ONC201 and imipridones: Anti-cancer compounds with clinical efficacy.

ONC201 was originally discovered as TNF-Related Apoptosis Inducing Ligand (TRAIL)-inducing compound TIC10. ONC201 appears to act as a selective antagonist of the G protein coupled receptor (GPCR) dopamine receptor D2 (DRD2), and as an allosteric agonist of mitochondrial protease caseinolytic protease P (ClpP). Downstream of target engagement, ONC201 activates the ATF4/CHOP-mediated integrated stress response leading to TRAIL/Death Receptor 5 (DR5) activation, inhibits oxidative phosphorylation via c-myc, and inactivates Akt/ERK signaling in tumor cells. This typically results in DR5/TRAIL-mediated apoptosis of tumor cells; however, DR5/TRAIL-independent apoptosis, cell cycle arrest, or antiproliferative effects also occur. The effects of ONC201 extend beyond bulk tumor cells to include cancer stem cells, cancer associated fibroblasts and immune cells within the tumor microenvironment that can contribute to its efficacy. ONC201 is orally administered, crosses the intact blood brain barrier, and is under evaluation in clinical trials in patients with advanced solid tumors and hematological malignancies. ONC201 has single agent clinical activity in tumor types that are enriched for DRD2 and/or ClpP expression including specific subtypes of high-grade glioma, endometrial cancer, prostate cancer, mantle cell lymphoma, and adrenal tumors. Synergy with radiation, chemotherapy, targeted therapy and immune-checkpoint agents has been identified in preclinical models and is being evaluated in clinical trials. Structure-activity relationships based on the core pharmacophore of ONC201, termed the imipridone scaffold, revealed novel potent compounds that are being developed. Imipridones represent a novel approach to therapeutically target previously undruggable GPCRs, ClpP, and innate immune pathways in oncology.

Biological activity of weekly ONC201 in adult recurrent glioblastoma patients.

BACKGROUND: ONC201 is a dopamine receptor D2 (DRD2) antagonist that penetrates the blood-brain barrier. ONC201 efficacy has been shown in glioblastoma animal models and is inversely correlated with dopamine receptor DRD5 expression. ONC201 is well tolerated in adult recurrent glioblastoma patients with dosing every 3 weeks and has achieved an objective radiographic response in a patient harboring the H3 K27M mutation. METHODS: In a window-of-opportunity arm, 6 adult subjects initiated ONC201 prior to re-resection of recurrent glioblastoma with intratumoral concentrations as the primary endpoint. An additional 20 adults with recurrent glioblastoma received single agent weekly oral ONC201 at 625 mg, with progression-free survival at 6 months (PFS6) by Response Assessment in Neuro-Oncology (RANO) criteria as the primary endpoint. RESULTS: The window-of-opportunity arm achieved its primary endpoint with intratumoral ONC201 concentrations at ~24 hours following the second weekly dose ranging from 600 nM to 9.3 µM. Intratumoral pharmacodynamics assessed by activating transcriptional factor 4, death receptor 5, and apoptosis induction relative to archival samples were observed with the strongest intensity and uniformity among patients with low DRD5 tumor expression. The primary endpoint of PFS6 by RANO was not achieved at 5% in this molecularly unselected cohort; however, 1 of 3 patients enrolled with the H3 K27M mutation had a complete regression of enhancing multifocal lesions that remained durable for >1.5 years. No treatment modifications or discontinuations due to toxicity were observed, including in those who underwent re-resection. CONCLUSIONS: Weekly ONC201 is well tolerated, and meaningful intratumoral concentrations were achieved. ONC201 may be biologically active in a subset of adult patients with recurrent glioblastoma.

A Bayesian machine learning approach for drug target identification using diverse data types.

