Abstract
Six sulfonamides derived from indanes and tetralines were synthesized. The human carbonic anhydrase isozymes hCA I and hCA II inhibition effects of the synthesized sulfonamides were determined. From these compounds, while N-(5,6-dimethoxy-2,3-dihydro-1H-inden-2-yl)methane sulfonamide showed the most potent inhibitory effect against hCA I (Ki = 46 ± 5.4 µM, r2 = 0.978), N-(1,2,3,4-tetrahydronaphthalene-2-yl)methanesulfonamide was found to have the best inhibitory effect against hCA II (Ki = 94 ± 7.6 µM, r2 = 0.982).
Introduction
Sulfonamides are prominent biologically active compounds. Most drugs contain this functional group in their structuresCitation1. A carbonic anhydrase (CA) inhibitor drug acetazolamide (diamox, 1) is used to treat glaucomaCitation2 and idiopathic intracranial hypertensionCitation3. A sulfonamide drug sultiame (sulthiame, 2) is an anticonvulsant that reported to be used in the treatment of epilepsy and West syndromeCitation4. A CA inhibitor diuretic ethoxzolamide (3) can be used in the treatment of glaucoma and duodenal ulcersCitation5 (Figure 1).
Figure 1. Some sulfonamide drugs.
The CAs (E.C: 4.2.1.1) form a family of enzymes that catalyze the rapid interconversion of carbon dioxide (CO2) and water to bicarbonate () and protons (H+), a reversible reaction that occurs rather slowly in the absence of a catalyst. The reaction catalyzed by CA is:
In the hydration direction, the first step is the nucleophilic attack of a Zn2+-bound hydroxide ion on CO2 with consequent formation of , which is then displaced from the active site by a water molecule. Finally, it regenerates the catalytically active Zn2+-bound hydroxide ion through a proton transfer reaction from the Zn2+-bound water molecule to an exogenous proton acceptor or to an active site residue. Sulfonamides are the most important CA inhibitors, bind in a tetrahedral geometry of the Zn2+ ions, in deprotonated stateCitation6.
clearly shows that the deprotonated sulfonamide is coordinated to the Zn2+ ion of the CA isoenzymes, and its NH moiety participates in hydrogen bond with Thr 199, which in turn is engaged in another hydrogen bond to the carboxylate group of Glu 106, whereas one of the oxygen atoms of the sulfonamide moiety also participated in hydrogen bond with backbone-NH moiety of Thr 199Citation7. These structures provide close insight into why the sulfonamide group appears to have unique properties for CA inhibition.
Scheme 1. Putative CA inhibition mechanism by sulfonamide inhibitors.
CAs are ubiquitous zinc enzymes and present in Archaea, prokaryotes and eukaryotes, encoded by three distinct and evolutionarily unrelated gene families: the α-CAs present in vertebrates, eubacteria, algae and cytoplasm of green plants. The β-CAs are predominantly in eubacteria, algae and chloroplasts of both mono- as well as dicotyledons. The γ-CAs are mainly in Archaea and some eubacteriaCitation1,Citation8. Up to now, in higher vertebrates, including humans, 16 different CA isozymes or CA-related proteins have been described, which possess different subcellular localization and tissue distributionCitation1,Citation7. Among CA isozymes are the cytosolic ones (CA I, CA II, CA III, CA VII); CA-I is found together with CA-II in erythrocytes. CA-II is the most widely distributed CA in the eye, kidney, central nervous system (CNS) and inner ear. There are also membrane-bound (CA IV, CA IX, CA XII, CA XIV), mitochondrial (CA V), secretory forms (CA VI) and several acatalytic forms (CA VIII, CA X, CA XI)Citation9,Citation10. Other CA isoforms are found in a variety of tissues where they participate in several important biologic processesCitation11–14. These enzymes catalyze a very simple physiological reaction, the interconversion between CO2 and , and are involved in crucial physiological processes connected with respiration and transport of
between metabolizing tissues and the lungs, pH and CO2 homeostasis, electrolyte secretion in a variety of tissues or organs, biosynthetic reactions such as gluconeogenesis, lipogenesis, and ureagenesis, bone resorption, calcification, tumorigenicity, and many other physiologic or pathologic processes. Thus, it is not surprising that many of these isozymes have been discovered as important targets for inhibitors with clinical applicationsCitation1,Citation7,Citation8,Citation15. Almost all of the most potent inhibitors of CAs contain a terminal sulfonamide as the anchoring group to coordinate the catalytic zincCitation16,Citation17. These sulfonamides are widely used clinically, mainly as antiglaucoma agents but also for the therapy of other diseases such as increased intracranial pressure, various neurological/neuromuscular pathologies such as epilepsy, genetic hemiplegic migraine, and ataxia, tardive dyskinesia, hypokalemic periodic paralysis, essential tremor and Parkinson’s disease, and mountain sickness. Accordingly, drugs of this pharmacological class are under constant developmentCitation1,Citation8,Citation13,Citation14.
