Page 5 of 7
ACS Medicinal Chemistry Letters
also like to acknowledge the staff at Cornell High Energy Synꢀ
chrotron Source (CHESS) for assisting with Xꢀray data collection.
most potent hCA XII inhibitor, and exhibited cytotoxicity toꢀ
1
2
3
4
5
6
7
8
wards three different cancer cell lines. Interestingly, this
chemotype behaves differently from other related nonꢀ
classical inhibitors. In fact, it does not directly bind to the
metal ion or to the zincꢀbound water/hydroxide ion: the zinc
ion−carboxylate distances in our model are 3.78 and 4.62 Å
(for contact with each oxygen atom), too much longer to supꢀ
port a favorable zinc ion coordination to carboxylate functionꢀ
ality of the ligand. Otherwise, this compound seems to estabꢀ
lish contacts with residues located in proximity of the catalytic
site, which displaces the ordered water network (shorter zinc
ionsꢀwater distance, i.e. 2.26 and 1.94 Å, for zinc–O1 and
zinc–O2, were revealed), thus definitely producing an indirect
interference with the zinc ion. However, 2c displayed imꢀ
proved antiproliferative effects when tested in simulated hyꢀ
poxia conditions, considering the putative expected involveꢀ
ment of hCA XII inhibition. These results could provide strucꢀ
tural determinants for the discovery of novel anticancer agents
with original mechanism of action, warranting further develꢀ
opment.
ABBREVIATIONS
hCA, human carbonic anhydrase; CAIs, carbonic anhydrase inhibꢀ
itors; Ki, inhibition constant; ZBG, zincꢀbinding group; PCꢀ3,
Hormoneꢀindependent prostate cells; HEK 293, human embryonic
kidney cells; SHꢀSY5Y, human neuroblastoma cells.
9
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ASSOCIATED CONTENT
Supporting Information
The Supporting Information is available free of charge on the
ACS Publications website. Experimental details for chemistry, Xꢀ
1
ray crystallography and molecular modeling. HꢀNMR spectra for
title compounds 1b-d and 2a-d, and their intermediates 4a-c and
5a-c. Mass spectra for 1b-d, 2a-d, and 5a-c. Xꢀray crystallogꢀ
raphy statistics for data processing and refinement of 4d in comꢀ
plex with hCA II (Table S1). Details of experimental biology proꢀ
cedure (CA inhibition assay, cell culture, assessment of cell viaꢀ
bility). Molecular modelling additional data (Table S2 and Figures
S1–S5). Antiproliferative activity of compounds 1a, 2a, 1c, and
2c (Figure S6).
AUTHOR INFORMATION
Corresponding Author
M.S.: phone, +39 079ꢀ228ꢀ753; fax: +39 079ꢀ229ꢀ559, eꢀmail:
C.T.S: phone, 39ꢀ055ꢀ4573005; fax, +39ꢀ055ꢀ4573385, eꢀmail:
Author Contributions
The manuscript was written through contributions of all authors.
All authors have given approval to the final version of the manuꢀ
script. ‡These authors contributed equally.
Notes
The content is solely the responsibility of the authors and does not
necessarily represent the official views of the National Institutes
of Health. PDB ID 5CJL: 1d in complex with hCA II (Table S1).
Authors will release the atomic coordinates and experimental data
upon article publication. The authors declare no competing finanꢀ
cial interest.
(14) Supuran, C. T. Drug Design of zinc-enzyme Inhibitors:
functional, structural, and disease applications, Part II;
Wiley, Hoboken, New Jersey, 2009.
(15) Supuran, C. T., Scozzafava, A., Conway, J., Eds. Carbonic
anhydrase: its inhibitors and activators; CRC Press: Boca
Raton, FL, 2004.
(16) Alterio, V.; Di Fiore, A.; D’Ambrosio, K.; Supuran, C. T.;
De Simone, G. Multiple binding modes of inhibitors to carꢀ
bonic anhydrases: how to design specific drugs targeting 15
different isoforms? Chem. Rev. 2012, 112, 4421ꢀ4468.
ACKNOWLEDGMENT
This research was financed in part the National Institutes of
Health, project CA165284. CL is supported by the National Cenꢀ
ter for Advancing Translational Sciences of the National Institutes
of Health under University of Florida Clinical and Translational
Science Awards TL1TR001428 and UL1TR001427.We would
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