Concise Synthesis of Heterocycle-Fused Naphthoquinones by Employing Sonogashira Coupling and Tandem Addition-Elimination/Intramolecular Cyclization

Abstract

A concise method for the synthesis of heterocycle-fused naphthoquinones such as naphtho[2,3-b]-furan-4,9-dione, 1H-benz[f]indole-4,9-dione, and naphtho[2,3-b]thiophene-4,9-dione was developed. This method employed Sonogashira coupling and tandem addition-elimination/intramolecular cyclization, and it enabled the preparation of versatile heterocycle-fused naphthoquinones from one substrate.

Heterocycle-fused naphthoquinones such as naphtho[2,3-b]furan-4,9-dione,^1–7) 1H-benz[f]indole-4,9-dione,^8–17) and naphtho[2,3-b]thiophene-4,9-dione,^18–27) which display a variety of biological activities, are found in various natural and artificial compounds. Despite the importance of these compounds, a survey of the literature revealed that very few efficient general methods have been developed to prepare heterocycle-fused naphthoquinones although many procedures have been established to prepare specific compounds. We are interested in studying heterocycle-fused naphthoquinones isolated from Tabebuia avellanedae Lorentz ex Griseb²⁸⁾ (Bignoniaceae) (syn. T. impetiginosa). Among these constituents, we previously reported that (−)-5-hydroxy-2-(1′-hydoxyethyl)naphtho[2,3-b]furan-4,9-dione (1aa) has a strong antiproliferative activity against various tumor cell lines^29–31) (Fig. 1). In addition, we reported the stereoselective synthesis of 1aa using Noyori reduction as a key step although tedious manipulations necessary in the synthesis and an inadequate enantiomeric excess from the final asymmetric hydrogenation prevented its further application. Recently, indolequinones were synthesized via Sonogashira coupling effectively, followed by copper-catalyzed intramolecular cyclization.^32,33) In this synthesis, halogenated quinones reacted smoothly with terminal acetylenes to give the coupling product. Therefore, we decided to use this methodology to synthesize 1aa and its related compounds. Here, we describe the development of a general and concise method for constructing heterocycle-fused naphthoquinone motifs involving Sonogashira coupling and tandem addition-elimination/intramolecular cyclization.

Fig. 1. Structure of 1aa

Results and Discussion

Chart 1 shows our strategies. The product 3 that is synthesized from the halogenated naphthoquinone 2 with the terminal acetylene 4 using Sonogashira coupling was a key intermediate in the synthesis of heterocycle-fused naphthoquinones 1 because the replacement of the dimethylamino group of 3 by another functional group (a hydroxyl, thiol, or amino group) followed by intramolecular cyclization would allow the construction of diverse heterocycle-fused naphthoquinone motifs.

Chart 1. Synthesis of 1 from 2

First, we investigated the Sonogashira coupling of 3-bromo-2-(dimethylamino)-5-hydroxynaphthalene-1,4-dione (2a) with but-3-yn-2-ol (4a) because we²⁹⁾ and another research group³⁴⁾ independently reported that the presence of the phenolic hydroxy group at C-5 considerably increases the antiproliferative effect of naphthoquinones on human-tumor cell lines. We used our previously reported procedure with a few modifications^33,35) and examined the reaction of 2a (1 eq) with racemic 4a (2 eq) in the presence of Cu₂O (0.2 eq), pyridine (100 eq, i.e., pyridine–copper=50 : 1), and K₂CO₃ (0.4 eq) in N,N-dimethylformamide (DMF) at room temperature. The conversion of 2a to 3aa occurred using our reaction conditions, and 3aa was obtained in 84% yield (Table 1, entry 1). Decreasing the amount of copper salt to 0.1 eq gave a lower yield of 3 (Table 1, entry 2). Next, using the reaction conditions shown in the entry 1 of Table 1, we tested the coupling reaction on a series of halogenated naphthoquinones 2 and terminal acetylenes 4. Alkynes bearing an aryl or alkyl substituent reacted smoothly with the halogenated naphthoquinone 2a to give the desired products 3ab and 3ac in good yields (Table 1, entries 3, 4). In addition, a satisfactory result was obtained using the halogenated naphthoquinone 2b which does not have a hydroxyl group on the aromatic ring as a substrate (Table 1, entry 5). The yield of 3ca decreased slightly when naphthoquinone 2c, which has a hydroxyl group at C-8, was used as the substrate (Table 1, entry 6).

