Hoock, J. G. F. | Search bioRxiv

bioRxiv2023

Identification of non-conventional small molecule degraders and stabilizers of squalene synthase

Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=184 SRC="FIGDIR/small/543387v1_figu1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@1645e05org.highwire.dtl.DTLVardef@58ad70org.highwire.dtl.DTLVardef@1e2c57aorg.highwire.dtl.DTLVardef@112fcbd_HPS_FORMAT_FIGEXP M_FIG C_FIG Squalene synthase (SQS) is an essential enzyme in the mevalonate pathway whose abundance and activity control cholesterol biosynthesis and homeostasis. Although catalytic inhibitors of SQS have been developed to attenuate cholesterol, none so far have been approved for therapeutic use. Herein we sought to develop SQS degraders using targeted protein degradation (TPD) as an approach to lower overall cellular cholesterol content. We found that KY02111, a small molecule ligand of SQS, could selectively cause SQS to degrade in a proteasome-dependent manner. In contrast, compounds based on the same scaffold linked to E3 ligase recruiting ligands led to SQS stabilization. Whole cell proteomic analysis found KY02111 to reduce only the levels of SQS, while lipidomic analysis determined that KY02111 treatment concomitantly reduced cellular cholesteryl ester content. SQS stabilizers were shown to shield SQS from its natural turnover without recruiting their matching E3 ligase. Our work shows that degradation of SQS is possible despite a challenging biological setting and lays the groundwork for future development of either SQS degrading or stabilizing probes.

cell biology↗

bioRxiv2023

Selective inhibition of OSBP blocks retrograde trafficking by inducing partial Golgi degradation

Sterol-binding proteins are important regulators of lipid homeostasis and membrane integrity; however, the discovery of selective small molecule modulators can be challenging due to structural similarities in the sterol binding domains. We report the discovery of highly potent and selective inhibitors of oxysterol binding protein (OSBP), which we term oxybipins. Sterol-containing chemical chimeras aimed at identifying new sterol binding proteins by targeted degradation, led to a significant reduction in Golgi-associated proteins. The degradation was found to occur at lysosomes, concomitant with changes in general protein glycosylation, indicating that the degradation of Golgi proteins was a downstream effect. By establishing a sterol transport protein biophysical assay panel, we discovered that the oxybipins potently inhibited OSBP, resulting in blockage of retrograde trafficking and attenuating Shiga toxin toxicity. As the oxybipins do not target any other sterol transporters tested, we advocate their use as chemical tools to study OSBP function and therapeutic relevance.

cell biology↗