Practical Strategy to 1,1′-Dideoxygossypol Derivatives from Gossypol and Its Antitumor Activities

Abstract

A practical and scalable route for the synthesis of 1,1′-dideoxygossypol from natural polyphenol product gossypol is described. The key step is the successful regioselective deacetylation of hexaacetyl apogossypol 9 and the following reductive removal of hydroxyl groups. The two steps of deacetylation occurred on the different sites under different conditions. The synthetic route follows a simple protection–deprotection strategy, and the yields of each step are over 85%. The total yield of this 9-step synthesis is over 40%, which is much better than the reported total synthesis method. The antitumor results illustrate that 1,1′-hydroxyl groups are not necessary for antitumor activities. In addition, 1,1′-dideoxygossypol has superior aqueous solubility (215 mg/L) compared to gossypol (64 mg/L).

1. Introduction

Gossypol 1 (Figure 1) has attracted people to investigate its chemical modification and biological properties with polyphenol structure.¹ Derivative types from gossypol were limited due to the unique chemical properties of polyphenols aldehyde.^1,2 Modification of functional groups on gossypol currently provided most products such as gossypol Schiff bases, gossylic lactones, apogossypol 2, and so forth.² For example, the aldehyde derivatives of gossypol were limited because the phenolic hydroxyl groups in the 1 and 1′ (peri) positions complicate the chemistry of functional groups at the 8 and 8′ positions. Functionalization of three hydroxyls on two naphthyl scaffolds was also less studied without suitable methods. The seemingly simple full functionalization derivatives of hydroxyls have been proven challenging because the product mixtures derived from alkylation or esterification were quite complex owing to gossypol’s various tautomeric forms. Although tetramethylation and hexamethylation gossypol products were only reported in 1939, no further research followed this route and different methylation mixtures were obtained by our several attempts.³ The roles of 1 and 1′ hydroxyl groups were proved unnecessary in some biological activity tests, such as the synthesized 1,1′-dideoxygossypol 3 which exhibited good anti-HIV activity.⁴ However, antitumor activities of 3 were not studied ever. 1,1′-Dideoxylgossypol could be considered as the substituted scaffold for gossypol without the 1,1′-hydroxyl interference. Identification of essential fragment parts from gossypol would be significant for the investigation of the structure–activity relationship (SAR) of gossypol derivatives.

Figure 1.

Structure of gossypol 1, apogossypol 2, and 1,1′-dideoxygossypol 3.

The direct removal of the hydroxyl groups was not reported due to the multiple hydroxyls of the parent molecule gossypol. The linear total synthesis of 1,1′-dideoxygossypol 3 was developed by Royer et al.(4) (Figure 2) The reported synthesis featured the incorporation of the carbon atoms for the second ring of the naphthalene system through a 10-step transformation including Grignard, reduction, cyclization, bromination, coupling reactions, and so forth starting from 1-bromo-2-isopropyl-3,4-dimethoxybenzene 4. The total yield was low because of the low yield of the coupling step from 5 to 6 (53% yield), and 1-bromo-2-isopropyl-3,4- dimethoxybenzene 4 is not a cheap starting material. What is more, the atom economy was bad because of the usage of Grignard reagent, bromination, and coupling reaction steps. A simple and reliable functionalization method from gossypol will benefit research on this field.

Figure 2.

Total synthesis of 1,1′-dideoxygossypol 3 from 1-bromo-2-isopropyl-3,4-dimethoxybenzene 4 by Royer et al.⁴

Herein, we reported a practical and scalable route that leads to the novel synthesis of 1,1′-dideoxygossypol through a regioselective deprotection of a phenolic hydroxyl group strategy.

