Molecular docking of fucoidan (a polysaccharide compound Turbinaria conoides) with carbonic anhydrase IX—A potent target molecule in breast cancer

Santhanam Revathi Chitra; Syed Ali Mohamed Yahoob; Anuradha Venkatraman

doi:10.7324/JABB.2021.95.1s4

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Abstract

Turbinaria conoides is a type of brown seaweed containing fucoidan, a complex polysaccharide. Fucoidan is known to have antiviral, antibacterial, antifungal, and anticancer activity. The current study is focused on the extraction of fucoidan from T. conoides and the extracted fucoidan (FE) was confirmed by high performance liquid chromatography using standard fucose and the anti-scavenging property of the FE was determined by ferric reducing antioxidant power assay (FRAP) and superoxide dismutase (SOD). The structure of fucoidan was retrieved from Pubchem (CID 92023653), and it was allowed to dock against the carbonic anhydrase IX enzyme retrieved from (PDB ID 2HKF) which is up-regulated by the cancer cells. FRAP and SOD showed significant anti-scavenging property compared to the standard and molecular docking analysis using the Molegro virtual docker (MVD 5.0) of fucoidan against the carbonic anhydrase IX (CA IX) showed significant binding affinity and hydrogen bond interactions with Molegro score −66.5818 with 8 H bonds having an H-bond score of −18.8865. Fucoidan as a carbonic anhydrase IX inhibitor and consequently as a lead molecule against breast cancer can be further validated by suitable in vitro or in vivo investigation.

Keyword: Carbonic anhydrase IX docking fucoidan insilico Molegro virtual docker Turbinaria conoides

Citation:

Chitra SR, Yahoob SAM, Anuradha V. Molecular docking of fucoidan (a polysaccharide compound Turbinaria conoides) with carbonic anhydrase IX—A potent target molecule in breast cancer. J Appl Biol Biotech, 2021;9(S1):20–25.

Copyright: Author(s). This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial-ShareAlike license.

