Dose intensification of TRAIL-inducing ONC201 inhibits metastasis and promotes intratumoral NK cell recruitment
Jessica Wagner, C. Leah Kline, Lanlan Zhou, Kerry S. Campbell, Alexander W. MacFarlane, Anthony J. Olszanski, Kathy Q. Cai, Harvey H. Hensley, Eric A. Ross, Marie D. Ralff, Andrew Zloza, Charles B. Chesson, Jenna H. Newman, Howard Kaufman, Joseph Bertino, Mark Stein, Wafik S. El-Deiry
Jessica Wagner, C. Leah Kline, Lanlan Zhou, Kerry S. Campbell, Alexander W. MacFarlane, Anthony J. Olszanski, Kathy Q. Cai, Harvey H. Hensley, Eric A. Ross, Marie D. Ralff, Andrew Zloza, Charles B. Chesson, Jenna H. Newman, Howard Kaufman, Joseph Bertino, Mark Stein, Wafik S. El-Deiry
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Research Article Oncology

Dose intensification of TRAIL-inducing ONC201 inhibits metastasis and promotes intratumoral NK cell recruitment

Abstract

ONC201 is a first-in-class, orally active antitumor agent that upregulates cytotoxic TRAIL pathway signaling in cancer cells. ONC201 has demonstrated safety and preliminary efficacy in a first-in-human trial in which patients were dosed every 3 weeks. We hypothesized that dose intensification of ONC201 may impact antitumor efficacy. We discovered that ONC201 exerts dose- and schedule-dependent effects on tumor progression and cell death signaling in vivo. With dose intensification, we note a potent anti-metastasis effect and inhibition of cancer cell migration and invasion. Our preclinical results prompted a change in ONC201 dosing in all open clinical trials. We observed accumulation of activated NK+ and CD3+ cells within ONC201-treated tumors and that NK cell depletion inhibits ONC201 efficacy in vivo, including against TRAIL/ONC201-resistant Bax–/– tumors. Immunocompetent NCR1-GFP mice, in which NK cells express GFP, demonstrated GFP+ NK cell infiltration of syngeneic MC38 colorectal tumors. Activation of primary human NK cells and increased degranulation occurred in response to ONC201. Coculture experiments identified a role for TRAIL in human NK-mediated antitumor cytotoxicity. Preclinical results indicate the potential utility for ONC201 plus anti–PD-1 therapy. We observed an increase in activated TRAIL-secreting NK cells in the peripheral blood of patients after ONC201 treatment. The results offer what we believe to be a unique pathway of immune stimulation for cancer therapy.

Authors

Jessica Wagner, C. Leah Kline, Lanlan Zhou, Kerry S. Campbell, Alexander W. MacFarlane, Anthony J. Olszanski, Kathy Q. Cai, Harvey H. Hensley, Eric A. Ross, Marie D. Ralff, Andrew Zloza, Charles B. Chesson, Jenna H. Newman, Howard Kaufman, Joseph Bertino, Mark Stein, Wafik S. El-Deiry

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Figure 7

ONC201-induced NK activation and accumulation play an important role in antitumor effect.

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ONC201-induced NK activation and accumulation play an important role in ...
(A) Final tumor volumes of Bax–/– (top) and CT26 (bottom) mice treated with 100 mg/kg ONC201 weekly for 4 weeks and GM1 every 5 days as prescribed. (B) NK cell enrichment by ONC201 and depletion by GM1 are confirmed with flow cytometry in the CT26-bearing mice. (C) Fluorescence microscopy from coculture of HCT116 p53–/––GFP and NK cells or NK media treated as indicated for 48 hours. (D) Quantitation of fluorescence microscopy. Multiplex analysis of (E) IP-10 and (F) IFN-α2a levels within conditioned media from ONC201-treated HCT116 cells. More cytokines were analyzed as indicated in Supplemental Figure 15. 10 μM ONC201, 2 mg/ml RIK-2. Green: GFP, blue: calcein AM; red: PI. Merge was performed by using ImageJ. Non-GFP blue+ cells are NK cells. Flow cytometry was performed on PI+ CD19–CD45+ cells. For mice, n = 10 for CT26 and Bax–/– GM1 studies. For cocultures, n = 3 and samples were run separately twice. For multiplex, n = 3 run in duplicate (6 samples total). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 compared with vehicle using 2-sided Wilcoxon’s rank-sum test; for multiplex ELISA study, 3 separate conditioned media samples per cohort were harvested and run in duplicate. The mean of each duplicate was taken and compared using a 2-sided Wilcoxon’s rank-sum test. Data represent mean ± SD.