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Journal of Enzyme Inhibition and Medicinal Chemistry Volume 31, 2016 - Issue sup2
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Research Article

Design, synthesis and biological evaluation of N-(5-methyl-isoxazol-3-yl/1,3,4-thiadiazol-2-yl)-4-(3-substitutedphenylureido) benzenesulfonamides as human carbonic anhydrase isoenzymes I, II, VII and XII inhibitors

Chandra Bhushan MishraBio-Organic Chemistry Laboratory, Dr. B. R. Ambedkar Centre for Biomedical Research, University of Delhi, Delhi, India,
,
Shikha KumariBio-Organic Chemistry Laboratory, Dr. B. R. Ambedkar Centre for Biomedical Research, University of Delhi, Delhi, India,
,
Andrea AngeliDipartimento Neurofarba, Università degli Studi di Firenze, Sezione di Scienze Farmaceutiche e Nutraceutiche, Florence, Italy,
,
Simona Maria MontiIstituto di Biostrutture e Bioimmagini (IBB) CNR, via Mezzocannone, Naples, Italy, and
,
Martina BuonannoIstituto di Biostrutture e Bioimmagini (IBB) CNR, via Mezzocannone, Naples, Italy, and
,
Amresh PrakashSchool of Computational and Integrative Sciences, Jawaharlal Nehru University, New Delhi, India
,
Manisha TiwariBio-Organic Chemistry Laboratory, Dr. B. R. Ambedkar Centre for Biomedical Research, University of Delhi, Delhi, India, Correspondence[email protected]
&
Claudiu T. SupuranDipartimento Neurofarba, Università degli Studi di Firenze, Sezione di Scienze Farmaceutiche e Nutraceutiche, Florence, Italy, Correspondence[email protected]
 show all
Pages 174-179 | Received 16 May 2016, Accepted 31 May 2016, Published online: 17 Jun 2016

Abstract

A series of N-(5-methyl-isoxazol-3-yl/1,3,4-thiadiazol-2-yl)-4-(3-substitutedphenylureido) benzenesulfonamide derivatives has been designed, synthesized and screened for their in vitro human carbonic anhydrase (hCA; EC 4.2.1.1) inhibition potential. These newly synthesized sulfonamide compounds were assessed against isoforms hCA I, II, VII and XII, with acetazolamide (AAZ) as a reference compound. The majority of these compounds were found quite weak inhibitor against all tested isoforms. Compound 15 showed a modest inhibition potency against hCA I (Ki = 73.7 μM) and hCA VII (Ki = 85.8 μM). Compounds 19 and 25 exhibited hCA II inhibition with Ki values of 96.0 μM and 87.8 μM, respectively. The results of the present study suggest that, although the synthesized derivatives have weak inhibitory potential towards all investigated isoforms, some of them may serve as lead molecules for the further development of selective inhibitors incorporating secondary sulfonamide functionalities, a class of inhibitors for which the inhibition mechanism is poorly understood.

Introduction

The carbonic anhydrases (CAs; EC 4.2.1.1) are a large group of ubiquitous zinc containing enzymes, present in all mammals, including humanCitation1. CAs are encoded by six distinct gene families known as alpha (α), beta (β), gamma (γ), delta (δ) zeta (ζ) and eta (η) and all human CAs belongs to class alphaCitation2–4. These enzymes are well described by their different molecular features, oligomeric arrangement as well as kinetic propertiesCitation5. Till date, a total number of 15 different human carbonic anhydrases (hCA) isoforms have been discovered, among which CA I-III, CA VII and CA XIII are present in cytosol; CA IV, CA IX, CA XII and CA XIV are membrane bound; CA VA and CA VB are restricted to the mitochondrion and CA VI is secreted in milk and salivaCitation6. Functionally, CAs are primarily involved in catalyzing the rapid interconversion of CO2 and H2O to bicarbonate (HCO3-) and protons (H+) using a metal hydroxide nucleophilic mechanismCitation7. The hCAs play a pivotal role in a variety of physiological processes such as CO2/bicarbonate transportation, respiration, electrolyte secretion, gluoconeogensis, lipogenesis, ureagenesis, bone resorption, neuronal excitability and tumorigenicityCitation8. CAs have profound involvement in many pathological conditions like obesity, glaucoma, kidney dysfunction, osteoporosis, gastric ulcers, migraine, epilepsy and cancerCitation9. For example CA II deficiency is the major defect in disease conditions such as osteopetrosis, cerebral calcification and renal tubular acidosisCitation10. Another isoform, CA VII is implicated in generating neuronal excitation leading to the development of seizures as well as in the neuropathic pain controlCitation11. hCA II, IV and XII are druggable targets for anti-glaucoma agents while, hCA VII and hCA XIV are known drug targets for antiepileptic drugs. hCA IX and XII are anti-tumor drug targets.At present, CAs are attractive therapeutic targets to control these aforementioned diseases. The CAs are also essential for other living organism including pathogens.