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Reference

1.Wang ZJ, Zheng L, Yang JM, Kang Y, Park YD. Proteomic analyses for profiling regulated proteins/enzymes by Fucusvesiculosus fucoidan in B16 melanoma cells: a combination of enzyme kinetics functional study. Int J Biol Macromol 2018;112:667-74. https://doi.org/10.1016/j.ijbiomac.2018.02.011
2. Deniaud-Bouët E, Hardouin K, Potin P, Kloareg B, Hervé C. A review about brown algal cell walls and fucose-containing sulfated polysaccharides: cell wall context, biomedical properties and key research challenges. Carbohydr Polym 2017;175:395-408. https://doi.org/10.1016/j.carbpol.2017.07.082
3. Sanjeewa KKA, Lee JS, Kim WS, Jeon YJ. The potential of brownalgae polysaccharides for the development of anticancer agents: an update on anticancer effects reported for fucoidan and laminaran. Carbohydr Polym 2017;177:451-9. https://doi.org/10.1016/j.carbpol.2017.09.005
4. Omar HE, Eldien HM, Badary MS, Al-Khatib BY, AbdElgaffar SK. The immunomodulating and antioxidant activity of fucoidan on the splenic tissue of rats treated with cyclosporine A. J Basic Appl Zool 2013;66:243-54. https://doi.org/10.1016/j.jobaz.2013.05.003
5. Ngo DH, Kim SK. Sulfated polysaccharides as bioactive agents from marine algae. Int J Biol Macromol 2013;62:70-5. https://doi.org/10.1016/j.ijbiomac.2013.08.036
6. Raghavendran HR, Srinivasan P, Rekha S. Immunomodulatory activity of fucoidan against aspirin-induced gastric mucosal damage in rats. Int Immunopharmacol 2011;11(2):157-63. https://doi.org/10.1016/j.intimp.2010.11.002
7. Synytsya A, Bleha R, Synytsya A, Pohl R, Hayashi K, Yoshinaga K, et al. Mekabufucoidan: structural complexity and defensive effects against avian influenza A viruses. Carbohydr Polym 2014;111:633- 44. https://doi.org/10.1016/j.carbpol.2014.05.032
8. Balboa EM, Conde E, Moure A, Falqué E, Domínguez H. In vitro antioxidant properties of crude extracts and compounds from brown algae. Food Chem 2013;138(Suppl 2-3):1764-85. https://doi.org/10.1016/j.foodchem.2012.11.026
9. Liu L, Heinrich M, Myers S, Dworjanyn SA. Towards a better understanding of medicinal uses of the brown seaweed Sargassum in traditional Chinese medicine: a phytochemical and pharmacological review. J Ethnopharmacol 2012;142:591-619. https://doi.org/10.1016/j.jep.2012.05.046
10. Chakraborty K, Praveen NK, Vijayan KK, SydaRao G. Evaluation of phenolic contents and antioxidant activities of brown seaweeds belonging to Turbinaria spp. (Phaeophyta, Sargassaceae) collected from Gulf of Mannar. Asian Pac J Trop Biomed 2013;3(1):8-16. https://doi.org/10.1016/S2221-1691(13)60016-7
11. Finkel T, Holbrook NJ. Oxidants, oxidative stress and the biology of aging. Nature 2000; 408:239-47. https://doi.org/10.1038/35041687
12. Wiseman H, Halliwell B. Damage to DNA by reactive oxygen and nitrogen species: role ininflammatory disease and progression to cancer. Biochem J 1996;313:17-29. https://doi.org/10.1042/bj3130017
13. Lee SE, Hwang HJ, Ha JS, Jeong HS, Kim JH. Screening of medicinal plant extracts for antioxidant activity. Life Sci 2003;73:167-79. https://doi.org/10.1016/S0024-3205(03)00259-5
14. Schinella GR, Tournier HA, Prieto JM, de Buschiazzo PM, Rios JL. Antioxidant activity of anti-inflammatory plant extracts. Life Sci 2002;70:1023-33. https://doi.org/10.1016/S0024-3205(01)01482-5
15. Di Matteo V, Esposito E. Biochemical and therapeutic effects of antioxidants in the treatment of Alzheimer's disease, Parkinson's disease, and amyotrophic lateral sclerosis. Curr Drug Target CNS Neurol Disord 2003;2:95-107. https://doi.org/10.2174/1568007033482959
