak for these protons in the HSQC spectrum (Supplementary Information, Figure S9). Additionally, upon heating a sample of 3 in D₂O at 100 °C for 24 h, partial H–D exchange (approximately 33%) is observed for these NH and NH₂ protons. All quinazoline protons appear as singlets in the ¹H NMR spectrum while the protons of the N-aryl group are coupled to the protons and the fluorine atom on the same ring. The methylene and methyl protons of the p-methoxylbenzyl group are visible as singlets at 4.30 and 3.72 ppm, respectively. Meanwhile, the aromatic protons of the p-methoxylbenzyl group are easily identified as the two doublets integrating to two protons each at 7.33 and 6.88 ppm, respectively.

The ¹³C NMR spectrum (Figure 3 and Figure S6) of N⁴-(3-chloro-4-fluorophenyl)-7-((4-methoxybenzyl)thio)quinazoline-4,6-diamine (3) was also in agreement with the molecular structure. Assignment of the individual carbon signals to the various aromatic components of 3 was aided by HSQC (Supplementary Information, Figure S9) and HMBC (Supplementary Information, Figure S10). Notable features include the five doublets for the fluorine-containing aromatic ring, of which the doublet at 152.9 ppm with a coupling constant of 242 Hz can be clearly identified as the carbon directly bound to fluorine. The p-methoxybenzyl group is visible at first glance by the presence of the methoxy carbon at 55.1 ppm and the methylene carbon at 35.5 ppm, as well as the aromatic carbons at 130.2 and 113.9 ppm, respectively.

As expected, the ¹⁹F NMR spectrum (Supplementary Information, Figure S7) shows one aromatic C-F peak (at −123.91 ppm). Additionally, HRMS (Supplementary Information, Figure S10) confirmed the expected atomic composition of 3. The IR spectrum (Supplementary Information, Figure S11) confirms the presence of the NH (medium absorption) which overlaps with the NH₂ (weak absorption).

3. Materials and Methods

3.1. General Information and Analyses

Reagents and solvents were purchased from Fisher Scientific or TCI Chemicals and used as supplied. Ethanol for the nucleophilic aromatic solution was used as supplied and not degassed prior to use.

Melting points were obtained using a MEL-TEMP apparatus (Cambridge, MA, USA) and are uncorrected. The ¹H NMR, ¹³C{¹H} NMR, and ¹⁹F{¹H} NMR spectra were all recorded on a 400 MHz Bruker Avance III spectrometer (Bruker, Billerica, MA, USA) with a 5 mm liquid-state Smart Probe at 298 K. Chemical shifts (δ_H, δ_C) are expressed in parts per million (ppm) and reported in the ¹H NMR spectra relative to the resonance of the residual protons of the DMSO-d₆ (δ_H = 2.50 ppm) or in the ¹³C{¹H} NMR spectra relative to the resonance of the deuterated solvent DMSO-d₆ (δ_C = 39.52 ppm). Chemical shifts in the ¹⁹F{¹H} NMR spectra are reported relative to the internal standard fluorobenzene (δ_F = −113.15). High-Resolution Mass Spectrometry (HRMS) data were obtained on an LTQ Orbitrap XL (Thermo Fisher Scientific, Waltham, MA, USA) in FT Orbitrap Mode at a resolution of 100′000. IR spectra were measured in solid form on a Cary 630 FTIR (Agilent, Santa Clara, CA, USA).

3.2. Synthesis of N-(3-Chloro-4-fluorophenyl)-7-((4-methoxybenzyl)thio)-6-nitroquinazolin-4-amine (2)

A 500 mL round-bottom flask equipped with a stir bar was loaded with 8.417 g of N-(3-chloro-4-fluorophenyl)-7-fluoro-6-nitroquinazolin-4-amine (25.00 mmol, 1 equiv) and 200 mL of ethanol and placed under an atmosphere of argon. A total of 4.241 g of (4-methoxyphenyl)methanethiol (27.5 mmol, 1.1 equiv) was added with a syringe, followed by a dropwise addition of a solution of 1.100 g of NaOH (27.5 mmol, 1.1 equiv) in 10 mL of H₂O. The reaction mixture was stirred at room temperature for 2 h, after which TLC indicated the completion of the reaction. The solid was filtered off, washed with H₂O, followed by ethanol and, finally, diethyl ether to yield the product in 99% yield (11.604 g, 24.64 mmol) in the form of a bright yellow powder, m.p. 210–211 °C.