2. Results and Discussion

2.1. Retrosynthetic Analysis

There are three hydroxyl groups in each half gossypol. Full modification of these hydroxyl groups had been only reported by reacting gossypol with methyl or acetyl protecting groups but without thorough characterization.^3,5 Based on our recent attempts, full substituted products were mixed with partial substituted products and various tautomeric forms. However, full functionalized hydroxyl on apogossypol 2 was found practical without the tautomer interference. The different reactive activities of hydroxyls were confirmed.⁶ When the disturbing aldehydes were converted into nitriles, gossylic nitrile-1,1′-diacetate 7 was obtained with 6,6′,7,7′-four acetyl groups selectively removed from hexaacetyl gossylic nitrile 8 (Figure 3). Then selective functionalization of the adjacent 6,6′,7,7′-hydroxyl groups of apogossypol lacking aldehyde was achieved.⁷⁻⁹ We proposed apogossypol without aldehyde, which would be possible to functionalize a specific hydroxyl site within several steps.¹⁰ Thus, this strategy could be applied in the synthesis of 1,1′-dideoxygossypol.

Figure 3.

6,6′,7,7′-four acetyl groups selectively removed from hexaacetyl gossylic nitrile 7.

Based on the abovementioned analysis, the preparation of 1,1′-dideoxygossypol 3 could be realized by formylation of methyl-1,1′-dideoxyapogossypol 6 (Scheme 1). We proposed that methyl-1,1′-dideoxyapogossypol 6 could be synthesized from intermediate 10 after reduction and protection steps. The key intermediate 10 would be obtained from selective deacetylation of apogossypol 2 based on the abovementioned method.

Scheme 1. Retrosynthesis of 1,1′-Dideoxygossypol 3.

2.2. Synthesis of 1,1′-Dideoxygossypol through a Regioselective Deprotection of Phenolic Hydroxyl Groups

From gossypol acetate 1 (Scheme 2), apogossypol 2 was prepared with a known procedure by treating gossypol acetate with NaOH aqueous at 90 °C and was then acetylated in the presence of acetic anhydride to afford compound 9 with 92% yield (two steps). Next, the selective acetyl deprotection condition was explored.^6,7 Directly refluxing compound 9 with potassium carbonate in methanol failed to give the deacetylation product 10 because of the insoluble of compound 9 in methanol; this problem could be solved by the methanol/dichloromethane mixture solvent. The yield of compound 10 was 71% because the oxidized catechol formed under air conditions.⁷ Thus, the first deacetylation yield could be improved up to 85% under a N₂ atmosphere. The compound 10 was readily prepared on a 10 g scale.

Scheme 2. Synthesis of 1,1′-Dideoxygossypol 3.

Reagents and conditions: (a) 40% NaOH aqueous, N₂, 90 °C; (b) H₂SO₄, 0 °C; (c) Ac₂O, DMAP, DCM, r.t.; (d) K₂CO₃, MeOH/DCM/H₂O (v/v/v = 4:2:1), N₂, 65 °C; (e) MeI, K₂CO₃, DMF, r.t.; (f) LiAlH₄, THF, 0 °C to r.t.; (g) Tf₂O, pyridine, THF, r.t.; (h) Pd(OAc)₂, dppp, Et₃SiH, DMF, N₂, 90 °C; (i) Cl₂CHOCH₃, TiCl₄, DCM, 0 °C; (j) BBr₃, DCM, −20 °C to r.t.

For intermediate 10, it needed the removal of 1,1′-hydroxyl to the target 1,1′-dideoxygossypol. It was necessary to protect the remaining hydroxyls of phenol before removing 1,1′-hydroxyl groups. Protection of the catechol moieties of compound 10 by methyl iodide gave the methylated intermediate 11 in 92% yield. There were no methods to get deoxygenated products from phenyl acetate scaffold directly. However, there were hydrogenolysis methods for phenol and phenol derivatives. The conditions for hydrogenolysis of phenols was too harsh for gossypol, and phenol triflate could be reduced into deoxygenated products with reducing reagents.¹¹ Then triflates of 1,1′-hydroxyl 13 were proposed to be obtained.