16. Mahdi-Pour B, Jothy SL, Latha LY, Chen Y, Sasidharan S. Antioxidant activity of methanol extracts of different parts of Lantana camara. Asian Pac J Trop Biomed 2012;2(12):960-5. https://doi.org/10.1016/S2221-1691(13)60007-6
17. Reddy ARK, Grace JR. Anticancer activity of methanolic extracts of selected mangrove plants. Int J Pharm Sci Res 2007;7(9):3852-6.
18. Brahimi-Horn MC, Chiche J, Pouyssegur J. Hypoxia and cancer. J Mol Med 2007;85:1301-7. https://doi.org/10.1007/s00109-007-0281-3
19. Chaudary N, Hill RP. Hypoxia and metastasis. Clin Cancer Res 2007;13:1947-9. https://doi.org/10.1158/1078-0432.CCR-06-2971
20. Finak G, Bertos N, Pepin F, Sadekova S, Souleimanova M, Zhao H, et al. Stromal gene expression predicts clinical outcome in breastcancer. Nat Med 2008;14:518-27. https://doi.org/10.1038/nm1764
21. Kroemer G, Pouyssegur J. Tumor cell metabolism: cancer's Achilles'heel. Cancer Cell 2008;13:472-82. https://doi.org/10.1016/j.ccr.2008.05.005
22. Chaudary N, Hill RP. Hypoxia and metastasis in breast cancer. Breast Dis 2006;26:55-64. https://doi.org/10.3233/BD-2007-26105
23. Alterio V, Hilvo M, Di Fiore A, Supuran CT, Pan P, Parkkila S, et al. Crystal structure of the catalytic domain of the tumor-associated human carbonic anhydrase IX. Proc Natl Acad Sci U S A 2009;106:16233-8. https://doi.org/10.1073/pnas.0908301106
24. Supuran CT. Carbonic anhydrases: novel therapeutic applications for inhibitors and activators. Nat Rev Drug Discov 2008;7:168-81. https://doi.org/10.1038/nrd2467
25. Akurathi V, Dubois L, Lieuwes NG, Chitneni SK, Cleynhens BJ, Vullo D, et al. Synthesis and biological evaluation of a 99mTc-labelled sulfonamide conjugate for in vivo visualization of carbonic anhydrase IX expression in tumor hypoxia. Nucl Med Biol 2010;37:557-64. https://doi.org/10.1016/j.nucmedbio.2010.02.006
26. Berry M, Fielding B, Gamieldien J. Practical considerations in virtual screening and molecular docking. In: Tran QN, Arabnia HR (eds.). Emerging trends in computational biology, bioinformatics, and systems biology. National Center for Biotechnology Information, U.S. National Library of Medicine, Rockville Pike, Bethesda MD, pp 487- 502, 2015. https://doi.org/10.1016/B978-0-12-802508-6.00027-2
27. Lengauer T, Rarey M. Computational methods for biomolecular docking. Curr Opin Struct Biol 1996;6(3):402-6. https://doi.org/10.1016/S0959-440X(96)80061-3
28. Chen X, Xing R, Yu H, Liu S, Li P. A new extraction method of fucoidan from the soaked water of brown seaweed (Laminaria japonica) Desalin Water Treat 2012;40:204-8. https://doi.org/10.5004/dwt.2012.2961
29. Santhanam RC, Yahoob SAM, Venkatraman A. Antiscavenging property and characterization using HPLC of the fucoidan extracted from Turbinaria conoides. Int J Sci Technol Res 2019;8(11):2201-6.
30. Oyaizu M. Studies on product of browning reaction prepared from glucose amine. Jap J Nutr 1986;44:307315. https://doi.org/10.5264/eiyogakuzashi.44.307
31. Nishimiki M, Rao NA, Yagi K. The occurrence of super-oxide anion in the reaction of reduced phenazinemethosulfate and molecular oxygen. Biochem Biophys Res Commun1972;46:849-53. https://doi.org/10.1016/S0006-291X(72)80218-3
32. Gfeller D, Grosdidier A, Wirth M, Daina A, Michielin O, Zoete V. Swiss target prediction: a web server for target prediction of bioactive small molecules. Nucleic Acids Res 2014;42(Web Server issue):W32-8. https://doi.org/10.1093/nar/gku293