Acetazolamide (AAZ) is considered as an excellent CA inhibitor (CAI) and is used for the treatment of glaucoma, epilepsy and altitude sicknessCitation12. Several other CAIs such as ethoxolamide (EZA), brinzolamide (BRZ), dorzolamide (DZA), methazolamide (MZA), and dichlorophenamide (DCP) are useful clinically as anti-glaucoma agents but, they are also used to treat various neurological as well as neuromuscular disorders such as epilepsy, genetic hemiplegic migraine and ataxia, tardive diskinesia, hypokalemic periodic paralysis, essential tremor and Parkinsons diseaseCitation13. Unfortunately, the presently used CAIs are non-selective to a particular CA isoform, which leads to the occurrence of peripheral side effects. Therefore, the development of novel CAIs having high potency and selectivity against specific isoform still represents a needful and challenging task to obtain newer compounds which could replace the “old ones”, in consequence avoiding side effects as well as improving therapeutic safety.

Sulfonamides containing molecules have presented a remarkable history in CA inhibition. The primary sulfonamide group (-SO2NH2) is present in these molecules which bind as anions to the Zn2+ ion in the enzyme active site with high affinity, and block catalysisCitation14. In the past, several research reports showed that the urea functional group along with sulfonamide also seemed to be beneficial to generate effective CA inhibitory activityCitation15. In the present study, we have used sulfamethoxazole/sulfamethiazole to develop potent CAIs and also phenyl urea moiety was also installed to this pharmacophore to develop new CAIs. showed some known CAIs and the molecular framework of our synthesized compound. The synthesized derivatives have been well characterized by using 1H NMR, 13C NMR and mass spectroscopy. The purity was analyzed by HPLC. The esterase activity was performed to study the potential of the newly synthesized compounds (15–26) towards hCA I, hCA II, hCA VII and hCA XII inhibition.

Figure 1. Sulfonamides CA inhibitors (acetazolamide, methazolamide and SLC-0111), sulfadrugs (sulfadiazine and sulfapyridine) and the designed novel compounds 15–26.

Figure 1. Sulfonamides CA inhibitors (acetazolamide, methazolamide and SLC-0111), sulfadrugs (sulfadiazine and sulfapyridine) and the designed novel compounds 15–26.

Materials and methods

Chemistry

All the chemicals and reagents were obtained from Sigma Aldrich (St. Louis, MO), Alfa Aesar (Massachusetts), S.D Fine Chemicals (India) and Merck (Darmstadt, Germany) and solvents for reaction medium were dried by standard methods. Analytical thin layer chromatography (TLC) was performed on commercially available silica gel (Kieselgel 60, F254) coated aluminum plates (Merck). Column chromatography purification was performed using silica gel Merck 100–200 mesh. Melting points were taken in open capillaries using model KSPII, KRUSS, (Germany). 1H NMR and 13C NMR were recorded on Jeol-400 MHz High Resolution NMR Spectrophotometer (JEOL USA, Inc., Peabody, MA). Chemical shifts in 1H NMR are reported in parts per million and coupling constant (J) in Hertz (Hz) using DMSO-d6 as solvent. The splitting patterns are designated as follows: s, singlet; d, doublet; t, triplet; q, quadruplet. Mass spectra were recorded on an Agilent 6310 Ion trap LC/MS and the purity of final compounds was analyzed by using reverse phase HPLC (Shimadzu, Kyoto, Japan) with C-18 column.

