Rapid Construction of a Chloromethyl-Substituted Duocarmycin-like Prodrug

Abstract

The construction of duocarmycin-like compounds is often associated with lengthy synthetic routes. Presented herein is the development of a short and convenient synthesis of a type of duocarmycin prodrug. The 1,2,3,6-tetrahydropyrrolo[3,2-e]indole-containing core is here constructed from commercially available Boc-5-bromoindole in four steps and 23% overall yield, utilizing a Buchwald–Hartwig amination followed by a sodium hydride-induced regioselective bromination. In addition, protocols for selective mono- and di-halogenations of positions 3 and 4 were also developed, which could be useful for further exploration of this scaffold.

1. Introduction

Duocarmycin A (1) and SA (2) are prominent members of the duocarmycin family that possess extreme cytotoxic properties (Figure 1) [1,2,3]. They were isolated from the Streptomyces sp. in Japan in 1988 and 1990, respectively [4,5]; in the early 1990s, their structures were confirmed by synthesis [6,7,8]. Since then, duocarmycin and its analogs have attracted a lot of attention among synthetic and medicinal chemists, owing to their structural complexity and interesting biological properties. Their mode of action is site-specific DNA alkylation, and their strongly alkylating properties can be attributed to the strained cyclopropane moiety (Figure 1). Unfortunately, the cytotoxicity is not only devoted to the cancer cells; therefore, a variety of duocarmycin analogs [1,2,3], prodrugs [9,10,11,12,13,14,15,16,17,18,19], and even antibody–drug conjugates [20] have been developed in the pursuit for more selective cancer treatments. In a medicinal chemistry project working with prodrugs that, upon site-selective CYP2W1 oxidation, form the phenolic counterpart and render the compound harmful [14,17] (3, Figure 1), we needed access to the chloromethyl-substituted 1,2,3,6-tetrahydropyrrolo[3,2-e]indole core 10 (Figure 2).

The existing synthetic pathways are elaborative and/or give the wrong substitution pattern (Figure 2). Furthermore, in our early attempts to use Boc-5-nitroindole 9 as starting material, we faced several problems, such as over-reduction when reducing the nitro group (i.e., the generation of indoline), the generation of complex mixtures when performing the halogenation reaction on the aniline, and problems with controlling the mono-Boc protection of the aniline.

Figure 2. Previous versus new routes from commercial starting materials [14,16,21,22].

Figure 2. Previous versus new routes from commercial starting materials [14,16,21,22].

In our approach, we envisioned that the desired di-Boc-protected 5-aminoindole intermediate 12 (Figure 3) could be synthesized from commercially available Boc-5-bromoindole 11 via a Buchwald–Hartwig amination with tBu-carbamate followed by a regioselective bromination. This strategy would considerably shorten the route and also overcome the problems related to the nitro reduction and mono-Boc protection of the aniline nitrogen; vide supra.

2. Results and Discussion

The Pd(OAc)₂/XPhos-catalyzed Buchwald–Hartwig amination of Boc-5-bromoindole (11) with tBu-carbamate performed well, and compound 13 could be isolated in 78% yield (Scheme 1). Performing the subsequent halogenation under acidic conditions (i.e., NXS/TsOH) on the Boc-protected aniline gave the wrong regioisomer, although with complete selectivity, and the 3-bromo (14) and 3-iodo (15) products could be isolated in 74% and 71% yields, respectively, using the two different halogen sources. We envisioned that the deprotonation of the Boc-protected aniline with NaH prior to the halogenation might render the aromatic ring sufficiently electron-rich to direct the halogenation to the right position (see Supporting Information). Gratifyingly, that strategy gave the desired 4-bromo analog 12 in 65% yield with complete regioselectivity. All attempts to introduce iodine in this position failed, even when using a more electrophilic I⁺ source (i.e., N-Iodosaccharin [23]), other solvents, or elevated temperatures.

To our delight, further halogenation of 12 to give 3-iodo-4-bromo compound 16 went smoothly under acidic conditions (NIS/TsOH) in 71% yield. To conclude the synthesis towards the duocarmycin-type prodrug, compound 12 smoothly underwent allylation with 1,3-dichloropropene to give 17 [14] in 82% yield, followed by a tris(trimethylsilyl)silane (TTMSS)/azaisobutyronitrile (AIBN)-induced radical 5-exo-trig cyclization according to published procedures to furnish compound 10 [14] in 56% yield (Scheme 2). After Boc deprotection and subsequent EDC/NaHCO₃ amide coupling with 5-fluoroindole-2-carboxylic acid, the desired prodrug rac—18 [17] was isolated in 65% yield over two steps. In addition, the enantiomers were separated by chiral supercritical fluid chromatography (SFC) to give (+)—18 and (−)—18 with ee ≥ 99%.

