Regioselective Suzuki–Miyaura reactions of 4,7-dichloro-N-methylisatin. Synthesis, anti-HIV activity and modeling study

Abstract

Suzuki–Miyaura reactions of 4,7-dichloro-N-methylisatin provide a convenient access to arylated methylisatins. The reactions proceed with excellent site-selectivity in favour of position 4, due to electronic reasons. All the new compounds were evaluated in vitro for their antiviral activity against the replication of HIV-1 and HIV-2 in MT4 cells using a MTT assay. 4-(4-Acetylphenyl)-7-chloro-1-methylindoline-2,3-dione (8l), containing an acetyl group located at carbon C(4) of the isatin backbone, showed an IC₅₀ value of >3.47 μM against HIV-2 with a therapeutic index (SI) of 4. This means that 8l was cytotoxic to MT-4 cells at a CC₅₀ value of 13.43 μM; also 4,7-dichloro-1-methylindoline-2,3-dione (5), 7-chloro-4-(4-chlorophenyl)-1-methylindoline-2,3-dione (8c), 7-chloro-4-(4-ethoxyphenyl)-1-methylindoline-2,3-dione (8g) and 7-chloro-1-methyl-4-(4-vinylphenyl)indoline-2,3-dione (8m) were cytotoxic to MT-4 cells within a 2.21–3.11 μM concentration range. In a docking study, 8l interacted with several amino acids in the reverse transcriptase (RT) binding site of HIV-2.

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Introduction

Isatin derivatives (1H-indole-2,3-diones) show diverse biological activities,¹ such as antibacterial,² antifungal,³ antiinflammatory⁴ and anticonvulsant activity.⁵ A variety of substituted isatin derivatives have been synthesized to date. The possibility of isatin to be attacked by nucleophiles at carbon C-3 allowed for the synthesis of a large number of 3-substituted isatins. This is also reflected by numerous biologically active 3-substituted indolin-2-ones that have been reported in the literature.^6–8 Most recently, Grewal and coworkers⁹ have extensively studied the synthesis and biological screening of isatin derivatives. Furthermore, isatin derivatives have received considerable attention, due to their potent anticancer activities.^10–12 Meanwhile, Liu and colleagues¹³ identified a class of isatin O-acyl oximes that selectively inhibit neuronal ubiquitin C-terminal hydrolase (UCH-L1) in a H1299 lung cancer cell line, which is proposed to be linked to tumor progression upon upregulation. Lee et al.¹⁴ reported a novel indirubin analog, indirubin-5-nitro-3′-monoxime (1) which inhibited cell proliferation against various human cancer cells, meanwhile sunitinib malate (Sutent®) (2) had been approved by FDA for the treatment of advanced renal carcinoma,¹⁵ and gastrointestinal stromal tumors.¹⁶ Vine et al.¹⁷ have reported that the introduction of electron withdrawing groups at the benzene ring of isatin are generally found to induce cancer cell death via apoptosis.

Owing to the broad spectrum of chemotherapeutic properties of isatin derivatives, several researchers^18–24 have found that such derivatives are ideal drugs for AIDS treatment, because they suppress HIV replication. Examples of such isatin derivatives are 3-[(4-amino-5(3,4,5-trimethoxybenzyl)pyrimidin-2-yl)imino)-5-bromo-1-(morpholinomethyl)indolin-2-one (3) and its N-acetyl derivative.²⁵ Furthermore, several isatin derivatives showed remarkable anti-HIV activity, like sulfonamide-benzene derivatives and Schiff and Mannich bases of isatin.^26–28 We report herein a convenient approach to arylated isatins by what are, to the best of our knowledge, the first Suzuki–Miyaura cross-coupling reactions of 4,7-dichloro-1-methylindoline-2,3-dione. The reactions proceed with very good regioselectivity in favor of position 4, due to electronic reasons. Besides, the new arylated isatin derivatives were evaluated for their anti-HIV activity. In addition, the molecular modeling structure was studied (Fig. 1).

Fig. 1 Some potentially active isatin derivatives.

Results and discussion

Chemistry

Commercially available 4,7-dichloroisatin (4) was converted, by a known procedure, to N-methyl-4,7-dichloroisatin (5) in 90% yield by using methyl iodide, DMF as a solvent and K₂CO₃ as the base (Scheme 1).²⁹ Treatment of 5 with arylboronic acids 6a–f (2.0 equiv.), applying a Suzuki–Miyaura reaction, afforded N-methyl-4,7-diarylisatins 7a–f in 52–82% yield (Scheme 1). Both electron-poor and electron-rich arylboronic acids could be successfully employed. The best yields were obtained by using Pd(PPh₃)₄ (6 mol%) as the catalyst and K₃PO₄ (3.0 equiv) as the base in dioxane at 120 °C for 8 h (Table 1).

Scheme 1 Reagent and conditions: (i) (1) K₂CO₃ (1.2 equiv.), DMF (1 mL per 0.1 mmol of isatin) 1 h, 4 to 20 °C; (2) CH₃I (1.1 equiv.), KI (cat., 0.2 equiv.), 80 °C, 5 h; (ii) ArB(OH)₂ (6a–f) (2.0 equiv.), K₃PO₄ (3.0 equiv.), Pd(PPh₃)₄ (6 mol%), 1,4-dioxane, 120 °C, 8 h.

Table 1 Synthesis of 7a–f

6, 7	Ar	7^a (%)
a Yields of isolated products.
6a, 7a	3,5-MeC6H3	61
6b, 7b	4-(MeO)C6H4	82
6c, 7c	4-ClC6H4	52
6d, 7d	4-MeC6H4	58
6e, 7e	4-EtC6H4	65
6f, 7f	4-FC6H4	72

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An optimization of the synthesis of 7b was carried out by using various reaction conditions, such as K₂CO₃, KF and NEt₃ as bases, various solvents, like toluene, DMF and THF, and Pb(OAc)₂ and Pd(PPh₃)₂Cl₂ as catalysts (Table 2). The use of standard conditions, K₃PO₄ (3.0 equiv.), Pd(PPh₃)₄ (6 mol%) in dioxane at 120 °C for 8 h proved to give the best yields of 7b (82%).

Table 2 Optimization of the synthesis of 7b

Entry	Base	Solvent	T (°C)	Catalysts	t (h)	Yield^a (%)
a Yields of isolated yields.
1	K3PO4	Toluene	100	Pd(OAc)2	8	25
2	K2CO3	DMF	130	Pd(PPh3)4	9	38
3	KF	Dioxane	80	Pd(OAc)2	10	47
4	NEt3	THF	65	Pd(PPh3)2Cl2	7	34
5	K3PO4	Dioxane	120	Pd(PPh3)4	8	82
6	K2CO3	Toluene	90	Pd(OAc)2	10	25

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The Suzuki–Miyaura reaction of 5 with arylboronic acids 6 (1.0 equiv.) afforded the 7-chloro-4-aryl-isatins 8a–d, f–m in 49–87% yields and with very good site-selectivity (Scheme 2). The best conditions, avoiding double-coupling, require the use of 1.2 equiv. of the arylboronic acid at 70 °C instead of 120 °C (reaction time 6 h). Both electron-poor and electron-rich arylboronic acids were successfully used (Table 3).

Scheme 2 Reagents and conditions: (i) 5 (1.0 equiv), ArB(OH)₂, 6a–d, f–m (1.2 equiv.), K₃PO₄ (1.5 equiv.), Pd(PPh₃)₄ (3 mol%), dioxane, 70 °C, 6 h.

