Suppression of the Ubiquitin Pathway by Small Molecule Binding to Ubiquitin Enhances Doxorubicin Sensitivity of the Cancer Cells

Abstract

Development of inhibitors for ubiquitin pathway has been suggested as a promising strategy to treat several types of cancers, which has been showcased by recent success of a series of novel anticancer drugs based on inhibition of ubiquitin pathways. Although the druggability of enzymes in ubiquitin pathways has been demonstrated, ubiquitin itself, the main agent of the pathway, has not been targeted. Whereas conventional enzyme inhibitors are used to silence the ubiquitination or reverse it, they cannot disrupt the binding activity of ubiquitin. Herein, we report that the scaffolds of sulfonated aryl diazo compounds, particularly Congo red, could disrupt the binding activity of ubiquitin, resulting in the activity equivalent to inhibition of ubiquitination. NMR mapping assay demonstrated that the chemical directly binds to the recognition site for ubiquitin processing enzymes on the surface of ubiquitin, and thereby blocks the binding of ubiquitin to its cognate receptors. As a proof of concept for the druggability of the ubiquitin molecule, we demonstrated that Congo red acted as an intracellular inhibitor of ubiquitin recognition and binding, which led to inhibition of ubiquitination, and thereby, could be used as a sensitizer for conventional anticancer drugs, doxorubicin.

In vivo modulation of ubiquitin chains by N-methylated non-proteinogenic cyclic peptides

“…10 An attractive alternative is to target the signal for degradation: find molecules to bind to and interfere with the recognition of Lys48-linked Ub chains. [11][12][13][14][15] However, given the breadth of cellular processes that rely on differently linked Ub chains, specificity is crucial 5 and also a challenge due to the following reasons: Ub chains are assembled from the same monomer, and the differences in structure and dynamics of differently linked-Ub chains can be subtle. 16,17 Moreover, chain length is also important.…”

“…Those discovered to date suffer from weak binding and poor specificity for linkage and length. [12][13][14][15] Typically, these are symmetric molecules that bind monomeric Ub weakly, and then achieve what affinity they have for Ub chains through multimeric interactions. However, true recognition of Ub chains with a specific linkage was achieved recently using an alternative modality -de novo cyclic peptides.…”

“…To achieve this, they first used solid-phase peptide synthesis (SPPS) coupled to Lys analogue native chemical ligation 20,21 to generate Lys48linked Ub chains. 11 These Ub chains were of sufficient purity for use as targets for the RaPID system, a method that uses reprogrammed ribosome peptide synthesis to synthesize, and then screen, extremely large (410 12 ) cyclic peptide libraries for binding molecules. 22 Tight-binding, having low nM K D , and linkage-specific cyclic peptides were discovered, and their novel Ub recognition abilities were explained by their binding modes.…”

Construction of a circRNA-miRNA-mRNA network based on competitive endogenous RNA reveals the function of circRNAs in osteosarcoma

“…Furthermore, the ubiquitination of proteins has been found to be involved in the cancer process. Suppression of the ubiquitin pathway by small molecule binding to ubiquitin enhances the doxorubicin sensitivity of cancer cells [33]. Epigenetic silencing of ubiquitin-specific protease 4 by snail1 contributes to macrophage-dependent inflammation and therapeutic resistance in lung cancer [34].…”

Protein Degradation and the Pathologic Basis of Phenylketonuria and Hereditary Tyrosinemia

“…Congo red inhibits the conjugation of K48- and K63-linked polyubiquitination. Inhibiting the ubiquitin itself inhibits all ubiquitin-mediated proteasomal/autophagic degradation, and it can therefore be used to enrich the population of all functional mutant proteins in a cell [ 135 ]. The ubiquitin inhibitor, Congo red, by itself or in combination with another strategy, could be important in supporting the survival of functional but misfolded PAH and FAH proteins.…”