3. Materials and Methods

General Methods: All solvents and reagents were used as received from commercial suppliers. N-Bromosuccinimide (NBS) was recrystallized from hot water and dried under vacuum for 24 h and then stored under cold and dark conditions. Sodium hydride was used as 60% dispersion in mineral oil. Column chromatography was employed on normal-phase silica gel (230–400 mesh, 60 Å; the eluents are given in brackets). ¹H- and ¹³C-NMR spectra were recorded on a 400 MHz spectrometer at 298 K and calibrated using the residual peak of the solvent as an internal standard [CDCl₃ (CHCl₃ δ_H 7.26 ppm, CDCl₃ δ_C 77.16 ppm)]. HRMS was performed using a microTOF instrument with electrospray ionization (ESI), and sodium formate was used as a calibration chemical. Optical rotations were measured on a polarimeter at 589 nm (D line of sodium) and 20 °C. Chiral chromatography was performed on supercritical fluid chromatography equipment, using mixtures of MeOH and supercritical CO₂ as eluents.

Di-tert-butyl 1-(chloromethyl)-1,2-dihydropyrrolo[3,2-e]indole-3,6-dicarboxylate (10): tert-Butyl-4-bromo-5-((tert-butoxycarbonyl)(3-chloroallyl)amino)-1H-indole-1-carboxylate 17 (600 mg, 1.24 mmol) was dissolved in dry toluene (40 mL), and the solution was degassed for 1 h (by bubbling N₂ gas through the solution under stirring). Azobisisobutyronitrile (AIBN) (49 mg, 0.30 mmol) and tris(trimethylsilyl)silane (TTMSS) (0.41 mL, 1.34 mmol) were added, and the reaction was heated to 90 °C (with a preheated oil bath) in a sealed tube for 5 h. The solvent was evaporated, and the crude material was dissolved in MeOH (12 mL) and stirred at rt for 10 min. The solvent was evaporated, and the crude product was purified by column chromatography on silica gel (hexanes:EtOAc 95:5) to give compound 10 as a colorless oil (280 mg, 56%). The spectral data agreed with the published data [14].

tert-Butyl 4-bromo-5-((tert-butoxycarbonyl)amino)-1H-indole-1-carboxylate (12): tert-Butyl 5-((tert-butoxycarbonyl)amino)-1H-indole-1-carboxylate 13 (200 mg, 0.60 mmol) was dissolved in dry DMF (2 mL) and cooled to 0 °C with an ice bath. NaH (60 mg, 60% in mineral oil, 1.5 mmol) was added, followed by NBS (129 mg, 0.72 mmol); the ice bath was removed, and the reaction was stirred for 30 min. The reaction mixture was poured onto saturated NaHCO₃ (aq) and extracted with EtOAc. The organic phase was dried (Na₂SO₄), filtered, and concentrated. The crude material was purified by column chromatography on silica gel (hexanes:EtOAc 95:5) to give compound 12 as a colorless foam (160 mg, 65%). The spectral data agreed with the published data [14].

tert-Butyl 5-((tert-butoxycarbonyl)amino)-1H-indole-1-carboxylate (13): N-Boc-5-bromoindole 11 (1.5 g, 5.06 mmol), tert-butyl carbamate (712 mg, 6.08 mmol), Pd(OAc)₂ (57 mg, 0.25 mmol), XPhos (241 mg, 0.50 mmol), and Cs₂CO₃ (2.31 g, 7.09 mmol) were mixed in dry 1,4-dioxane (45 mL), and the vessel was flushed with N₂ gas, sealed, and heated to 90 °C for 20 h. The reaction mixture was diluted with EtOAc, filtered through Celite, and concentrated. The crude material was purified by column chromatography on silica gel (hexanes:EtOAc 95:5) to give compound 13 as a colorless foam (1.32 g, 78%). ¹H-NMR (CDCl₃, 400 MHz) δ 8.01 (brd, J = 8.0 Hz, 1H), 7.75 (brs, 1H), 7.55 (brd, J = 4.0 Hz, 1H), 7.14 (dd, J = 8.0, 4.0 Hz, 1H), 6.70 (brs, 1H, NH), 6.48 (dd, J = 3.7, 0.8 Hz, 1H), 1.65 (s, 9H), 1.52 (s, 9H); ¹³C-NMR (CDCl₃, 100 MHz) δ 153.3, 149.8, 133.7, 131.5, 131.1, 126.6, 116.4, 115.3, 110.9, 107.4, 83.6, 80.3, 28.5 (3C), 28.3 (3C); HRMS (ESI/TOF) m/z: [M + Na]⁺ Calcd for C₁₈H₂₄N₂O₄Na 355.1634; Found 355.1633.