Table 3 Synthesis of 8a–d, f–m

6, 8	Ar	8^a (%)
a Yields of isolated products.
6a, 8a	3,5-(Me)2C6H3	63
6b, 8b	4-(MeO)C6H4	83
6c, 8c	4-ClC6H4	52
6d, 8d	4-MeC6H4	79
6f, 8f	4-FC6H4	87
6g, 8g	4-(EtO)C6H4	85
6h, 8h	4-iProC6H4	73
6i, 8i	4-tBuC6H4	78
6j, 8j	3-MeC6H4	51
6k, 8k	3,5-(MeO)2C6H3	87
6l, 8l	4-(Acetyl)C6H4	53
6m, 8m	4-(Vinyl)C6H4	49

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The structures of the newly prepared compounds were confirmed by their IR, ¹H, ¹³C NMR and mass spectra, where 7a–f, 8a–m and 9a–c exhibited additional signals for the protons of the newly introduced aromatic ring. The aromatic protons have been assigned (cf. Experimental section). In the ¹³C NMR spectra, the carbonyl carbon atom at C-2 of the isatin ring resonated at δ 158.4–168.3 ppm, while the lower field resonances at δ 178.2–181.9 ppm were assigned to the carbonyl carbon atoms at C-3 of isatin moiety. C-5 of the isatin ring were observed at δ 124.1–127.4 ppm, while the resonances at δ 144.4–148.3 ppm were assigned to C-7a of the fused nucleus. In addition, the NMe group appeared at δ 28.6–33.7 ppm. Compound 8b was selected for further study via the HMBC³⁰ and NOESY³¹ NMR spectroscopic measurements. The gradient-selected HMBC spectrum of 8b showed a ³J_C,H heteronuclear correlation of C-4 of isatin ring (δ_C 133.9 ppm) to the H-2′ proton (δ_H 7.32 ppm) of the aromatic moiety at C-4 of isatin. The ¹H, ¹H NOESY spectrum was characterized by two correlations: one indicated by correlation of protons of methoxy group at δ_H 3.72 ppm with H-5′ of the same aromatic ring at δ_H 6.87 ppm, while the other one was observed between the H-6′ of the aromatic ring at δ_H 7.32 ppm with H-5 of the isatin ring at δ_H 6.87 ppm (Fig. 2).

Fig. 2 J_C,H correlations in the HMBC (single head arrow) and NOESY (double head arrow) of 8b.

The structures of 8b and 8d were independently confirmed by X-ray crystal structure analyses (Fig. 3 and 4 and Experimental section).

Fig. 3 Molecular structure of 7-chloro-4-(4-methoxyphenyl)-1-methylindoline-2,3-dione (8b) in the crystal. Displacement ellipsoids are drawn at 50% probability level.

Fig. 4 Molecular structure of 7-chloro-1-methyl-4-(p-tolyl)indoline-2,3-dione (8d) in the crystal. Displacement ellipsoids are drawn at 50% probability level.

It was interesting for us to study the regioselectivity of the Suzuki–Miyaura reaction of the dichloroisatin moiety, as this substrate contains two different electron deficient centers (C-4 and C-7). Thus, the one-pot Suzuki–Miyaura reaction of 5 with two different arylboronic acids 6 (sequential addition of 1.2 equiv. of each boronic acid) afforded the N-methyl-4,7-diarylisatins 6a–d in 73–81% yields (Scheme 3). The reactions were carried out at 70 °C in the first step (to avoid double coupling), followed by 120 °C in the second step (Scheme 3).

Scheme 3 Reagents and Conditions: (i) 5 (1.0 equiv.), Ar¹B(OH)₂, 6m, k, d (1.2 equiv.), K₃PO₄ (1.5 equiv.), Pd(PPh₃)₄ (3 mol%), dioxane, 70 °C, 6 h; (ii) Ar²B(OH)₂, 6d, b (1.2 equiv.), K₃PO₄ (1.5 equiv.), Pd(PPh₃)₄ (3 mol%), 1,4-dioxane, 120 °C, 6 h.

The results shown above (Scheme 3) reveal that the chlorine residue at C-4 of the isatin ring has been initially replaced by the aryl group, while the second replacement took place in the second stage (Fig. 5). The result can be explained by the fact that position 4, located next to the carbonyl group, is more electron deficient than position 7. In addition, the selectivity might be explained by complexation of the catalyst by the carbonyl group (catalyst-directing effect) (Table 4).

Fig. 5 Possible explanation for the reaction.

Table 4 Synthesis of 9a–c

6	9	Ar^1	Ar^2	9^a (%)
a Yields of isolated products.
6i, d	9a	4-tBuC6H4	4-MeC6H4	57
6k, b	9b	3,5-(MeO)2C6H3	4-(MeO)C6H4	70
6d, b	9c	4-MeC6H4	4-(MeO)C6H4	68

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In vitro anti-HIV assay

Compounds 5, 7a–f, 8a–d, 8f–m and 9a–c, were tested for their in vitro anti-HIV-1 (strain III_B) and HIV-2 (strain ROD) activity in human T-lymphocyte (MT-4) cells, using MTT method.³² The results are summarized in Table 5, in which the data for azidothymidine (AZT)³³ and lamivudine (3TC)³⁴ were included for comparison purposes. Compound 8l was found to be the only compound from the series inhibiting HIV-2 replication in cell culture at a concentration of 100 μM. Compound 8l showed an IC₅₀ value of >3.47 μM and a CC₅₀ value of 13.43 μM, resulting in a selectivity index (SI) of 4, meanwhile no antiretroviral activity was observed (SI < 1) against HIV-1. However, the activity of the compound is much lower than those of the corresponding reference compounds, AZT, and 3TC. Derivatives 5, 8c, 8g and 8m demonstrated low CC₅₀ values of >2.21 μM, >2.24 μM, 3.11 μM and 2.24 μM, respectively, but with a selectivity index (SI) < 1 in comparison to the other analogues. This suggests that these compounds might be used as lead for the development of anticancer drugs.

Table 5 In vitro anti-HIV-1^a and HIV-2^b of new N-methylisatin derivatives

Entry	HIV-1 (IIIB) av. IC50 (μM)^c	HIV-1 (IIIB) av. IC50 (μM)^c	av. CC50 (μM)^d	SI^e (IIIB)	SI^e (ROD)	SD
a Anti-HIV-1 activity measured with strain IIIB.b Anti-HIV-2 activity measured with strain ROD.c Compound concentration required to achieve 50% protection of MT-4 cells from the HIV-1 and HIV-2 induced cytopathogenic effect.d Average CC50: compound concentration that reduces the viability of mock-infected MT-4 cells by 50%.e SI: selectivity index (CC50/IC50). Measurements were done in duplicate. Data represent mean values ± standard deviations for at least two separate experiments. The value >XXX means that no IC50 value was reached at concentrations as high as the measured CC50 value for a specific compound.
5	>2.21	>2.21	2.21	<1	<1	0.33
7a	>61.75	>61.75	61.75	<1	<1	4.20
7b	>35.28	>35.28	35.28	<1	<1	18.77
7c	>10.92	>10.92	10.92	<1	<1	2.42
7d	>12.30	>12.30	12.30	<1	<1	0.89
7e	>4.88	>4.88	4.88	<1	<1	3.66
7f	>12.75	>12.75	12.75	<1	<1	0.65
8a	>30.35	>30.35	30.35	<1	<1	14.48
8b	>125.0	>125.0	125.0	<1	<1	—
8c	>2.24	>2.24	2.24	<1	<1	0.36
8d	>15.12	>15.12	15.12	<1	<1	0.93
8f	>10.85	>10.85	10.85	<1	<1	3.91
8g	>3.11	>3.11	3.11	<1	<1	0.50
8i	>9.96	>9.96	9.96	<1	<1	2.85
8j	>12.93	>12.93	12.93	<1	<1	1.16
8k	>68.33	>68.33	68.33	<1	<1	3.69
8l	>13.43	>13.43	13.43	<1	4	1.06
8m	>2.24	>2.24	2.24	<1	<1	0.34
9a	>33.16	>33.16	33.16	<1	<1	21.22
9b	>42.13	>42.13	42.13	<1	<1	17.64
9c	>12.28	>12.28	12.28	<1	<1	0.89
AZT	0.0019	0.0018	>25	>13 144	>14 245
3TC	0.51	2.02	>20	>39	>10

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Appropriate reference compounds are always included in the experiments assessing the anti-HIV activity of new compounds. Our choice for AZT and 3TC as reference compounds is based on the fact that NRTIs are equipotent against HIV-1 and HIV-2.