Because of the important biological activities of sulfonamides, the synthesis and CA inhibitory properties of novel sulfonamides 28–33 will be useful for further synthetic and biological purposes. In this context, here we report the first synthesis of sulfonamides 28, 30–33. We also evaluate CA isoenzymes (hCA I and hCA II) inhibitory affects of compounds 28–33.
Experimental
All chemicals and solvents are commercially available. All solvents were distilled and dried according to standard procedures. Silica gel (SiO2, 60 mesh; Merck, Darmstadt, Germany) was used for column chromatography (CC). In this study, 1 mm of SiO2 60 PF (Merck) on glass plates was used for preparative thick layer chromatography. The m.p. of all compounds was determined with cap. melting-point apparatus (BUCHI 530; Flawil, Switzerland); uncorrected. IR spectra were recorded as solutions in 0.1 mm cells with a Mattson 1000 FT-IR spectrophotometer (Cambridge, England). 1H- and 13C-NMR spectra were recorded on 400 (100)-MHz Varian spectrometer (Danbury, CT) in deuterated solvents (CDCl3 and D2O) with tetramethylsilane (TMS, SiMe4) as an internal standard for protons and solvent signals as internal standard for carbon spectra. Chemical shift values were mentioned δ in ppm. Elemental analyses were recorded on Leco CHNS-932 apparatus (Saint Joseph, MI). CA inhibitory properties of samples were determined on a spectrophotometer (UV-1208, Shimadzu, Japan).
Synthesis
Synthesis of alcohols 7Citation18, 8Citation19 and 9Citation20, compound 1621, amine hydrochloride salt 21Citation22 and amine 22Citation2Citation3 has been described in the literature.
Standard procedure for the synthesis of azides: 1-azido-5,6-dimethoxyindane (10)
Alcohol 7 (4.47 g, 23.16 mmol) was dissolved in dry THF (100 ml). To this solution, diphenylphosphoryl azide (DPPA; 7.62 , 27.68 mmol) and DBU (4.19 g, 27.68 mmol) were added at 0 °C under Ar(g). The mixture was stirred for 2 h at the same temperature, then at 25 °C for 12 h. After the solvent was evaporated, CH2Cl2 (80 ml) and 10%HCl (30 ml) were added to the residue. Organic phase was separated and washed with 10%HCl (2 × 20 ml). Drying of the organic layer over Na2SO4, evaporation of the solvent and the CC of the residue on silica gel (30 g) with 10% EtOAc–hexane afforded azide 10 (3.60 g, 71% yield).
1-Azido-5,6-dimethoxyindane (10)
Colorless oil. 1H-NMR (400 MHz, CDCl3) δ 6.88 (s, 1H, Ar-H), 6.77 (s, 1H, Ar-H), 4.79 (dd, 1H, CH-N3, J = 4.2 and J = 7.1 Hz), 3.88 (s, 3H, OCH3), 3.86 (s, 3H, OCH3), 3.02 (A part of AB, m, 1H, Ha of CH2), 2.80 (B part of AB, ddd, 1H, Hb of CH2, J = 4.8, J = 8.4 and J = 15.4 Hz), 2.50–2.41 (m, 1H, H of CH2), 2.17–2.10 (m, 1H, H of CH2). Citation13C-NMR (100 MHz, CDCl3) δ 150.3 (C), 148.7 (C), 136.0 (C), 132.4 (C), 107.7 (CH), 107.4 (CH), 66.7 (CH–N3), 56.3 (OCH3), 56.2 (OCH3), 33.1 (CH2), 30.7 (CH2). IR (CH2Cl2, cm−1): 3070, 2999, 2938, 2857, 2833, 2092, 1607, 1505, 1465, 1413, 1339, 1311, 1260, 1223, 1188, 1092. Anal. Calcd for (C11H13N3O2): C 60.26, H 5.98 and N 19.17; Found C 60.30, H 5.97 and N 19.13.
1-Azidoindane (11)
A yellowish oily azide 11 was synthesized from 8 by applying the standard procedure described above for the synthesis of azides (87% yield). 1H- and 13C-NMR data of 11 are in good agreement with data given in the literatureCitation24.
1-Azido-5-methoxy-1,2,3,4-tetrahydronaphthalene (12)
Compound 12 was synthesized from 9 as described in “Standard Procedure for the synthesis of azides: 1-azido-5,6-dimethoxyindane (10)” section, 80% yield, oily. 1H-NMR (400 MHz, CDCl3) δ 7.24 (t, 1H, Ar-H, J = 8.1 Hz), 6.95 (d, 1H, Ar-H, J = 8.1 Hz), 6.82 (d, 1H, Ar-H, J = 8.1 Hz), 4.58–4.57 (m, 1H, CH–N3), 3.95 (s, 3H, OCH3), 2.96–2.79 (m, 1H, H of CH2), 2.60–2.54 (m, 1H, H of CH2) and 2.02–1.83 (m, 4H, 2CH2). Citation13C-NMR (100 MHz, CDCl3) δ 157.5 (C), 135.0 (C), 126.7 (CH), 126.6 (C), 121.4 (CH), 109.5 (CH), 59.7 (CH–N3), 55.7 (OCH3), 28.9 (CH2), 22.9 (CH2) and 18.5 (CH2). IR (CH2Cl2, cm−1): 2940, 2937, 2098, 1587, 1470, 1438, 1331, 1253, 1102 and 1065. Anal. Calcd for (C11H13N3O): C 65.01, H 6.45 and N 20.68; Found C 65.04, H 6.49 and N 20.60.