However, the second deacetylation hydrolysis of 11 was found to be unusual. The inorganic bases such as K₂CO₃ in first deacetylation failed to get the free hydroxyl 12 (Table 1, entry 1), while stronger inorganic base LiOH under heat conditions provided 12 with low yield along with the same portion side product 12a (Table 1, entry 2). The formed product 12 was oxidized to the by-product mono-quinone 12a under basic conditions, which was unstable and decomposed. That was the reason why the yields were still unsatisfactory with NaOH and KOH (Table 1, entry 3 and 4). We therefore turned our attention to reduction reagents. To our delight, intermediate 12 was obtained in good yield using LiAH₄ (Table 1, entry 5), and no oxidized by-product 12a was detected without question (Table 1, entry 5). Next, the 1,1′-hydroxyl groups were protected by Tf₂O to give intermediate 13 in 93% yield with pyridine as base after screening a series of organic bases (TEA, DIPEA, TBD, DABCO, DBU, and pyridine) (See Supporting Information Table S1). The following hydrogenation of OTf groups was accomplished by treating intermediate 13 with Et₃SiH in the presence of Pd(OAc)₂, and then Met-1,1′-dideoxyapogossypol 6 was produced with 93% yield.^12,13 The NMR spectroscopic data of our synthetic Met-1,1′-dideoxyapogossypol 6 were identical to the reported data.⁴

Table 1. Reaction Condition Optimization of the Deacetylation of Compound 11^a.

entry	reagents	solvent	T (°C)	12 (% yield)b (%)	12a (% yield)b (%)
1	K2CO3c	MeOH/DCM	r.t. to 40 °Cd	0	0
2	10% LiOH	MeOH/DCM	r.t. to 40 °Cd	30	25
3	10% NaOH	MeOH/DCM	r.t. to 40 °Cd	51	19
4	10% KOH	MeOH/DCM	r.t. to 40 °Cd	47	23
5	LiAlH4	THF	0 °C to r.t.	81	0

^aReaction conditions: 11 (0.1 mmol), reagents (1.0 mmol, 10.0 equiv), solvent (3.0 mL, MeOH/DCM, v/v = 2:1), 16 h.

^bIsolate yield.

^cSaturated aqueous.

^dStirred at room temperature for 10 h (no reaction), then rise to 40 °C and stirred for another 6 h.

Methyl-1,1′-dideoxygossypol 14 could be made from formylation of methyl-1,1′-dideoxyapogossypol 6 with known chemistry.⁴ The yield of compound 14 could be up to 95%. Finally, removing the methyl groups on compound 14 with BBr₃ provided the target 1,1′-dideoxygossypol 3 in 86% yield. The characterization data from our synthesized 1,1′-dideoxygossypol were identical to the reported data.⁴ Thus, 1,1′-dideoxygossypol was obtained via a 9-step protection–deprotection strategy from gossypol; the total yield was over 40%, which was 2.9-fold better than the reported method.

To explore the antitumor activity contribution of each functional group from gossypol, the corresponding group could be removed separately. For example, apogossypol without aldehyde has shown similar activities as gossypol.¹⁴ Thus the aldehydes were proved less useful in contributing to the antitumor activities. The compound 3 was tested without one pair hydroxyl group compared with gossypol. From the results in Table 2, seven different type of cells were tested for both gossypol 1 and compound 3. The IC₅₀ values varied from 0.94 to 5.99 μM for compound 3, which exhibited similar antitumor activities as gossypol, Therefore, 1,1′-hydroxyl groups of gossypol might not significantly contribute to most of the biological activities. In addition, the aqueous solubility of compound 3 without 1,1′-hydroxyl groups was found to be 3–4 fold higher than that of gossypol (215 mg/L vs 64 mg/L), which was very important physicochemical property for ADME.