Our data demonstrated that compound 8l is not active against HIV-1, but has an activity to HIV-2, in contrast to what was observed in the NNRTIs assay.³⁵ However, the above data suggested that the halo group binding the benzene ring of isatin backbone (e.g.: 8) considerably increase the anti-HIV activity, meanwhile substitution of the benzene ring of isatin by an arylketo group, as present in 8l, would enhance such inhibition for HIV, in comparison to the effectiveness of other functional groups. In conclusion, the activity of 8l against HIV-2 may be considered as an important lead for the development of a more potent and selective molecules which could be used in combination with other drugs to treat individuals infected with HIV-2.

Molecular modeling analysis

Our molecular docking analysis of the new analogs is based on the modeling studies which were performed to understand the binding mode of these analogs with the HIV-2 RT binding pocket (NIBP) (PDB code: 1MU2 (ref. 36)). The molecular docking was performed using SYBYL-X 1.1, and the results were visualized with PYMOL.³⁷ HIV-2 reverse transcriptase (RT) demonstrates an intrinsic resistance to non-nucleoside RT inhibitors (NNRTIs), one of two classes of anti-AIDS drugs that target the viral RT, however, HIV-2 RT has a similar overall fold to HIV-1 RT but has structural differences within the “NNRTI pocket” at both conserved and nonconserved residues.³⁶ Compound 8l has been selected for the docking modeling study, since it showed the lowest binding energy score (−9.2 kcal mol⁻¹) (Fig. 6). As shown in Fig. 6, the isatin backbone is located in the middle of the binding pocket, anchoring the two carbonyl groups at C-2 and C-3 in a favourable position for hydrogen bonding with the Lys102 and Thr107 of the reverse transcriptase (RT) enzyme, respectively. Further, the amino acids in the binding pocket of RT enzyme are mainly lipophilic with aromatic residues.³⁶ Therefore, not only hydrogen bonding but hydrophobic interactions also play vital role in deciding anti-HIV activity.³⁸ The aromatic ring (PhCOMe) of 8l fitted into an aromatic rich subpocket surrounded by the aromatic side chain of Tyr188. Detailed analysis of the binding mode showed that the aromatic ring of PhCOMe group pointed toward the aromatic ring of Tyr188 residue apparently developing π–π stacking interactions, where the electrostatic interaction is stabilized by these stacking type interactions. Overall, the combination of hydrophobic interaction and hydrogen bondings appears to govern the binding of 8l with HIV-2 RT.

Fig. 6 Docked conformation of 8l showing two hydrogen bonds: Thr107 with oxygen atom at C-3 of isatin ring and Lys102 with oxygen atom at C-2 of the same ring. In addition, a hydrophobic interaction was observed between the phenyl group of acetophenone moiety at C-4 of isatin backbone and Tyr188 of reverse transcriptase (RT) enzyme residues of HIV-2.

Experimental section

Melting points are uncorrected and were measured on a Büchi melting point apparatus B-545 (BÜCHI Labortechnik AG, Switzerland). NMR data were obtained at 400 and 600 MHz (¹H) as well as 100 MHz and 150.91 MHz (¹³C) spectrometers (Avance III, Bruker, Germany) with TMS as internal standard and on δ scale in ppm. Heteronuclear assignments were verified by HMQC and NOESY NMR experiments. Microanalytical data were obtained with a Vario, elemental apparatus (Shimadzu, Japan). Mass spectra were recorded on 70 eV EI and FAB MAT 8200 spectrometers (Finnigana MAT, USA). Silica gel (0.040–0.063 mm) used for column chromatography and analytical silica gel TLC plates 60 F254 were purchased from Merck. In a number of cases, the solvent heptane (mixture of isomers) could not be completely removed. The corresponding signals were marked in the copies of NMR spectra.

4,7-Dichloro-1-methylindoline-2,3-dione (5)

Isatin 4 (216 mg, 1.00 mmol) was taken up in anhydrous DMF (1 mL per 0.1 mmol isatin) and cooled on ice with stirring. Solid K₂CO₃ (166 mg, 1.2 mmol) was added in one portion, and the dark colored suspension was brought to room temperature and stirred for a further 1 h. The appropriate MeI (156 mg, 1.1 mmol) and KI (33.2 mg, 0.20 mmol) were added, and the reaction material had been consumed (TLC). The reaction mixture was poured into 0.5 M HCl (50 mL) and extracted with ethyl acetate (50 mL). The organic layer was washed with brine and dried (Mg₂SO₄) and filtered. The filtrate was evaporated, and the crude product was purified on a flash column chromatography using CH₂Cl₂ as eluent to give 5 as an orange solid (195 mg, 90%); mp 173–175 °C. ¹H NMR (300 MHz, CDCl₃): δ 3.57 (s, 3H, CH₃), 6.95 (d, J = 8.8 Hz, 1H, ArH), 7.37 (d, J = 8.8 Hz, 1H, ArH). ¹³C-NMR (75.4 MHz, CDCl₃): δ 29.8 (NCH₃), 115.7, 116.6 (C_arom.), 126.6 (C-4), 132.8 (C-6), 140.3 (C-5), 147.7 (C-7a), 157.7 (C₂=O), 179.3 (C₃=O). IR (KBr, cm⁻¹): v 3450, 3075, 3063, 2952, 2921, 2851, 1788 (w), 1728, 1584 (s), 1584, 1494 (w). GC-MS (EI, 70 eV): m/z (%) 227/229 ([M]⁺, 2 × [³⁵Cl], 100), 203 (21), 201 (21), 200 (10), 174 (78), 160 (12), 158 (16), 151 (12), 111 (17). HRMS (EI, 70 eV): calcd for C₉H₅³⁵Cl₂NO₂ ([M]⁺): 228.96973, found 228.96932.

General procedure for the preparation of biaryl-N-methylisatin analogues (7a–f)

To a suspension of 5 (70 mg, 0.304 mmol), dioxane (3 mL), Pd(PPh₃)₄ (11 mg, 6 mol%, 0.0092 mmol), and arylboronic acid (0.669 mmol) was added K₃PO₄ (98 mg, 0.46 mmol) in dioxane (3 mL). The mixture was heated at 120 °C under Argon atmosphere in a pressure tube for 8 h. The reaction mixture was diluted with water and extracted with CH₂Cl₂ (3 × 25 mL). The combined organic layers were dried (Na₂SO₄), filtered and the filtrate was concentrated in vacuo. The residue was purified by flash chromatography (silica gel, EtOAc/heptane).

4,7-Bis(3,5-dimethylphenyl)-1-methylindoline-2,3-dione (7a). From 3,5-dimethylphenylboronic acid 6a (100 mg). Yield: 78 mg (69%); as a red solid; mp 164–166 °C. ¹H NMR (300 MHz, CDCl₃): δ 2.29 (s, 12H, 4 × CH₃), 3.20 (s, 3H, CH₃), 6.70 (d, J = 8.6 Hz, 2H, ArH), 6.97 (d, J = 8.3 Hz, 2H, ArH), 7.16–7.19 (m, 4H, ArH), 7.57 (t, J = 8.3 Hz, 2H, ArH). ¹³C NMR (75.46 MHz, CDCl₃): δ 21.3 (4 × CH₃), 33.7 (NCH₃), 125.7 (C-5), 126.7, 129.7, 130.8, 131.8, 137.5, 139.2, 143.3, 145.0 (C_arom.), 147.6 (C-7a), 147.8, 152.2, 157.3, 157.5, 158.2 C–OMe, 166.4 (C₂=O), 180.9 (C₃=O). IR (KBr, cm⁻¹): ν 3075, 2953, 2919, 2852 (w), 1742, 1723 (s), 1605, 1585, 1564, 1499 (m). GC-MS (EI, 70 eV): m/z (%) 369 ([M]⁺, 100), 344 (23), 332 (11), 220 (16), 163 (10), 120 (10). HRMS (EI, 70 eV) calcd for C₂₅H₂₃NO₂ ([M]⁺): 369.17288, found: 369.17233.

