The X-ray data for the five CA II adducts of sulfonamides
a CAI may contribute significantly to the overall potency of
1
the compound.
6-22
1
–5 presented above show that the benzenesulfonamide part of
the molecules is very much superposable between the five
adducts, whereas the ureido and aryl/cycloalkyl moieties are
much less so. Data of Fig. 3 clearly show that 3 of the
In conclusion, this study demonstrates the importance of the
hydrophobic pockets/regions within the CA II active site for
the binding of a series of structurally related sulfonamides
possessing various ureido substitutions. These findings can be
extended to other classes of CAIs, and probably also to other
CA isoforms, less well investigated than CA II, with the
possibility of designing inhibitors with a better selectivity for
the various isoforms with medicinal chemistry applications.
5
compounds (1, 2 and 5) bind in a rather similar manner,
with the 4-fluorophenyl, pentafluorophenyl and cyclopentyl
moieties observed in the same active site region. This is a
rather well defined hydrophobic pocket observed in other CA
1
2,13
II–sulfonamide adducts,
which we have defined in a recent
1
2
paper hydrophobic pocket 2. In contrast to these three
inhibitors, the 3-nitrophenyl substituted compound 4 binds
in a less utilized hydrophobic pocket, which we have defined
Notes and references
1
2
pocket 1, these two pockets being separated by Phe131, an
1 C. T. Supuran, Nat. Rev. Drug Discovery, 2008, 7, 168.
2
J. F. Domsic, B. S. Avvaru, C. U. Kim, S. M. Gruner,
M. Agbandje-McKenna, D. N. Silverman and R. McKenna,
J. Biol. Chem., 2008, 283, 30766.
amino acid residue critical for orientating inhibitors in most
1
CA isoforms. Few inhibitors observed to date to bind in the
4
hydrophobic pocket 1, compound 4 being one of the 6 cases
1
3 D. W. Christianson and C. A. Fierke, Acc. Chem. Res., 1996, 29,
31.
2
3
detected so far.
What is even more interesting, the
4
5
V. Alterio, A. Di Fiore, K. D’Ambrosio, C. T. Supuran and G. De
Simone, X-Ray crystallography of CA inhibitors and its importance
in drug design in Drug Design of Zinc-Enzyme Inhibitors:
Functional, Structural, and Disease Applications, ed. C. T. Supuran
and J. Y. Winum, Wiley, Hoboken, 2009, pp. 73–138.
2
-iso-propylphenyl derivative, 3, exploits yet another hydro-
phobic patch, which is an intermediate location between
pockets 1 and 2 mentioned above, which we tentatively assign
the name hydrophobic region 3. At this point, by comparing
the inhibition constants of the five inhibitors (Fig. 1) with their
superposition when bound to the enzyme (Fig. 3), a very
interesting fact emerges: the three inhibitors binding in the
K. Ko
A. Heine, C. T. Supuran and G. Klebe, Angew. Chem., Int. Ed.,
007, 46, 7697.
¨
hler, A. Hillebrecht, J. Schulze Wischeler, A. Innocenti,
2
6 A. E. Eriksson, T. A. Jones and A. Liljas, Proteins: Struct., Funct.,
Genet., 1988, 4, 274.
7
hydrophobic pocket 2 are the weaker ones (K s in the range of
I
C. Temperini, A. Scozzafava and C. T. Supuran, Bioorg. Med.
Chem. Lett., 2010, 20, 474.
5
0–226 nM) in the small series of derivatives investigated
here. The compound located in the hydrophobic pocket 1,
8 S. K. Nair, P. A. Ludwig and D. W. Christianson, J. Am. Chem.
Soc., 1994, 116, 3659.
compound 4, is a much more effective CA II inhibitor
9
A. Maresca, C. Temperini, H. Vu, N. B. Pham, S. A. Poulsen,
A. Scozzafava, R. J. Quinn and C. T. Supuran, J. Am. Chem. Soc.,
(
K
I
of 15 nM) whereas the one binding in the intermediate
hydrophobic patch, between the two pockets, observed for the
first time here, is an excellent CAI, with a K of 3.3 nM against
2009, 131, 3057.