General procedure for the synthesis of novel N-(5-methyl-isoxazol-3-yl/1,3,4-thiadiazol-2-yl)-4–(3-substitutedphenylureido) benzenesulfonamides (15–26)

Sulfamethoxazole/sulfamethiazole (10 mmol) was fully dissolved in dried DMF and then subsequent isocyanate (10 mmol) was added dropwise to the reaction mixture and heated at 90–100 °C for 5–6 h. The appeared precipitate was filtered and washed with hot petroleum ether as well as water. The obtained crude products were purified by column chromatography using chloroform/methanol (98:02) as an eluent to furnish target compounds (15–26).The purity of the synthesized compounds was analyzed by HPLC using acetonitrile/methanol (98:02) as mobile phase ().

Scheme 1. Synthesis of N-(5-methyl-isoxazol-3-yl/1,3,4-thiadiazol-2-yl)-4-(3-substituted phenylureido)benzenesulfonamide derivatives. Reagent and conditions: A. Dried DMF reflux, 5–6 h.

Scheme 1. Synthesis of N-(5-methyl-isoxazol-3-yl/1,3,4-thiadiazol-2-yl)-4-(3-substituted phenylureido)benzenesulfonamide derivatives. Reagent and conditions: A. Dried DMF reflux, 5–6 h.

N-(5-Methyl-isoxazol-3-yl)-4–(3-phenyl-ureido)-benzenesulfonamide (15)

White solid; yield 1.74 g; mp 240–242 °C; 1H NMR (DMSO-d6,400 MHz): δ 2.28 (s, 3H, CH3), 6.13 (s, 1H, CH), 6.98 (t, 1H, Ar-H, J = 7.2 Hz), 7.27 (t, 2H, Ar-H, J = 7.6 Hz), 7.44 (d, 2H, Ar-H, J = 7.6 Hz), 7.63 (d, 2H, Ar-H, J = 9.1 Hz), 7.75 (d, 2H, Ar-H, J = 7.7 Hz), 8.82 (s, 1H, NH), 9.15 (s, 1H, NH), 11.29 (s, 1H, NH). 13C NMR (DMSO-d6): 12.08, 95.3, 117.7, 118.4, 122.3, 128.1, 128.8, 131.4, 139.1, 144.3, 152.1, 157.6, 170.2. LC–MS: m/e; 372 (M+). HPLC purity: 97.6%.

4-[3–(4-Fluoro-phenyl)-ureido]-N-(5-methyl-isoxazol-3-yl)-benzenesulfonamide (16)

White solid; yield 1.92 g; mp 228–230 °C; 1H NMR (DMSO-d6,400 MHz): δ 2.28 (s, 3H, CH3), 6.12 (s, 1H, CH), 7.12 (t, 2H, Ar-H, J = 8.7 Hz), 7.43–7.47 (m, 2H, Ar-H), 7.61 (d, 2H, Ar-H, J = 9.1 Hz), 7.76 (d, 2H, Ar-H, J = 8.4 Hz), 8.85 (s, 1H, NH), 9.10 (s, 1H, NH), 11.28 (s, 1H, NH). 13C NMR (DMSO-d6): 12.07, 95.3, 115.2, 115.5, 117.7, 120.2, 120.3, 128.1, 131.5, 135.4, 144.3, 152.2, 156.4, 157.6, 158.7, 170.2. LC–MS: m/e; 391 (M + 1). HPLC purity: 97.1%.