4,7-Bis(4-methoxyphenyl)-1-methylindoline-2,3-dione (7b). From 4-methoxyphenylboronic acid 6b (102 mg). Yield: 93 mg (82%) as a red solid; mp 220–222 °C. ¹H NMR (300 MHz, CDCl₃): δ 3.20 (s, 6H, 2 × OCH₃), 3.79 (s, 3H, CH₃), 6.73 (dd, J = 8.3, 2.4 Hz, 2H, ArH), 6.97 (d, J = 8.0 Hz, 2H, ArH), 7.16–7.19 (m, 4H, ArH), 7.55 (d, J = 8.0 Hz, 2H, ArH). ¹³C NMR (75.46 MHz, CDCl₃): δ 33.7 (NCH₃), 38.1 (2xOMe), 119.3 (C_arom.-3′ + C_arom.-5′), 125.1 (C-5), 128.6, 128.9, 129.6, 130.6, 130.8, 132.4, 132.4 (C_arom.), 134.9 (C-7), 144.3 (C-7a), 146.5 (2 × C-OMe), 160.6 (C₂=O), 181.2 (C₃=O). IR (KBr, cm⁻¹): ν 3074, 2952, 2918, 2851 (w), 1743, 1724 (m), 1605, 1586, 1564, 1497, 1488 (s). GC-MS (EI, 70 eV): m/z (%) 373 ([M]⁺, 100), 364 (23), 365 (11), 310 (16), 311 (32), 210 (34), 174 (10), 134 (23). HRMS (EI, 70 eV) calcd for C₂₃H₁₉NO₄ ([M]⁺): 373.13141, found: 373.13100.

4,7-Bis(4-chlorophenyl)-1-methylindoline-2,3-dione (7c). From 4-chlorophenylboronic acid 6c (105 mg). Yield: 60 mg (52%) as a red solid; mp 231–232 °C. ¹H NMR (300 MHz, CDCl₃): δ 3.20 (s, 3H, CH₃), 6.81 (d, J = 8.2 Hz, 2H, ArH), 7.10 (d, J = 8.1 Hz, 2H, ArH), 7.22–7.26 (m, 4H, ArH), 7.10 (t, J = 7.8 Hz, 2H, ArH). ¹³C NMR (75.46 MHz, CDCl₃): δ 33.0 (NCH₃), 124.5 (C-5), 127.2, 128.4, 129.2, 130.9, 132.5, 134.9 (C_arom.), 140.7 (C_arom.-1′), 146.6 (C-7a), 156.4 (C₂=O), 180.9 (C₃=O). IR (KBr, cm⁻¹): ν 3074, 2952, 2919, 2851 (w), 1743, 1724 (s), 1605, 1586, 1565, 1499 (m). GC-MS (EI, 70 eV): m/z (%) 381 ([M]⁺, 2 × [³⁵Cl], 100), 352 (23), 252 (11), 250 (16). HRMS (EI, 70 eV) calcd for C₂₁H₁₃³⁵Cl₂NO₂ ([M]⁺): 381.03233, found: 381.03211.

1-Methyl-4,7-(di-p-tolyl)indoline-2,3-dione (7d). From 4-methylphenylboronic acid 6d (91 mg). Yield: 62 mg (58%) as a red solid; mp154–155 °C. ¹H NMR (300 MHz, CDCl₃): δ 2.27 (s, 6H, 2 × CH₃), 3.19 (s, 3H, CH₃), 6.73 (d, J = 8.3 Hz, 2H, ArH), 6.99 (d, J = 8.0 Hz, 2H, ArH), 7.16 (t, J = 7.8 Hz, 4H, ArH), 7.57 (t, J = 8.3 Hz, 2H, ArH). ¹³C NMR (75.46 MHz, CDCl₃): δ 26.8 (2 × CH₃), 33.7 (NCH₃), 120.0 (C_arom.), 125.7 (C-5), 129.7, 131.8, 137.5, 139.2 (C_arom.), 143.3 (C_arom.-1′), 147.6 (C-7a), 165.4 (C₂=O), 181.9 (C₃=O). IR (KBr, cm⁻¹): ν 3074, 2952, 2918, 2851 (w), 1743, 1724 (s), 1605, 1586, 1564, 1497 (m). GC-MS (EI, 70 eV): m/z (%) 341 ([M]⁺, 100), 244 (23), 232 (11), 210 (16), 164 (10). HRMS (EI, 70 eV) calcd for C₂₃H₁₉NO₂ ([M]⁺): 341.14158, found: 341.14124.

4,7-Bis(4-ethylphenyl)-1-methylindoline-2,3-dione (7e). From 4-ethylphenylboronic acid 6e (100 mg). Yield: 73 mg (65%) as a red solid; mp 123–125 °C. ¹H NMR (300 MHz, CDCl₃): δ 1.30 (m, 6H, 2 × CH₃), 2.22 (m, 4H, 2 × CH₂), 3.19 (s, 3H, CH₃), 6.73 (dd, J = 8.3, 2.4 Hz, 2H, ArH), 6.99 (d, J = 8.0 Hz, 2H, ArH), 7.16 (d, J = 7.8 Hz, 4H, ArH), 7.57 (t, J = 8.3 Hz, 2H, ArH). ¹³C NMR (75.46 MHz, CDCl₃): δ 20.8 (2 × CH₂CH₃), 24.6 (2 × CH₂CH₃), 33.7 (NCH₃), 124.8 (C-5), 125.0, 128.7, 128.8, 129.6, 130.7, 130.9, 132.4, 132.5, 134.9 (C_arom.), 144.1 (2 × C_arom.-Et), 146.6 (C-7a), 160.4 (C₂=O), 180.2 (C₃=O). IR (KBr, cm⁻¹): ν 3074, 2952, 2918, 2851 (w), 1743, 1724 (s), 1605, 1586, 1564, 1497 (m). GC-MS (EI, 70 eV): m/z (%) 369 ([M]⁺, 100), 344 (23), 332 (11), 210 (16), 164 (10), 154 (23). HRMS (EI, 70 eV) calcd for C₂₅H₂₃NO₂ ([M]⁺): 369.17288, found: 369.17255.

4,7-Bis(4-fluorophenyl)-1-methylindoline-2,3-dione (7f). From 4-fluorophenylboronic acid 6f (94 mg). Yield: 76 mg (72%) as a red solid; mp 190–192 °C. ¹H NMR (300 MHz, CDCl₃): δ 3.10 (s, 3H, CH₃), 6.75–6.79 (m, 2H, ArH), 6.94–7.05 (m, 4H, ArH), 7.31 (t, J = 8.2 Hz, 2H, ArH), 7.46–7.50 (m, 2H, ArH). ¹³C NMR (75.46 MHz, CDCl3): δ 29.6 (NCH₃), 100.6 (d, J_3′,F = 26.4 Hz, C_arom.-3′), 100.7 (d, J_3′′,F = 22.6 Hz, C_arom.-3′′), 100.8 (d, J_5′,F = 20.6 Hz, C_arom.-5′), 110.9 (d, J_5′′,F = 30.6 Hz, C_arom.-5′′), 115.2 (q, J_3′,F = 32.2 Hz, C_arom.-3′), 115.5 (q, J_3′′,F = 28.0 Hz, C_arom.-3′′), 124.4 (C-5), 125.7, 128.9 (C_arom.), 130.2 (q, J_4′,F = 280.1 Hz, C_arom.-4′), 130.9 (q, J_4′′,F = 280 Hz, C_arom.-4′′), 146.2 (C-7a), 160.3 (C₂=O), 180.1 (C₃=O). ¹⁹F NMR (282 MHz, CDCl₃): δ −114.4, −112.0. IR (KBr, cm⁻¹): ν 3273, 3065, 2957, 2922, 2851 (w), 1782 (s), 1727, 1716, 1687, 1603, 1580 (w). GC-MS (EI, 70 eV): m/z (%) 394 ([M]⁺, 100), 370 (17), 369 (30), 265 (10), 255 (23), 188 (34), 165 (12). HRMS (EI, 70 eV) calcd for C₂₁H₁₃F₂NO₂ [M]⁺: 349.09144; found: 349.09100.

