Bedard, N. | bioRxiv Science

bioRxiv2025

Dyr726, a brain-penetrant inhibitor of PI3Kα, Type III receptor tyrosine kinases, and WNT signalling

The vast majority of clinical small molecule multi-kinase inhibitors (mKI) report abject failures in targeting cancers with high stem cell contents like high-grade glioma and colorectal cancers. The FDA-approved mKIs to date ablate receptor tyrosine kinase signaling but do not target the paradoxical WNT signaling which is a key survival driver for the self-renewing cancer stem cells. The WNT pathway enhances cancer plasticity and triggers relapse of highly heterogenous tumours. Using de novo synthesis and structure-activity-relationship (SAR) studies with blood-brain-barrier (BBB) penetrant mKI scaffolds, we designed a highly potent and selective small molecule inhibitor of PI3K, PDGFR/KIT, and the WNT pathway denoted Dyr726. Dyr726 is superior to clinical mKIs and inhibits PI3K-AKT-mTOR and WNT-pathway signaling at multiple nodes thereby impeding proliferation, invasion, and tumour growth. Phospho-proteomic, structural, and target engagement analyses, combined with in vitro, in vivo efficacy, and pharmacokinetic studies reveal that Dyr726 is a brain-penetrant small molecule which effectively reduces tumour volume and extends survival of murine orthotopic models. Our current work establishes a first-in-class brain penetrant small molecule mKI which simultaneously antagonize the PI3K-AKT-mTOR and WNT pathways in preclinical cancer stem cell cultures, adult and pediatric primary organoids, and orthotopic murine models with positive efficacy in combination with clinical standard of care.

cancer biology↗

bioRxiv2022

Genetic inactivation of the USP19 deubiquitinase regulates a-synuclein ubiquitination and inhibits accumulation of Lewy body like aggregates in mice

The USP19 deubiquitinase is found in a locus associated with Parkinsons Disease (PD), interacts with heat shock proteins and promotes secretion of a-synuclein (a-syn) through the misfolding associated protein secretion (MAPS) pathway. Since these processes might modulate the processing of a-syn aggregates during the progression of PD, we tested the effect of USP19 knockout (KO) in mice expressing the A53T mutation of a-syn and in whom a-syn preformed fibrils (PFF) had been injected in the striatum. Compared to WT, KO brains showed decreased accumulation of phospho-synuclein (pSyn) positive aggregates. The improved pathology was associated with less activation of microglia, higher levels of synaptic marker proteins and improved performance in a tail suspension test. Exposure of primary neurons from WT and KO mice to PFF in vitro also led to decreased accumulation of pSyn aggregates. KO did not affect uptake of PFF in the cultured neurons. It also did not affect the propagation of aggregates as assessed by exposing WT or KO neurons to PFF and measuring pSyn positive aggregates in non-exposed adjacent neurons separated using a microfluidics device. We conclude that USP19 instead modulates intracellular dynamics of aggregates. Indeed, at the early time following PFF injection when the number of pSyn positive neurons were similar in WT and KO brains, the KO neurons contained less aggregates. KO brain aggregates stained more intensely with anti-ubiquitin antibodies. Immunoprecipitation of soluble proteins from primary neurons exposed to PFF with antibodies to ubiquitin or pSyn showed higher levels of ubiquitinated a-syn oligomeric species in the KO neurons. We propose that the improved pathology in USP19 KO brains may arise from decreased formation or enhanced clearance of the more ubiquitinated aggregates and/or enhanced disassembly towards more soluble oligomeric species. USP19 inhibition may represent a novel therapeutic approach that targets the intracellular dynamics of a-syn complexes.

neuroscience↗