Ординатура / Офтальмология / Английские материалы / Carbonic Anhydrase Its Inhibitors and Activators_Supuran, Scozzafava, Conway_2004

ized and tested as antibacterial agents, with many such derivatives currently being used to treat bacterial infections (Mandell and Petri 1996). Sulfonamide derivatives, widely used in clinical medicine as pharmacological agents with a wide variety of biological actions, were designed from the simple sulfanilamide lead molecule (Scozzafava et al. 2003; Casini et al. 2002). In addition to the antibacterials mentioned previously, the unsubstituted aromatic/heterocyclic sulfonamides act as carbonic anhydrase inhibitors (CAIs) (Supuran and Scozzafava 2001; Supuran et al. 2003), whereas other types of derivatives show diuretic activity (high-ceiling diuretics or thiadiazine diuretics), hypoglycemic activity, anticancer properties (Supuran 2003) or inhibitory effects on the aspartic HIV protease, being used to treat AIDS and HIV infection (Scozzafava et al. 2003). Historically, the first sulfonamide metal complex reported was the silver(I) derivative of sulfanilamide, prepared from the sulfonamide sodium salt and silver nitrate by Braun and Towle (1941). The metal complexes of substituted sulfanilamides were then investigated in detail by Bult (1983). However, few crystal structures of such complexes were reported, whereas metal complexes of other types of sulfonamides (except the CAIs, discussed later) have not been investigated.

6.2 ACETAZOLAMIDE COMPLEXES

Among the sulfonamide CAIs, acetazolamide [(H2acm); 5-acetamido-1,3,4,-thiadi- azole-2-sulfonamide; see Scheme 6.1) has been extensively used clinically (under the trademark Diamox) as a diuretic drug, and is still used at present to treat glaucoma, epilepsy and other neuromuscular diseases and as a diagnostic tool (Maren 1967; Supuran and Scozzafava 2000; Supuran et al. 2003).

The crystal structure of H2acm was reported by Mathew and Palenik (1974). Neutral sulfonamides are expected to behave as poor ligands toward metal ions,

SCHEME 6.1

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because of the withdrawal of electron density from the nitrogen atom onto the electronegative oxygen atoms. However, if the sulfonamide nitrogen atom bears a dissociable hydrogen atom, this same electron-withdrawing effect increases its acidity, and in the deprotonated form, sulfonamidate anions can act as effective σ-donor ligands for cations. Acetazolamide has several donor atoms that can bind metal ions; however, according to Mathew and Palenik (1974), H2acm is not a good ligand, even though acetazolamide was earlier used for the gravimetric determination of Ag(I) (Malecki et al. 1984). As a consequence, complexes of sulfonamides have only recently been investigated in detail, mainly by our groups.

When does acetazolamide behave as a ligand through the thiadiazole ring and when through the sulfonamidate group? This depends on the presence of the two acidic groups in this ligand: the sulfonamido and the acetamido moieties. A search in the literature for research on the acidic character of acetazolamide indicates that this property was controversial until Ferrer et al. (1990a) studied potentiometrically the determination of acid constants of acetazolamide by using the SUPERQUAD program. As acetazolamide contains two acidic protons, four species can be present simultaneously in solution (Scheme 6.1). The pKa1 and pKa2 values determined in aqueous and in aqueous–ethanol media were 7.19 and 7.52, and 8.65 and 9.41, respectively. Which value corresponds to the monodeprotonated species through the sulfonamido group and which to the monodeprotonated species through the acetamido group? Probably the most acidic group is the sulfonamido one, as considered by several researchers, including Kimura et al. (1990), but there is little experimental evidence to confirm it. Independent of the assignment of pKa values, the most important point is that at neutral pH the two monodeprotonated and dideprotonated species coexist in concentrations high enough to allow metal ions to bind to any of them (Ferrer et al. 1990a). Probably, the final result is not only a consequence of the affinity of the metal ion for this ligand but also an ensemble of factors such as crystal packing and hydrogen bonds of the coordination compound.