Standard procedure for the synthesis of amine hydrochlorides: 1-amino-5,6-dimethoxyindane hydrochloride (13)
Pd-C (50 mg) and azide 10 (2.00 g, 12 mmol) in MeOH (50 ml) and CHCl3 (4 ml) were placed in a 100-ml flask. A balloon filled with H2 gas (3 L) was fitted to the flask. The mixture was deoxygenated by flushing with H2 and then hydrogenated at RT for 20 h. The catalyst was removed by filtration. Recrystallization of the residue from MeOH–Et2O gave 1-amino-5,6-dimethoxyindan hydrochloride (13) (1.98 g, 94% yield).
1-Amino-5,6-dimethoxyindane hydrochloride (13)
Colorless crystal. M.p.: 247–249 °C. 1H-NMR (400 MHz, D2O) δ 6.91 (s, 1H, Ar-H), 6.88 (s, 1H, Ar-H), 4.64–4.58 (m, 4H, CH–N, NH3 and H2O), 3.67 (s, 6H, 2OCH3), 2.95–2.87 (A part of AB, m, 1H, Ha of CH2), 2.77–2.69 (B part of AB, m, 1H, Hb of CH2), 2.42–2.35 (m, 1H, H of CH2) and 2.00–1.93 (m, 1H, H of CH2). Citation13C-NMR (100 MHz, D2O) δ 149.7 (C), 147.6 (C), 137.6 (C), 130.0 (C), 108.3 (CH), 107.6 (CH), 56.2 (OCH3), 56.0 (OCH3), 55.8 (CH–N), 30.4 (CH2) and 29.7 (CH2).
1-Aminoindane hydrochloride (14)
Standard procedure described in “Standard procedure for the synthesis of amine hydrochlorides: 1-amino-5,6-dimethoxyindane hydrochloride (13)” section was applied to azide 11 to give 14, 91% yield, white solid. M.p.: 207–209 °C (literatureCitation25 m.p.: 207–208 °C). 1H-NMR (400 MHz, CDCl3) δ 7.31 (d, 1H, Ar-H, J = 7.5 Hz), 7.24–7.15 (m, 3H, Ar-H), 4.69–4.59 (m, 4H, CH–N, NH3 and H2O), 3.00–2.93 (A part of AB, m, 1H, H of CH2), 2.82 (B part of AB, ddd, 1H, H of CH2, J = 5.7, J = 8.7 and J = 16.0 Hz), 2.46–2.37 (m, 1H, H of CH2), 1.98–1.90 (m, 1H, H of CH2). Citation13C-NMR (100 MHz, D2O) δ 144.6 (C), 138.1 (C), 129.8 (CH), 127.2 (CH), 125.5 (CH), 124.5 (CH), 55.8 (CH-N), 30.2 (CH2) and 29.7 (CH2).
1-amino-5-methoxy-1,2,3,4-tetrahydronaphthalene hydrochloride (15)
Applying the procedure described in “1-Azido-5-methoxy-1,2,3,4-tetrahydronaphthalene (12)” section to azide 12 gave white solid 15 with 93% yield. M.p.: 248–250 °C (literatureCitation26 m.p.: >250 °C). 1H-NMR (400 MHz, D2O) δ 7.08 (t, 1H, Ar-H, J = 8.1 Hz), 6.79 (d, 1H, Ar-H, J = 8.1 Hz), 6.78 (d, 1H, Ar-H, J = 8.1 Hz), 4.64 (bs, 1H of NH3 and H of H2O), 4.32 (dd, 1H, CH–N, J = 5.0 and J = 9.9 Hz), 3.61 (s, 3H, OCH3), 2.54 (dt, 1H, H of CH2, J = 5.5 and J = 17.8 Hz), 2.33 (dt, 1H, H of CH2, J = 7.3 and J = 17.8 Hz), 1.92–1.74 (m, 2H, CH2), 1.72–1.60 (m, 2H, CH2). Citation13C-NMR (100 MHz, D2O) δ 157.0 (C), 132.4 (C), 127.3 (CH), 126.8 (C), 120.6 (CH), 111.0 (CH), 55.6 (OCH3), 49.0 (CH–N), 26.7 (CH2), 22.0 (CH2) and 17.0 (CH2).