Table 2. Evaluation of Gossypol Derivatives Using Cell Viability Assays.

comp.	H460 IC50α (μM)	HCT116 IC50α (μM)	PC-3 IC50α (μM)	L02 IC50α (μM)	Jekio-1 IC50α (μM)	Daji IC50α (μM)	Daudi IC50α (μM)
1	3.49	5.4	4.57	1.31	0.55	1.27	7.84
3	2.58	1.88	5.98	2.26	0.94	6.24	5.99

3. Conclusions

In general, we have developed a concise and practical route for the synthesis of 1,1′-dideoxygossypol from natural product gossypol. The nine-step synthesis features expedient construction of 1,1′-dideoxygossypol via judiciously combining various simple protection–deprotection methodologies, and the total yield was over 40%, which was better than reported total synthesis strategy. Using the reported simple method, we have obtained gram scale 1,1′-dideoxygossypol. People have already known that 8,8′-aldehyde is unnecessary. For the first time, 1,1′-hydroxyl was proved useless for the antitumor activities. The solubility was improved with less hydroxyl. So researchers can use compound 3 as the starting molecule to investigate the properties of gossypol scaffold. Undoubtedly, this strategy would benefit design of other convenient routes in the future for the synthesis of other gossypol derivatives and explore the SAR of gossypol. The existence of the exposed catechol moieties of 1,1′-dideoxygossypol provides opportunities to investigate the effect of the individual hydroxyl groups on the biological activity of gossypol, which has been seldom explored. Several hydroxyl-removed gossypol products on different positions have been investigated with this strategy, and then the chemistry part and SAR will be reported in due course.

4. Experimental Section

4.1. General Chemical Methods

Unless otherwise noted, all purchased reagents were used as received without further purification. All anhydrous solvents were dried and purified by standard techniques before use. Reactions were monitored by thin-layer chromatography (TLC) on silica gel GF254, and visualization was achieved by UV light (254 and 365 nm). Column chromatography purifications were performed using a 230–400 mesh silica gel. NMR spectra were measured on a Bruker AVANCE 600 MHz for ¹H NMR spectra and 150 MHz for ¹³C NMR spectra and calibrated from the residual solvent signal. In reported spectral data, the format (δ) chemical shift (multiplicity, J values in Hz, integration) was used with the following abbreviations: s = singlet, d = doublet, t = triplet, q = quartet, m = multiplet. The purity and low-resolution mass spectrometry (LRMS) of all the synthesized compounds were determined by a liquid chromatography tandem mass spectrometer (SHIMADZU LC-2030C LT/LCMS-2020, InertSustain C18 column, 4.6 × 150 mm, 5 μm, 40 °C, flow rate = 0.8 mL/min) with aqueous CH₃CN and water. Purity was confirmed by analytical reversed phase HPLC using an Inersil ODS-P C18 column (5 μm, 4.6 × 250 mm) eluted with a water-acetonitrile gradient moving from 0 to 100% CH₃CN in 30 min.

4.2. Chemistry

4.2.1. Synthesis of 5,5′-Diisopropyl-3,3′-dimethyl-[2,2′-binaphthalene]-1,1′,6,6′,7,7′-Hexaol Apogossypol 2

A solution of gossypol acetate 1 (1 g, 1.93 mmol) in 10 mL of 40% NaOH was heated under N₂ at 90 °C for 2 h in the dark. The reaction mixture was cooled and poured slowly onto ice (50 mL) and concentrated H₂SO₄ (10 mL) mixture to form a white precipitation, and the mixture was extracted with EA (3*200 mL). The combined organic phase was dried over Na₂SO₄, filtered, and concentrated. The residual was used directly without further purification.

4.2.2. Synthesis of 5,5′-Diisopropyl-3,3′-dimethyl-[2,2′-binaphthalene]-1,1′,6,6′,7,7′- Hexayl Hexaacetate 9

To a solution of the abovementioned crude apogossypol 2 in 20 mL of DCM were added DMAP (24 mg, 0.20 mmol) and Ac₂O (3.6 mL, 38.6 mmol). After the addition was completed, the reaction mixture was stirred at room temperature and monitored by TLC and LC–MS. After the reaction was completed, the solvent was removed and the residual was purified by flash chromatography to afford 1.14 g (92% yield for two steps) of compound 9 as a yellow solid.

mp 281.5–282.4 °C, ¹H NMR (600 MHz, DMSO-d₆): δ 8.18 (s, 2H), 7.58 (s, 2H), 4.13–3.77 (m, 2H), 2.43 (s, 6H), 2.31 (s, 6H), 2.13 (s, 6H), 1.95 (s, 6H), 1.44–1.38 (m, 12H). MS (ESI) calcd for C₄₀H₄₂O₁₂, 714.27; not found.