Acetazolamide is the sulfonamide that has been most extensively studied as a ligand (Alzuet et al. 1994c). The interest in its coordination chemistry comes from its properties as an inhibitor of CA and also from the multitude of coordination possibilities that it offers. Acetazolamide has several potential donor positions (at the heterocyclic ring, acetamide group, sulfonamide moiety), and, as mentioned previously, two acidic groups that afford four different species: (1) H2acm, (2) sulfona- midate-Hacm–, (3) acetamidate-Hacm– and (4) the dideprotonated species acm2– (Scheme 6.1). Therefore, acetazolamide can act as a very versatile ligand (Ferrer et al., 1990a; Alzuet et al. 1994c).

Ferrer et al. (1987) reported the first complexes of acetazolamide, [Co(Hacm)2(NH3)2] and [Zn(Hacm)2(NH3)2]. Since then, many compounds with transition metal ions and a few with main-group metal ions have been reported (Alzuet et al. 1994a, 1994c; Ferrer et al. 1989a, 1989b; Supuran et al. 2003). Table 6.1 lists most of the acetazolamide complexes described in the literature. Most of them have been investigated by the research groups of Borrás and Supuran (previously reviewed by Alzuet et al. 1994c). In all the reported complexes, acetazolamide acts as a ligand in anionic form, either monoor dideprotonated, which correlates well with the syntheses being performed: by adding a strong base (KOH, Bu4NOH); or

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186 Carbonic Anhydrase

TABLE 6.1

Reported Acetazolamide Complexes

a ea: ethylamine, dea: diethylamine, tea: triethylamine, L: tris[2-(1-methylbenzimidazol-2- yl)ethyl]nitromethane, L′: [12]aneN3=1,5,9-triazacyclododecane, en: 1,2-ethanediamine, tn: 1,3-pro- panediamine, dien: diethylenetriamine, dipn: dipropylenetriamine.

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O(2)

FIGURE 6.1 Crystal structure of [Ni(Hacm)2(NH3)4] (6.1). (Reprinted from Ferrer, S. et al. (1989b) Inorganic Chemistry 28, 160–163. With permission from ACS.)

in the presence of a weak one, such as ammonia, aliphatic monoamines (ea, dea and tea), chelating diamines (en and tn) or tridentate triamines (dien and dipn; see Table 6.1 for amine abbreviations); or by using the sodium salt of the ligand (NaHacm). The only exception is the silver complex Ag2(acm), prepared directly from the acetazolamide without a base (Ferrer et al. 1989a).

Despite the extensive study of acetazolamide complexes, only five have been analyzed by crystallographic methods. The first published crystal structure corresponds to that of the purple compound [Ni(Hacm)2(NH3)4] (6.1, Figure 6.1; Ferrer et al. 1989b). It consists of discrete units that contain a NiN6 chromophore with the metal center in a slightly elongated rhombically distorted octahedral environment formed by four ammonia molecules in equatorial positions [average d(Ni – NH3) = 2.105 Å] and two trans Hacm– ligands in the apical ones [d(Ni – N(2)) = 2.150 Å]. The structure shows acetazolamide deprotonated at the acetamido group and coordinating through N(2), the N-thiadiazole atom closest to the group with the negative charge. The changes in the ligand, mainly by the deprotonation rather than by the coordination, are clearly reflected on the IR spectrum of the complex (Table 6.2), with the carbonyl band shifted and split but with the sulfonamido bands not suffering significant modification as compared with the corresponding bands of the ligand.

The next compound for which the crystal structure was described was the dark blue [Cu(acm)(NH3)2(OH2)2]2 · 2H2O (6.2; Figure 6.2; Ferrer et al. 1990b). Its structure, made of dinuclear entities and crystallization water molecules, corresponds to

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TABLE 6.2

Significant IR Bands (cm1) for Acetazolamide and Its Complexesa

a s: strong; m: medium; w: weak; sh: shoulder, b: broad, d: doublet.