Methyl 5,6-dimethoxyindane-2-carboxylate (17)
Et3SiH (7.43, 63.94 mmol) was added to a solution of 16 (4.00 g, 15.98 mmol) in TFA (15 ml) under N2 at room temperature. After the reaction mixture was refluxed for 2.5 h, TFA was evaporated. EtOAc (80 ml) and a concentrated solution of Na2CO3 in H2O (50 ml) were added to the residue. Organic phase was separated and again washed with a solution concentration Na2CO3 in H2O (50 ml). The organic phase was dried over Na2SO4 and solvent was evaporated. Chromatography of the residue on silica gel (25 g) column with 20% EtOAc–hexane yielded yellowish oily 17 (3.02 g, 80%). 1H-NMR (400 MHz, CDCl3) δ 6.72 (s, 2H, Ar-H), 3.70 (s, 3H, OCH3), 3.696 (s, 3H, OCH3), 3.692 (s, 3H, OCH3), 3.35–3.32 (m, 1H, CH–CO), 3.21–3.09 (m, 4H, 2CH2). Citation13C-NMR (100 MHz, CDCl3) δ 176.0 (CO), 148.4 (2C), 133.2 (2C), 107.7 (2CH), 56.2 (2OCH3), 52.1 (OCH3), 44.0 (CH-CO) and 36.4 (2CH2).
5,6-Dimethoxy-indane-2-carboxylic acid (18)
Compound 17 (2.50 g, 10.58 mmol) was dissolved in MeOH (60 ml) and to this solution, 4 M NaOH (10 ml) was added at room temperature. The reaction mixture was stirred at room temperature for 20 h. After most of MeOH was evaporated, CH2Cl2 (20 ml) and H2O (20 ml) were added to the residue. Organic layer was dispatched and the aqueous phase was acidified with 37% HCl (pH < 2). EtOAc (40 ml) was added to this acidified solution and organic layer was separated. H2O phase was extracted with EtOAc (2 × 50 ml) and combined organic layers were dried over Na2SO4. After evaporation of the solvent and crystallization of the residue with CH2Cl2–hexane, acid 18 was obtained (2.25 g, 96%). M.p.: 124–126 °C, 1H-NMR (400 MHz, CDCl3) δ 11.4 (bs, 1, OH), 6.74 (s, 2H, Ar-H), 3.85 (s, 6H, 2OCH3), 3.41–3.25 (m, 1H, CH–CO), 3.23–3.14 (m, 4H, 2CH2). Citation13C-NMR (100 MHz, CDCl3) δ 182.1 (CO), 148.6 (2C), 133.0 (2C), 107.7 (2CH), 56.3 (2OCH3), 43.9 (CH-CO) and 36.2 (2CH2).
Benzyl 5,6-dimethoxyindane-2-ylcarbamate (19)
To a stirred solution of 18 (2.10 g, 9.45 mmol) in anhydrous benzene (60 ml), DPPA (3.12 g, 11.33 mmol) and Et3N (1.15 g, 11.33 mmol) were added. The mixture was heated at reflux temperature for 4 h. Then, benzyl alcohol (3.07 g, 28.48 mmol) was added, and refluxing of the mixture was continued for 30 h. The mixture was cooled to room temperature, the solvent was evaporated. Purifying of the resulting residue by CC on silica gel (20 g) with 20% EtOAc–hexane afforded 2.45 g (79%) of 19, white solid. M.p.: 126–128 °C. 1H-NMR (400 MHz, CDCl3) δ 7.33 (bs, 5H, Ph-H), 6.73 (s, 2H, Ar-H), 5.08 (s, 2H, OCH2), 4.52 (bs, 1H, NH), 3.88–3.79 (m, 7H, H of CH–N and 6H of 2OCH3), 3.23 (A part of AB, dd, 2H, 2Ha-CH, J = 6.6 and J = 15.5 Hz), 2.73 (B part of AB, dd, 2H, 2Hb-CH, J = 4.1 and J = 15.5 Hz). Citation13C-NMR (100 MHz, CDCl3) δ 156.2 (CO), 148.6 (2C), 136.7 (C), 132.5 (2C), 128.8 (2CH), 128.4 (2CH), 108.2 (3CH), 66.9 (OCH2), 56.3 (2OCH3), 52.9 (CH–N) and 40.5 (2CH2). IR (CH2Cl2, cm−1): 3339, 3063, 3033, 2997, 2939, 2834, 1697, 1610, 1530, 1504, 1465, 1454, 1313, 1259, 1227, 1188, 1100 and 1044. Anal. Calcd for (C19H21NO4): C 69.71, H 6.47 and N 4.28; Found C 69.68, H 6.48 and N 4.28.
2-Amino-5,6-dimethoxyindane hydrocloride (20)
The hydrogenolysis reaction of carbamate 19 was performed by the similar procedure described above for the reduction of azides to amine hydrochlorides in “1-Azido-5-methoxy-1,2,3,4-tetrahydronaphthalene (12)” section. But, here instead of azides, carbamate 19 was used to give 20 (95%) as colorless crystals m.p. 287–289 °C (literatureCitation22 m.p. 287–289 °C). 1H- and Citation13C-NMR data are in agreement with data given previouslyCitation22.