4.2.3. Synthesis of 6,6′,7,7′-Tetrahydroxy-5,5′-diisopropyl-3,3′-dimethyl-[2,2′-binaphth-alene]-1,1′-diyl Diacetate 10

Compound 9 (970 mg, 1.36 mmol) was dissolved in 40 mL of MeOH, 20 mL of DCM, and 10 mL of water, and then K₂CO₃ (1.88 g, 13.6 mmol) was added. The resulting reaction mixture was heated to 65 °C and monitored by TLC and LC–MS. After the reaction was completed, the mixture was poured into 100 mL of 2 M HCl and extracted with EA (3*200 mL). The combined organic phase was dried over Na₂SO₄, filtered, and concentrated. The residual was purified by flash chromatography to give 630 mg (85%) of compound 10 as a yellow solid. (Note: compound 10 was not stable).

mp: decomposed, ¹H NMR (400 MHz, DMSO-d₆): δ 7.80 (s, 2H), 6.88 (s, 2H), 3.87 (m, 2H), 2.03 (s, 6H), 1.87 (s, 6H), 1.46 (dd, J = 6.9, 5.1 Hz, 12H). MS (ESI) calcd for C₃₂H₃₄O₈, 546.22; found, m/z^– = 545.20.

4.2.4. Synthesis of 5,5′-Diisopropyl-6,6′,7,7′-tetramethoxy-3,3′-dimethyl-[2,2′-binaphth-alene]-1,1′-diyl Diacetate 11

To a solution of compound 10 (630 mg, 1.2 mmol) in DMF (15 mL) was added K₂CO₃ (1.6 g, 11.5 mmol), after the addition was completed, and the resulting solution was stirred for additional 15 min at room temperature. Then MeI (1.4 mL, 23.1 mmol) was added, and the reaction mixture was stirred at ambient temperature and monitored by TLC and LC–MS. After the reaction was completed, the mixture was poured into 50 mL of water and extracted with EA (3*100 mL). The combined organic phase was washed with brine, dried over Na₂SO₄, filtered, and concentrated. The residual was purified by flash chromatography to give 639 mg (92%) of compound 11 as a yellow solid.

mp 234.3–235.5 °C, ¹H NMR (600 MHz, CDCl₃): δ 7.93 (s, 2H), 6.91 (s, 2H), 3.99–3.93 (m, 8H), 3.91 (s, 6H), 2.18 (s, 6H), 1.94 (s, 6H), 1.54 (t, J = 6.3 Hz, 12H). ¹³C NMR (150 MHz, CDCl₃): δ 169.3, 152.4, 144.0, 135.2, 132.7, 128.5, 125.8, 123.4, 122.6, 99.2, 61.0, 55.3, 29.7, 26.9, 22.1, 20.5, 20.2. MS (ESI) calcd for C₃₆H₄₂O₈, 602.29; found, m/z⁺ = 603.30.

4.2.5. 5,5′-Diisopropyl-6,6′,7,7′-tetramethoxy-3,3′-dimethyl-[2,2′-binaphthalene]-1,1′-diol 12

A solution of compound 11 (639 mg, 1.1 mmol) in THF (15 mL) was cooled to 0 °C. LiAlH₄ (101 mg, 2.7 mmol) was added to the solution in batches, and the resulting mixture was stirred for 1 h. The reaction was quenched with 2 M HCl and extracted with EA (3*100 mL). The combined organic phase was washed with brine, dried over Na₂SO₄, filtered, and concentrated. The residual was purified by column chromatography to give 445 mg (81%) of compound 12 as a white solid.

mp 225.8–226.6 °C, ¹H NMR (600 MHz, CDCl₃): δ 7.71 (s, 2H), 7.49 (s, 2H), 5.26 (s, 2H), 4.09–3.97 (m, 8H), 3.94 (s, 6H), 2.15 (s, 6H), 1.57 (s, 12H). ¹³C NMR (150 MHz, CDCl₃): δ 151.5, 149.4, 134.9, 132.7, 129.1, 120.2, 117.0, 113.0, 100.5, 61.1, 55.5, 27.0, 22.1, 20.7. MS (ESI) calcd for C₃₂H₃₈O₆, 518.27; found, m/z^– = 517.30.