N(2)*

N(1)*

FIGURE 6.2 Crystal structure of [Cu(acm)(NH3)2(OH2)]2 ◊ 2H2O (6.2). (Reprinted from Ferrer, S. et al. (1990b) Inorganic Chemistry 29, 206–210. With permission from ACS.)

the only polynuclear derivative of acetazolamide currently available. In the centrosymmetric dimer, the two Cu(II) ions are connected by a double N(1) – N(2) thiadiazole bridge. The metallic centers form CuN3O + N2 chromophores with a tetragonally elongated octahedral geometry around each copper ion. Each acetazolamide ligand, in its doubly deprotonated form (acm2–), acts as a bidentate chelate toward one Cu(II) ion via the N(1)thiadiazole and the N(4)sulfonamidate atoms and as a monodentate ligand toward the other Cu(II) ion via the N(2)thiadiazole atom, with the following bonding distances: d(Cu – N(2)) = 2.020 Å, d(Cu – N(4)) = 2.028 Å and d(Cu – N(1)) = 2.546 Å. Despite its negative charge, the tridentate acm2– does not bind through the acetamidate group. The IR spectrum of the complex is in agreement with the deprotonation of both the acetamido and sulfonamido groups, with significant shifts of the corresponding bands to lower frequencies (Table 6.2).

The third crystal structure, determined by Hartmann and Vahrenkamp (1991), was that of the complex [Zn(Hacm)2(NH3)2] (6.3), already obtained by Ferrer et al.

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(1987). The structure is formed by a mononuclear species that contains the Zn(II) ion tetrahedrically surrounded by four nitrogen atoms, two of them from two ammonia molecules, with d(Zn–N) = 2.021 Å, and the other two from two sulfonamidate groups, with d(Zn–N) = 1.977 Å. Despite the apparent similarity with compound 6.1, [Ni(Hacm)2(NH3)4], the behavior of acetazolamide as ligand in 6.3 is completely different. In the present case, acetazolamide is also monodeprotonated, but deprotonation takes place on a different group (at the sulfonamido moiety), and it also acts as a monodentate ligand, but through the nitrogen sulfonamidate atom. These differences are clearly indicated by the IR spectrum of 6.3 (Table 6.2), with shifts of the SO2 bands to lower frequencies, which, together with the electronic spectrum of the [Co(Hacm)2 (NH3)2] compound, allowed Borrás’s group to predict the right structure before the x-ray study was available. (The only aspect not solved by the spectral evidences was whether the coordination was produced via the N or the O sulfonamidate atoms; Ferrer et al. 1987.) Finally, it is remarkable that the structural details of 6.3 compare very favorably to those of the acetazolamide bound to CA II, with similar findings for the coordination geometry and for the donor atom of the inhibitor (Vidgren et al. 1990).

The complexes described previously were synthesized working with an excess of ammonia. The next studies involved syntheses in the presence of chelating diamines and led to the isolation of two new compounds for which the crystal structures could be determined: the purple [Cu(Hacm)2(en)2] (6.4; en = 1,2-ethanedi- amine; Figure 6.3) and the dark blue [Cu(Hacm)2(tn)2] (6.5; tn = 1,3-propanediamine;

N(6)*

C(6)

N(6) Cu

N(5)*

FIGURE 6.3 ORTEP drawing of [Cu(Hacm)2(en)2] (6.4) (en = 1,2-ethanediamine). (Reprinted from Ferrer, S. et al. (1992) Inorganica Chimica Acta 192, 129–138. With permission from Elsevier.)

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FIGURE 6.4 ORTEP drawing of [Cu(Hacm)2(tn)2] (6.5) (tn = 1,3-propanediamine). (Reprinted from Ferrer et al. (1992) Inorganica Chimica Acta 192, 129–138. With permission from Elsevier.)