Table 1. Reaction of 3-Bromo-2-(dimethylamino)-naphthalene-1,4-dione (2) with Acetylene (4)^a)

Entry	Substrate 2 X, Y	Acetylene 4 R	Time (h)	Product	Yield (%)b)
1	OH, H (2a)	CH(OH)CH3 (4a)	17	3aa	84
2c)	OH, H (2a)	CH(OH)CH3 (4a)	22	3aa	59 (24)d)
3	OH, H (2a)	Ph (4b)	17	3ab	93
4e)	OH, H (2a)	CH2CH2Ph (4c)	5	3ac	92
5	H, H (2b)	CH(OH)CH3 (4a)	17	3ba	91
6	H, OH (2c)	CH(OH)CH3 (4a)	21	3ca	62

a) Substrate 1 (0.5 mmol), Pd(OAc)₂ (3 mol%), Cu₂O (0.1 mmol), acetylene (1.0 mmol), pyridine (8.0 mL), and K₂CO₃ (0.2 mmol) were stirred in DMF at rt. b) Isolated yield. c) Cu₂O (0.05 mmol) was used. d) The numbers in parentheses are the yields of 2a recovered. e) At 50°C.

Table 2. Conversion of the Coupling Product 3aa to the Cyclized Product 1aa^a)

Entry	Solvent	Volume ratio H2O : Solvent	Time (h)	(±)-1aa Yield (%)b)
1	MeOH	2 : 1	1.5	56
2	EtOH	2 : 1	5	55
3	BuOH	2 : 1	12	61
4	MeCN	2 : 1	12	41
5	THF	2 : 1	12	No reaction
6	Acetone	2 : 1	12	37
7	Dioxane	2 : 1	12	0
8	DMF	2 : 1	4	39
9	MeOH	MeOH only	36	Trace
10 c)	MeOH	2 : 1	2	52
11d)	MeOH	2 : 1	4	52
12	MeOH	1 : 1	4	39
13	MeOH	1 : 2	20	44
14e)	MeOH	2 : 1	5	74
15  f–h)	MeOH	2 : 1	5	66
16  f,h,i)	MeOH	2 : 1	5	77

a) A solution of the substrate 3 (0.11 mmol) in H₂O and solvent (total volume: 30 mL) were refluxed. b) Isolated yield. c) Total volume was 9 mL. d) Total volume was 90 mL. e) DMF (500 µL) was used. f) Substrate 3 (0.50 mmol) was used. g) Total volume was 45 mL. h) DMF (2.5 mL) was used. i) Total volume was 144 mL.

We tested a range of reaction conditions to evaluate the potential of the general synthetic methods for heterocycle-fused naphthoquinones. The reaction of the substrate 3aa with Na₂S in H₂O and MeOH (H₂O–MeOH=1 : 10) proceeded smoothly to give thiophene-fused naphthoquinone 1bb in 88% yield (Table 3, entry 1). However, the replacement of the dimethylamino group of 3aa by an amino group, without a second cyclization, occurred when aqueous NH₃ solution was used. Adding an excess of K₂CO₃ to the reaction mixture resulted in conversion to the cyclized product 1cc in 57% yield (Table 3, entry 2). Moreover, these reaction conditions were suitable for the reaction with methylamine although sodium hydroxide was used as the base, giving N-methylated pyrrole-fused naphthoquinones 1dd in 72% yield. We conducted further investigations using various substrates and reacted the coupling products 3ba, 3ab, 3ac, and 3ca under conditions A–D (detailed in Table 3). The reaction of 3ba, which contains a more general naphthoquinone motif, using reaction conditions A–C gave the cyclized products 1ee, 1ff, and 1gg, respectively, with rather lower chemical yields than those obtained for 1bb or 1dd. (Table 3, entries 4–6). The substrate 3ab, which has a phenyl substituent, gave the desired product 1hh in moderate yield, whereas a lower yield was surprisingly found when the substrate 3ac, which has a 2-phenylethyl substituent, was used (Table 3, entries 7, 8). Unsatisfactory yields were obtained when the naphthoquinone 3ca, which has a hydroxyl group at C-8, was used as the substrate (Table 3, entry 9). TLC analysis during the reaction shown in the entry 9 of the Table 3 revealed that the consumption of the substrate was slower in this reaction than that while using the substrate shown in the entry 14 of the Table 2, under the same reaction conditions. Thus, we speculated that the lower reactivity of the substrate 3ca in the first addition-elimination step causes degradation to unknown compounds during the reaction, and the intramolecular hydrogen bond between carbonyl oxygen at C-4 and hydroxyl group at C-5 plays a crucial role in the addition-elimination step.