Standard procedure for hydrolysis of amine hydrochloride salts to amines: 1-amino-5,6-dimethoxyindane (23)
Amine hydrochloride salt 13 (1.98 g, 12.00 mmol) was dissolved in MeOH (40 ml) and cooled to 0 °C. To this solution, solution of 10% NaOH (20 ml) was added. The reaction mixture was stirred at room temperature for 3 h. After most of MeOH was evaporated, CH2Cl2 (50 ml) and H2O (20 ml) were added to the residue. Organic layer was separated and H2O layer was extracted with CH2Cl2 (2 × 30 ml). Combined organic layers were dried over Na2SO4 and CH2Cl2 was evaporated. Oily aminoindane 23 (1.45 g, 94%) was synthesized and used without further purification in the next step. Amines 24–27 were synthesized from their corresponding salts 14, 15, 20 and 21 by the same procedure with yields of 92%, 95%, 95% and 94%, respectively.
Standard procedure for the synthesis of sulfonamides: N-(5,6-dimethoxyindane-1-yl)methanesulfonamide (28)
Amine 23 (0.47 g, 2.44 mmol) was dissolved in CH2Cl2 (30 ml) and this solution was cooled to 0 °C. To this solution, Et3N (0.23 g, 2.93 mmol) and MeSO2Cl (0.34 g, 2.93 mmol) were added. The reaction mixture was stirred at room temperature for 15 h. After the solvent was evaporated, the residue was chromatographed on silica gel (30 g) column with 30% EtOAc–hexane. Sulfonamide 28 was synthesized as white solid (0.53 g, 80% yield).
N-(5,6-dimethoxyindane-1-yl)methanesulfonamide (28)
Melting point (m.p.): 144–146 °C. 1H-NMR (400 MHz, CDCl3) δ 6.90 (s, 1H, Ar-H), 6.72 (s, 1H, Ar-H), 4.93–4.88 (m, 1H, CH-N), 4.66 (bd, 1H, NH, J = 9.2 Hz), 3.85 (s, 3H, OCH3), 3.84 (s, 3H, OCH3), 3.03 (s, 3H, CH3), 2.94 (A part of AB, ddd, 1H, CH2, J = 3.9, J = 8.3 and J = 15.5 Hz), 2.81–2.73 (B part of AB, m, 1H, CH2), 2.61 (dddd, 1H, CH2, J = 4.0, J = 7.8, J = 12.1 and J = 16.8 Hz), 1.96–1.87 (m, 1H, CH2). Citation13C-NMR (100 MHz, CDCl3) δ 149.9 (C), 148.9 (C), 134.9 (C), 133.5 (C), 107.7 (CH), 107.2 (CH), 59.4 (CH–N), 56.4 (OCH3), 56.2 (OCH3), 42.1 (CH3), 35.5 (CH2), 30.1 (CH2). IR (CH2Cl2, cm−1): 3284, 3002, 2935, 2857, 2834, 1608, 1504, 1465, 1412, 1309, 1264, 1221, 1188, 1148, 1097. Anal. Calcd for (C12H17NO4S): C 53.12, H 6.32, N 5.16 and S 11.82; Found C 53.08, H 6.34, N 5.12 and S 11.80.
N-(Indane-1-yl)methanesulfonamide (29)
Yield 82%; m.p.: 93–95 °C. 1H-NMR (400 MHz, CDCl3) δ 7.37–7.19 (m, 4H, Ar-H), 5.06 (bd, 1H, NH, J = 9.0 Hz), 4.89–4.83 (m, 1H, CH–N), 3.00–2.93 (m, 4H, 1H of CH2 and CH3), 2.84–2.76 (m, 1H of CH2), 2.55 (dddd, 1H of CH2, J = 3.6, J = 7.8, J = 11.3 and J = 15.8 Hz), 1.89 (dddd, 1H of CH2, J = 3.6, J = 8.1, J = 12.8 and J = 15.8 Hz). Citation13C-NMR (100 MHz, CDCl3) δ 143.0 (C), 142.4 (C), 128.5 (CH), 127.1 (CH), 125.1 (CH), 124.5 (CH), 59.0 (CH-N), 41.9 (CH3), 35.1 (CH2), 30.2 (CH2). Anal. Calcd for (C10H13NO2S): C 56.85, H 6.20, N 6.63 and S 15.18; Found C 56.82, H 6.18, N 6.66 and S 15.21.
N-(5-metoksi-1,2,3,4-tetrahydronaphthalene-1-yl)methanesulfonamide (30)
Yield 83%; m.p.: 142–144 °C. 1H-NMR (400 MHz, CDCl3) δ 7.18 (dd, 1H, Ar-H, J = 7.7 and J = 8.1 Hz), 7.02 (d, 1H, Ar-H, J = 7.7 Hz), 6.75 (d, 1H, Ar-H, J = 8.1 Hz), 4.64 (ddd, 1H, CH–N, J = 5.5, J = 7.7 and J = 13.0 Hz), 4.50 (d, 1H, NH, J = 7.7 Hz), 3.82 (s, 3H, OCH3), 3.07 (s, 3H, CH3), 2.72 (A part of AB, dt, 1H, Ha of CH2, J = 6.1 and J = 18.0 Hz), 2.57 (B part of AB, dt, 1H, Hb of CH2, J = 7.1 and J = 18.0 Hz), 2.07–1.79 (m, 4H, 2CH2). Citation13C-NMR (100 MHz, CDCl3) δ 157.4 (C), 137.0 (C), 127.0 (CH), 126.9 (C), 120.8 (CH), 109.0 (CH), 55.6 (OCH3), 52.5 (CH-N), 42.4 (CH3), 31.1 (CH2), 22.9 (CH2) and 18.8 (CH2). IR (CH2Cl2): 3647, 3280, 3008, 2935, 2862, 1587, 1470, 1438, 1408, 1319, 1252, 1149, 1101, 1078 cm−1. Anal. Calcd for (C12H17NO3S): C 56.45, H 6.71, N 5.49 and S 12.56; Found C 56.42, H 6.72, N 5.49 and S 12.57.