Drug target identification is a crucial step in development, yet is also among the most complex. To address this, we develop BANDIT, a Bayesian machine-learning approach that integrates multiple data types to predict drug binding targets. Integrating public data, BANDIT benchmarked a ~90% accuracy on 2000+ small molecules. Applied to 14,000+ compounds without known targets, BANDIT generated ~4,000 previously unknown molecule-target predictions. From this set we validate 14 novel microtubule inhibitors, including 3 with activity on resistant cancer cells. We applied BANDIT to ONC201-an anti-cancer compound in clinical development whose target had remained elusive. We identified and validated DRD2 as ONC201's target, and this information is now being used for precise clinical trial design. Finally, BANDIT identifies connections between different drug classes, elucidating previously unexplained clinical observations and suggesting new drug repositioning opportunities. Overall, BANDIT represents an efficient and accurate platform to accelerate drug discovery and direct clinical application.

Pediatric and adult H3 K27M-mutant diffuse midline glioma treated with the selective DRD2 antagonist ONC201.

BACKGROUND: H3 K27M-mutant diffuse midline glioma is a fatal malignancy with no proven medical therapies. The entity predominantly occurs in children and young adults. ONC201 is a small molecule selective antagonist of dopamine receptor D2/3 (DRD2/3) with an exceptional safety profile. Following up on a durable response in the first H3 K27M-mutant diffuse midline glioma patient who received ONC201 (NCT02525692), an expanded access program was initiated. METHODS: Patients with H3 K27M-mutant gliomas who received at least prior radiation were eligible. Patients with leptomeningeal spread were excluded. All patients received open-label ONC201 orally once every week. Safety, radiographic assessments, and overall survival were regularly assessed at least every 8 weeks by investigators. As of August 2018, a total of 18 patients with H3 K27M-mutant diffuse midline glioma or DIPG were enrolled to single patient expanded access ONC201 protocols. Among the 18 patients: seven adult (> 20 years old) and seven pediatric (< 20 years old) patients initiated ONC201 with recurrent disease and four pediatric patients initiated ONC201 following radiation, but prior to disease recurrence. FINDINGS: Among the 14 patients with recurrent disease prior to initiation of ONC201, median progression-free survival is 14 weeks and median overall survival is 17 weeks. Three adults among the 14 recurrent patients remain on treatment progression-free with a median follow up of 49.6 (range 41-76.1) weeks. Among the 4 pediatric patients who initiated adjuvant ONC201 following radiation, two DIPG patients remain progression-free for at least 53 and 81 weeks. Radiographic regressions, including a complete response, were reported by investigators in a subset of patients with thalamic and pontine gliomas, along with improvements in disease-associated neurological symptoms. INTERPRETATION: The clinical outcomes and radiographic responses in these patients provide the preliminary, and initial clinical proof-of-concept for targeting H3 K27M-mutant diffuse midline glioma with ONC201, regardless of age or location, providing rationale for robust clinical testing of the agent.

Imipridone ONC212 activates orphan G protein-coupled receptor GPR132 and integrated stress response in acute myeloid leukemia.

Imipridones constitute a novel class of antitumor agents. Here, we report that a second-generation imipridone, ONC212, possesses highly increased antitumor activity compared to the first-generation compound ONC201. In vitro studies using human acute myeloid leukemia (AML) cell lines, primary AML, and normal bone marrow (BM) samples demonstrate that ONC212 exerts prominent apoptogenic effects in AML, but not in normal BM cells, suggesting potential clinical utility. Imipridones putatively engage G protein-coupled receptors (GPCRs) and/or trigger an integrated stress response in hematopoietic tumor cells. Comprehensive GPCR screening identified ONC212 as activator of an orphan GPCR GPR132 and Gαq signaling, which functions as a tumor suppressor. Heterozygous knock-out of GPR132 decreased the antileukemic effects of ONC212. ONC212 induced apoptogenic effects through the induction of an integrated stress response, and reduced MCL-1 expression, a known resistance factor for BCL-2 inhibition by ABT-199. Oral administration of ONC212 inhibited AML growth in vivo and improved overall survival in xenografted mice. Moreover, ONC212 abrogated the engraftment capacity of patient-derived AML cells in an NSG PDX model, suggesting potential eradication of AML initiating cells, and was highly synergistic in combination with ABT-199. Collectively, our results suggest ONC212 as a novel therapeutic agent for AML.