Table 3. Conversion of the Coupling Product 3 to the Cyclized Product 1

Entry	Substrate	Time (h)	Conditions	Product	Yield (%)a)
1	3aa	15	Na2S, H2O–MeOH (1 : 10), rt (condition B)	1bb	88
2	3aa	3	28% NH3 aq, K2CO3, DMF, at 80°C (condition C)	1cc	57
3	3aa	3	CH3NH2·HCl, Et3N, NaOH, NMP–H2O (10 : 1), at 80°C (Condition D)	1dd	72
4	3ba	20	H2O–MeOH (2 : 1), reflux (condition A)	1ee	47
5	3ba	24	Condition B	1ff	56
6	3ba	3	Condition C	1gg	38
7	3ab	24	Condition A	1hh	70
8	3ac	24	Condition A	1ii	40
9	3ca	24	Condition A	1jj	30

a) Isolated yield. Condition A: Substrate 3 (0.5 mmol) were refluxed in H₂O–MeOH (2 : 1) (total volume: 144 mL). Condition B: Substrate 3 (0.5 mmol) and Na₂S (1.0 mmol) were stirred in H₂O–MeOH (1 : 10) (total volume: 11 mL) at rt. Condition C: Substrate 3 (0.5 mmol), 28% NH₃ aq. (5.0 mmol) and K₂CO₃ (2.5 mmol) were stirred in the solvent (30 mL) at 80–100°C. Condition D: Substrate 3 (0.5 mmol), CH₃NH₂·HCl (1.5 mmol), Et₃N (1.5 mmol) and NaOH (5.0 mmol) were stirred in the solvent (33 mL) at 80°C.

Finally, the ability of (S)-1bb, which was synthesized from 3aa and commercially available (S)-4a using the developed method, to suppress the growth of human tumor cell lines, including A549 (lung) and MCF-7 (breast), was tested. The compound (S)-1bb exhibited comparable antiproliferative effects to that of β-lapachone against both cell lines, with the IC₅₀ of (S)-1bb and β-lapachone against A549 being 5.57 and 4.21 µm, respectively, and the IC₅₀ of (S)-1bb and β-lapachone against MCF-7 being 5.57 and 9.96 µm, respectively. β-Lapachone is known as a potent anticancer compound contained in Tabebuia avellanedae similarly to 1aa and β-lapachone-based anticancer drug is recently developing in U.S.A.³⁸⁾ Therefore, heterocycle-fused naphthoquinone analogues are promising anticancer compounds.

Conclusion

In conclusion, we developed a general method to construct heterocycle-fused naphthoquinones utilizing Sonogashira coupling and tandem addition-elimination/intramolecular cyclization. The future applications of this strategy and the biological activities of the various naphthoquinones synthesized are currently being investigated and will be reported in due course.

Experimental

General

All melting points are uncorrected. ¹H- and ¹³C-NMR spectra (500 MHz for ¹H and 125 MHz for ¹³C) were obtained in CDC1₃, unless otherwise noted. The chemical shift values are expressed in ppm relative to internal tetramethylsilane. Abbreviations are as follows: s, singlet; d, doublet; t, triplet; m, multiplet; br, broad. IR is expressed in cm⁻¹. Purification was performed using silica gel column chromatography. All reagents were purchased from chemical companies and used as received. All reactions were conducted under an argon atmosphere, unless otherwise stated.

3-Bromo-2-(dimethylamino)-5-hydroxynaphthalene-1,4-dione (2a)³⁹⁾: To a solution of 2-(dimethylamino)-5-hydroxynaphthalene-1,4-dione (500 mg, 2.30 mmol) in DMF (15 mL) was added a solution of N-bromosuccinimide (NBS) (491 mg, 2.76 mmol) in DMF (6 mL), and the mixture was stirred for 0.5 h at rt. The mixture was quenched with H₂O at 0°C and extracted with EtOAc. The organic extracts were washed with H₂O and brine, dried over Na₂SO_4, and then concentrated. The column chromatography (hexane–EtOAc=12 : 1) gave 2a (656 mg, 96% yield) as red solid with mp 90–91°C. ¹H-NMR δ: 3.29 (6H, s), 7.18–7.20 (1H, m), 7.46–7.50 (2H, m), 12.45 (1H, s). ¹³C-NMR δ: 45.0, 109.7, 113.7, 119.5, 124.4, 131.3, 134.6, 154.3, 160.5, 181.2, 183.1.