4-[3–(4-Chloro-phenyl)-ureido]-N-(5-methyl-isoxazol-3-yl)-benzenesulfonamide (17)

White solid; yield 2.35 g; mp 238–240 °C; 1H NMR (DMSO-d6,400 MHz): δ 2.28 (s, 3H, CH3), 6.12 (s, 1H, CH), 7.33 (d, 2H, Ar-H, J = 8.4 Hz), 7.47 (d, 2H, Ar-H, J= 8.4 Hz), 7.62 (d, 2H, Ar-H, J = 9.1 Hz), 7.75 (d, 2H, Ar-H, J = 8.4 Hz), 8.97 (s, 1H, NH), 9.20 (s, 1H, NH), 11.28 (s, 1H, NH). 13C NMR (DMSO-d6): 12.09, 95.4, 117.8, 120.0, 125.9, 128.1, 131.6, 138.2, 144.1, 152.1, 157.6, 170.2. LC–MS: m/e; 406 (M+). HPLC purity: 97.9%.

N-(5-Methyl-isoxazol-3-yl)-4-(3-p-tolyl-ureido)-benzenesulfonamide (18)

White solid; yield 1.49 g; mp 233–235 °C; 1H NMR (DMSO-d6,400 MHz): δ 2.23 (s, 3H, CH3), 2.28 (s, 3H, CH3), 6.12 (s, 1H, CH), 7.08 (d, 2H, Ar-H, J = 7.6 Hz), 7.32 (d, 2H, Ar-H, J = 8.4 Hz), 7.61 (d, 2H, Ar-H, J = 9.1 Hz), 7.74 (d, 2H, Ar-H, J = 8.4 Hz), 8.71 (s, 1H, NH), 9.11 (s, 1H, NH), 11.20 (s, 1H, NH). 13C NMR (DMSO-d6): 12.0, 20.3, 95.3, 117.3, 118.5, 128.1, 129.2, 131.2, 131.3, 136.6, 144.4, 152.1, 157.6, 170.2. LC–MS: m/e; 386 (M+). HPLC purity: 99.4%.

4-[3–(4-Methoxy-phenyl)-ureido]-N-(5-methyl-isoxazol-3-yl)-benzenesulfonamide (19)

White solid; yield 1.84 g; mp 212–214 °C; 1H NMR (DMSO-d6,400 MHz): δ 2.28 (s, 3H, CH3), 3.70 (s, 3H, CH3), 6.12 (s, 1H, CH), 6.86 (d, 2H, Ar-H, J = 8.4 Hz), 7.34 (d, 2H, Ar-H, J = 8.4 Hz), 7.61 (d, 2H, Ar-H, J = 8.4 Hz), 7.73 (d, 2H, Ar-H, J = 9.1 Hz), 8.63 (s, 1H, NH), 9.08 (s, 1H, NH), 11.21 (s, 1H, NH). 13C NMR (DMSO-d6): 12.0, 55.1, 95.3, 114.0, 117.5, 120.3, 128.1, 131.2, 132.1, 144.5, 152.3, 154.8, 157.6, 170.2. LC–MS: m/e; 402 (M+). HPLC purity: 99.7%.

4-(3-Benzyl-ureido)-N-(5-methyl-isoxazol-3-yl)-benzenesulfonamide (20)

White solid; yield 2.12 g; mp 218–220 °C; 1H NMR (DMSO-d6,400 MHz): δ 2.27 (s, 3H, CH3), 4.29 (d, 2H, CH2, J = 6.1 Hz), 6.10 (s, 1H, CH), 6.82 (t, 1H, NH, J = 5.7 Hz), 7.21–7.33 (m, 5H, Ar-H), 7.57 (d, 2H, Ar-H, J = 9.1 Hz), 7.69 (d, 2H, Ar-H, J = 8.4 Hz), 9.08 (s, 1H, NH), 11.22 (s, 1H, NH). 13C NMR (DMSO-d6): 12.0, 42.7, 95.3, 117.1, 126.8, 127.1, 128.1, 128.3, 130.7, 139.9, 145.0, 154.7, 157.7, 170.2. LC–MS: m/e; 387 (M + 1). HPLC purity: 99.4%.

