Abstract 5223: Differentiated receptor pharmacology of imipridone ONC201: The first DRD2 antagonist in clinical development for oncology
Authors: Madhukar Neel , Prabhu Varun Vijay , Anantharaman Lakshmi , Sulli Chidananda , Davidson Edgar , Deacon Sean , Tarapore Rohinton , Rucker Joseph , Charter Neil , Doranz Banjamin , Oster Wolfgang , Elemento Olivier , Allen Joshua 
Published: July 2017

Abstract
ONC201 is the founding member of the imipridone class of compounds which selectively target G protein-coupled receptors (GPCRs). ONC201, currently in clinical trials, possesses an exceptional safety profile combined with anti-cancer effects that are driven by activation of the integrated stress response and inhibition of Ras signaling. In this study, we identified and characterized the previously unknown binding target of ONC201.
Following its phenotypic discovery, a series of experiments indicated that ONC201 does not directly interact with many known cancer drug targets. BANDIT - a machine learning based drug target identification platform - predicted that ONC201 selectively antagonizes the GPCR dopamine receptor D2 (DRD2). DRD2 is overexpressed in many cancers, controls various pro-survival mechanisms including Ras signaling and stress pathways, and its antagonism causes anti-proliferative and pro-apoptotic effects in malignant cells.
PathHunter® β-Arrestin and cAMP assays determined that ONC201 selectively antagonizes DRD2 and DRD3. Antipsychotics antagonize multiple dopamine receptor family members that belong to either the D1-like or D2-like subfamilies that cause opposing downstream effects. Consistent with BANDIT, in contrast to antipsychotics, ONC201 did not antagonize other dopamine receptors or other GPCRs with known ligands. Schild analysis and radioligand competition assays revealed a DRD2 affinity of ~3uM, consistent with ONC201 anticancer activity. In accordance with superior selectivity of ONC201 among the GPCR superfamily, ONC201 exhibited a wide therapeutic window in tumor versus normal cell viability assays compared to antipsychotics. In support of the hypothesis that selectively targeting D2-like receptors yields superior anti-cancer efficacy, combined DRD2/DRD1 inhibition with tool compounds was inferior to DRD2 inhibition alone.
Further characterization revealed that ONC201 had a very slow association rate for DRD2 relative to antipsychotics, whereas the dissociation rate was similar to atypical antipsychotics that are well tolerated. Shotgun mutagenesis alanine scan mapping across 350 amino acids of DRD2 identified 8 residues critical for ONC201-mediated antagonism of dopamine-induced calcium flux. Several of these residues were not conserved among the dopamine receptor family and the residue with the largest effect is not conserved in any other family member, suggesting the basis of ONC201 specificity. Consistent with competitive inhibition, several residues were within the orthosteric binding site, however, two allosteric residues were also identified.
In summary, ONC201 is the first DRD2 antagonist under clinical development for oncology and its differentiated receptor pharmacology explains its unique selectivity, anti-cancer activity, and safety that has been observed in clinical trials.
Citation Format: Neel Madhukar, Varun Vijay Prabhu, Lakshmi Anantharaman, Chidananda Sulli, Edgar Davidson, Sean Deacon, Rohinton Tarapore, Joseph Rucker, Neil Charter, Banjamin Doranz, Wolfgang Oster, Olivier Elemento, Joshua Allen. Differentiated receptor pharmacology of imipridone ONC201: The first DRD2 antagonist in clinical development for oncology [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5223. doi:10.1158/1538-7445.AM2017-5223