General procedure for the preparation compounds (8a–d, f–m)

To a dioxane suspension (3 mL) of 5 (70 mg, 0.304 mmol), Pd(PPh₃)₄ (5 mg, 3 mol%, 0.005 mmol), and arylboronic acid (0.365 mmol) was added K₃PO₄ (49 mg, 0.23 mmol). The mixture was heated at 70 °C under argon atmosphere in a pressure tube for 6 h. The reaction mixture was diluted with water and extracted with CH₂Cl₂ (3 × 25 mL). The combined organic layers were dried (Na₂SO₄), filtered and the filtrate was concentrated in vacuo. The residue was purified by flash chromatography (silica gel, EtOAc/heptane).

7-Chloro-4-(3,5-dimethylphenyl)-1-methylindoline-2,3-dione (8a). From 3,5-dimethylphenylboronic acid 6a (53 mg). Yield: 77 mg (85%) as a red solid; mp 124–125 °C. ¹H NMR (300 MHz, CDCl₃): δ 2.29 (s, 6H, 2 × CH₃), 3.59 (s, 3H, CH₃), 6.87 (d, J = 8.4 Hz, 1H, ArH), 7.01 (d, J = 8.7 Hz, 3H, ArH), 7.41 (d, J = 8.3 Hz, 1H, ArH). ¹³C NMR (62.9 MHz, CDCl₃): δ 20.2 (2 × CH₃), 28.9 (NCH₃), 125.5 (C-5), 125.9, 130.0, 134.0, 136.8, 138.5 (C_arom.), 141.5, 142.3 (C_arom.-3′ + C_arom.-5′), 146.1 (C-7a), 157.2 (C₂=O), 180.0 (C₃=O). IR (KBr, cm⁻¹): ν 3446, 3073, 3047, 3032, 2950, 2922, 2853 (w), 1727 (s), 1637 (w), 1585, 1581, 1557 (w). GC-MS (EI, 70 eV): m/z (%) 299 ([M]⁺, [³⁵Cl], 100), 272 (12), 255 (23), 250 (30), 246 (11), 244 (16), 228 (15), 222 (18), 221 (25), 199 (17). HRMS (EI, 70 eV) calcd for C₁₇H₁₄³⁵ClNO₂ ([M]⁺): 299.07131; found: 299.07122.

7-Chloro-4-(4-methoxyphenyl)-1-methylindoline-2,3-dione (8b). From 4-methoxyphenylboronic acid 6b (54 mg). Yield: 78 mg (85%) as a red solid; mp 114–115 °C. ¹H NMR (300 MHz, CDCl₃): δ 3.47 (s, 3H, CH₃), 3.72 (s, 3H, OCH₃), 6.87 (dd, J = 4.3, 8.7 Hz, 3H, ArH), 7.32 (d, J = 8.6 Hz, 2H, ArH), 7.40 (d, J = 8.2 Hz, 1H, ArH). ¹³C NMR (62.9 MHz, CDCl₃): δ 28.9 (NCH₃), 54.5 (OCH₃), 112.7 (C_arom.-3′ + C_arom.-5′), 126.3 (C-5), 127.0, 129.4, 128.6 (C_arom.), 133.9 (C-4), 138.5 (C-6), 146.2 (C-7a), 157.4 (C₂=O), 159.5 (C–OMe), 180.5 (C₃=O). IR (KBr, cm⁻¹): ν 3068, 3044, 3019, 2999, 2956, 2845 (w), 1741, 1728, 1605, 1591, 1561 (m). GC-MS (EI, 70 eV): m/z (%) 303 ([M]⁺, [³⁷Cl], 34), 301 ([M]⁺, [³⁵Cl], 100), 275 (12), 273 (33), 272 (15), 245 (13), 244 (17), 242 (37), 238 (11), 230 (21), 210 (46), 173 (12), 167 (16). HRMS (EI, 70 eV), calcd for C₁₆H₁₂³⁷ClNO₃ ([M]⁺): 303.04783; found: 303.04707; calcd for C₁₆H₁₂³⁵ClNO₃ ([M]⁺): 301.04983; found: 301.05002.

7-Chloro-4-(4-chlorophenyl)-1-methylindoline-2,3-dione (8c). From 4-chlorophenylboronic acid 6c (56 mg). Yield: 79 mg (87%) as a red solid; mp 183–184 °C. ¹H NMR (300 MHz, CDCl₃): δ 3.14 (s, 3H, CH₃), 6.87 (d, J = 8.4 Hz, 1H, ArH), 7.28–7.34 (m, 4H, ArH), 7.47 (d, J = 8.7 Hz, 1H, ArH). ¹³C NMR (62.9 MHz, CDCl₃): δ 30.0 (NCH₃), 125.2 (C-5), 127.7, 128.9, 132.2, 138.3, (C_arom.), 146.2 (C-7a), 160.5 (C₂=O), 179.0 (C₃=O). IR (KBr, cm⁻¹): ν 3077, 2952, 2921, 2852 (w), 1730, 1580, 1558 (s), 1610 (w). GC-MS (EI, 70 eV): m/z (%) 305 ([M]⁺, 2 × [³⁵Cl], 97), 279 (13), 259 (16), 277 (25), 250 (10), 249 (12), 248 (14), 242 (38), 216 (14), 164 (32). HRMS (EI, 70 eV): calcd for 287.05245; C₁₅H₉³⁵Cl₂NO₂ ([M]⁺): 305.00103; found: 305.00144.

7-Chloro-1-methyl-4-(p-tolyl)indoline-2,3-dione (8d). From 4-methylphenylboronic acid 6d (48.5 mg). Yield: 74 mg (85%) as a red solid; mp 141–142 °C. ¹H NMR (300 MHz, CDCl₃): δ 1.53 (s, 3H, CH₃), 2.34 (s, 3H, CH₃), 6.82 (d, J = 8.4 Hz, 1H, ArH), 7.09 (d, J = 8.2 Hz, 2H, ArH), 7.18 (d, J = 8.0 Hz, 2H, ArH), 7.38 (d, J = 8.0 Hz, 1H, ArH). ¹³C NMR (62.9 MHz, CDCl₃): δ 21.3 (C_arom.-Me), 28.8 (NCH₃), 125.2 (C-5), 127.7, 127.9, 131.2, 138.4, 138.6 (C_arom.), 146.8 (C-7a), 157.5 (C₂=O), 180.0 (C₃=O). IR (KBr, cm⁻¹): ν 3469, 3451, 3089, 3068, 3043, 3026, 3003, 2952, 2919, 2854 (w), 1731 (s), 1610 (w), 1589, 1580, 1559 (w). GC-MS (EI, 70 eV): m/z (%) 287 ([M]⁺, [³⁷Cl], 35), 285 ([M]⁺, [³⁵Cl], 97), 270 (13), 259 (10), 257 (33), 256 (23), 244 (16), 229 (15), 228 (18), 227 (14), 195 (16). HRMS (EI, 70 eV) calcd for C₁₆H₁₂³⁷ClNO₂ ([M]⁺): 287.05271; found: 287.05245; calcd for C₁₆H₁₂³⁵ClNO₂ ([M]⁺): 285.05566; found: 285.05522.

