Inhibitors of cullin-RING E3 ubiquitin ligase 4 with antitumor potential

Abstract

Cullin-RING (really intersting new gene) E3 ubiquitin ligases (CRLs) are the largest E3 family and direct numerous protein substrates for proteasomal degradation, thereby impacting a myriad of physiological and pathological processes including cancer. To date, there are no reported small-molecule inhibitors of the catalytic activity of CRLs. Here, we describe high-throughput screening and medicinal chemistry optimization efforts that led to the identification of two compounds, 33-11 and KH-4-43, which inhibit E3 CRL4 and exhibit antitumor potential. These compounds bind to CRL4’s core catalytic complex, inhibit CRL4-mediated ubiquitination, and cause stabilization of CRL4’s substrate CDT1 in cells. Treatment with 33-11 or KH-4-43 in a panel of 36 tumor cell lines revealed cytotoxicity. The antitumor activity was validated by the ability of the compounds to suppress the growth of human tumor xenografts in mice. Mechanistically, the compounds’ cytotoxicity was linked to aberrant accumulation of CDT1 that is known to trigger apoptosis. Moreover, a subset of tumor cells was found to express cullin4 proteins at levels as much as 70-fold lower than those in other tumor lines. The low-cullin4–expressing tumor cells appeared to exhibit increased sensitivity to 33-11/KH-4-43, raising a provocative hypothesis for the role of low E3 abundance as a cancer vulnerability.

[6]-Paradol suppresses proliferation and metastases of pancreatic cancer by decreasing EGFR and inactivating PI3K/AKT signaling

“…Ubiquitination plays an important role in protein localization, metabolism, function, regulation and degradation [ 22 ]. At the same time, it is also involved in the regulation of cell cycle, proliferation, apoptosis, differentiation, metastasis, gene expression, transcriptional regulation, signal transduction, injury repair, inflammation and immunity, and almost all life activities [ 23 ]. Zhang et al found Ginsenoside compound K inhibited the proliferation of liver cancer via promoting the degradation of HIF-1α ubiquitination [ 24 ].…”

Targeting Cullin-RING E3 Ubiquitin Ligase 4 by Small Molecule Modulators

“…These results have provided proof-of-principle evidence that an E2-E3 interface can be perturbed with small molecule modulators. Using this approach, largescale HTS with extensive follow-up hit-to-lead studies, led to identification of a class of small-molecule inhibitors against E3 CRL4 [82].…”

“…In an effort to discover small molecule modulators that target E3 CRL, HTS and follow up hit-to-lead studies were carried out, resulting in identification of two compounds, 33-11 and KH-4-43, which inhibit E3 CRL4 and exhibit antitumor potential [82]. These compounds bind to CRL4's core catalytic complex, inhibit E3 CRL4A CRBN -dependent ubiquitination of CK1α in vitro, and cause stabilization of CRL4's substrate Cdt1 in cells (Figure 4).…”

“…According to the Structural Genomics Consortium, chemical probes are required to minimally have in vitro potency of the target protein at <100 nM, possess >30X selectivity relative to other sequence-related proteins of the same target family, and have demonstrated on-target effects at <1 μM [84]. KH-4-43 appears close to these criteria because it has a binding K d to E3 ROC1-CUL4A CTD or the highly related ROC1-CUL1 CTD at 83 nM or 9.4 μM, respectively [82], which represents a difference of two orders of magnitude. KH-4-43 exhibits cytotoxicity in a subset of tumor cell lines with EC 50 approaching ~2 μM.…”

“…Based on preliminary structure activity relationship (SAR) studies, the 33-11/KH-4-43 scaffolds are highly amenable to medicinal chemistry optimization with several substituents available for modification and building blocks that have been easily incorporated into the scaffold [82]. Continued characterization of the key pharmacophores of validated leads will provide improved CRL4 potency and selectivity, as well as drug-like properties and pharmacokinetic parameters.…”