2-Bromo-3-(dimethylamino)naphthalene-1,4-dione (2b)³⁵⁾: To the solution of 2-bromonaphthalene-1,4-dione (2.00 g, 8.44 mmol) in toluene (100 mL) was added dimethylamine (12.6 mL, 2.0 m solution in THF, 25.3 mmol) at 0°C. After the mixture was stirred for 12 h, the solvent was removed under reduced pressure. The residue was purified by column chromatography (hexane–EtOAc=4 : 1) to give 2-(dimethylamino)naphthalene-1,4-dione (2.13 g, 90% yield) as red solid. To a solution of 2-(dimethylamino)naphthalene-1,4-dione (1.50 g, 7.50 mmol) in DMF (30 mL) was added a solution of NBS (1.60 g, 9.00 mmol) in DMF (15 mL), and the mixture was stirred for 0.5 h at rt. The mixture was quenched with H₂O at 0°C and extracted with EtOAc. The organic extracts were washed with H₂O and brine, dried over Na₂SO_4, and then concentrated. The column chromatography (hexane–EtOAc=12 : 1) gave 2b (1.60 g, 76% yield) as red solid with mp 110–112°C. ¹H-NMR δ: 3.25 (6H, s), 7.61–7.69 (2H, m), 7.96–7.98 (1H, m), 8.07–8.09 (1H, m). ¹³C-NMR δ: 44.7, 112.9, 126.6, 126.7, 131.2, 131.3, 132.7, 133.9, 153.5, 177.8, 181.8.

2-Bromo-3-(dimethylamino)-5-hydroxynaphthalene-1,4-dione (2c)³⁹⁾: To a solution of 2-(dimethylamino)-8-hydroxynaphthalene-1,4-dione (1.0 g, 4.61 mmol) in DMF (30 mL) was added a solution of NBS (980 mg, 5.53 mmol) in DMF (12 mL), and the mixture was stirred for 0.5 h at rt. The mixture was quenched with H₂O at 0°C and extracted with EtOAc. The organic extracts were washed with H₂O and brine, dried over Na₂SO₄, and then concentrated. The column chromatography (hexane–EtOAc=12 : 1) gave 2c (980 mg, 72% yield) as red solid with mp 109–111°C. ¹H-NMR δ: 3.25 (6H, s), 7.17 (1H, dd, J= 1.1, 8.3 Hz), 7.54–7.58 (1H, m), 7.65 (1H, dd, J=1.1, 7.5 Hz), 11.81 (1H, s). ¹³C-NMR δ: 44.9, 114.3, 114.8, 119.5, 123.3, 131.5, 136.6, 153.0, 161.8, 177.4, 186.9.

General Procedure for Sonogashira Reaction of Naphthoqionone 2 with Acetylene 4

Under Ar atmosphere, a mixture of Cu₂O (14 mg, 0.10 mmol), acetylene 4 (1.00 mmol), K₂CO₃ (28 mg, 0.20 mmol), and pyridine (8.0 mL, 100 mmol) was stirred for 2 h at rt. A solution of compound 2 (0.50 mmol) and Pd(OAc)₂ (3.4 mg, 0.015 mmol) in DMF (5.0 mL) was added to this suspension, and the reaction mixture was stirred at rt. The mixture was quenched with H₂O at 0°C and extracted with CHCl₃. The organic extracts were washed with H₂O and brine, dried over Na₂SO₄, and then concentrated.

2-(Dimethylamino)-5-hydroxy-3-(3-hydroxybut-1-yn-1-yl)naphthalene-1,4-dione (3aa): Starting from 2a and 4a, this compound was prepared according to the general procedure. The column chromatography (hexane–EtOAc=3 : 1) gave 3aa (120 mg, 84% yield) as purple solid with mp 126–128°C. ¹H-NMR δ: 1.57 (3H, d, J=6.6 Hz), 3.42 (6H, s), 4.86 (1H, q, J=6.6 Hz), 7.18–7.20 (1H, m), 7.45–7.47 (2H, m), 12.68 (1H, s). ¹³C-NMR δ: 24.0, 45.3, 59.1, 78.3, 103.0, 104.2, 114.5, 119.0, 124.6, 132.2, 134.5, 155.5, 160.4, 182.6, 187.7. IR (KBr): 3464, 1624, 1547, 1473, 1068, 935, 895, 779. High resolution (HR)-MS (electrospray ionization (ESI)) m/z: [M+H]⁺ Calcd for [C₁₆H₁₆NO₄]⁺, 286.1079; Found, 286.1077.