Figure 6.4; Ferrer et al. 1992). In both these mononuclear complexes, acetazolamide is deprotonated at the acetamido group, but its binding mode to the metal ion is completely different from those of previously investigated derivatives 6.1 to 6.3. In both 6.4 and 6.5, the Cu(II) ions, on the symmetry centers, exhibit an elongated octahedral geometry, with the four diamine nitrogen atoms in an approximately square coplanar arrangement [d(Cu–N) = 2.001 and 2.009 Å for 6.4; d(Cu–N) = 2.044 and 2.048 Å for 6.5]; two O-sulfonamido atoms in 6.4 and two N(2)-thiadiazole atoms in 6.5 (from two trans acetazolamidate ligands) complete the octahedra at longer distances [2.652 Å and 2.457 Å, respectively]. Again the donor positions of acetazolamide do not belong to the group that bears the deprotonation in either of the two complexes. In 6.5, the ligand binds in the expected way; its structure is similar to that of 6.1 [although in 6.5 the distortion on apical positions is more pronounced, in agreement with the different nature of the metal ion, Cu(II) instead of Ni(II)]. For 6.4, however, the coordination mode of acetazolamide is unexpected: it binds through one oxygen sulfonamido atom (and at a semicoordinating distance), even though the negative charge is on the other substituent of the ring (in 6.1 and 6.5, the donor atom, N(2), participates on the negative charge of N(3) due to electronic delocalization). This unusual behavior of 6.4 must be related to the Cu(II) Jahn–Teller effect and to the existence of a network of strong hydrogen bonds that stabilize the excess of negative charge at N(3) through pairing of acetazolamidate anions. Therefore, in the presence of these chelating amines, acetazolamide behaves as an ambidentate, unusually weak anionic ligand. The major IR changes are in

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TABLE 6.3

Coordination Ways of Acetalozamide Determined by x-Ray Crystallography

agreement with the deprotonation of the acetamido group (6.4, 6.5); the weak O-sul- fonamido coordination has small effects on the sulfonamide IR bands of 6.4 (Table 6.2).

Some conclusions were derived from these five structures of acetazolamide complexes. First, acetazolamide always binds in an anionic form, with three possibilities: as acetamidate, as sulfonamidate or dideprotonated. Second, coordination is not necessarily achieved through the deprotonated group: the acetamido/-ate group never interacts directly with the metal ion. Third, acetazolamide can exhibit at least four different ways of coordination, presented in Table 6.3, which confirm its versatility as a ligand. Finally, the IR spectrum can be used as a diagnostic tool to propose the binding mode in cases when a crystal structure is not available (Table 6.2).

Apart from the previously mentioned structures, the binding of acetazolamide to model complexes that mimic the CA active site has also been investigated. Kimura et al. (1990) reported the complex [Zn(Hacm)([12]aneN3)](ClO4) ([12]aneN3 = 1,5,9- triazacyclododecane) as a model of the interaction of acetazolamide with the active metal center of CA. Although good crystals could not be obtained for x-ray studies, the spectroscopic investigation indicated that the monodeprotonated sulfonamide is coordinated to Zn(II) through the sulfonamido nitrogen atom. Casella’s group prepared Zn(II), Co(II) and Cu(II) complexes with the ligand tris[2-(1-methylbenzimidazol- 2-yl)ethyl]nitromethane (L), whose donor positions mimic the environment of the Zn(II) site of CA, and then studied the formation of adducts of such complexes with acetazolamide (Alzuet et al. 1994a). Two ternary complexes [Zn(Hacm)L](ClO4) and [Co(Hacm)L](ClO4)· H2O were obtained. In the case of Cu(II), it was not possible to isolate the pure ternary complex, but mixed-ligand complexes with the triamines dien and dipn, with the formula [Cu(Hacm)(dien)](ClO4) · H2O and [Cu(Hacm)(dipn)](ClO4), were obtained. The available spectral evidences indicated

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that in these cases acetazolamide binds in its monoanionic form through the deprotonated sulfonamido group. These ternary complexes can be considered analogues of the adducts formed by CA and the inhibitor acetazolamide. These complexes are the only acetazolamide compounds so far reported that contain a noncoordinating anion.