N-(5-methyl-[1,3,4]thiadiazol-2-yl)-4-(3-phenyl-ureido)-benzenesulfonamide (21)

White solid; yield 2.16 g; mp 226–228 °C; 1H NMR (DMSO-d6,400 MHz): δ 2.45 (s, 3H, CH3), 6.98 (t, 1H, Ar-H, J = 7.6 Hz), 7.28 (t, 2H, Ar-H, J = 7.9 Hz), 7.44 (d, 2H, Ar-H, J = 8.4 Hz), 7.58 (d, 2H, Ar-H, J = 9.1 Hz), 7.68 (d, 2H, Ar-H, J = 9.1 Hz), 8.77 (s, 1H, NH), 9.09 (s, 1H, NH), 13.81 (s, 1H, NH). 13C NMR (DMSO-d6): 16.1, 117.6, 118.4, 122.2, 127.1, 128.8, 134.3, 139.2, 143.5, 152.2, 154.3, 167.7. LC–MS: m/e; 390 (M + 1). HPLC purity: 96.8%.

4-[3-(4-Fluoro-phenyl)-ureido]-N-(5-methyl-[1,3,4]thiadiazol-2-yl)-benzene sulfonamide (22)

White solid; yield 1.64 g; mp 252–254 °C; 1H NMR (DMSO-d6,400 MHz): δ 2.45 (s, 3H, CH3), 7.10–7.14 (m, 2H, Ar-H), 7.43–7.47 (m, 2H, Ar-H), 7.58 (d, 2H, Ar-H, J = 8.3 Hz), 7.68 (d, 2H, Ar-H, J = 8.4 Hz), 8.81 (s, 1H, NH), 9.10 (s, 1H, NH), 13.82 (s, 1H, NH). 13C NMR (DMSO-d6): 16.1, 115.2, 115.5, 117.6, 120.2, 120.3, 127.0, 134.4, 135.6, 143.5, 152.3, 154.4, 156.4, 158.8, 167.7. LC–MS: m/e; 408 (M + 1). HPLC purity: 99.5%.

4-[3–(4-Chloro-phenyl)-ureido]-N-(5-methyl-[1,3,4]thiadiazol-2-yl)-benzene sulfonamide (23)

White solid; yield 2.0 g; mp 275–277 °C; 1H NMR (DMSO-d6,400 MHz): δ 2.44 (s, 3H, CH3), 7.32 (d, 2H, Ar-H, J = 9.1 Hz), 7.48 (d, 2H, Ar-H, J = 9.1 Hz), 7.59 (d, 2H, Ar-H, J = 8.4 Hz), 7.69 (d, 2H, Ar-H, J = 9.1 Hz), 8.93 (s, 1H, NH), 9.14 (s, 1H, NH), 13.8 (s, 1H, NH).13C NMR (DMSO-d6): 16.1, 117.7, 120.0, 125.8, 127.1, 128.7, 134.5, 138.3, 143.3, 152.1, 154.4, 167.7. LC–MS: m/e; 423 (M+). HPLC purity: 98.6%.

N-(5-methyl-[1,3,4]thiadiazol-2-yl)-4-(3-p-tolyl-ureido)-benzenesulfonamide (24)

White solid; yield 1.75 g; mp 258–260 °C; 1H NMR (DMSO-d6,400 MHz): δ 2.23 (s, 3H, CH3), 2.45 (s, 3H, CH3), 7.08 (d, 2H, Ar-H, J = 8.4 Hz), 7.32 (d, 2H, Ar-H, J = 8.4 Hz), 7.57 (d, 2H, Ar-H, J = 9.9 Hz), 7.67 (d, 2H, Ar-H, J = 9.9 Hz), 8.66 (s, 1H, NH), 9.05 (s, 1H, NH), 13.8 (s, 1H, NH).13C NMR (DMSO-d6): 16.09, 20.3, 117.5, 118.5, 127.0, 129.2, 131.1, 134.1, 136.6, 143.5, 152.2, 154.3, 167.7. LC–MS: m/e; 403 (M+). HPLC purity: 96.7%.