10 F. Carta, C. Temperini, A. Innocenti, A. Scozzafava, K. Kaila and
C. T. Supuran, J. Med. Chem, 2010, 53, 5511.
I
CA II. As above, the differences of activity between the
most active (derivative 3) and the least active (compound 5)
sulfonamides investigated here are significant, a factor of 68, and
this difference can be directly attributed to the rather diverse
binding of the substituted ureido tails of these compounds in the
various hydrophobic pockets/regions of the enzyme.
Another aspect that should be stressed here, which probably
explains the rather variable binding patterns of these structurally
similar compounds to CA II, is related to the presence of the
ureido fragment (NHCONH) which connects the benzene-
sulfonamide part of the molecule to the aryl/cycloalkyl tails.
As seen from the crystallographic data, the torsion angles
between these two fragments of the scaffold are different in the
five compounds investigated here. This probably allows the
flexibility of the inhibitor to select the most energetically
favourable hydrophobic pocket to bind into and avoid steric
1
1 The synthesis of inhibitors 1–5 was done by reaction of
sulfanilamide and the corresponding aryl/alkyl isocyanate as
described earlier in C. T. Supuran, A. Scozzafava, B. C. Jurca
and M. A. Ilies, Eur. J. Med. Chem., 1998, 33, 83.
2 B. S. Avvaru, J. M. Wagner, A. Maresca, A. Scozzafava,
A. H. Robbins, C. T. Supuran and R. McKenna, Bioorg. Med.
Chem. Lett., 2010, 20, 4376.
1
13 J. Wagner, B. S. Avvaru, A. H. Robbins, A. Scozzafava,
C. T. Supuran and R. McKenna, Bioorg. Med. Chem., 2010, 18,
4873.
4 V. Menchise, G. De Simone, V. Alterio, A. Di Fiore, C. Pedone,
A. Scozzafava and C. T. Supuran, J. Med. Chem., 2005, 48, 5721.
1
15 J. Y. Winum, C. Temperini, K. El Cheikh, A. Innocenti, D. Vullo,
S. Ciattini, J. L. Montero, A. Scozzafava and C. T. Supuran,
J. Med. Chem., 2006, 49, 7024.
1
6 Z. Otwinowski and W. Minor, Methods Enzymol, 1997, 276, 307.
17 S. Z. Fisher, C. M. Maupin, M. Budayova-Spano,
L. Govindasamy, C. K. Tu, M. Agbandje-McKenna,
D. N. Silverman, G. A. Voth and R. McKenna, Biochemistry,
2
8 P. D. Adams, P. V. Afonine, G. Bunkoczi, V. B. Chen, I. W. Davis,
007, 42, 2930.
1
5
clashes and/or to make as many as possible favorable
interactions with amino acid residues within the enzyme
cavity. Most of the CAIs of sulfonamide type investigated
1
´
N. Echols, J. J. Headd, L.-W. Hung, G. J. Kapral, R. W.
Grosse-Kunstleve, A. J. McCoy, N. W. Moriarty, R. Oeffner,
R. J. Read, D. C. Richardson, J. S. Richardson, T. C. Terwilliger
and P. H. Zwart, Acta Crystallogr., 2010, D66, 213.
2
earlier contained CONH or SO NH linkers instead of the
ureido one present in 1–5. These two different linkers allow
less flexibility for the inhibitor scaffold, and probably this is
the reason why most of those compounds bind in the cannonical
hydrophobic pocket 2. It is thus rather obvious that even a
very minor moiety (in this case the linker) from the scaffold of
19 A. W. Schuettelkopf and D. M. F. van Aalten, Acta Crystallogr.,
004, D60, 1355.
2
2
2
0 P. Emsley and K. Cowtan, Acta Crystallogr., 2004, 60, 2126.
1 R. A. Laskowski, M. W. MacArthur, D. S. Moss and
J. M. Thornton, J. Appl. Crystallogr., 1993, 26, 283.
22 R. G. Khalifah, J. Biol. Chem., 1971, 246, 2561.
This journal is c The Royal Society of Chemistry 2010
Chem. Commun., 2010, 46, 8371–8373 8373