tert-Butyl 3-bromo-5-((tert-butoxycarbonyl)amino)-1H-indole-1-carboxylate (14): tert-Butyl 5-((tert-butoxycarbonyl)amino)-1H-indole-1-carboxylate 13 (200 mg, 0.60 mmol) was dissolved in DMF (2 mL), NBS (118 mg, 0.66 mmol) and TsOH·H₂O (23 mg, 0.12 mmol) were added, and the reaction was stirred at rt for 10 min. The reaction mixture was poured onto saturated NaHCO₃ (aq) and extracted with EtOAc. The organic phase was dried (Na₂SO₄), filtered, and concentrated. The crude material was purified by column chromatography on silica gel (hexanes:EtOAc 95:5) to give compound 14 as a colorless foam (183 mg, 74%). ¹H-NMR (CDCl₃, 400 MHz) δ 8.02 (brd, J = 8.0 Hz, 1H), 7.65 (brs, 1H), 7.60 (brs, 1H), 7.24 (brd, J = 8.0 Hz, 1H), 6.67 (brs, 1H, NH), 1.65 (s, 9H), 1.54 (s, 9H); ¹³C-NMR (CDCl₃, 100 MHz) δ 153.1, 148.9, 134.5, 130.9, 130.0, 125.5, 117.5, 115.6, 109.2, 97.9, 84.4, 80.6, 28.5 (3C), 28.3 (3C); HRMS (ESI/TOF) m/z: [M + Na]⁺ Calcd for C₁₈H₂₃BrN₂O₄Na 433.0739; Found 433.0755.

tert-Butyl 5-((tert-butoxycarbonyl)amino)-3-iodo-1H-indole-1-carboxylate (15): tert-Butyl 5-((tert-butoxycarbonyl)amino)-1H-indole-1-carboxylate 13 (1.3 g, 3.91 mmol) was dissolved in DMF (14 mL), NIS (1.06 g, 4.71 mmol) and TsOH·H₂O (149 mg, 0.78 mmol) were added, and the reaction was stirred at rt for 15 h. The reaction mixture was poured onto saturated NaHCO₃ (aq) and extracted with EtOAc. The organic phase was washed with 10 wt% Na₂S₂O₅ (aq), dried (Na₂SO₄), filtered, and concentrated. The crude material was purified by column chromatography on silica gel (hexanes:EtOAc 95:5) to give compound 15 as a colorless foam (1.28 g, 71%). ¹H-NMR (CDCl₃, 400 MHz) δ 8.00 (brd, J = 8.0 Hz, 1H), 7.69 (brs, 1H), 7.53–7.46 (m, 1H), 7.27 (brd, J = 8.0 Hz, 1H), 6.73 (brs, 1H, NH), 1.65 (s, 9H), 1.54 (s, 9H); ¹³C-NMR (CDCl₃, 100 MHz) δ 153.1, 148.7, 134.6, 132.7, 131.1, 130.8, 117.5, 115.5, 111.3, 84.3, 80.6, 65.4, 28.5 (3C), 28.2 (3C); HRMS (ESI/TOF) m/z: [M + Na]⁺ Calcd for C₁₈H₂₃IN₂O₄Na 481.0601; Found 481.0595.