N-(1,2,3,4-tetrahydronaphthalene -2-yl)methanesulfonamide (31)
85% yield; m.p.: 126–128 °C. 1H-NMR (400 MHz, CDCl3) δ 7.16–7.05 (m, 4H, Ar-H), 4.52 (d, 1H, NH, J = 7.5 Hz), 3.87–3.81 (m, 1H, CH–N), 3.17 (A part of AB, dd, 1H, Ha of CH2, J = 4.7 and J = 16.2 Hz), 3.01 (s, 3H, CH3) 2.92 (t, 2H, CH2, J = 6.6 Hz), 2.77 (B part of AB, dd, 1H, Hb of CH2, J = 8.2 and J = 16.2 Hz), 2.18–2.11 (m, 1H, CH2), 1.90–1.81 (m, 1H, CH2). Citation13C-NMR (100 MHz, CDCl3) δ 135.2 (C), 133.5 (C), 129.6 (CH), 129.1 (CH), 126.7 (CH), 126.4 (CH), 50.0 (CH-N), 42.2 (CH3), 37.2 (CH2), 30.3 (CH2) and 27.2 (CH2). IR (CH2Cl2): 3647, 3272, 3020, 2928, 2852, 1494, 1439, 1316, 1150, 1113, 1072 cm−1. Anal. Calcd for (C11H15NO2S): C 58.64, H 6.71, N 6.22 and S 14.23; Found C 58.63, H 6.71, N 6.28 and S 14.25.
N-(5,6-dimethoxy-2,3-dihydro-1H-inden-2-yl)methanesulfonamide (32)
78% yield; mp.: 162–164 °C. 1H-NMR (400 MHz, CDCl3) δ 6.72 (s, 2H, Ar-H), 4.89 (bd, 1H, NH, J = 8.4 Hz), 4.31–4.22 (m, 1H, CH–N), 3.83 (s, 6H, 2OCH3), 3.24 (A part of AB, dd, 2H of CH2, J = 7.2 and J = 15.5 Hz), 2.98 (s, 3H, CH3), 2.83 (B part of AB, dd, 2H of CH2, J = 5.8 and J = 15.5 Hz). Citation13C-NMR (100 MHz, CDCl3) δ 148.7 (2C), 131.8 (2C), 108.1 (2CH), 56.3 (2OCH3), 55.3 (CH–N), 41.7 (CH3) and 41.0 (2CH2). IR (CH2Cl2): 3249, 2958, 2930, 2840, 1605, 1560, 1504, 1443, 1404, 1311, 1219, 1188, 1169, 1152, 1096 cm−1. Anal. Calcd for (C12H17NO4S): C 53.12, H 6.32, N 5.16 and S 11.82; Found C 53.10, H 6.31, N 5.18 and S 11.85.
N-(Indane-2-yl)methanesulfonamide (33)
80% yield; m.p.: 134–136 °C. 1H-NMR (400 MHz, CDCl3) δ 7.23–7.17 (m, 4H, Ar-H), 4.64 (ddd, 1H, CH–N, J = 5.5, J = 7.7 and J = 13.0 Hz), 4.70 (bd, 1H, NH, J = 7.7 Hz), 4.31 (m, 1H, CH–N), 3.30 (A part of AB, dd, 2H, 2Ha of CH2, J = 7.1 and J = 15.7 Hz), 3.01 (s, 3H, CH3) 2.91 (B part of AB, dd, 2H, 2Hb of CH2, J = 5.9 and J = 15.7 Hz). Citation13C-NMR (100 MHz, CDCl3) δ 140.15 (2C), 127.3 (2CH), 124.95 (2CH), 55.1 (CH–N), 41.81 (2CH2) and 41.02 (CH3). IR (CH2Cl2): 3230, 3064, 2964, 2930, 2901, 1459, 1436, 1306, 1228, 1135, 1103, 1018 cm−1. Anal. Calcd for (C10H13NO2S): C 56.85, H 6.20, N 6.63 and S 15.18; Found C 56.88, H 6.23, N 6.60 and S 15.21.
Purification of hCA I and hCA II by affinity chromatography
CA isoenzymes were purified affinity chromatography using a column packed with Sepharose 4B-L-Tyrosine sulphanilamide resin via a simple one-step method using Sepharose-4B-L tyrosine-sulphanilamide affinity gel chromatography; described previouslyCitation27,Citation28.