4-[3-(4-Methoxy-phenyl)-ureido]-N-(5-methyl-[1,3,4]thiadiazol-2-yl)-benzenesulfonamide (25)

White solid; yield 2.0 g; mp 238–240 °C; 1H NMR (DMSO-d6,400 MHz): δ 2.48 (s, 3H, CH3), 3.70 (s, 3H, CH3), 6.86 (d, 2H, Ar-H, J = 9.1 Hz), 7.34 (d, 2H, Ar-H, J = 9.1 Hz), 7.57 (d, 2H, Ar-H, J = 8.4 Hz), 7.65 (d, 2H, Ar-H, J = 8.4 Hz), 8.58 (s, 1H, NH), 9.02 (s, 1H, NH), 13.8 (s, 1H, NH).13C NMR (DMSO-d6): 16.0, 55.1, 114.0, 117.4, 120.2, 127.0, 132.2, 134.0, 143.6, 152.3, 154.7, 167.7. LC–MS: m/e; 419 (M+). HPLC purity: 99.3%.

4-(3-Benzyl-ureido)-N-(5-methyl-[1,3,4]thiadiazol-2-yl)-benzenesulfonamide (26)

White solid; yield 2.2 g; mp 270–272 °C; 1H NMR (DMSO-d6,400 MHz): δ 2.44 (s, 3H, CH3), 4.28 (q, 2H, CH2, J = 6.0 Hz), 6.76 (t, 1H, NH, J = 6.1 Hz), 7.20–7.33 (m, 5H, Ar-H), 7.53 (d, 2H, Ar-H, J = 9.1 Hz), 7.62 (d, 2H, Ar-H, J = 8.4 Hz), 9.02 (s, 1H, NH), 13.8 (s, 1H, NH).13C NMR (DMSO-d6): 16.0, 42.7, 117.0, 126.8, 127.0, 127.1, 128.3, 133.5, 140.0, 144.2, 154.2, 154.8, 167.6. LC–MS: m/e; 404 (M + 1). HPLC purity: 99.3%.

Carbonic anhydrase inhibition assay

An applied photophysics stopped-flow instrument has been used for assaying the CA catalyzed CO2 hydration activityCitation16. Phenol red (at a concentration of 0.2 mM) has been used as indicator, working at the absorbance maximum of 557 nm, with 20 mM Hepes (pH 7.4) as buffer, and 20 mM Na2SO4 for maintaining constant the ionic strength (this anion is not inhibitory and has a KI> 200 mM against these enzymes), following the initial rates of the CA-catalyzed CO2 hydration reaction for a period of 10–100 s. The CO2 concentrations ranged from 1.7 to 17 mM for the determination of the kinetic parameters and inhibition constants. For each measurement at least six traces of the initial 5–10% of the reaction have been used for determining the initial velocity, working with 10-fold decreasing inhibitor concentrations ranging between 0.1 nM and 10–100 μM (depending on the inhibitor potency, but at least five points at different inhibitor concentrations were employed for determining the inhibition constants). The uncatalyzed rates were determined in the same manner and subtracted from the total observed rates. Stock solutions of inhibitor (0.1 mM) were prepared in distilled-deionized water and dilutions up to 0.1 nM were done thereafter with the assay buffer. Inhibitor and enzyme solutions were preincubated together for 15 min at room temperature prior to assay, in order to allow for the formation of the E-I complex. The inhibition constants were obtained by non-linear least-squares methods using the Cheng–Prusoff equation, and represent the mean from at least three different determinations. The human isoforms hCA I, II, VII and XII were recombinant enzymes produced as described earlier in our laboratoryCitation17–32.

Result and discussion

Chemistry

The synthetic route utilized for the preparation of N-(5-methyl-isoxazol-3-yl/1,3,4-thiadiazol-2-yl)-4–(3-substitutedphenylureido) benzenesulfonamide derivativesis depicted in .

