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Membrane-associated carbonic anhydrase CA isoform IV participates in carbon metabolism and pH homeostasis and is implicated in the development of eye diseases such as retinitis pigmentosa and glaucoma. A series of substituted benzenesulfonamides were designed and their binding affinity to CA IV was determined by fluorescent thermal shift assay and isothermal titration calorimetry ITC A series of substituted benzenesulfonamides were designed and their binding affinity to CA IV was determined by fluorescent thermal shift assay and isothermal titration calorimetry ITC. Compound [ 4-chlorophenylsulfanylsulfamoyl-benzoyl amino]propyl acetate 19 bound CA IV with the K d of 1. The compound could be developed as a drug targeting CA IV.