33. Daina A, Michielin O, Zoete V. Swiss target prediction: updated data and new features for efficient prediction of protein targets of small molecules. Nucleic Acids Res. 2019;47(W1):W357-64. https://doi.org/10.1093/nar/gkz382
34. Alterio V, Hilvo M, Di Fiore A, Supuran CT, Pan P, Parkkila S, et al. Crystal structure of catalytic domain of the tumor-associated human carbonic anhydrase IX. Proc Natl Acad Sci [Internet] 2009;106(38):16233-8. https://doi.org/10.1073/pnas.0908301106
35. Dahl MK, Richardson T. Photo generation of superoxide anion in serum of bovine milk and in model systems containing riboflavin and amino acids. J Dairy Sci 1978;61:400-7. https://doi.org/10.3168/jds.S0022-0302(78)83613-3
36. Bandaranayake WM. Bioactivities, bioactive compounds and chemical constituents of mangrove plants. Wetlands Ecol Manage 2002;10(6):421-52. https://doi.org/10.1023/A:1021397624349
37. Ponnan A, Ramu K, Marudhamuthu M, Marimuthu R, Siva K, Kadarkarai M. Antibacterial, antioxidant and anticancerproperties of Turbinaria conoides (J. Agardh) Kuetz clinical phytoscience. Clin Phytosci 2017;3:5. https://doi.org/10.1186/s40816-017-0042-y
38. Gopidas SK, Subramani N. In vitro antioxidant and cytotoxic properties of fucoidan from three Indian brown seaweeds, Asian J Pharm Clin Sci 2019;12(9):99-105. https://doi.org/10.22159/ajpcr.2019.v12i9.34164
39. Pastorek, J, Pastorekova S, Callebaut I, Mornon JP, Zelník V, Opavský R, et al. Cloning and characterization of MN, a human tumorassociated protein with a domain homologous to carbonic anhydrase and a putative helix-loop-helix DNA binding segment. Oncogene 1999;9:2877-88.
40. Jang M, Kim SS, Lee J. Cancer cell metabolism, implication for therapeutic targets, Exp Mol Med 2013;45:e45. https://doi.org/10.1038/emm.2013.85
41. Petrou A, Geronikaki A, Terzi E, Guler OO, Tuccinardi T, Supuran CT. Inhibition of carbonic anhydrase isoforms I II, IX and XII with secondary sulfonamides incorporating benzothiazole scaffolds. J Enzyme Inhib Med Chem 2016;31:1306-11. https://doi.org/10.3109/14756366.2015.1128427
42. Neri D, Supuran CT. Interfering with pH regulation in tumour as therapeutic strategy. Nat Rev Drug Discov 2011;10:767-77. https://doi.org/10.1038/nrd3554
43. Winum JY, Rami M, Scozzafava A, Montero JL, Supuran C. Carbonic anhydrase IX: a new druggable target for the design of antitumor agents. Med Res Rev 2008;28:445-63. https://doi.org/10.1002/med.20112
44. Winum JY, Pastorekova S, Jakubickova L, Montero JL, Scozzafava A, Pastorek J, et al. Carbonic anhydrase inhibitors synthesis and inhibiyion of cytosolic/tumour- associated carbonic anhudraseisozyme I,II, and IX with bis-sulfamates. Bioorg Med Chem Lett 2005;15:579- 84. https://doi.org/10.1016/j.bmcl.2004.11.058
45. Lomelino CL, Mahon BP, McKenna R, Carta F, Supuran CT. Kinetic and X-ray crystallographic investigations on carbonic anhydrase isoforms I, II, IX and XII of a thioureido analog of SLC-0111. Bioorg Med Chem 2016;24:976-81. https://doi.org/10.1016/j.bmc.2016.01.019
46. Pastorek J, Pastorekova S. Hypoxia-induced carbonicanhydrase IX as a target for cancer therapy: from biology to clinicaluse. Semin Cancer Biol 2015;31:52-64. https://doi.org/10.1016/j.semcancer.2014.08.002
47. Ozensoy Guler O, Supuran CT, Capasso C. Carbonic anhydrase IX as a novel candidate in liquid biopsy. J Enzyme Inhib Med Chem 2020;35:255-60. https://doi.org/10.1080/14756366.2019.1697251

