Recent progress in understanding the role of catalytically active carbonic anhydrases in tumors has opened new possibilities for diagnostic and/or therapeutic applications of carbonic anhydrase inhibitors selectively blocking the enzyme activity of cancer-related isoforms, namely CA IX and CA XII. Different classes of inhibitors have been investigated in order to evaluate their usefulness as in vivo imaging tools, as modulators of intratumoral pH that influences uptake of conventional chemotherapeutics, or as drugs impeding survival of tumor cells exposed to physiological stresses including hypoxia and acidosis. Here we summarize the most important data related to expression, regulation and functional aspects of cancer-related carbonic anhydrases and discuss advances in synthesis and preclinical studies of isozyme-selective and highly efficient carbonic anhydrase inhibitors.