Orpinolide disrupts a leukemic dependency on cholesterol transport by inhibiting the oxysterol-binding protein OSBP

Abstract

Metabolic alterations in cancer precipitate in associated dependencies that can be therapeutically exploited. To meet this goal, natural product inspired small molecules can provide a resource of invaluable chemotypes. Here, we identify orpinolide, a synthetic withanolide analog with pronounced anti-leukemic properties via orthogonal chemical screening. Through multi-omics profiling and genome-scale CRISPR/Cas9 screens, we identify that orpinolide disrupts Golgi homeostasis via a mechanism that requires active phosphatidylinositol 4-phosphate (PI4P) signaling at the endoplasmic reticulum (ER)- Golgi membrane interface. Thermal proteome profiling and genetic validation studies reveal the oxysterol-binding protein OSBP as the direct and phenotypically relevant target of orpinolide. Collectively, these data reaffirm sterol transport as a therapeutically actionable dependency in leukemia and motivate ensuing translational investigation via the probe-like compound orpinolide.

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

“…Following this, we aimed to determine target engagement of OSBP and related ORPs in a cellular setting. For this, a cellular thermal shift assay (CETSA) in a split nanoluciferase (HiBiT-LgBiT) system in KBM7 cells overexpressing HiBiT-tagged OSBP, as well as the closely related ORP4L and ORPs 2 and 9 was carried out 49 . Consistently with the biochemical data, oxybipin-1 and oxybipin-2 exhibited a significant stabilization effect on OSBP with ∆T > 5 °C while C5 only exhibited an insignificant stabilization effect on OSBP with ∆T ~ 1 °C, which further supports the interaction between the oxybipins and OSBP in live cells.…”