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1.Wang ZJ, Zheng L, Yang JM, Kang Y, Park YD. Proteomic analyses for profiling regulated proteins/enzymes by Fucusvesiculosus fucoidan in B16 melanoma cells: a combination of enzyme kinetics functional study. Int J Biol Macromol 2018;112:667-74. https://doi.org/10.1016/j.ijbiomac.2018.02.011

2. Deniaud-Bouët E, Hardouin K, Potin P, Kloareg B, Hervé C. A review about brown algal cell walls and fucose-containing sulfated polysaccharides: cell wall context, biomedical properties and key research challenges. Carbohydr Polym 2017;175:395-408. https://doi.org/10.1016/j.carbpol.2017.07.082

3. Sanjeewa KKA, Lee JS, Kim WS, Jeon YJ. The potential of brownalgae polysaccharides for the development of anticancer agents: an update on anticancer effects reported for fucoidan and laminaran. Carbohydr Polym 2017;177:451-9. https://doi.org/10.1016/j.carbpol.2017.09.005

4. Omar HE, Eldien HM, Badary MS, Al-Khatib BY, AbdElgaffar SK. The immunomodulating and antioxidant activity of fucoidan on the splenic tissue of rats treated with cyclosporine A. J Basic Appl Zool 2013;66:243-54. https://doi.org/10.1016/j.jobaz.2013.05.003

5. Ngo DH, Kim SK. Sulfated polysaccharides as bioactive agents from marine algae. Int J Biol Macromol 2013;62:70-5. https://doi.org/10.1016/j.ijbiomac.2013.08.036

6. Raghavendran HR, Srinivasan P, Rekha S. Immunomodulatory activity of fucoidan against aspirin-induced gastric mucosal damage in rats. Int Immunopharmacol 2011;11(2):157-63. https://doi.org/10.1016/j.intimp.2010.11.002

7. Synytsya A, Bleha R, Synytsya A, Pohl R, Hayashi K, Yoshinaga K, et al. Mekabufucoidan: structural complexity and defensive effects against avian influenza A viruses. Carbohydr Polym 2014;111:633- 44. https://doi.org/10.1016/j.carbpol.2014.05.032

8. Balboa EM, Conde E, Moure A, Falqué E, Domínguez H. In vitro antioxidant properties of crude extracts and compounds from brown algae. Food Chem 2013;138(Suppl 2-3):1764-85. https://doi.org/10.1016/j.foodchem.2012.11.026

9. Liu L, Heinrich M, Myers S, Dworjanyn SA. Towards a better understanding of medicinal uses of the brown seaweed Sargassum in traditional Chinese medicine: a phytochemical and pharmacological review. J Ethnopharmacol 2012;142:591-619. https://doi.org/10.1016/j.jep.2012.05.046

10. Chakraborty K, Praveen NK, Vijayan KK, SydaRao G. Evaluation of phenolic contents and antioxidant activities of brown seaweeds belonging to Turbinaria spp. (Phaeophyta, Sargassaceae) collected from Gulf of Mannar. Asian Pac J Trop Biomed 2013;3(1):8-16. https://doi.org/10.1016/S2221-1691(13)60016-7

11. Finkel T, Holbrook NJ. Oxidants, oxidative stress and the biology of aging. Nature 2000; 408:239-47. https://doi.org/10.1038/35041687

12. Wiseman H, Halliwell B. Damage to DNA by reactive oxygen and nitrogen species: role ininflammatory disease and progression to cancer. Biochem J 1996;313:17-29. https://doi.org/10.1042/bj3130017

13. Lee SE, Hwang HJ, Ha JS, Jeong HS, Kim JH. Screening of medicinal plant extracts for antioxidant activity. Life Sci 2003;73:167-79. https://doi.org/10.1016/S0024-3205(03)00259-5

14. Schinella GR, Tournier HA, Prieto JM, de Buschiazzo PM, Rios JL. Antioxidant activity of anti-inflammatory plant extracts. Life Sci 2002;70:1023-33. https://doi.org/10.1016/S0024-3205(01)01482-5

15. Di Matteo V, Esposito E. Biochemical and therapeutic effects of antioxidants in the treatment of Alzheimer's disease, Parkinson's disease, and amyotrophic lateral sclerosis. Curr Drug Target CNS Neurol Disord 2003;2:95-107. https://doi.org/10.2174/1568007033482959

16. Mahdi-Pour B, Jothy SL, Latha LY, Chen Y, Sasidharan S. Antioxidant activity of methanol extracts of different parts of Lantana camara. Asian Pac J Trop Biomed 2012;2(12):960-5. https://doi.org/10.1016/S2221-1691(13)60007-6

17. Reddy ARK, Grace JR. Anticancer activity of methanolic extracts of selected mangrove plants. Int J Pharm Sci Res 2007;7(9):3852-6.

