Program: Oral and Poster Abstracts
Session: 604. Molecular Pharmacology and Drug Resistance in Myeloid Diseases: Poster III
Tumor necrosis factor related apoptosis inducing ligand (TRAIL) can initiate apoptosis through interaction with its death receptors. TRAIL has also shown selectivity for tumor cells over normal cells, prompting interest as an anticancer agent. ONC201 is a small molecule that was first identified as a TRAIL-inducing agent and is the founding member of the imipridone family of anticancer agents that selectively target G protein-coupled receptors. Its downstream mechanism of action involves induction of the integrated stress response pathway and inactivation of the AKT and ERK signaling pathways. Interestingly, these pathways are constitutively active in a majority of AML patients. Preclinical studies show that ONC201 induces apoptosis in LICs and shows promising clinical activity. In this study, we investigated the effects of a novel ONC201 analog, ONC213, in preclinical AML models.
To begin, we assessed ONC213 sensitivity in AML cell lines and primary AML patient samples. ONC213 sensitivity, as measured by MTT assays, varied among the AML cell lines (n=9) and primary AML patient samples (n=24), with IC50s ranging from 91.7 nM to 626.1 nM in the AML cell lines and 150 nM to 2.4 µM in the primary AML patient samples (median IC50 was 445 nM). While ONC213 showed anti-proliferative effects in the nanomolar range, annexin V/propidium iodide staining and flow cytometry analysis revealed heterogeneous responses in 9 AML cell lines and 15 primary AML patient samples treated with ONC213 for 48 h. Striking induction of apoptosis (about 72% after treatment with 500 nM for 48 h) was observed in MOLM-13 and MV4-11 cells, while little to no induction of cell death (about 6-11% after treatment with 2 µM ONC213 for 48 h) occurred in THP-1 and U937 cells. However, prolonged exposure of THP-1 and U937 cells which showed little to no response to the 48 h treatment, showed robust induction of apoptosis following 120 h ONC213 treatment (about 50-70% after treatment with 2 µM for 120 h).
Next, we tested the effects of ONC213 on LICs. Primary AML patient samples were treated with ONC213 for 48 h and then plated in methylcellulose. The number of surviving AML cells after 2 weeks revealed significant reduction in the number of AML cells capable of generating leukemia cell colonies (AML-CFUs). Treatment with 500 nM decreased the number AML-CFUs to less than 5% compared to vehicle control in 6 primary patient samples. Normal human hematopoietic stem/progenitor cells (CD34+ cells) were treated in the same manner and showed that ONC213 had no significant effect on colony formation, suggesting that ONC213 spares normal hematopoietic cells. To begin to assess in vivo efficacy, MOLM-13 cells were injected into immunocompromised NSG mice through the tail vein. Two days post-injection the mice were randomly placed into vehicle control or 50 mg/kg ONC213 cohorts (7 mice per cohort). The mice were treated on a daily schedule for 2 weeks. The mice were sacrificed 2 days after the last treatment. Vehicle-treated control mice had significantly larger spleens than ONC213 treated mice (p = 0.007) and showed a trend of higher AML cell engraftment (p = 0.094), as assessed by anti-CD45 antibody staining and flow cytometry analysis, compared to ONC213 treated mice. ONC213 treatment was not associated with body weight loss. Preliminary mechanistic studies suggest that cell death induced by ONC213 could be associated with downregulation of Mcl-1 and inhibition of AKT in AML cell lines. Since the in vitro data shows that ONC213 has activity against LICs and bulk leukemia cells, further molecular studies to understand the heterogeneity in responsiveness to ONC213 are warranted. Taken together, our preclinical studies strongly support further development of ONC213 as a potential therapeutic agent for the treatment of AML.
Disclosures: Allen: Oncoceutics: Employment, Equity Ownership, Patents & Royalties. Stogniew: Oncoceutics: Employment. Prabhu: Oncoceutics: Employment, Equity Ownership.
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