2-(Dimethylamino)-5-hydroxy-3-(phenylethynyl)naphthalene-1,4-dione (3ab): Starting from 2a and 4b, this compound was prepared according to the general procedure. The column chromatography (hexane–EtOAc=12 : 1) gave 3ab (147 mg, 93% yield) as purple solid with mp 128–129°C. ¹H-NMR δ: 3.47 (6H, s), 7.17–7.19 (1H, m), 7.31–7.35 (3H, m), 7.43–7.48 (2H, m), 7.50–7.53 (2H, m), 12.78 (1H, s). ¹³C-NMR δ: 45.1, 84.2, 102.4, 104.1, 114.5, 118.8, 123.4, 124.4, 128.3, 128.4, 130.9, 132.2, 134.4, 154.9, 160.4, 182.4, 187.4. IR (KBr): 1680, 1618, 1338, 1161, 1067, 922, 880, 754. HR-MS (ESI) m/z: [M+Na]⁺ Calcd for [C₂₀H₁₅NO₃Na]⁺, 340.0950; Found, 340.0939.

2-(Dimethylamino)-5-hydroxy-3-(4-phenylbut-1-yn-1-yl)naphthalene-1,4-dione (3ac): Starting from 2a and 4c, this compound was prepared according to the general procedure at 50°C. The column chromatography (hexane–EtOAc=12 : 1) gave 3ac (159 mg, 92% yield) as purple solid with mp 88–90°C. ¹H-NMR δ: 2.85–2.89 (2H, dd), 2.93–2.97 (2H, dd), 3.17 (6H, s), 7.15–7.32 (6H, m), 7.43–7.44 (2H, m), 12.80 (1H, s). ¹³C-NMR δ: 22.1, 34.7, 44.7, 75.5, 103.0, 104.7, 114.4, 118.7, 124.3, 126.3, 128.4, 128.5, 132.1, 134.3, 140.5, 155.2, 160.3, 182.6, 187.9. IR (KBr): 2920, 1666, 1616, 1475, 1383, 1066, 947, 845, 783, 762. HR-MS (ESI) m/z: [M+Na]⁺ Calcd for [C₂₂H₁₉NO₃Na]⁺, 368.1263; Found, 368.1255.

2-(Dimethylamino)-3-(3-hydroxybut-1-yn-1-yl)naphthalene-1,4-dione (3ba): Starting from 2b and 4a, this compound was prepared according to the general procedure. The column chromatography (hexane–EtOAc=2 : 1) gave 3ba (122 mg, 91% yield) as purple solid with mp 117–118°C. ¹H-NMR δ: 1.55 (3H, d, J=6.6 Hz), 3.35 (6H, s), 4.86 (1H, q, J=6.6 Hz), 7.56 (1H, ddd, J=1.4, 7.5, 7.5 Hz), 7.63 (1H, ddd, J=1.4, 7.5, 7.5 Hz), 7.88 (1H, dd, J=1.4, 7.5 Hz), 8.00 (1H, dd, J=1.4, 7.5 Hz). ¹³C-NMR δ: 23.8, 44.9, 58.8, 78.9, 104.5, 104.6, 125.8, 126.3, 132.0, 132.3, 132.4, 133.9, 154.6, 182.2, 183.0. IR (KBr): 3423, 1670, 1547, 1340, 1279, 1016, 978, 723. HR-MS (ESI) m/z: [M+H]⁺ Calcd for [C₁₆H₁₆NO₃]⁺, 270.1130; Found, 270.1136.

3-(Dimethylamino)-5-hydroxy-2-(3-hydroxybut-1-yn-1-yl)naphthalene-1,4-dione (3ca): Starting from 2c and 4a, this compound was prepared according to the general procedure. The column chromatography (hexane–EtOAc=3 : 1) gave 3ca (88 mg, 62% yield) as purple solid with mp >135°C (dec). ¹H-NMR δ: 1.57 (3H, d, J=6.6 Hz), 3.42 (6H, s), 4.87 (1H, q, J=6.6 Hz), 7.13 (1H, dd, J=1.5, 7.9 Hz), 7.55 (1H, dd, J=7.5, 7.9 Hz), 7.59 (1H, dd, J=1.5, 7.5 Hz), 11.87 (1H, s). ¹³C-NMR δ: 23.9, 45.3, 59.0, 78.8, 105.5, 106.5, 114.9, 118.6, 123.0, 132.4, 136.9, 153.9, 161.7, 181.6, 187.8. IR (KBr): 1680, 1618, 1550, 1477, 1386, 1296, 1066, 922, 754. HR-MS (ESI) m/z: [M+H]⁺ Calcd for [C₁₆H₁₆NO₄]⁺, 286.1079; Found, 286.1077.