18. Brahimi-Horn MC, Chiche J, Pouyssegur J. Hypoxia and cancer. J Mol Med 2007;85:1301-7. https://doi.org/10.1007/s00109-007-0281-3

19. Chaudary N, Hill RP. Hypoxia and metastasis. Clin Cancer Res 2007;13:1947-9. https://doi.org/10.1158/1078-0432.CCR-06-2971

20. Finak G, Bertos N, Pepin F, Sadekova S, Souleimanova M, Zhao H, et al. Stromal gene expression predicts clinical outcome in breastcancer. Nat Med 2008;14:518-27. https://doi.org/10.1038/nm1764

21. Kroemer G, Pouyssegur J. Tumor cell metabolism: cancer's Achilles'heel. Cancer Cell 2008;13:472-82. https://doi.org/10.1016/j.ccr.2008.05.005

22. Chaudary N, Hill RP. Hypoxia and metastasis in breast cancer. Breast Dis 2006;26:55-64. https://doi.org/10.3233/BD-2007-26105

23. Alterio V, Hilvo M, Di Fiore A, Supuran CT, Pan P, Parkkila S, et al. Crystal structure of the catalytic domain of the tumor-associated human carbonic anhydrase IX. Proc Natl Acad Sci U S A 2009;106:16233-8. https://doi.org/10.1073/pnas.0908301106

24. Supuran CT. Carbonic anhydrases: novel therapeutic applications for inhibitors and activators. Nat Rev Drug Discov 2008;7:168-81. https://doi.org/10.1038/nrd2467

25. Akurathi V, Dubois L, Lieuwes NG, Chitneni SK, Cleynhens BJ, Vullo D, et al. Synthesis and biological evaluation of a 99mTc-labelled sulfonamide conjugate for in vivo visualization of carbonic anhydrase IX expression in tumor hypoxia. Nucl Med Biol 2010;37:557-64. https://doi.org/10.1016/j.nucmedbio.2010.02.006

26. Berry M, Fielding B, Gamieldien J. Practical considerations in virtual screening and molecular docking. In: Tran QN, Arabnia HR (eds.). Emerging trends in computational biology, bioinformatics, and systems biology. National Center for Biotechnology Information, U.S. National Library of Medicine, Rockville Pike, Bethesda MD, pp 487- 502, 2015. https://doi.org/10.1016/B978-0-12-802508-6.00027-2

27. Lengauer T, Rarey M. Computational methods for biomolecular docking. Curr Opin Struct Biol 1996;6(3):402-6. https://doi.org/10.1016/S0959-440X(96)80061-3

28. Chen X, Xing R, Yu H, Liu S, Li P. A new extraction method of fucoidan from the soaked water of brown seaweed (Laminaria japonica) Desalin Water Treat 2012;40:204-8. https://doi.org/10.5004/dwt.2012.2961

29. Santhanam RC, Yahoob SAM, Venkatraman A. Antiscavenging property and characterization using HPLC of the fucoidan extracted from Turbinaria conoides. Int J Sci Technol Res 2019;8(11):2201-6.

30. Oyaizu M. Studies on product of browning reaction prepared from glucose amine. Jap J Nutr 1986;44:307315. https://doi.org/10.5264/eiyogakuzashi.44.307

31. Nishimiki M, Rao NA, Yagi K. The occurrence of super-oxide anion in the reaction of reduced phenazinemethosulfate and molecular oxygen. Biochem Biophys Res Commun1972;46:849-53. https://doi.org/10.1016/S0006-291X(72)80218-3

32. Gfeller D, Grosdidier A, Wirth M, Daina A, Michielin O, Zoete V. Swiss target prediction: a web server for target prediction of bioactive small molecules. Nucleic Acids Res 2014;42(Web Server issue):W32-8. https://doi.org/10.1093/nar/gku293

33. Daina A, Michielin O, Zoete V. Swiss target prediction: updated data and new features for efficient prediction of protein targets of small molecules. Nucleic Acids Res. 2019;47(W1):W357-64. https://doi.org/10.1093/nar/gkz382