4.2.6. Synthesis of 5,5′-Diisopropyl-6,6′,7,7′-tetramethoxy-3,3′-dimethyl-[2,2′-binaphtha-lene]-1,1′-diyl Bis(trifluoromethanesulfonate) 13

A solution of compound 12 (401 mg, 0.77 mmol) and pyridine (0.3 mL, 3.9 mmol) in DCM (10 mL) was cooled to 0 °C. Tf₂O (0.4 mL, 2.3 mmol) was added to the solution dropwise, and the reaction mixture was stirred at ambient temperature and monitored by TLC and LC–MS. After the reaction was completed, the reaction was quenched with MeOH. The solvent was removed, and the residual was purified by column chromatography to give 563 mg (93%) of compound 13 as a white solid.

mp 212.5–214.8 °C, ¹H NMR (600 MHz, CDCl₃): δ 8.10 (s, 2H), 7.43 (s, 2H), 4.03–4.01 (m, 1H), 4.00 (s, 6H), 3.99–3.97 (m, 1H), 3.95 (s, 6H), 2.29 (s, 6H), 1.57–1.52 (m, 12H). ¹³C NMR (150 MHz, CDCl₃): δ 153.5, 142.7, 135.3, 133.2, 129.4, 125.2, 123.9, 118.0 (C–F, q, J = 320.7), 99.6, 77.2, 77.0, 76.8, 61.1, 55.5, 27.1, 22.1, 20.4. MS (ESI) calcd for C₃₄H₃₆F₆O₁₀S₂, 782.17; not found.

4.2.7. Synthesis of 5,5′-Diisopropyl-6,6′,7,7′-tetramethoxy-3,3′-dimethyl-2,2′-binaphthal-ene 6

To a solution of compound 13 (563 mg, 0.72 mmol) in DMF (10 mL) under a N₂ atmosphere were added Pd(OAc)₂ (31 mg, 0.14 mmol) and dppp (58 mg, 0.14 mmol), and the resulting mixture was stirred under N₂ for 5 min; then Et₃SiH (501 mg, 4.3 mmol) was added. The resulting mixture was sealed and heated to 90 °C and monitored by TLC and LC–MS. After the reaction was completed, the mixture was poured into water and extracted with EA (3*200 mL). The combined organic phase was washed with brine, dried over Na₂SO₄, filtered, and concentrated. The residual was purified by column chromatography to give 350 mg (93%) of compound 6 as a white solid.

mp 216.2–217.0 °C, HPLC purity: 96.2% (80% CH₃CN/H₂O), t_R = 6.67 min. ¹H NMR (600 MHz, CDCl₃): δ 8.05 (s, 2H), 7.57 (s, 2H), 7.06 (s, 2H), 4.10–4.01 (m, 2H), 3.99 (s, 6H), 3.96 (s, 6H), 2.27 (s, 6H), 1.62 (d, J = 6.9, 12H). ¹³C NMR (150 MHz, CDCl₃): δ 151.9, 147.4, 139.2, 134.8, 131.9, 130.0, 127.6, 126.9, 124.3, 105.4, 77.2, 77.0, 76.8, 61.1, 55.4, 26.8, 22.2, 20.8. MS (ESI) calcd for C₃₂H₃₈O₄, 486.28; not found.