General Procedure for Tandem Addition-Elimination/Intramolecular Cyclization Reaction

Condition A: A solution of compound 3 (0.50 mmol) in DMF (2.5 mL) was added to the solution of H₂O (96 mL) and MeOH (48 mL) and was stirred at reflux. The mixture was extracted with EtOAc and the organic extracts were washed with brine, dried over Na₂SO₄, and then concentrated.

Condition B: To a suspension of Na₂S (78 mg, 1.00 mmol) in MeOH (10 mL) and H₂O (1 mL) was added compound 3 (0.50 mmol), then stirred at rt. The mixture was quenched with H₂O at 0°C and extracted with EtOAc. The organic extracts were washed with H₂O and brine, dried over Na₂SO_4, and then concentrated.

Condition C: To a solution of compound (0.50 mmol) in DMF (30 mL) was added 28% aqueous NH₃ (0.40 mL, 5.00 mmol), and then, the mixture was stirred at 80°C. TLC of the mixture (hexane–EtOAc=1 : 1) showed absence of starting material. After adding K₂CO₃ (345 mg, 2.50 mmol), the reaction mixture was stirred at 80°C. The mixture was extracted with CHCl₃. The organic extracts were washed with H₂O and brine and dried over Na₂SO₄ and then concentrated.

Condition D: A mixture of compound 3 (0.50 mmol), MeNH₂· HCl (101 mg, 1.50 mmol) and Et₃N (0.21 mL, 1.50 mmol) in NMP (30 mL) was stirred for 0.5 h at 80°C. A solution of NaOH (200 mg, 5.00 mmol) in H₂O (3 mL) was added to this mixture and was stirred at 80°C. The mixture was extracted with CHCl₃. The organic extracts were washed with H₂O and brine and dried over Na₂SO₄ and then concentrated.

5-Hydroxy-2-(1-hydroxyethyl)naphtho[2,3-b]furan-4,9-dione (1aa)²⁾: Starting from 3aa, this compound was prepared according to the general procedure (Condition A). The column chromatography (hexane–EtOAc=4 : 1) gave 1aa (100 mg, 77% yield) as yellow solid with mp 164–165°C. ¹H-NMR δ (CDCl₃): 1.66 (d, 3H, J=6.6 Hz), 2.23 (d, 1H, J=5.3 Hz), 5.05 (m, 1H), 6.85 (d, 1H, J=0.7 Hz), 7.28 (dd, 1H, J=1.2, 8.5 Hz), 7.62 (dd, 1H, J=7.6, 8.5 Hz), 7.76 (dd, 1H, J=1.2, 7.6 Hz), 12.18 (s, 1H). ¹³C-NMR (CDCl₃) δ: 21.5, 63.8, 103.4, 115.2, 120.0, 125.3, 131.0, 132.7, 136.3, 152.1, 162.3, 165.4, 172.7, 186.5.

5-Hydroxy-2-(1-hydroxyethyl)naphtho[2,3-b]thiophene-4,9-dione (1bb): Starting from 3aa, this compound was prepared according to the general procedure (Condition B). The column chromatography (hexane–EtOAc=4 : 1) gave 1bb (121 mg, 88% yield) as yellow solid with mp 179–181°C. ¹H-NMR (CDCl₃–MeOD) δ: 1.62 (3H, d, J=6.5 Hz), 5.14 (1H, q, J=6.5 Hz), 7.26 (1H, dd, J=1.0, 8.3 Hz), 7.47 (1H, s), 7.62 (1H, dd, J=7.5, 8.3 Hz), 7.74 (1H, dd, J=1.0, 7.5 Hz), 12.33 (1H, s). ¹³C-NMR (CDCl₃–MeOD) δ: 25.1, 65.9, 115.7, 120.0, 121.1, 124.8, 133.9, 136.3, 142.6, 144.0, 162.4, 162.6, 177.7, 185.3. IR (KBr): 3290, 1634, 1454, 1296, 1223, 1093, 752, 702. HR-MS (ESI) m/z: [M+Na]⁺ Calcd for [C₁₄H₁₀SO₄Na]⁺, 297.0198; Found, 297.0207.

(S)-1bb: Pale yellow needles with mp 209–211°C. [α]_D²⁵ −10.8 (c=0.12, CHCl₃) for >99% ee (HPLC, Daicel Chiralpak AD-H, hexane–i-PrOH=9 : 1, 1.0 mL/min, 254 nm, minor: 24.9 min and major: 30.4 min).