34. Alterio V, Hilvo M, Di Fiore A, Supuran CT, Pan P, Parkkila S, et al. Crystal structure of catalytic domain of the tumor-associated human carbonic anhydrase IX. Proc Natl Acad Sci [Internet] 2009;106(38):16233-8. https://doi.org/10.1073/pnas.0908301106

35. Dahl MK, Richardson T. Photo generation of superoxide anion in serum of bovine milk and in model systems containing riboflavin and amino acids. J Dairy Sci 1978;61:400-7. https://doi.org/10.3168/jds.S0022-0302(78)83613-3

36. Bandaranayake WM. Bioactivities, bioactive compounds and chemical constituents of mangrove plants. Wetlands Ecol Manage 2002;10(6):421-52. https://doi.org/10.1023/A:1021397624349

37. Ponnan A, Ramu K, Marudhamuthu M, Marimuthu R, Siva K, Kadarkarai M. Antibacterial, antioxidant and anticancerproperties of Turbinaria conoides (J. Agardh) Kuetz clinical phytoscience. Clin Phytosci 2017;3:5. https://doi.org/10.1186/s40816-017-0042-y

38. Gopidas SK, Subramani N. In vitro antioxidant and cytotoxic properties of fucoidan from three Indian brown seaweeds, Asian J Pharm Clin Sci 2019;12(9):99-105. https://doi.org/10.22159/ajpcr.2019.v12i9.34164

39. Pastorek, J, Pastorekova S, Callebaut I, Mornon JP, Zelník V, Opavský R, et al. Cloning and characterization of MN, a human tumorassociated protein with a domain homologous to carbonic anhydrase and a putative helix-loop-helix DNA binding segment. Oncogene 1999;9:2877-88.

40. Jang M, Kim SS, Lee J. Cancer cell metabolism, implication for therapeutic targets, Exp Mol Med 2013;45:e45. https://doi.org/10.1038/emm.2013.85

41. Petrou A, Geronikaki A, Terzi E, Guler OO, Tuccinardi T, Supuran CT. Inhibition of carbonic anhydrase isoforms I II, IX and XII with secondary sulfonamides incorporating benzothiazole scaffolds. J Enzyme Inhib Med Chem 2016;31:1306-11. https://doi.org/10.3109/14756366.2015.1128427

42. Neri D, Supuran CT. Interfering with pH regulation in tumour as therapeutic strategy. Nat Rev Drug Discov 2011;10:767-77. https://doi.org/10.1038/nrd3554

43. Winum JY, Rami M, Scozzafava A, Montero JL, Supuran C. Carbonic anhydrase IX: a new druggable target for the design of antitumor agents. Med Res Rev 2008;28:445-63. https://doi.org/10.1002/med.20112

44. Winum JY, Pastorekova S, Jakubickova L, Montero JL, Scozzafava A, Pastorek J, et al. Carbonic anhydrase inhibitors synthesis and inhibiyion of cytosolic/tumour- associated carbonic anhudraseisozyme I,II, and IX with bis-sulfamates. Bioorg Med Chem Lett 2005;15:579- 84. https://doi.org/10.1016/j.bmcl.2004.11.058

45. Lomelino CL, Mahon BP, McKenna R, Carta F, Supuran CT. Kinetic and X-ray crystallographic investigations on carbonic anhydrase isoforms I, II, IX and XII of a thioureido analog of SLC-0111. Bioorg Med Chem 2016;24:976-81. https://doi.org/10.1016/j.bmc.2016.01.019

46. Pastorek J, Pastorekova S. Hypoxia-induced carbonicanhydrase IX as a target for cancer therapy: from biology to clinicaluse. Semin Cancer Biol 2015;31:52-64. https://doi.org/10.1016/j.semcancer.2014.08.002

47. Ozensoy Guler O, Supuran CT, Capasso C. Carbonic anhydrase IX as a novel candidate in liquid biopsy. J Enzyme Inhib Med Chem 2020;35:255-60. https://doi.org/10.1080/14756366.2019.1697251