4.2.8. Synthesis of 5,5′-Diisopropyl-6,6′,7,7′-tetramethoxy-3,3′-dimethyl-[2,2′-binaphtha-lene]-8,8′-Dicarbaldehyde 14

To a solution of compound 6 (350 mg, 0.72 mmol) in dry DCM (10 mL) at 0 °C was added TiCl₄ (410 mg, 2.16 mmol) under a N₂ atmosphere. After addition was completed, the mixture was stirred for an additional 30 min at 0 °C. Then dichloromethyl methyl ether (414 mg, 3.6 mmol) in dry DCM (5 mL) was added dropwise over 15 min. The resulting reaction mixture was stirred at ambient temperature and monitored by TLC and LC–MS. After the reaction was completed, the reaction mixture was poured onto ice-water and extracted with DCM (3*200 mL). The combined organic phase was washed with brine, dried over Na₂SO_4, filtered, and concentrated. The residual was purified by column chromatography to give 371 mg (95%) of compound 14 as a yellow solid.

mp: 255.4–265.2 °C, HPLC purity: 97.6% (80% CH₃CN/H₂O), t_R = 6.77 min. ¹H NMR (600 MHz, CDCl₃): δ 10.74 (s, 2H), 9.07 (s, 2H), 8.08 (s, 2H), 4.12–4.06 (m, 2H), 4.05 (s, 6H), 3.96 (s, 6H), 2.26 (s, 6H), 1.59 (d, J = 6.4, 12H). ¹³C NMR (150 MHz, CDCl₃): δ 191.8, 160.1, 144.5, 141.9, 134.1, 128.9, 126.8, 126.0, 124.3, 121.8, 77.2, 77.0, 76.8, 62.5, 61.1, 27.5, 22.1, 22.0, 21.0. MS (ESI) calcd for C₃₄H₃₈O₆, 542.27; found m/z⁺ = 543.25.

4.2.9. Synthesis of 6,6′,7,7′-Tetrahydroxy-5,5′-diisopropyl-3,3′-dimethyl-[2,2′-binaphtha-lene]-8,8′-dicarbaldehyde 3

To a solution of compound 14 (271 mg, 0.5 mmol) in dry DCM (10 mL) was added BBr₃ (2.5 g, 10.0 mmol) at −5 °C. The reaction was allowed to warm to room temperature and monitored by TLC and LC–MS. After the reaction was completed, the reaction was quenched with 2 M HCl/ice mixture and extracted with EA (3*100 mL). The combined organic phase was dried over Na₂SO₄, filtered, and concentrated. The residual was crystallized by EA/PE to give 209 mg (86%) of compound 3 as a gray solid.

mp: decomposed, HPLC purity: 96.1% (80% CH₃CN/H₂O), t_R = 4.87 min. ¹H NMR (600 MHz, DMSO-d₆): δ 10.74 (s, 2H), 8.43 (s, 2H), 8.07 (s, 2H), 4.13–3.85 (m, 2H), 2.19 (s, 6H), 1.52–1.44 (m, 12H). ¹³C NMR (150 MHz, DMSO-d₆): δ 195.4, 156.0, 143.6, 139.9, 135.6, 132.6, 126.4, 126.0, 124.3, 121.3, 110.8, 40.3, 40.2, 40.0, 39.9, 39.8, 39.6, 39.5, 31.5, 30.2, 27.1, 20.7. MS (ESI) calcd for C₃₀H₃₀O₆, 486.20; found m/z^– = 485.20.

4.2.10. Cell Viability Assays

The antitumor activities of compounds were determined as previously described by cell viability assays.¹⁵

4.3. Shake-Flask Aqueous Solubility Assay

Aqueous solubility assay (kinetic assay conditions) is applied to measure the compound solubility. Solubility measurement is performed starting from DMSO stock solution of the test compound. The full procedure is added in the Supporting Information..

Supporting Information Available

The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acsomega.2c02955.

• Base screening of synthesis compound 13 results, copies of ¹H, ¹³C NMR, LCMS, and HPLC spectra of all products, and aqueous solubility assay (PDF)

The authors declare the following competing financial interest(s): This research was supported by private funding from Jiangsu Dowe Biological Engineering Technology Co., Ltd. (Dowe). Dowe has filed a patent application on this synthetic process, in which W.W., Y.-Z.L. and J.-C.G. are co-inventors. W.W. is paid and owns stock in Dowe. Y.-Z.L. and J.-C.G. are employees of Dowe.