Safety and enhanced immunostimulatory activity of the DRD2 antagonist ONC201 in advanced solid tumor patients with weekly oral administration.

BACKGROUND: ONC201 is a small molecule antagonist of DRD2, a G protein-coupled receptor overexpressed in several malignancies, that has prolonged antitumor efficacy and immunomodulatory properties in preclinical models. The first-in-human trial of ONC201 previously established a recommended phase II dose (RP2D) of 625 mg once every three weeks. Here, we report the results of a phase I study that evaluated the safety, pharmacokinetics (PK), and pharmacodynamics (PD) of weekly ONC201. METHODS: Patients ≥ 18 years old with an advanced solid tumor refractory to standard treatment were enrolled. Dose escalation proceeded with a 3 + 3 design from 375 mg to 625 mg of ONC201. One cycle, also the dose-limiting toxicity (DLT) window, was 21 days. The primary endpoint was to determine the RP2D of weekly ONC201, which was confirmed in an 11-patient dose expansion cohort. RESULTS: Twenty patients were enrolled: three at 375 mg and 17 at 625 mg of ONC201. The RP2D was defined as 625 mg with no DLT, treatment discontinuation, or dose modifications due to drug-related toxicity. PK profiles were consistent with every-three-week dosing and similar between the first and fourth dose. Serum prolactin and caspase-cleaved cytokeratin-18 induction were detected, along with intratumoral integrated stress response activation and infiltration of granzyme B+ Natural Killer cells. Induction of immune cytokines and effectors was higher in patients who received ONC201 once weekly versus once every three weeks. Stable disease of > 6 months was observed in several prostate and endometrial cancer patients. CONCLUSIONS: Weekly, oral ONC201 is well-tolerated and results in enhanced immunostimulatory activity that warrants further investigation. TRIAL REGISTRATION: NCT02250781 (Oral ONC201 in Treating Patients With Advanced Solid Tumors), NCT02324621 (Continuation of Oral ONC201 in Treating Patients With Advanced Solid Tumors).

Mitochondrial ClpP-Mediated Proteolysis Induces Selective Cancer Cell Lethality.

The mitochondrial caseinolytic protease P (ClpP) plays a central role in mitochondrial protein quality control by degrading misfolded proteins. Using genetic and chemical approaches, we showed that hyperactivation of the protease selectively kills cancer cells, independently of p53 status, by selective degradation of its respiratory chain protein substrates and disrupts mitochondrial structure and function, while it does not affect non-malignant cells. We identified imipridones as potent activators of ClpP. Through biochemical studies and crystallography, we show that imipridones bind ClpP non-covalently and induce proteolysis by diverse structural changes. Imipridones are presently in clinical trials. Our findings suggest a general concept of inducing cancer cell lethality through activation of mitochondrial proteolysis.

Preclinical evaluation of the imipridone family, analogs of clinical stage anti-cancer small molecule ONC201, reveals potent anti-cancer effects of ONC212.