5-Hydroxy-2-(1-hydroxyethyl)-1H-benzo[f]indole-4,9-dione (1cc): Starting from 3aa, this compound was prepared according to the general procedure (Condition C). The column chromatography (hexane–EtOAc=4 : 1) gave 1cc (73 mg, 57% yield) as yellow solid with mp >215°C (dec). ¹H-NMR (dimethyl sulfoxide (DMSO)) δ: 1.49 (3H, d, J=6.5 Hz), 4.83 (1H, dq, J=5.3, 6.5 Hz), 5.50 (1H, d, J=5.3 Hz), 6.63 (1H, s), 7.29 (1H, dd, J=1.2, 8.3 Hz), 7.63 (1H, dd, J=1.2, 7.4 Hz), 7.71 (1H, dd, J=7.4, 8.3 Hz), 12.69 (1H, s), 12.93 (1H, s). ¹³C-NMR (DMSO) δ: 23.9, 62.6, 104.5, 115.7, 119.1, 124.4, 126.8, 132.4, 134.1, 136.4, 148.8, 161.8, 174.1, 187.5. IR (KBr): 3290, 1628, 1253, 1207, 1097, 827, 764, 704. HR-MS (ESI) m/z: [M+Na]⁺ Calcd for [C₁₄H₁₁NO₄Na]⁺, 280.0586; Found, 280.0575.

5-Hydroxy-2-(1-hydroxyethyl)-1-methyl-1H-benzo[f]indole-4,9-dione (1dd)³³⁾: Starting from 3aa, this compound was prepared according to the general procedure (Condition D). The column chromatography (hexane–EtOAc=3 : 1) gave 1ee (98 mg, 72% yield) as yellow solid with mp 219°C. ¹H-NMR δ: 1.69 (3H, d, J=6.5 Hz), 4.13 (3H, s), 4.95 (1H, q, J=6.5 Hz), 6.68 (1H, s), 7.18 (1H, dd, J=1.1, 8.4 Hz), 7.54 (1H, dd, J=7.3, 8.4 Hz), 7.65 (1H, dd, J=1.1, 7.3 Hz), 12.58 (1H, s). ¹³C-NMR δ: 22.2, 33.4, 62.1, 105.1, 115.5, 119.2, 124.1, 126.8, 131.8, 134.4, 135.4, 145.4, 162.0, 175.7, 186.8.

2-(1-Hydroxyethyl)naphtho[2,3-b]furan-4,9-dione (1ee): Starting from 3ba, this compound was prepared according to the general procedure (Condition A). The column chromatography (hexane–EtOAc=2 : 1) gave 1ee (57 mg, 47% yield) as yellow solid with mp 153–155°C. ¹H-NMR δ: 1.59 (d, 3H, J=6.7 Hz), 4.98 (1H, q, J=6.7 Hz), 6.79 (1H, s), 7.66–7.71 (2H, m), 8.10–8.16 (2H, m). ¹³C-NMR δ: 21.5, 63.9, 103.8, 126.9, 127.0, 131.3, 132.4, 133.1, 133.8, 134.0, 152.0, 165.1, 173.5, 180.7. IR (KBr): 3350, 1680, 1591, 1537, 1365, 1219, 1196, 1103, 955, 713. HR-MS (ESI) m/z: [M+H]⁺ Calcd for [C₁₄H₁₁O₄]⁺, 243.0657; Found, 243.0670.

2-(1-Hydroxyethyl)naphtho[2,3-b]thiophene-4,9-dione (1ff): Starting from 3ba, this compound was prepared according to the general procedure (Condition B). The column chromatography (hexane–EtOAc=2 : 1) gave 1ff (72 mg, 56% yield) as yellow solid with mp 175–176°C. ¹H-NMR δ: 1.67 (3H, d, J=6.3 Hz), 5.21 (1H, q, J=6.3 Hz), 7.74–7.76 (2H, m), 8.15–8.31 (2H, m). ¹³C-NMR δ: 25.3, 66.5, 121.9, 126.9, 127.2, 133.3, 133.6, 133.7, 133.8, 142.9, 144.0, 160.4, 178.1, 179.6. IR (KBr): 3280, 1668, 1593, 1460, 1325, 1301, 1265, 1096, 710. HR-MS (ESI) m/z: [M+Na]⁺ Calcd for [C₁₄H₁₀SO₃Na]⁺, 281.0248; Found, 281.0242.

