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Majkut, J.

Publications and source records attributed to Majkut, J..

2 recordsLinked to original sources

Targeted degradation of GSPT1 and NEK7 by a molecular glue prodrug for treatment of HCC

Targeted Protein Degradation (TPD) technology, in the form of CRBN-modulating molecular glues, offers numerous unprecedented therapeutic benefits as evidenced by the success of approved high-value immunomodulatory imide drugs (IMiDs) such as lenalidomide and pomalidomide. Building upon these successes, we employed a small CRBN-focused library of molecular glues in a phenotypic screen against hepatocellular carcinoma (HCC) cell lines. While the original library was primarily designed to target SALL4, we identified additional CRBN substrates, including GSPT1, NEK7, and CK1, whose degradation potently induced cell death in HCC cell lines. Subsequent lead optimization efforts yielded a compound, ABS-752, which demonstrated superior in vitro and in vivo activity through the potent degradation of GSPT1. Notably, ABS-752 does not form ternary complexes with CRBN and the neosubstrates. Further investigations revealed that ABS-752 is a prodrug activated by the monoamine oxidase, VAP-1, to an aldehyde intermediate and subsequently to the active molecule, ABT-002. VAP-1, which is overexpressed in cirrhotic liver, was identified as the primary monoamine oxidase responsible for the conversion of ABS-752. ABS-752 is currently in clinical trials for the treatment of HCC.

molecular biology↗

Regulation of FLIP(L) and TRAIL-R2 signalling by the SCFSkp2 Ubiquitin Ligase Complex

Depending on its expression levels, the long splice form of the pseudo-caspase FLIP (FLIP(L)) can act as an inhibitor (high expression) or activator (low expression) of apoptosis induction by the TRAIL-R2 death-inducing signalling complex (DISC); its expression levels are therefore tightly regulated. Here, we demonstrate that the Skp1-Cullin-1-F-box (SCF) Cullin-Ring E3 Ubiquitin Ligase complex containing Skp2 (SCFSkp2) regulates the stability of FLIP(L) (but not the short splice form FLIP(S)), and, unusually, this is mediated by direct binding of FLIP(L) to Cullin-1 rather than via Skp2. By fine mapping the interaction of FLIP(L) with Cullin-1 to the large subunit of its pseudo-caspase domain, we found that the interaction is significantly stronger with FLIP(L)s DISC-processed p43-form. Importantly, this interaction disrupts the ability of p43-FLIP to interact with FADD, caspase-8 and another DISC component, TRAF2. Moreover, we find that SCFSkp2 associates with TRAIL-R2 constitutively and does so independently of FLIP(L) and other canonical DISC components. Inhibition of Cullin-1 expression (using siRNA) or activity (using a NEDDylation inhibitor, MLN4924) enhanced FLIP(L) and TRAF2 levels at the TRAIL-R2 DISC and enhanced caspase-8 processing. This suggests that processing of FLIP(L) to p43-FLIP at the TRAIL-R2 DISC enhances its interaction with co-localised SCFSkp2, leading to disruption of p43-FLIPs association with the DISC thereby altering caspase-8 processing. These findings provide important new insights into how FLIP(L) expression and TRAIL-R2 signaling is controlled.

molecular biology↗