7-Chloro-4-(4-fluorophenyl)-1-methylindoline-2,3-dione(8f). From 4-flourophenylboronic acid 6f (50 mg). Yield: 75 mg, (85%) as a red solid; mp 202–203 °C. ¹H NMR (300 MHz, CDCl₃): δ 3.59 (s, 3H, CH₃), 6.90 (d, J = 8.9 Hz, 1H, ArH), 7.05 (t, J = 8.3 Hz, 2H, ArH), 7.33–7.35 (m, 2H, ArH), 7.45 (d, J = 8.5 Hz, 1H, ArH). ¹³C NMR (75.46 MHz, CDCl₃): δ 28.9 (NCH₃), 114.2 (d, J_3′,5′,F = 21.6 Hz, C_arom.-3′ + C_arom.-5′), 115.3 (d, J = 8.2 Hz, C_arom.-2′ + C_arom.-6′ + C-5), 125.2 (C-5), 125.6 (C-Cl), 129.8 (C-4 + C-3a), 138.7 (C-6), 139.9 (d, J_1′,F = 3.3 Hz, C_arom.-1′), 147.1 (C-7a), 157.1 (q, J_4′,F = 245 Hz, C_arom.-4′), 164.0 (C₂=O), 180.1 (C₃=O). ¹⁹F NMR (282 MHz, CDCl₃): δ −111.5. IR (KBr, cm⁻¹): ν 3458, 3081, 2956, 2916, 2849 (w), 1736, 1727 (s), 1637 (w), 1596, 1569 (M). GC-MS (EI, 70 eV): m/z (%) 291 ([M]⁺, [³⁷Cl], 22), 289 ([M]⁺, [³⁵Cl], 62), 261 (23), 260 (10), 233 (13), 232 (20), 199 (15), 182 (13). HRMS (EI, 70 eV): calcd for C₁₅H₉³⁵ClFNO₂ ([M]⁺): 289.03004; found: 289.03004; calcd for C₁₅H₉³⁷ClFNO₂ ([M]⁺): 291.02709; found: 291.02803.

7-Chloro-4-(4-ethoxyphenyl)-1-methylindoline-2,3-dione (8g). From 4-ethoxyphenylboronic acid 6g (59 mg). Yield: 81 mg (84%) as a red solid; mp 94–95 °C. ¹H NMR (300 MHz, CDCl₃): δ 1.14 (t, J = 8.0 Hz, 3H, CH₃), 3.18 (s, 3H, CH₃), 3.32 (q, J = 6.5, 2.8 Hz 2H, CH₂), 7.28 (t, J = 8.0 Hz, 3H, ArH), 7.36–7.38 (m, 3H, ArH). ¹³C NMR (62.9 MHz, CDCl₃): δ 13.9 (CH₂CH₃), 28.8 (NCH₃), 62.9 (CH₂CH₃), 113.1 (C_arom.-3′ + C_arom.-5′), 126.5 (C-5), 127.0, 127.4, 127.5, 131.9, 132.2, 134.4, 131.2 (C_arom.), 146.8 (C-7a), 157.5 (C₂=O + C_arom.-OEt), 180.0 (C₃=O). IR (KBr, cm⁻¹): ν 3056, 3021, 2972, 2962, 2923, 2898, 2852 (w), 1726 (s), 1608, 1581, 1559, 1514, 1501 (w). GC-MS (EI, 70 eV): m/z (%) 317 ([M]⁺, [³⁷Cl], 34), 315 ([M]⁺, [³⁵Cl], 100), 287 (21), 259 (27), 258 (21), 242 (23), 230 (20), 224 (20), 196 (67). HRMS (EI, 70 eV) calcd for C₁₇H₁₄³⁷ClNO₃ ([M]⁺): 317.06327; found: 317.06310; calcd for C₁₇H₁₄³⁵ClNO₃ ([M]⁺): 315.06622; found: 315.06644.

7-Chloro-4-(4-isopropoxyphenyl)-1-methylindoline-2,3-dione(8h). From 4-isopropylphenylboronic acid 6h (58 mg). Yield: 85 mg (84%) as a red solid; mp 83–85 °C. ¹H NMR (300 MHz, CDCl₃): δ 1.19 (d, J = 6.4 Hz, 6H, 2 × CH₃), 3.32 (s, 3H, CH₃), 3.61–3.65 (m, 1H, CH), 6.80 (d, J = 8.2 Hz, 1H, ArH), 6.91 (d, J = 8.5 Hz, 2H, ArH), 7.29–7.35 (m, 3H, ArH). ¹³C NMR (62.9 MHz, CDCl₃): δ 21.0 (2 × CH₃), 28.9 (NCH₃), 68.9 (OCH), 114.3, 115.3 (C_arom.-3′ + C_arom.-5′), 125.9 (C-5), 126.3, 138.5, 139.5, 141.0 (C_arom.), 146.6 (C-7a), 156.5 (C_arom.-OⁱPr), 159.5 (C₂=O), 178.2 (C₃=O). IR (KBr, cm⁻¹): ν 3444, 3062, 3022, 2978, 2951, 2945 (w), 1728, 1579 (s), 1563, 1514, 1479 (m). GC-MS (EI, 70 eV): m/z (%) = 331 ([M]⁺, [³⁷Cl], 24), 329 ([M]⁺, [³⁵Cl], 100), 289 (23), 288 (14), 287 (77), 245 (13), 261 (23), 260 (18), 259 (74), 258 (22), 242 (24), 197 (14). HRMS (EI, 70 eV), calcd for C₁₈H₁₆³⁷ClNO₃ ([M]⁺): 331.07895; found: 331.07837; calcd for C₁₈H₁₆³⁵ClNO₃ ([M]⁺): 329.08116; found: 329.08132.

4-(4-tert-Butylphenyl)-7-chloro-1-methylindoline-2,3-dione (8i). From tert-butylphenylboronic acid 6m (63 mg). Yield: 84 mg (84%) as a red solid; mp 104–105 °C. ¹H NMR (300 MHz, CDCl₃): δ 1.53 (s, 9H, 3 × CH₃), 2.34 (s, 3H, CH₃), 6.81 (d, J = 8.6 Hz, 2H, ArH), 7.02 (d, J = 8.0 Hz, 2H, ArH), 7.18 (d, J = 8.0 Hz, 2H, ArH). ¹³C NMR (62.9 MHz, CDCl₃): δ 19.3 (C(Me)₃), 28.6 (NMe), 35.8 (C(Me)₃), 125.2 (C-5), 127.9, 131.2, 138.4 (C_arom.), 141.1 (C_arom.-1′), 146.1 (C_arom.-4′), 146.8 (C-7a), 157.5 (C₂=O), 180.0 (C₃=O). IR (KBr, cm⁻¹): ν 3469, 3451, 3089, 3068, 3043, 3026, 3003, 2952, 2919, 2854 (w), 1731 (s), 1610 (w), 1589, 1580, 1559 (w). GC-MS (EI, 70 eV): m/z (%) 327 ([M]⁺, [³⁵Cl], 97), 270 (13), 259 (10), 257 (33), 256 (23), 244 (16), 229 (15), 228 (18), 227 (14), 195 (16). HRMS (EI, 70 eV) calcd for C₁₉H₁₈³⁵ClNO₂ ([M]⁺): 327.10261; found: 327.1024.

7-Chloro-1-methyl-4-(m-tolyl)indoline-2,3-dione (8j). From 4-isopropylphenylboronic acid 6j (58 mg). Yield: 74 mg (85%) as a red solid; mp 124–125 °C. ¹H NMR (300 MHz, CDCl₃): δ 2.30 (s, 3H, CH₃), 3.38 (s, 3H, CH₃), 6.87 (d, J = 8.4 Hz, 1H, ArH), 7.12–7.23 (m, 4H, ArH), 7.41 (d, J = 8.3 Hz, 1H, ArH). ¹³C NMR (62.9 MHz, CDCl₃): δ 20.3 (Me), 28.9 (NCH₃), 124.8 (C-5), 125.9, 127.1, 128.4, 129.0, 134.0, 136.9 (C_arom.), 140.1 (C_arom.-1′), 141.2 (C_arom.-3′), 146.1 (C-7a), 157.2 (C₂=O), 180.0 (C₃=O). IR (KBr, cm⁻¹): ν 3447, 3074, 3048, 3033, 2952, 2921, 2854 (w), 1728 (s), 1637 (w), 1588, 1581, 1558 (w). GC-MS (EI, 70 eV): m/z (%) 287 ([M]⁺, [³⁷Cl], 35), 285 ([M]⁺, [³⁵Cl], 97), 270 (13), 259 (10), 257 (33), 256 (23), 244 (16), 229 (15), 228 (18), 227 (14), 195 (16). HRMS (EI, 70 eV) calcd for C₁₆H₁₂³⁷ClNO₂ ([M]⁺): 287.05271; found: 287.05245; calcd for C₁₆H₁₂³⁵ClNO₂ ([M]⁺): 285.05566; found: 285.05522.