2-(1-Hydroxyethyl)-1H-benzo[f]indole-4,9-dione (1gg): Starting from 3ba (0.19 mmol), this compound was prepared according to the general procedure (Condition C). The column chromatography (hexane–EtOAc=4 : 1) gave 1gg (17 mg, 38% yield) as yellow solid with mp >188°C (dec). ¹H-NMR (CDCl₃–MeOD) δ: 1.58 (3H, d, J=6.6 Hz), 4.94 (1H, q, J=6.6 Hz), 6.60 (1H, s), 7.67–7.70 (2H, m), 8.10–8.15 (2H, m). ¹³C-NMR (CDCl₃–MeOD) δ: 22.9, 63.3, 104.7, 126.4, 126.9, 128.2, 132.2, 133.3, 133.4, 133.7, 134.2, 146.2, 175.8, 181.9. IR (KBr): 3340, 1630, 1513, 1371, 1200, 920, 723. HR-MS (ESI) m/z: [M+H]⁺ Calcd for [C₁₄H₁₂NO₃]⁺, 242.0817; Found, 242.0805.

5-Hydroxy-2-phenylnaphtho[2,3-b]furan-4,9-dione (1hh): Starting from 3ab, this compound was prepared according to the general procedure (Condition A). The column chromatography (hexane–EtOAc=6 : 1) gave 1hh (101 mg, 70% yield) as red solid with mp >237°C (dec). ¹H-NMR (pyridine) δ: 7.36 (1H, dd, J=1.2, 8.5 Hz), 7.42–7.48 (3H, m), 7.57 (1H, s), 7.60–7.62 (1H, m), 7.88 (1H, dd, J=1.2, 7.5 Hz), 7.91–7.94 (2H, m), 12.55 (1H, s). ¹³C-NMR (pyridine) δ: 103.3, 115.6, 119.5, 122.7, 124.8, 125.6, 128.4, 129.3, 130.3, 133.4, 136.5, 150.0, 160.2, 162.2, 172.1, 193.0. IR (KBr): 3120, 1635, 1535, 1483, 1452, 1221, 823, 762. HR-MS (ESI) m/z: [M+Na]⁺ Calcd for [C₁₈H₁₀O₄Na]⁺, 313.0477; Found, 313.0485.

5-Hydroxy-2-phenethylnaphtho[2,3-b]furan-4,9-dione (1ii): Starting from 3ac, this compound was prepared according to the general procedure (Condition A). The column chromatography (hexane–EtOAc=6 : 1) gave 1ii (64 mg, 40% yield) as yellow solid with mp 167–168°C. ¹H-NMR δ: 3.06–3.16 (4H, m), 6.54 (1H, s), 7.19–7.32 (6H, m), 7.60 (1H, dd, J=7.4, 8.3 Hz), 7.74 (1H, dd, J=1.1, 7.4 Hz), 12.17 (1H, s). ¹³C-NMR δ: 30.1, 33.5, 104.4, 115.2, 119.8, 125.0, 126.6, 128.2, 128.6, 131.4, 132.8, 136.1, 139.7, 151.8, 162.2, 163.7, 172.4, 186.7. IR (KBr): 1678, 1635, 1472, 1379, 1043, 845, 762. HR-MS (ESI) m/z: [M+Na]⁺ Calcd for [C₂₀H₁₄O₄Na]⁺, 341.0790; Found, 341.0789.

8-Hydroxy-2-(1-hydroxyethyl)naphtho[2,3-b]furan-4,9-dione (1jj)²⁾: Starting from 3ca, this compound was prepared according to the general procedure (Condition A). The column chromatography (hexane–EtOAc=4 : 1) gave 1jj (39 mg, 30% yield) as yellow solid with mp 173–175°C. ¹H-NMR δ: 1.66 (3H, d, J=6.6 Hz), 5.05 (1H, q, J=6.6 Hz), 6.86 (1H, s), 7.26 (1H, dd, J=1.0, 8.4 Hz), 7.60 (1H, m), 7.71 (1H, dd, J=1.0, 7.4 Hz), 12.02 (1H, s). ¹³C-NMR δ: 21.5, 63.8, 104.2, 114.7, 120.1, 125.2, 131.9, 133.2, 136.3, 151.2, 162.6, 165.8, 178.5, 179.7.

Antiproliferative Effect Assay

The antiproliferative effects of (S)-1bb was examined in cancer cell lines. These cells were maintained in usual 10% fetal serum Dulbecco’s minimum essential medium (DMEM) through experiments and exposed to four dose concentrations of (S)-1bb in a humidified atmosphere (37°C, 5% CO₂) for 72 h. After the reaction, cells were further incubated with 0.25% trypan blue dye for 20 min and counted for viable cells under light microscopic apparatus. IC₅₀ values were calculated from separate experiments performed in triplicate.

Acknowledgment

This study was supported in part by Grants-in-Aid for Young Scientists (B) (KAKENHI: 24790116) to MY. The authors are grateful to Taheebo Japan Co., Ltd. and SANSHIN METAL WORKING Co., Ltd. for their generous financial support to this project.

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