Discovery of a New Binding Site on Human Choline Kinase α1: Design, Synthesis, Crystallographic Studies, and Biological Evaluation of Asymmetrical Bispyridinium Derivatives

Abstract

Human choline kinase α (CKα) is a validated drug target for the treatment of cancer. In recent years, a large number of CK inhibitors have been synthesized, and one of them is currently being evaluated in Phase I clinical trials as a treatment for solid tumors. Here we have evaluated a new series of asymmetrical biscationic CK inhibitors by means of enzymatic, crystallographic, and antitumor studies. We demonstrate that one of these structures adopts a completely new binding mode not observed before inducing the aperture of an adjacent binding site. This compound shows antiproliferative and apoptotic effects on cancer cells through activation of caspase-3. Therefore, this study not only provides fruitful insights into the design of more efficient compounds that may target different regions in CKα1 but also explains how these compounds induce apoptosis in cancer cells.

Choline kinase alpha—Putting the ChoK-hold on tumor metabolism

“…bis-pyridinium, bis-quinolinium) and cytostatic ( e.g. V-11-0711) ChoKα inhibitors may be the length of the molecule and the ability to extend from the choline-binding site to one of the adjacent clefts formed by residues which open or close depending on enzyme conformation [134]. The possibility of undetected off-target binding, however, remains a concern until these discrepancies are resolved.…”

Choline metabolism-based molecular diagnosis of cancer: an update

“…Combined treatment with 5-fluorouracil and siRNA silencing [30] or Chk-α inhibition [31] demonstrated synergistic effects of both treatments in breast and colorectal cancer models, respectively. Advances have also been made in characterizing the structure of Chk-α through the identification of a new binding site that may result in the design of more effective compounds [32,33]. …”

Pharmacophore‐Based Virtual Screening to Discover New Active Compounds for Human Choline Kinase α1

“…Overall, this compound contains 3 of the 5 moieties present in our proposed pharmacophore (Figure S4) that in turn are required to interact with the protein as described above (Figure 2e and 4a). Owing to these interactions, hit 10 shows two binding modes that allow on one hand to compete with the magnesium atom (this metal is coordinated by Asp330 and Asn311 and the phosphate groups) and the ADP phosphate groups (Figure 4b), and on the other hand to mimic the positive charge present in HC‐3,31 choline/phosphocholine, and HC‐3 derivatives such as compound 12 ,8c compound 13 8b and compound 14 8a (Figures 4b–f). In all these compounds the positive charge is key to maintain a cation‐π interaction with Trp420.…”

“…Overall, the results of crystallization studies of Hs CKα1 in complex with hit 10, combined with previous structural data,8 contribute to support a scenario in which the majority of Hs CKα1 inhibitors generally occupy the choline‐binding site, using only one positive charged moiety of the molecule to engage aromatic residues in cation‐π interactions. Although the second positive charge of biscationic compounds is usually exposed to the solvent, for one particular compound inducing the aperture of an adjacent new binding site to the choline binding site, it was found that both positive charges are engaged in cation‐π interactions 8a. Remarkably, a second copy of hit 10 has been identified in the ATP binding site, mimicking a magnesium atom that is coordinated by the ATP γ‐phosphate and making electrostatic and hydrogen bond interactions with polar residues.…”

“…6a–e One of these molecules, TCD‐717 ( 2 ), has even passed Phase I under clinical trials for advanced solid tumors 7. Very recently the first crystal structures of several complexes of Hs CKα1 bound to some of these inhibitors have enlightened their binding mode majorly through interactions to the choline‐binding site 8a–c. Inhibitors have been also developed using not only a rational approach but also virtual and high‐throughput screening of small‐molecule libraries 9…”