An equimolar ratio of sulfamethoxazole/sulfamethiazole and substituted phenyl isocyantes were reacted in dry DMF to afford N-(5-methyl-isoxazol-3-yl/1,3,4-thiadiazol-2-yl)-4–(3-substituted phenylureido)benzenesulfonamide derivatives. All compounds were purified by using column chromatography using chloroform/methanol as eluent. The purity of the final compounds was analyzed by HPLC analysis using acetonitrile/methanol (98:02) as mobile phase. All compounds presented percent purity more than 95% and were fully characterized by NMR and mass spectroscopy.

We have used SLC-0111 as lead molecule for designing these compoundsCitation33,Citation34, as this derivative recently completed successfully the Phase I clinical trials as an anti-tumor agent. In addition, a range of secondary and tertiary sulfonamides has been investigated as CAIs recently, some of them showing effective and selective inhibition of several important isoformsCitation35–49. However, the inhibition mechanism with secondary and tertiary sulfonamides is unknown, as no X-ray crystal structures of such derivatives bound to the CA are available to date. Thus, the sulfonamides reported here incorporate in their molecule the urea fragment found in SLC-0111 and the secondary sulfonamide moiety present in sulfa drugs and several recently investigated CAIsCitation35–49.

In vitro carbonic anhydrase activity

All the synthesized compounds (15–26) were studied for the inhibition of four CA isozymes of human origin, i.e. hCA I, hCA II, hCA VII and hCA XII (). The following structure activity relationship (SAR) was obtained by analyzing CA inhibition data of :

  1. In the isoxazole subseries (15–20) compound substituted phenyl ring (compound 15) at the terminal end has mild hCA inhibitory activity for both hCA I (Ki = 73.7 μM)and hCA VII (Ki = 85.8 μM) isoforms.

  2. In this subseries, compound substituted with para-methoxy phenyl (compound 19) also exhibited low inhibition against hCA II (Ki = 96.0 μM). Moreover, this compound displayed a Ki value of >100 μM towards hCA I, VII and XII, thus seems to inactive for these isoforms. The remaining compounds in this subseries did not produce remarkable inhibitory potential against hCA I, II, VII and XII isoforms.

  3. In thiadiazole subseries (21–26), compound 25 comprising para-methoxy phenyl ring at its terminal end has shown a modest inhibitory potential against hCA II with Ki value of 87.8 μM. This compound was inactive against hCA I, VII and XII (Ki= >100 μM).

  4. Overall, the SAR study indicates that substitutions on the phenyl ring did not produce remarkable inhibitory potential against tested hCA isoforms except para-methoxy substitution. Therefore, further structural modifications are necessarily required to achieve more potent CAIs.

Table 1. hCA I, II, VII and XII inhibition with compounds 15–26, with AAZ as standard.

Table

Conclusions

A set of N-(5-methyl-isoxazol-3-yl/1,3,4-thiadiazol-2-yl)-4–(3-substitutedphenylureido) benzenesulfonamide derivatives were synthesized in a good yield and were characterized by using NMR and mass spectroscopy techniques. All compounds were subjected for their in-vitro CA inhibitory activity and only compound 15 showed CA inhibition against hCA I and hCA VII isoform. The selectivity ratio of compounds 19 and 25 for hCA II isoform makes them interesting leads for the development of potent and selective hCA II inhibitor.

Declaration of interest

Chandra Bhushan Mishra (DSK-PDF) and Shikha Kumari are thankful to the University Grant Commission for providing financial support. Author Amresh Prakash is thankful to SERB-YSS for financial support. Manisha Tiwari deeply acknowledges the financial support to carry out this research work. Work from Supuran laboratory was financed by two EU grants (Dynano and Metoxia).

Acknowledgements

The University Science Instrumentation Center (USIC), University of Delhi is acknowledged for providing NMR and Mass spectral characterization of the synthesized compounds.

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