Anti-cancer small molecule ONC201 upregulates the integrated stress response (ISR) and acts as a dual inactivator of Akt/ERK, leading to TRAIL gene activation. ONC201 is under investigation in multiple clinical trials to treat patients with cancer. Given the unique imipridone core chemical structure of ONC201, we synthesized a series of analogs to identify additional compounds with distinct therapeutic properties. Several imipridones with a broad range of in vitro potencies were identified in an exploration of chemical derivatives. Based on in vitro potency in human cancer cell lines and lack of toxicity to normal human fibroblasts, imipridones ONC206 and ONC212 were prioritized for further study. Both analogs inhibited colony formation, and induced apoptosis and downstream signaling that involves the integrated stress response and Akt/ERK, similar to ONC201. Compared to ONC201, ONC206 demonstrated improved inhibition of cell migration while ONC212 exhibited rapid kinetics of activity. ONC212 was further tested in >1000 human cancer cell lines in vitro and evaluated for safety and anti-tumor efficacy in vivo. ONC212 exhibited broad-spectrum efficacy at nanomolar concentrations across solid tumors and hematological malignancies. Skin cancer emerged as a tumor type with improved efficacy relative to ONC201. Orally administered ONC212 displayed potent anti-tumor effects in vivo, a broad therapeutic window and a favorable PK profile. ONC212 was efficacious in vivo in BRAF V600E melanoma models that are less sensitive to ONC201. Based on these findings, ONC212 warrants further development as a drug candidate. It is clear that therapeutic utility extends beyond ONC201 to include additional imipridones.

First-in-Human Clinical Trial of Oral ONC201 in Patients with Refractory Solid Tumors.

Purpose: ONC201 is a small-molecule selective antagonist of the G protein-coupled receptor DRD2 that is the founding member of the imipridone class of compounds. A first-in-human phase I study of ONC201 was conducted to determine its recommended phase II dose (RP2D).Experimental Design: This open-label study treated 10 patients during dose escalation with histologically confirmed advanced solid tumors. Patients received ONC201 orally once every 3 weeks, defined as one cycle, at doses from 125 to 625 mg using an accelerated titration design. An additional 18 patients were treated at the RP2D in an expansion phase to collect additional safety, pharmacokinetic, and pharmacodynamic information.Results: No grade >1 drug-related adverse events occurred, and the RP2D was defined as 625 mg. Pharmacokinetic analysis revealed a Cmax of 1.5 to 7.5 µg/mL (∼3.9-19.4 µmol/L), mean half-life of 11.3 hours, and mean AUC of 37.7 h·µg/L. Pharmacodynamic assays demonstrated induction of caspase-cleaved keratin 18 and prolactin as serum biomarkers of apoptosis and DRD2 antagonism, respectively. No objective responses by RECIST were achieved; however, radiographic regression of several individual metastatic lesions was observed along with prolonged stable disease (>9 cycles) in prostate and endometrial cancer patients.Conclusions: ONC201 is a selective DRD2 antagonist that is well tolerated, achieves micromolar plasma concentrations, and is biologically active in advanced cancer patients when orally administered at 625 mg every 3 weeks. Clin Cancer Res; 23(15); 4163-9. ©2017 AACR.

Discovery and clinical introduction of first-in-class imipridone ONC201.

ONC201 is the founding member of a novel class of anti-cancer compounds called imipridones that is currently in Phase II clinical trials in multiple advanced cancers. Since the discovery of ONC201 as a p53-independent inducer of TRAIL gene transcription, preclinical studies have determined that ONC201 has anti-proliferative and pro-apoptotic effects against a broad range of tumor cells but not normal cells. The mechanism of action of ONC201 involves engagement of PERK-independent activation of the integrated stress response, leading to tumor upregulation of DR5 and dual Akt/ERK inactivation, and consequent Foxo3a activation leading to upregulation of the death ligand TRAIL. ONC201 is orally active with infrequent dosing in animals models, causes sustained pharmacodynamic effects, and is not genotoxic. The first-in-human clinical trial of ONC201 in advanced aggressive refractory solid tumors confirmed that ONC201 is exceptionally well-tolerated and established the recommended phase II dose of 625 mg administered orally every three weeks defined by drug exposure comparable to efficacious levels in preclinical models. Clinical trials are evaluating the single agent efficacy of ONC201 in multiple solid tumors and hematological malignancies and exploring alternative dosing regimens. In addition, chemical analogs that have shown promise in other oncology indications are in pre-clinical development. In summary, the imipridone family that comprises ONC201 and its chemical analogs represent a new class of anti-cancer therapy with a unique mechanism of action being translated in ongoing clinical trials.