Abstract 4028: High-dose short-exposure of osimertinib robustly inhibits growth of patient-derived metastatic colorectal cancer organoids
Authors: Iyer Kirti K. , Poel Dennis , Miggelenbrink Anne , Kerkhof Wouter , van den Hombergh Erik , de Jong Loek A. , van Erp Nielka P. , Tauriello Daniele V. , Verheul Henk M. 
Published: April 2023

Abstract
Despite major interest in tyrosine kinase inhibitors (TKIs) as a treatment option for metastatic colorectal cancer (mCRC), almost all TKIs tested for mCRC fail in early-phase clinical trials. Although showing specific target inhibition at low concentrations, TKIs have a much broader kinase inhibitory potency at higher concentrations. Alternative high dose regimens have been proposed to explore if efficacy can be improved with acceptable toxicity. We used 3D matrix-embedded tumor organoids as a preclinical platform to determine optimal drug exposure, i.e. preclinical pharmacology, and to dissect the mechanisms of action to potentially convert the dismal translational success rate of TKIs for mCRC. We established patient-derived tumor organoids (PDTOs) from mCRC biopsies and, based on favorable physicochemical and pharmacokinetic properties, selected 3 TKIs (sunitinib, cediranib and osimertinib). Following standard IC50 assessment using continuous dosing with a concentration range, we investigated the cytotoxic antitumor effect of high-dose, short-exposure (HDSE) treatment. Five PDTOs were exposed to 20 µM TKI for 1-24h, washed and given normal medium, and PDTO-outgrowth was determined 1 week later. At exposures of 1, 3 and 6h, we measured intra-tumoroid TKI concentrations using a clinically validated LC/MS-MS method. PDTO cell death was observed using live-cell microscopy, and quantified by both caspase 3/7 enzyme activity assay and cleaved caspase-3 immunofluorescent staining. While PDTOs could be categorized for their sensitivity across tested TKIs, all were highly sensitive for osimertinib (IC50 values 0.40-3.8 µM). Lower sensitivity was observed for sunitinib (2.0-10.5 µM) and cediranib (2.5-7.1 µM). Only for osimertinib exposure to 20 µM for 3h was sufficient to block proliferation in all PDTOs. Interestingly, peak intra-tumoroid TKI concentration measurements across PDTOs revealed marked cellular accumulation, indicating an expanded potential for target inhibition. The concentrations correlated with sensitivity: for sunitinib from 1.5 mM for the most sensitive PDTO to 0.72 mM for the least sensitive PDTO. Likewise, the corresponding cediranib concentrations were 0.15 mM vs 0.062 mM. All PDTOs had high intra-tumoroid osimertinib concentrations (0.90-1.6 mM). Lastly, we detected a significant increase in apoptosis after 3h of HDSE with osimertinib. Whereas our HDSE regimen shows promising results for all 3 TKIs, very short exposure with high-dose osimertinib effectively reduces proliferation and induces cell death in all mCRC PDTOs. While this is likely due to high intra-tumoroid concentrations reached by osimertinib—the mechanism of action at these concentrations remains unknown and is subject to further studies. In parallel, we propose that a HDSE osimertinib regimen warrants clinical exploration as a potential new treatment option for mCRC.
Citation Format: Kirti K. Iyer, Dennis Poel, Anne Miggelenbrink, Wouter Kerkhof, Erik van den Hombergh, Loek A. de Jong, Nielka P. van Erp, Daniele V. Tauriello, Henk M. Verheul. High-dose short-exposure of osimertinib robustly inhibits growth of patient-derived metastatic colorectal cancer organoids. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4028.