Abstract
The membrane-bound human carbonic anhydrase (hCA) IX is widely recognized as a marker of tumor hypoxia and a prognostic factor within several human cancers. Being undetected in most normal tissues, hCA-IX implies the pharmacotherapeutic advent of reduced off-target adverse effects. We assessed the potential anticancer activity of bumetanide-based analogues to inhibit the hCA-IX enzymatic activity and cell proliferation of two solid cancer cell lines, namely kidney carcinoma (A-498) and bladder squamous cell carcinoma (SCaBER). Bumetanide analogues efficiently inhibit the target hCA-IX in low nanomolar activity (IC50 = 4.4–23.7 nM) and have an excellent selectivity profile (SI = 14.5–804) relative to the ubiquitous hCA-II isoform. Additionally, molecular docking studies provided insights into the compounds’ structure–activity relationship and preferential binding of small-sized as well as selective bulky ligands towards the hCA-IX pocket. In particular, 2,4-dihydro-1,2,4-triazole-3-thione derivative 9c displayed pronounced hCA-IX inhibitory activity and impressive antiproliferative activity on oncogenic A-498 kidney carcinoma cells and is being considered as a promising anticancer candidate. Future studies will aim to optimize this compound to fine-tune its anticancer activity as well as explore its potential through in-vivo preclinical studies.
Highlights
As a worldwide healthcare burden, cancer has enormous financial and psychological impacts on patients, families, and healthcare systems [1]
Compounds 8a, 8h, 9b, and 9c were selected to be investigated for their potential antiproliferative activity on two different solid cancer cell lines, A-498/kidney carcinoma and SCaBER/bladder squamous cell carcinoma, and acetazolamide as the positive control standard
Cells were washed in 1× binding buffer (20× binding buffer: 0.1 M HEPES, pH 7.4; 1.4 M NaCl; 25 mM CaCl2diluted in dH2O) and incubated in the dark for 30 min on ice in Annexin V-containing binding buffer [1:100]
Summary
As a worldwide healthcare burden, cancer has enormous financial and psychological impacts on patients, families, and healthcare systems [1]. Bumetanide showed low nanomolar inhibition activity on the tumor-associated hCA isoforms (Ki = 21.1–25.8 nM) and is a promising effective anticancer agent because it showed moderate (250–700 nM) or weak (2.57–6.98 μM) inhibition for mitochondrial or cytosolic hCAs, respectively [39,41] Such a preferential pattern may be related to the sterically hindering phenoxy moiety being ortho to the sulphamoyl-Zn(II)-binding group. The bumetanide-containing compounds displayed weak to potent inhibitory activity toward the physiologically dominant off-target isoform hCA-II, with IC50 ranging from high nanomolar to low micromolar concentrations (IC50 = 205.08 and 5117 nM) with statistical significance (p < 0.05) as compared to reference control. Such favored activities suggest a different binding mode; the Series IV members may anchor their sterically hindered hydrazone-based extremities in a way favoring contacts with the pocket residues
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