TMIC-14. A novel IDO1 PROTAC decreases immunosuppressive extracellular IDO1 levels

Abstract BACKGROUND Indoleamine 2,3-dioxygenase 1 (IDO1) is an immunosuppressive enzyme expressed in >90% of patient-resected glioblastoma (GBM). IDO1 expression is inversely correlated with GBM patient survival. Extracellular IDO1 contributes to IDO1-mediated immunosuppression. We developed a novel protein degrader/PROTAC (NU227326) with a 4.7 nM DC50 for human IDO1 and compared its effects on extracellular IDO1 with a potent IDO1 enzyme inhibitor (BMS-986205; IC50=1.7nM) in cultured human GBM cells and in mice with human GBM. METHODS U87 or GBM43 were treated with NU227326, BMS-986205, ±human IFNg in vitro. Native PAGE, Western blotting, ELISA, and mass spectrometry evaluated IDO1 in the whole cell lysate, microvesicles, exosomes, and the vesicle-depleted extracellular fraction (VDF). NSG mice with intracranial GBM were intraperitoneally treated with IDO1 PROTAC and evaluated for intratumoral IDO1 protein levels. RESULTS BMS-986205 treatment led to multiple novel IDO1 bands in the VDF under Native PAGE conditions as compared to untreated- or NU227326 treated-GBM cells. BMS-986205 treatment dramatically increased extracellular IDO1 protein levels as compared to untreated- or NU227326 treated-GBM cells. Mass spec. identified multiple differences between BMS-986205- and NU227326-treated GBM cells. Systemic IDO1 PROTAC treatment led to a significant decrease of intra-GBM IDO1 protein in vivo. CONCLUSIONS Pharmacological IDO1 enzyme inhibitor treatment increases both intracellular and extracellular IDO1 protein levels. A novel IDO1 protein degrader decreases both intracellular and extracellular IDO1 protein levels. Ongoing studies aim to understand IDO1 PROTAC effects on the anti-GBM immune response in humanized mice with intracranial human GBM PDX and with a human immune system.