Hydratase and esterase activity assays
The effect of novel sulfonamides 28–33 on HCA isozyme activity was determined colorimetrically using CO2-hydration method of Wilbur and AndersonCitation29 described previouslyCitation9. Esterase activity was determined according to the method described by Verpoorte et alCitation30.
Protein determination
The yield of protein during the purification steps was determined spectrophotometrically at 595 nm according to the Bradford methodCitation31, explained previously using bovine serum albumin as the standard proteinCitation32.
SDS polyacrylamide gel electrophoresis
SDS polyacrylamide gel electrophoresis (SDS-PAGE) by the method of LaemmliCitation33 was performed for determination of isoenzymes purity as described previouslyCitation34.
Results and discussion
Chemistry
Reduction of ketones 4–6 with NaBH4 in MeOH at 0–25 °C gave corresponding alcohols 7,Citation18 8Citation19 and 9Citation20. Mitsunobu reactions of sec-alcohols with HN3 have been given in the literature clearlyCitation35,Citation36. By applying this method to compounds 7–9, we failed to synthesize azides 10–12. In the literature, the synthesis of benzylazides from benzylalcohols has been managed with DPPA in the presence of DBU, which is known as an alternative Mitsunobu reactionCitation37. By a similar approach, the reaction of alcohols 7–9 with DPPA under mild conditions afforded azides 10, 11Citation24 and 12 in good yields. Reduction of azides to their corresponding amine hydrochloride salts via Pd-C catalyzed hydrogenation in MeOH–CHCl3 has been reported.Citation22,Citation38 Following the same synthetic methodology, we synthesized amine hydrochloride salts 13, 14 and 15 ().
Scheme 2. Synthesis of benzylamine hydrochloride salts: (i) NaBH4, MeOH, 0–25 °C, 3 h; (ii) DPPA/DBU, THF, 0 °C, 2 h, then 25 °C, 12 h, Ar(g); and (iii) H2/Pd-C, CHCl3–MeOH, 25 °C, 20 h.
The reaction of 4 with CO(OMe)2 in the presence of NaH in THF at reflux temperature afforded 16. The reduction of ketone group of 16 with Et3SiH in CF3CO2H gave 17. Compound 18 was synthesized from the hydrolysis of 17 with aqueous NaOH in MeOH for 15 h then by acidification with concentrated HCl. The Curtius reactions of carboxylic acids with DPPA in benzene to alkyl isocyanates, the conversion of alkyl isocyanates to their corresponding carbamates and hydrogenolysis of carbamates to amine hydrochloride salts have been described previously.Citation39–41 By following this procedure, refluxing of acid 18 with DPPA in the presence of Et3N in benzene for 6 h, then addition of PhCH2OH and hitting of the reaction mixture at the same temperature for 30 furnished a new carbamate 19. Pd-C catalyzed hydrogenolysis of 19 in MeOH–CHCl3 gave dopamine analogue 20Citation2Citation4 ().
Scheme 3. Synthesis of 2-aminoindane hydrochloride 20: (i) (MeO)2CO/NaH, THF, 66 °C, 12 h; (ii) Et3SiH, TFA, 72 °C, 2.5 h; (iii) a) 4 M NaOH solution, MeOH, 25 °C, 20 h; (b) 37% HCl; (iv) (PhO)2PON3, Et3N, C6H6, reflux, 4 h then PhCH2OH, reflux, 30 h; and (v) H2/Pd-C, CHCl3–MeOH, 25 °C, 20 h.
Compounds 21Citation22 and 22Citation2Citation3 were synthesized according to the literature procedure. Amine hydrochloride salts 13–15, 20 and 21 were hydrolyzed to amines 23–27 with aqueous NaOH in MeOH. The synthesized amines 22–27 were used without further purification and characterization. All synthesized amines were converted to their sulfonamides 28–33 with MeSO2Cl in the presence of Et3N in CH2Cl2 at 0–25 °C (). The structures of all synthesized compounds were characterized by 1H- and Citation13C-NMR. Azide functional groups were characterized by IR.
Scheme 4. Synthesis of sulfonamides: (i) 10% NaOH solution, MeOH, 0–25 °C, 3 h and (ii) MeSO2Cl/NEt3, CH2Cl2, 15 h.
CA isoenzyme inhibition studies
CA isoenzyme purification and activity assay
In order to remind CA inhibition mechanism by sulfonamide inhibitors () should be noted that the active domain of CA isoenzymes contains an active zinc ion (Zn2+) site; a strong Lewis acid that binds to, and activates a substrate H2O molecule to catalyze the reversible hydration reaction of carbon dioxide. The metal ion is situated at the bottom of active site, being coordinated by three histidine residues (His 94, His 96 and His 119) and water molecule/hydroxide ionCitation42 (). Also, it was known that sulfonamides were the most important CA inhibitors, bind in a tetrahedral geometry of the Zn2+ ion, in deprotonated stateCitation1,Citation7 ().
