Coumarin carbonic anhydrase inhibitors from natural sources

Abstract

Coumarins constitute a relatively new class of inhibitors of the zinc enzyme carbonic anhydrase (CA, EC 4.2.1.1), possessing a unique inhibition mechanism, acting as "prodrug inhibitors." They undergo the hydrolysis of the lactone ring mediated by the esterase activity of CA. The formed 2-hydroxy-cinnamic acids thereafter bind within a very particular part of the enzyme active site, at its entrance, where a high variability of amino acid residues among the different mammalian CA isoforms is present, and where other inhibitors classes were not seen bound earlier. This explains why coumarins are among the most isoform-selective CA inhibitors known to date among the many chemotypes endowed with such biological activity. As coumarins are widespread secondary metabolites in some bacteria, plants, fungi, and ascidians, many such compounds from various natural sources have been investigated for their CA inhibitory properties and for possible biomedical applications, mainly as anticancer agents targeting hypoxic tumours.

Keywords: Carbonic anhydrase; coumarin; natural product; prodrug inhibitor.

Figure 1.

The simplest coumarin, compound 1, its hydrolysis product 2, and the natural product coumarin 3 (and its hydrolysis product 4) for which the CA inhibitory activity was first reported.

Figure 2.

CA inhibition mechanisms (A–D) and the CA activation mechanism (E). The CA modulators of activity incorporate various scaffolds and tails in their molecule, as well as characteristic functionalities for all category: (A) the zinc binders possess a zinc-binding group (ZBG) which is coordinated to the metal ion; (B) the compounds which anchor to the zinc-coordinated water incorporate an anchoring group (AG) which hydrogen bonds with the water coordinated to the metal and the OH of Thr199; (C) AGs are also present in compounds which occlude the entrance of the active site cavity; (D) some inhibitors which bind out of the active site; (E) the activators incorporate proton-shuttling moieties (PSMs) and bind in the same active site region as the inhibitors shown at (C). Only α-class CAs are considered here (as they are the enzymes present in mammals, including humans^12,14) although these mechanisms may be valid to other CA classes.

Figure 3.

X-ray crystal structure of adducts of coumarins 1 and 3 bound to hCA II. The hydrolysis products of the two coumarins, cis-2-hydroxycinnamic acid 2 (in yellow) and trans-2-hydroxy-cinnamic acid 4 (in magenta) were observed bound wat the entrance of the CA active site. The catalytic Zn(II) ion is the central violet sphere, its three protein ligands (His94, 96 and 119, CPK colors) and the proton shuttle residue His64 (in red) are evidenced. The hCA II backbone is shown as the green ribbon, with its various α-helices, β-sheets, and loops represented in a canonical manner.

Figure 4.

Natural product coumarins 5–35 investigated as CAIs.

Figure 5.

NPCs from the Sardinian plant Magydaris pastinacea investigated as CAIs.