6.3 METHAZOLAMIDE COMPLEXES

From the coordination point of view, methazolamide [N-(4-methyl-2-sulfamoyl- 2- 1,3,4-thiadiazolin-5-ylidene)acetamide] (Hmacm) behavior is much simpler than that of the structurally related acetazolamide, because methylation on the thiadiazole N-2 atom (probably the best donor atom in acetazolamide) makes it unable to interact with metal ions. Furthermore, methazolamide incorporates only one acidic proton, at the sulfonamide group, with a pKa of 7.4 (Alzuet et al. 1994c) and therefore can be used as a simpler model for understanding the interaction of sulfonamides with metal ions (Scheme 6.2). The synthesis, crystal structure and chemical characterization of several methazolamide metal complexes also containing ammonia or pyridine as ligands have been described (Table 6.4).

[Ni(macm)2(NH3)4] (6.6) was the first reported metal complex of methazolamide (Figure 6.5; Alzuet et al. 1993a). The geometry around the metal ion can be described as a slightly elongated rhombic octahedron. Methazolamide occupies the axial positions,

SCHEME 6.2

TABLE 6.4

Methazolamide Complexes with Structures Determined by x-Ray Crystallography

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N3*

FIGURE 6.5 Crystal structure of [Ni(macm)2(NH3)4] (6.6). (Reprinted from Alzuet, G. et al. (1993a) Inorganica Chimica Acta 203, 257–261. With permission from Elsevier.)

acting as a monodentate ligand coordinating through the nitrogen atom of the deprotonated sulfonamide group. A comparison with the related acetazolamide compound [Ni(Hacm)2(NH3)4] shows that the axial Ni-N(sulfonamide) distance (2.216Å) in the [Ni(macm)2(NH3)4] complex is slightly longer than that for Ni-N(thiadiazole) (2.150 Å) in [Ni(Hacm)2(NH3)4]. This difference in axial distances has been connected with the different ligand fields created by the donor nitrogen atoms. The higher bond strength of the N(thiadiazole) atom seems to be the result of the strong

delocalization in the thiadiazole ring of the Hacm– anion.

The complex [Co(macm)(NH3)(aib)2(NO3)2]·2H2O (6.7; Figure 6.6) was prepared by reacting Co(NO3)2 with methazolamide and ammonia in acetone (Alzuet et al. 1993b). It contains two molecules of 2-methyl-2-amino-4-imino pentane (aib), which acts as a bidentate ligand. The aib ligand is obtained in situ by condensation of two acetone and two ammonia molecules in a template-type reaction described by Curtis (1963). In addition, the air spontaneous oxidation of Co(II) to Co(III) occurs. Co(III) exhibits a nearly regular octahedral geometry. Methazolamide exhibits its usual coordination behavior, binding to the metal ion through the nitrogen atom of the sulfonamidate group (Alzuet et al. 1993b).

More recently, the compound [Zn(macm)2(NH3)2] (6.8; Figure 6.7) has been shown to be a good model of the interaction of methazolamide with the metal center in the active site of CA (Alzuet et al. 1995). The coordination sphere of the Zn(II) ion is a distorted tetrahedron. The Zn–N(sulfonamido) bond distances and the coordination angles compare well with those reported for the hCAII–acetazolamide complex (Vidgren et al. 1990). Analysis based on extended Huckel calculations applied to this compound support the results obtained by Vidgren et al. (1990), which indicated the importance of interactions of acetamido and thiadiazole ring in the complex formation between enzyme and the sulfonamide bound within the active site. Furthermore the isostructural [Co(macm)2(NH3)2] derivative has been obtained

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