Carbonic Anhydrase Inhibitors:  X-ray and Molecular Modeling Study for the Interaction of a Fluorescent Antitumor Sulfonamide with Isozyme II and IX

Abstract

The X-ray crystal structure of the fluorescent antitumor sulfonamide carbonic anhydrase (CA, EC, 4.2.1.1) inhibitor (4-sulfamoylphenylethyl)thioureido fluorescein (1) in complex with the cytosolic isoform hCA II is reported, together with a modeling study of the adduct of 1 with the tumor-associated isoform hCA IX. Its binding to hCA II is similar to that of other benzesulfonamides, with the ionized sulfonamide coordinated to the Zn2+ ion within the enzyme active site, and also participating in a network of hydrogen bonds with residues Thr199 and Glu106. The scaffold of 1 did not establish polar interactions within the enzyme active site but made hydrophobic contacts (<4.5 A) with Gln92, Val121, Phe131, Val135, Leu198, Thr199, Thr200, and Pro202. The substituted 3-carboxy-amino-phenyl functionality was at van der Waals distance from Phe131, Gly132, and Val135. The bulky tricyclic fluorescein moiety was located at the rim of the active site, on the protein surface, and strongly interacted with the alpha-helix formed by residues Asp130-Val135. All these interactions were preserved in the hCA IX-1 adduct, but the carbonyl moiety of the fluorescein tail of 1 participates in a strong hydrogen bond with the guanidine moiety of Arg130, an amino acid characteristic of the hCA IX active site. This may account for the roughly 2 times higher affinity of 1 for hCA IX over hCA II and may explain why in vivo the compound specifically accumulates only in hypoxic tumors overexpressing CA IX and not in the normal tissues. The compound is in clinical studies as an imaging tool for acute hypoxic tumors.

Structure-based drug discovery of carbonic anhydrase inhibitors

“…The same NH group made a hydrogen bond with the OH of Thr199 (of 2.86 Å). Furthermore, the two endocyclic nitrogen of the 1,3,4-thidiazole ring participates in two hydrogen bonds (of 2.50-2.80 Å) with the OH of Thr200, as reported earlier for a structurally related compounds [50][51][52][53][54][55][56][57][58][59] . One oxygen of the secondary SO 2 moiety of inhibitor 18 made a hydrogen bond (of 2.97 Å) with the NH 2 of Gln92.…”

“…As for other hCA II-sulphonamide adducts investigated earlier [50][51][52][53][54][55][56][57][58][59] , the deprotonated sulphonamide moiety of the inhibitor 18 was found coordinated to the Zn(II) ion at a distance of 1.96 Å 49 . The same NH group made a hydrogen bond with the OH of Thr199 (of 2.86 Å).…”

“…However, as seen from Figure 3, although the benzenesulphonamide fragments of the two compounds bind in a superposable manner to the enzyme (and in a similar fashion to most benzenesulphonamides crystallised up until now in complex with CAs [50][51][52][53] ), the orientation of the two thienylcarboxamide fragments is variable, as it also is the inhibition profile of these compounds against various CAs. For example, 20 is a weak hCA II inhibitor (K I of 390 nM), whereas 19 is much more effective one (K I of 50 nM) 60,61 .…”

Interfering with pH regulation in tumours as a therapeutic strategy

“…2), which are used for imaging purposes and for determining the role of CA9 in tumour acidification 28,39 which is a promiscuous, potent inhibitor of most carbonic anhydrase isoforms. Acetazolamide has been an important lead for generating other inhibitors with a better selectivity profile (compounds 4 and 5; FIG.…”

Carbonic anhydrase I and II inhibition with natural products: caffeine and piperine

“…These are the α-CAs (present in vertebrates, bacteria, algae and cytoplasm of green plants); the β-CAs (predominantly in bacteria, algae and chloroplasts of monodicotyledons and dicotyledons); the γ-CAs (mainly in archaea and some bacteria); and the δ-CAs and ζ-CAs (present in some marine diatoms) [1][2][3][4][5][6][7] . In mammals, 16 α-CA isozymes or CA-related proteins with different catalytic activity, subcellular localization and tissue distribution are there [8][9][10][11][12][13][14][15][16][17][18][19][20][21][22][23][24][25] . CAs catalyze a simple physiological reaction the conversion of CO 2 to the bicarbonate ion and protons.…”