¹H NMR (400 MHz, DMSO-d₆, 298 K): δ = 10.39 (s, 1H), 9.51 (s, 1H), 8.67 (s, 1H), 8.14 (d, J = 4.8 Hz, 1H), 7.82–7.79 (m, 2H), 7.45 (t, J = 9.1 Hz, 1H), 7.40 (d, J = 8.6 Hz, 2H), 6.92 (d, J = 8.6 Hz, 2H), 4.39 (s, 2H), 3.74 (s, 3H); ¹³C{¹H} NMR (100 MHz, DMSO-d₆, 298 K): δ = 158.8, 157.9, 157.8, 153.6 (d, J = 244.1 Hz), 151.4, 142.3, 140.7, 135.8, 130.5, 126.4, 124.5, 123.7, 123.0, 122.4 (d, J = 6.9 Hz), 118.9 (d, J = 18.4 Hz), 116.5 (d, J = 21.7 Hz), 114.1, 110.8, 55.1, 35.9; ¹⁹F{¹H} NMR (376 MHz, DMSO-d₆, 298 K, referenced to C₆H₅F): δ = −121.73. HRMS (ESI) m/z calculated for [M + H]⁺ = [C₂₂H₁₇ClFN₄O₃S]⁺ 471.0687; observed, 471.0687 (0 ppm).

3.3. Synthesis of N⁴-(3-Chloro-4-fluorophenyl)-7-((4-methoxybenzyl)thio)quinazoline-4,6-diamine (3)

A 500 mL round-bottom flask equipped with a stir bar was loaded with 11.106 g of N-(3-chloro-4-fluorophenyl)-7-((4-methoxybenzyl)thio)-6-nitroquinazolin-4-amine (23.58 mmol, 1 equiv), 6.308 g of NH₄Cl (117.9 mmol, 5.0 equiv), and 250 mL of EtOH/H₂O (4:1). The reaction flask was placed into an oil bath set to 80 °C, and 6.595 g of iron power (117.92 mmol, 5.0 equiv) was added while stirring. Then, the reaction flask was fitted with a reflux condenser and the reaction was stirred under argon at 80 °C until TLC indicated a complete reduction after 2 h of reaction time. After cooling to room temperature, the reaction mixture was filtered through celite and concentrated under reduced pressure. The residue was then basified with 1M NaOH and extracted twice with dichloromethane, dried over MgSO₄, and evaporated. After washing with hexanes, the product was obtained quantitatively in the form of a pale-yellow powder, m.p. 187 °C.

¹H NMR (400 MHz, DMSO-d₆, 298 K): δ = 9.54 (s, 1H), 8.36 (s, 1H), 8.19 (dd, J = 6.9, 2.6 Hz, 1H), 7.82–7.78 (m, 1H), 7.58 (s, 1H), 7.44–7.38 (m, 2H), 7.33 (d, J = 8.7 Hz, 2H), 6.88 (d, J = 8.7 Hz, 2H), 5.47 (s, 2H), 4.30 (s, 2H), 3.72 (s, 3H); ¹³C{¹H} NMR (100 MHz, DMSO-d₆, 298 K): δ = 158.5, 155.6, 152.9 (d, J = 242.1 Hz), 150.3, 144.9, 142.9, 137.2 (d, J = 3.0 Hz), 131.0, 130.2, 128.3, 126.8, 122.8, 121.7 (d, J = 6.7 Hz), 118.7 (d, J = 18.1 Hz), 116.5 (d, J = 21.6 Hz), 114.6, 113.9, 101.8, 55.1, 35.5; ¹⁹F{¹H} NMR (376 MHz, DMSO-d₆, 298 K, referenced to C₆H₅F): δ = −123.91. HRMS (ESI) m/z calculated for [M + H]⁺ = [C₂₂H₁₉ClFN₄OS]⁺ 441.0947; observed, 441.0948 (0.2 ppm).

4. Conclusions

In summary, we have prepared the target compound N⁴-(3-chloro-4-fluorophenyl)-7-((4-methoxybenzyl)thio)quinazoline-4,6-diamine (3) in two steps from N-(3-chloro-4-fluorophenyl)-7-fluoro-6-nitroquinazolin-4-amine (1) by a high-yielding nucleophilic aromatic substitution with a subsequent reduction. Both the intermediate N-(3-chloro-4-fluorophenyl)-7-((4-methoxybenzyl)thio)-6-nitroquinazolin-4-amine (2) and the target molecule N⁴-(3-chloro-4-fluorophenyl)-7-((4-methoxybenzyl)thio)quinazoline-4,6-diamine (3) were characterized by ¹H NMR, ¹³C NMR, ¹⁹F NMR, and HRMS. Additionally, the HSQC and HMBC of compound 3 allowed for the assignment of the ¹H and ¹³C signals, while FTIR showed the presence of the N-H bonds. Future work could include studying the antibacterial properties of compounds 2 and 3.