Novel isocoumarin–pyrazole hybrids: synthesis, characterization, and investigation of their carbonic anhydrase inhibitory activities

Aim Selective inhibition of tumor-associated carbonic anhydrase (CA) isoforms IX and XII is a key challenge in anticancer drug discovery, as most inhibitors cause off-target inhibition of cytosolic enzymes. We addressed this by designing and synthesizing a series of isocoumarin – pyrazole hybrids from isocoumarin – chalcone scaffolds and evaluating their potential as selective CA IX/XII inhibitors.Materials and methods Hybrids (5a–5o) were prepared via cyclization with substituted phenylhydrazines under reflux. Structures were confirmed by FT-IR and NMR analyses. Inhibition of hCA I, II, IX, and XII was determined using a stopped-flow CO2 hydration assay following enzyme – inhibitor preincubation.Results All compounds were obtained in good yields. No significant activity was observed against cytosolic hCA I and II, whereas all derivatives inhibited tumor-associated hCA IX and XII at low-to-medium micromolar levels. Electron-donating substituents improved potency, with compounds 5a and 5g being the most active.Conclusion This study shows that isocoumarin – pyrazole hybrids achieve selective inhibition of hCA IX and XII without affecting off-target isoforms. The results expand the chemical space of non-classical CA inhibitors and offer insights for the rational design of selective anticancer agents. Future work will focus on optimizing potency and evaluating efficacy in cellular tumor models.