7-Chloro-4-(3,5-dimethoxyphenyl)-1-methylindoline-2,3-dione(8k). From 3,5-dimethoxyphenylboronic acid 6k (65 mg). Yield: 86 mg (85%) as a red solid; mp 124–125 °C. ¹H NMR (300 MHz, CDCl₃): δ 3.58 (3H, CH₃), 3.72 (6H, 2 × OCH₃), 6.43–6.49 (m, 3H, ArH), 6.92 (d, J = 8.7 Hz, 1H, ArH), 7.42 (d, J = 8.4 Hz, 1H, ArH). ¹³C NMR (62.9 MHz, CDCl₃): δ 30.0 (NCH₃), 55.5 (2 × OCH₃), 101.6 (C_arom.-4′), 106.9 (C_arom.-3′ + C_arom.-5′), 126.8 (C-5), 128.0, 129.5, 134.4, 134.6, 135.0, 137.0 (C_arom.), 141.3 (C_arom.-1′), 147.3 (C-7a), 158.5 (C₂=O), 160.5 (2 × C-OMe), 179.2 (C₃=O). IR (KBr, cm⁻¹): ν 3455, 3094, 3077, 3053, 3011, 2947, 2841 (w), 1731 (s), 1695, 1684, 1652, 1646, 1635, 1601 (w). GC-MS (EI, 70 eV): m/z (%) 331 ([M]⁺, [³⁵Cl], 100), 316 (13), 303 (11), 302 (21), 288 (11), 252 (16), 245 (10). HRMS (EI, 70 eV) calcd for C₁₇H₁₄³⁵ClNO₄ ([M]⁺): 331.06114; found: 331.06122.

4-(4-Acetylphenyl)-7-chloro-1-methylindoline-2,3-dione (8l). From 4-acetylphenylboronic acid 6l (58 mg). Yield: 81 mg (85%) as a red solid; mp 124–125 °C. ¹H NMR (300 MHz, CDCl₃): δ 2.52 (s, 3H, COCH₃), 3.56 (s, 3H, CH₃), 6.95 (d, J = 8.4 Hz, 1H, ArH), 7.49 (t, J = 8.7 Hz, 3H, ArH), 7.93 (d, J = 8.8 Hz, 2H, ArH). ¹³C NMR (62.9 MHz, CDCl₃): δ 26.7 (COCH₃), 30.7 (NCH₃), 127.4 (C-5), 128.1, 128.3, 129.1130.8, 133.2 (C_arom.), 137.4 (C_arom.-4′), 139.9 (C_arom.-1′), 146.9 (C-7a), 158.5 (C₂=O), 181.1 (C₃=O), 197.5 (COMe). IR (KBr, cm⁻¹): ν 3072, 3058, 3006, 2958, 2921, 2850 (w), 1731, 1681, 1583, 1556 (s). GC-MS (EI, 70 eV): m/z (%) 315 ([M]⁺, [³⁷Cl], 30), 313 ([M]⁺, [³⁵Cl], 88), 298 (28), 285 (11), 272 (33), 271 (16), 244 (14), 242 (40), 214 (13), 207 (11), 180 (36), 164 (22), 150 (25). HRMS (EI, 70 eV), calcd for C₁₇H₁₂³⁷ClNO₃ ([M]⁺): 315.04707; found: 315.04762; calcd for C₁₇H₁₂³⁵ClNO₃ ([M]⁺): 313.05002; found: 313.04983.

7-Chloro-1-methyl-4-(4-vinylphenyl)indoline-2,3-dione (8m). From 4-vinylphenylboronic acid 6i (53 mg). Yield: 77 mg (86%) as a red solid; mp 124–125 °C. ¹H NMR (300 MHz, CDCl₃): δ 3.56 (3H, CH₃), 5.09 (d, J = 1.8 Hz, 1H, CH), 5.76 (d, J = 1.9 Hz, 1H, CH), 6.67 (q, J = 7.1, 2.2 Hz, 1H, CH), 6.91 (d, J = 8.9 Hz, 2H, ArH), 7.33–7.42 (m, H, ArH). ¹³C NMR (62.9 MHz, CDCl₃): δ 29.9 (NCH₃), 115.1 (CH₂ = CH), 126.1 (C-5), 126.3, 126.9, 129.2, 134.5 (C_arom.), 136.2 (CH₂=CH), 139.8 (C_arom.-4′), 140.4 (C_arom.-1′), 141.7 (C-7a), 158.3 (C₂=O), 180.1 (C₃=O). IR (KBr, cm⁻¹): ν 3449, 3067, 2944, 2922, 2851 (w), 1734 (s), 1627 (w), 1581 (s), 1554, 1478, 1458, 1438 (w). GC-MS (EI, 70 eV): m/z (%) 299 ([M]⁺, [³⁷Cl], 34), 297 ([M]⁺, [³⁵Cl], 100), 271 (14), 270 (17), 269 (42), 268 (30), 252 (16), 242 (21), 240 (40), 239 (11), 234 (18). HRMS (EI, 70 eV), calcd for C₁₇H₁₂³⁷ClNO₂ ([M]⁺): 299.05216; found: 299.05303; calcd for C₁₇H₁₂³⁵ClNO₂ ([M]⁺): 297.05511; found: 297.0545.

General procedure for the preparation of compounds (9a–c)

To a dioxane suspension (3 mL) of 5 (70 mg, 0.304 mmol), Pd(PPh₃)₄ (5 mg, 3 mol%, 0.005 mmol), and arylboronic acid Ar¹B(OH)₂ 6m, k, d (0.365 mmol) was added K₃PO₄ (49 mg, 0.230 mmol). The mixture was heated at 70 °C under argon atmosphere for 6 h. The mixture was cooled to 20 °C. Arylboronic acid Ar²B(OH)₂ 6d, b (0.365 mmol) was added K₃PO₄ (49 mg, 0.230 mmol). The mixture was heated at 120 °C under argon atmosphere in a pressure tube for 6 h. The reaction mixture was diluted with water and extracted with CH₂Cl₂ (3 × 25 mL). The combined organic layers were dried (Na₂SO₄), filtered and the filtrate was concentrated in vacuo. The residue was purified by flash chromatography (silica gel, EtOAc/heptane).

4-(4-tert-Butylphenyl)-1-methyl-7-p-tolylindoline-2,3-dione (9a). From tert-butylphenylboronic acid 6m (65 mg), 4-methylphenylboronic acid 6d (50 mg). Yield: 85 mg (73%) as a red solid; mp 172–174 °C. ¹H NMR (300 MHz, CDCl₃): δ 1.34 (s, 9H, 3 × CH₃), 2.31 (s, 3H, CH₃), 3.19 (s, 3H, CH₃), 6.71 (d, J = 8.4 Hz, 2H, ArH), 6.66 (d, J = 8.1 Hz, 2H, ArH), 7.11 (t, J = 7.3 Hz, 4H, ArH), 7.57 (d, J = 8.3 Hz, 2H, ArH). ¹³C NMR (75.46 MHz, CDCl₃): δ 19.1 (3xCH₃), 20.2 (C_arom.-CH₃), 33.7 (NCH₃ + C(Me)₃), 124.1 (C-5), 124.5, 124.7, 127.7, 127.9, 128.5, 132.0, 133.4, 137.2, 139.6, 141.2 (C_arom.), 148.3 (C-7a), 158.3 (C₂=O), 181.9 (C₃=O). IR (KBr, cm⁻¹): ν 3074, 2952, 2918, 2851 (w), 1743, 1724 (s), 1605, 1586, 1564, 1497 (m). GC-MS (EI, 70 eV): m/z (%) 383 ([M]⁺, 100), 369 (27), 368 (94), 340 (18), 327 (22), 326 (30), 313 (12), 312 (43), 310 (10), 299 (10), 296 (11), 284 (10), 283 (12), 282 (10), 141 (22). HRMS (EI, 70 eV) calcd for C₂₆H₂₅NO₂ ([M]⁺): 383.18853, found: 383.18788.

