Design of CK2β‐Mimicking Peptides as Tools To Study the CK2α/CK2β Interaction in Cancer Cells

Abstract

The ubiquitously expressed Ser/Thr kinase CK2 is a key regulator in a variety of key processes in normal and malignant cells. Due to its distinctive anti‐apoptotic and tumor‐driving properties, elevated levels of CK2 have frequently been found in tumors of different origin. In recent years, development of CK2 inhibitors has largely been focused on ATP‐competitive compounds; however, targeting the CK2α/CK2β interface has emerged as a further concept that might avoid selectivity issues. To address the CK2 subunit interaction site, we have synthesized halogenated CK2β‐mimicking cyclic peptides modified with the cell‐penetrating peptide sC18 to mediate cellular uptake. We investigated the binding of the resulting chimeric peptides to recombinant human CK2α using a recently developed fluorescence anisotropy assay. The iodinated peptide sC18‐I‐Pc was identified as a potent CK2α ligand (Ki=0.622 μm). It was internalized in cells to a high extent and exhibited significant cytotoxicity toward cancerous HeLa cells (IC50=37 μm) in contrast to non‐cancerous HEK‐293 cells. The attractive features and functionalities of sC18‐I‐Pc offer the opportunity for further improvement.

Interdependence of charge and secondary structure on cellular uptake of cell penetrating peptide functionalized silica nanoparticles

“…Although the latter might be disadvantageous with respect to future applications, in which endosomal release is important, we recently demonstrated that bioactivity was maintained for other CPP-based conjugates that were also taken up partly by endocytosis. [31][32][33] The highest intracellular intensity levels of the uorophore were visible in the case of HK15 followed by HK17 and HK3, which conrmed the previously observed trend during ow cytometry measurements of the free peptides ( Fig. 1A and B).…”

Synthesis and SAR of Tetracyclic Inhibitors of Protein Kinase CK2 Derived from Furocarbazole W16

“…Although 2 was able to affect this PPI, it did not significantly inhibit CK2 activity . The same behavior was observed for the cyclic 13 amino acid peptide Pc and its derivatives, which were designed as CK2β mimetics to inhibit CK2 subunit association …”

“…In addition to analyzing the influence of the test compounds on the CK2α/CK2β interaction, the inhibition of the enzymatic activity of the holoenzyme (CK2α 2 β 2 ), the CK2α subunit, and the mutated CK2α’ C336S subunit was investigated in a capillary electrophoresis (CE) assay. In this assay, the decapeptide RRRDDDSDDD, a known class I substrate suitable for assaying the kinase activity of both the CK2 holoenzyme and the catalytically active CK2 subunits, was reacted with ATP in the presence of enzyme, which transfers a phosphate group to Ser7 of the decapeptide. Due to the additional negative charge of the phosphate group, the decapeptide and the phosphorylated decapeptide can be separated by CE.…”

“…The proteins CK2α 1−335 and CK2β 1−193 were recombinantly expressed and purified as described in literature, with the exception of the first purification step on phosphocellulose, where the protein was eluted using high salt buffer (1 M NaCl, 25 mM Tris/HCl, pH 8.5) without applying a gradient. CK2β 1−193 was fluorescently labelled using the Nanotemper Monolitz TM NT.115 Protein Labeling Kit RED‐NHS according to the manufacturer's manual.…”

Discovery of holoenzyme-disrupting chemicals as substrate-selective CK2 inhibitors

“…Comparative molecular dynamics simulations performed on this complex highlighted, among the hydrophobic residues, the prominent role of Phe190 for a stable and active conformation of Pc 19 . Recently, improved versions of Pc were characterized with halogens in meta-position of Phe190 20 or a novel covalent linker 21 . Notably, none of the Pc derivatives are cell-permeable, and thus they all require a cell-penetrating peptide for delivery.…”