In this study, CA isoenzymes I and II (hCA I and hCA II) were purified from human erythrocytes. The purification of both CA isozymes was performed using a simple one-step method with a Sepharose-4B-l-thyrosine-sulfanilamide affinity CCCitation43. Human erythrocyte CA I isoenzyme was purified, 336.1-fold with a specific activity of 1445.2 EU mg−1 and overall yield of 62.4%; CA II isoenzyme was purified, 298.9-fold with a specific activity of 1285.4 EU mg−1 and an overall yield of 27.1%. We report here the first study on the inhibitory effects of novel sulfonamides 28–33 on the esterase activity of hCA I and hCA II. The inhibitor concentration that caused 50% inhibition (IC50) was determined from activity versus (%)-(sulfonamides) plots and the average inhibition constant (Ki values) was calculated from Lineweaver–Burk plots (). The data of show the following inhibition of hCA I and hCA II with novel sulfonamides 28–33, by an esterase assay, with 4-NPA as substrate.
Table 1. Human CA isoenzymes (hCA I and hCA II) inhibition data with some novel sulfonamides 28-33 by an esterase assay with 4-nitrophenylacetate as substrate.
The metal-complexing anions and the sulfonamides with a terminal SO2NH2 group that coordinates to the Zn2+ ion. Simple anions such as HS-, CN-, NCO-, N3-, , I− and HCOO− may bind with tetrahedral geometry or form trigonal–bipyramidal coordinationCitation1,Citation44, whereas the sulfonamides replace the water coordinated to zinc and the “deep-water” hydrogen-bonded to Thr199NH. Both waters are present in the uncomplexed stateCitation45. It had been shown that sulfamide and sulfamic acid act as moderate hCA II inhibitors, with inhibition constants of 1130 μM for sulfamide and 390 μM for sulfamic acid at the physiological pH (7.4), respectively. This contrasts remarkably to the strongly reduced affinity of the sulfate ion (1–2 M) toward hCA IICitation7.
CA isoenzyme inhibition effects
hCA I and hCA II inhibitory effects of the novel sulfonamides 28–33 were tested under in vitro conditions and IC50 and Ki values were calculated and given in .
We report here the initial study on the inhibitory effects of synthesized novel sulfonamides 28–33 and on the hydratase activity of hCA I and hCA II. The data in show the following regarding the inhibition of hCA I and hCA II activity by novel sulfonamides 28–33. The strongest inhibitory activity has been observed with compound 32 (IC50 = 266 µM, Ki = 46 ± 5.4 µM, r2 = 0.978), investigated here for the inhibition of the rapid cytosolic isozymes hCA I. We speculated that two methyl groups of both molecules sterically facilitate the inhibition of hCA I. Similarly, the compound 28 (IC50 = 487 µM, Ki = 52 ± 05.4 , r2 = 0.982) with the two methyl groups demonstrated same hCA I inhibitory activity. On the other hand, the most powerful hCA II isoenzyme inhibitory effect found in the compound 31 with Ki: 94 ± 7.6 µM (IC50 = 338 µM, r2 = 0.982, ). The half maximal inhibitory concentration (IC50) is a measure of the effectiveness of novel sulfonamides 28–33 in inhibiting CA isoenzyme function. A lower IC50 value reflects strong CA isoenzyme inhibition effect. Also, the remaining other sulfonamides were quite effective cytosolic isozyme hCA I inhibitors with IC50s within the range of 52 ± 9.2–228 ± 6.43 µM and hCA II inhibitors with IC50s in the range of 112 ± 28–372 ± 12.3 µM (). It is widely known that sulfonamides compounds are CA isoenzymesCitation46,Citation47. The results obtained from clearly showed that the physiologically dominant cytosolic isozyme hCA I and hCA II were effectively inhibited by compounds 22–28, with Kis within the range of 52 ± 9.2–372 ± 12.3 µM. The structure–activity relationship and inhibition properties of hCA II isoenzyme is particularly comparable to that what is outlined above for hCA I. Due to the two enzymes having a high sequence homology of amino acid present within the active siteCitation48.
The synthesized novel sulfonamides 28–33 have one aromatic ring. These new synthesized compounds have been investigated as CA isoenzymes inhibitors in this study. The rationale of investigating these compounds as CA inhibitors exists in the fact that the compounds with aromatic rings have been shown to be the only competitive inhibitor with CO2 as the substrate for the main isoform of CA, i.e. human CA I and II.
Conclusions
In summary, a series of novel 28, 30–33 and a known sulfonamide 29 were synthesized starting from convenient reagents. The synthesized compounds may be important for further synthetic and biological purposes. In addition, CA inhibitory properties of the synthesized compounds 28–33 have also been evaluated. These compounds 28, 30–33 are selective hCA I and hCA II inhibitors with selectivity ratios in the range of 52 ± 9.2–372 ± 12.3 µM.
Declaration of interest
We are greatly indebted to The Scientific and Technological Research Council of Turkey (TUBITAK, Grant no. TBAG-109T241) and Ataturk University (BAP2012/152) for their financial support of this study for (YA, AA, SG).
The authors report no conflicts of interest.
Notice of Correction:
The iFirst version of this article published online ahead of print on 12 January 2013 contained an error on page 6. In Scheme 4 “R1” should have read “R2”. The corrected version is shown in this issue.
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