Abstract
Abstract The quantification of Target Occupancy (TO) in cells after application of an inhibitor is an important parameter to correlate the biochemical features of a drug with its in vivo efficacy. Experimental evidence of TO is a robust support of the target hypothesis and helps to identify optimal dosing of the drug. The sensitive correlation spectroscopy methods (FCS and FCCS) have been used successfully in the past to analyze drug- target interactions. The technology is fast, specific and allows detection of drug target interactions in smallest sample volumes. Here we adopted the FCCS method to quantify the occupancy of the XPO1 target in cells and tumors treated with the Selective Inhibitor of Nuclear Export (SINE) compound Selinexor (KPT-330), currently in advanced clinical trials in patients with hematological and solid cancers. We used a fluorescently labeled tracer (Leptomycin B -Cy5; LMB-Cy5) to quantify the loading of XPO1 with Selinexor and confirmed target specificity of the signal using a fluorescent labeled antibody detecting native XPO1 protein. The amount of Antibody-XPO1-tracer complex was determined by FCCS. Occupancy of XPO1 target with Selinexor decreases the amount of tracer-target complexes in a dose and time dependent manner. Target occupancy studies of XPO1 were carried out in different cancer relevant cell lines and xenograft tumors retrieved from treated animals. As such the approach can be applied to study target occupancy in vitro and in vivo. Citation Format: Frank Becker, Kerrin Hansen, Marsha Crochiere, Yosef Landesmann, Stefan Hannus. Application of Fluorescence Correlation Spectroscopy as a novel tool to quantify target occupancy in cells and tumor tissue [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr LB-B09.
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