Carbonic anhydrase inhibitors. Inhibition of isozymes I, II, IV, V and IX with complex fluorides, chlorides and cyanides.

The inhibition of five human carbonic anhydrase (hCA, EC 4.2.1.1) isozymes, the cytosolic hCA I and II, the membrane-bound hCA IV, the mitochondrial hCA V and the tumour associated, transmembrane hCA IX, with complex anions incorporating fluoride, chloride and cyanide, as well as B(III), Si(IV), P(V), As(V), Al(III), Fe(II), Fe(III), Pd(II), Pt(II), Pt(IV), Cu(I), Ag(I), Au(I) and Nb(V) species has been investigated. Apparently, the most important factors influencing activity of these complexes are the nature of the central metal ion/element, and its charge. Geometry of these compounds appears to be less important, since both linear, tetrahedral, octahedral as well as pentagonal bipyramidal derivatives led to effective inhibitors. However, the five isozymes showed very different affinities for these anion inhibitors. The best hCA I inhibitors were cyanide, dicyanocuprate and dicyanoaurate (K(I)s in the range of 0.5-7.7 microM), whereas the least effective were fluoride and hexafluoroarsenate. The best hCA II inhibitors were cyanide, hexafluoroferrate and tetrachloroplatinate (K(I)s in the range of 0.02-0.51 mM), whereas the most ineffective ones were fluoride, hexafluoroaluminate and chloride. The best hCA IV inhibitors were dicyanocuprate (K(I) of 9.8 microM) and hexacyanoferrate(II) (K(I) of 10.0 microM), whereas the worst ones were tetrafluoroborate and hexafluoroaluminate (K(I)s in the range of 124-126 mM). The most effective hCA V inhibitors were cyanide, heptafluoroniobate and dicyanocuprate (K(I)s in the range of 0.015-0.79 mM), whereas the most ineffective ones were fluoride, chloride and tetrafluoroborate (K(I)s in the range of 143-241 mM). The best hCA IX inhibitors were on the other hand cyanide, heptafluoroniobate and dicyanoargentate (K(I)s in the range of 4 microM-0.33 mM), whereas the worst ones were hexacyanoferrate(III) and hexacyanoferrate(II).

Carbonic anhydrase inhibitors. Inhibition of the membrane-bound human and bovine isozymes IV with sulfonamides.

An inhibition study of the human and bovine membrane-associated isozymes of carbonic anhydrase (CA, EC 4.2.1.1), hCA IV and bCA IV, with a series of sulfonamides and sulfamates, some of which are widely clinically used, such as acetazolamide, methazolamide, ethoxzolamide, topiramate, dorzolamide, dichlorophenamide, celecoxib, and valdecoxib among others, is reported. In contrast to bCA IV, which is generally strongly inhibited by most of these derivatives, hCA IV has a rather different inhibition profile. Several of these compounds such as acetazolamide, ethoxzolamide, and bromosulfanilamide are potent hCA IV inhibitors (K(i)'s of 74-93 nM), others, such as celecoxib and some halogenated sulfanilamides are medium potency inhibitors (K(i)'s of 450-880 nM) whereas most of them are weak hCA IV inhibitors (methazolamide: 6.2 microM; dorzolamide 8.5 microM; topiramate 4.9 microM; dichlorophenamide: 15.3 microM). The hCA IV/bCA IV inhibition ratios for all the investigated compounds ranged between 1.05 (for acetazolamide) and 198.37 (for dorzolamide). Based on these results, we doubt that hCA IV is indeed one of the main contributors to the intraocular pressure (IOP) lowering effects of sulfonamide CA inhibitors, in addition to hCA II, as hypothesized earlier by Maren et al. (Mol. Pharmacol.1993, 44, 901-906). Indeed, both the very good hCA IV inhibitors (acetazolamide and ethoxzolamide) as well as the quite weak hCA IV inhibitors (methazolamide, dorzolamide, or dichlorophanamide) are effective in lowering IOP when administered either systemically or topically. The membrane-associated isozyme which probably is critical for aqueous humor secretion is hCA XII and not hCA IV.

Carbonic anhydrase inhibitors: inhibition of the membrane-bound human isozyme IV with anions.

The membrane-associated human isozyme of carbonic anhydrase, hCA IV, has been investigated for its interaction with anion inhibitors, for the CO(2) hydration reaction catalyzed by this enzyme. Surprisingly, halides were observed to act as potent hCA IV inhibitors, with inhibition constants in the range of 70-90 microM, although most of these ions, and especially fluoride, the best hCA IV inhibitor among the halides, are weak inhibitors of other isozymes, such as hCA I, II and V. The metal poisons cyanate, cyanide and hydrogen sulfide were weaker hCA IV inhibitors (K(i)'s in the range of 0.6-3.9 mM), whereas thiocyanate, azide, nitrate and nitrite showed even weaker inhibitory properties (K(i)'s in the range of 30.8-65.1 mM). Sulfate was a good hCA IV inhibitor (K(i) of 9 mM), although it is a much weaker inhibitor of isozymes I, II, V and IX. Excellent hCA IV inhibitory properties showed sulfamic acid, sulfamide, phenylboronic acid and phenylarsonic acid, with K(i)'s in the range of 0.87-0.93 microM, whereas their affinities for the other investigated isozymes were in the millimolar range. The interaction of some anions with the mitochondrial isozyme hCA V has also been investigated for the first time here. It has been observed that among all these isozymes, hCA V has the lowest affinity for bicarbonate and carbonate (K(i)'s in the range of 82-95 mM), which may represent an evolutionary adaptation of this isozyme to the rather alkaline environment (pH 8.5) within the mitochondria, where hCA V plays important functions in some biosynthetic reactions involving carboxylating enzymes (pyruvate carboxylase and acetyl coenzyme A carboxylase). There are important differences of affinity for anions between the two membrane-associated isozymes, hCA IV and hCA IX.