tert-Butyl 4-bromo-5-((tert-butoxycarbonyl)amino)-3-iodo-1H-indole-1-carboxylate (16): tert-Butyl 4-bromo-5-((tert-butoxycarbonyl)amino)-1H-indole-1-carboxylate 12 (140 mg, 0.34 mmol) was dissolved in DMF (1.4 mL), NIS (114 mg, 0.51 mmol) and TsOH·H₂O (16 mg, 0.08 mmol) were added, and the reaction was stirred at rt for 16 h. The reaction mixture was poured onto saturated NaHCO₃ (aq) and extracted with EtOAc. The organic phase was washed with 10 wt% Na₂S₂O₅ (aq), dried (Na₂SO₄), filtered, and concentrated. The crude material was purified by column chromatography on silica gel (hexanes:EtOAc 95:5) to give compound 16 as a colorless foam (130 mg, 71%). ¹H-NMR (CDCl₃, 400 MHz) δ 8.11 (m, 2H), 7.77 (s, 1H), 7.08 (brs, 1H, NH), 1.65 (s, 9H), 1.54 (s, 9H); ¹³C-NMR (CDCl₃, 100 MHz) δ 153.0, 148.2, 134.0, 132.7, 131.6, 126.5, 118.5, 114.5, 105.3, 85.0, 81.1, 61.2, 28.5 (3C), 28.2 (3C); HRMS (ESI/TOF) m/z: [M + Na]⁺ Calcd for C₁₈H₂₂BrIN₂O₄Na 558.9706; Found 558.9700.

tert-Butyl-4-bromo-5-((tert-butoxycarbonyl)(3-chloroallyl)amino)-1H-indole-1-carboxylate (17): tert-Butyl 4-bromo-5-((tert-butoxycarbonyl)amino)-1H-indole-1-carboxylate 12 (650 mg, 1.58 mmol) was dissolved in dry DMF (12 mL) and cooled to 0 °C, NaH (190 mg, 60% in mineral oil, 4.74 mmol) was added, and the reaction was stirred at 0 °C for 5 min. 1,3-Dichloropropene was added, the ice bath was removed, and the reaction was stirred at rt for 1 h. The reaction mixture was poured onto saturated NaHCO₃ (aq) and extracted with EtOAc. The organic phase was dried (Na₂SO₄), filtered, and concentrated. The crude material was purified by column chromatography on silica gel (hexanes:EtOAc 95:5) to give compound 17 as a colorless oil (630 mg, 82%). The spectral data agreed with the published data [14].

(1-(chloromethyl)-1,6-dihydropyrrolo[3,2-e]indol-3(2H)-yl)(5-fluoro-1H-indol-2-yl)methanone (18): Di-tert-butyl 1-(chloromethyl)-1,2-dihydropyrrolo[3,2-e]indole-3,6-dicarboxylate 10 (280 mg, 0.69 mmol) was dissolved in 4 M HCl in 1,4-dioxane (15 mL, 60 mmol), and the reaction was stirred at rt for 22 h. The solvent was evaporated, and the crude material was coevaporated from EtOAc two times. The crude material, together with 5-fluoro-1H-indole-2-carboxylic acid 19 (148 mg, 0.83 mmol), N-ethyl-N′-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC) (396 mg, 2.07 mmol), and NaHCO₃ (289 mg, 3.45 mmol), were mixed in dry DMF (10 mL), and the reaction was stirred at rt for 5 h. The reaction mixture was poured onto saturated NaHCO₃ (aq) and extracted with EtOAc. The organic phase was dried (Na₂SO₄), filtered, and concentrated. The crude material was purified by column chromatography on silica gel (hexanes:EtOAc 60:40 to 50:50) to give compound 18 (253 mg, 65%) as an off-white solid. The spectral data agreed with the published results [17]. The racemic product was separated by chiral supercritical fluid chromatography (SFC) to give (+)—18, [α]_D (c = 1.0, acetone) +17 and (−)—18, [α]_D (c = 1.0, acetone) -17, both with ee ≥ 99% (for chromatographic conditions and chromatograms, see Supporting Information).

4. Conclusions

In conclusion, we developed a four-step route to the desired chloromethyl-substituted 1,2,3,6-tetrahydropyrrolo[3,2-e]indole core 10, utilizing an unconventional NaH promoted site-selective bromination of Boc-protected amino indole 13 as the key step. Additionally, 3-iodo-4-bromo indole 16 constitutes an interesting starting point for further diversification. Closely related 3-iodo-4-bromo-indoles have been used in Pd-catalyzed cross-couplings such as the Mizoroki-Heck [24,25,26], Negishi [27], and Suzuki-Miyaura [28,29] reactions in various natural products and heterocyclic syntheses. Finally, the racemate of compound 18 was separated with chiral supercritical fluid chromatography for further investigation of this interesting prodrug.