4-(3,5-Dimethoxyphenyl)-7-(4-methoxyphenyl)-1-methylindoline-2,3-dione (9b). From 3,5-dimethoxyphenylboronic acid 6k (65 mg), 4-methoxyphenylboronic acid 6b (54 mg). Yield: 89 mg (73%) as a red solid; mp 172–174 °C. ¹H NMR (300 MHz, CDCl₃): δ 2.40 (s, 3H, CH₃), 3.59 (s, 9H, 3 × OCH₃), 6.70 (d, J = 8.4 Hz, 2H, ArH), 7.12 (t, J = 7.3 Hz, 5H, ArH), 7.55 (d, J = 8.3 Hz, 2H, ArH). ¹³C NMR (75.46 MHz, CDCl₃): δ 29.2 (NCH₃), 33.7 (3 × OCH₃), 124.1 (C-5), 124.5, 124.7, 127.7, 127.9, 128.5, 132.0, 133.4, 137.2, 139.6, 140.1, 141.2 (C_arom.), 147.8 (C-7a), 148.3, 151.1, 152.3 (3 × COMe), 168.3 (C₂=O), 180.9 (C₃=O). IR (KBr, cm⁻¹): ν 3070, 2951, 2917, 2851 (w), 1747, 1723 (s), 1604, 1585, 1564, 1497 (m). GC-MS (EI, 70 eV): m/z (%) 403 ([M]⁺, 100), 379 (27), 378 (94), 350 (18), 337 (22), 323 (12), 322 (43), 320 (10), 289 (10), 286 (11), 284 (13), 283 (12), 282 (10), 141 (22). HRMS (EI, 70 eV) calcd for C₂₄H₂₁NO₅ ([M]⁺): 403.14197, found: 403.14145.

7-(4-Methoxyphenyl)-1-methyl-4-p-tolylindoline-2,3-dione (9c). From 4-methylphenylboronic acid 6d (50 mg), 4-methoxyphenylboronic acid 6b (54 mg). Yield: 79 mg (73%); mp 172–174 °C. ¹H NMR (300 MHz, CDCl₃): δ = 2.35 (s, 3H, CH₃), 3.54 (s, 3H, OCH₃), 6.71 (d, J = 8.1 Hz, 2H, ArH), 6.64 (d, J = 8.6 Hz, 2H, ArH), 7.11–7.15 (m, 4H, ArH), 7.50 (d, J = 8.0 Hz, 2H, ArH). ¹³C NMR (75.46 MHz, CDCl₃): δ 20.2 (C_arom.-CH₃), 33.7 (NMe), 54.2 (OMe), 124.1 (C-5), 124.5, 124.7, 127.7, 127.9, 128.5, 132.0, 133.4, 137.2, 139.6, 141.2 (C_arom.), 147.8 (C-7a), (C), 158.3 (C₂=O), 181.9 (C₃=O). IR (KBr, cm⁻¹): ν 3174, 2952, 2818, 2751 (w), 1843, 1824 (s), 1705, 1686, 1564, 1497 (m). GC-MS (EI, 70 eV): m/z (%) 357 ([M]⁺, 100), 349 (27), 348 (94), 340 (18), 326 (30), 310 (17), 283 (13), 280 (10), 145 (22). HRMS (EI, 70 eV) calcd for C₂₃H₁₉NO₃ ([M]⁺): 357.13649, found: 357.13623.

Crystal structure data (8b). C₁₆H₁₂ClNO₃, M_r = 301.72, red plates, crystal dimensions = 0.30 × 0.19 × 0.17 mm³, orthorhombic, space group Pbca, a = 7.3328 (3), b = 9.2003 (4), c = 19.7779 (8) Å, Z = 4, D_calcd = 1.38 mg cm⁻³, μ(MoK_α) = 0.1 mm⁻¹, F(000) = 1.760e, T = 173(2) K. MoK_α radiation, λ = 0.71073 Å, 56 177 collected refls. (hkl – 21/20, ±16, ±27), 5261 unique refls. (R_int = 0.062), 290 refined parameters, R₁/wR₂ = 0.0771/0.1363 (all data), GOF = 1.023, Δρ_fin (max/min) = 0.33/−0.23e Å⁻³.

Crystal structure data (8d). C₁₆H₁₂ClNO₂, M_r = 285.72, red plates, crystal dimensions = 0.99 × 0.03 × 0.03 mm³, orthorhombic, space group Pbca, a = 3.8951 (6), b = 24.185 (3), c = 13.5874 (17) Å, Z = 4, D_calcd = 1.38 g cm⁻³, μ(MoKα) = 0.1 mm⁻¹, F(000) = 1.760e, T = 173(2) K. MoK_α radiation, λ = 0.71073 Å, 56 177 collected refls. (hkl – 21/20, ±16, ±27), 5261 unique refls. (R_int = 0.062), 290 refined parameters, R₁/wR₂ = 0.0771/0.1363 (all data), GOF = 1.023, Δρ_fin (max/min) = 0.36/−0.39e Å⁻³

CCDC 1410098 contains the supplementary crystallographic data for this paper.†

Crystal structure data (8d). C₁₆H₁₂ClNO₂, M_r = 285.72, red plates, crystal dimensions = 0.99 × 0.03 × 0.03 mm³, orthorhombic, space group Pbca, a = 3.8951 (6), b = 24.185 (3), c = 13.5874 (17) Å, Z = 4, D_calcd = 1.38 g cm⁻³, μ(MoKα) = 0.1 mm⁻¹, F(000) = 1.760e, T = 173(2) K. MoK_α radiation, λ = 0.71073 Å, 56 177 collected refls. (hkl – 21/20, ±16, ±27), 5261 unique refls. (R_int = 0.062), 290 refined parameters, R₁/wR₂ = 0.0771/0.1363 (all data), GOF = 1.023, Δρ_fin (max/min) = 0.36/−0.39e Å⁻³.

CCDC 1410099 contains the supplementary crystallographic data for this paper.†^,38

In vitro HIV assay

Evaluation of the antiviral activity of compounds 5, 7a–f, 8a–d, 8f–m and 9a–c against the HIV-1 strain (III_B) and the HIV-2 strain (ROD) in MT-4 cells was performed using an MTT assay as described previously.³² In brief, stock solutions (10 times final concentration) of test compounds were added in 25 μL volumes to two series of triplicate wells to allow simultaneous evaluation of their effects on mock and HIV infected cells at the beginning of each experiment. Stock solutions of the compounds were made in DMSO (10 mg mL⁻¹) as described in ref. 32. Serial 5-fold dilutions of test compounds were made directly in flat-bottomed 96-well microtiter trays using a Biomek 3000 robot (Beckman instruments). Untreated control, HIV and mock-infected cell samples, were included for each sample. HIV-1 (III_B)³⁹ or HIV-2 (ROD)⁴⁰ stock (50 μL) at 100–300 CCID50 (50% cell culture infectious dose) or culture medium was added to either of the infected or mock-infected wells of the microtiter tray. Mock-infected cells were used to evaluate the effect of test compound on uninfected cells in order to assess the cytotoxicity of the test compounds. Exponentially growing MT-4 cells³⁵ were centrifuged for 5 min at 1000 rpm, and the supernatant was discarded. The MT-4 cells were resuspended at 6 × 105 cells per ml, and 50 μL volumes were transferred to the microtiter tray wells. Five days after infection, the viability of the mock- and HIV-infected cells was examined spectrophotometrically.

Viruses

The origins of virus stocks were as described previously: The HIV-1 (III_B) strain was originally provided by Prof. R. C. Gallo and Dr M. Popovic (at that time at the NIH, Bethesda, MD, USA);³⁹ HIV-2 (ROD)⁴⁰ was provided by Dr L. Montagnier and was obtained from the culture supernatant of infected MT-4 cells.

Acknowledgements

Financial support by the DAAD (scholarships for A. M. H.) and by the State of Iraq is gratefully acknowledged.

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Footnote

† Electronic supplementary information (ESI) available. CCDC 1410098 and 1410099. For ESI and crystallographic data in CIF or other electronic format see DOI: 10.1039/c5ra16841d

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