Abstract B18: Quantification of BTK engagement by ibrutinib in peripheral blood mononuclear cells in a phase I clinical study.
Authors: Chaturvedi Shalini , Davis Cuc , Chang Stella , Sirisawad Mint , Jong Jan de , Chang Betty , Younes Anas , Fourneau Nele , Balasubramanian Sriram , Hall Brett 
Published: August 2015

Introduction: Bruton's tyrosine kinase (BTK) is a TEC-family receptor tyrosine kinase essential for B-cell receptor signaling and is critical for the growth and survival of several types of B-cell malignancies. Ibrutinib (PCI-32765) is a selective, oral, small molecule inhibitor that binds covalently to the active site of BTK (Cys481). Ibrutinib has been designated as a breakthrough targeted therapy for certain types of B-cell malignancies and is approved by the US Food and Drug Administration for patients with previously treated mantle cell lymphoma and chronic lymphocytic leukemia. As ibrutinib is being tested in various malignancies and multiple patient populations, it is important to understand its target engagement at different doses. Therefore, a novel, sensitive, and robust assay was developed to study target engagement on the Meso Scale Discovery (MSD) platform and used to study BTK occupancy in a Phase 1 clinical study of ibrutinib in combination with R-CHOP in B-cell Non-Hodgkin lymphoma patients (DBL1002; NCT01569750) which is described in detail elsewhere (Younes et al ASH 2013).

Description: This target engagement assay determines the relative amount of free unbound BTK by using a probe. The BTK molecules that are not already bound to ibrutinib bind to the probe while the remaining sample is washed off. Free BTK is assessed using an anti-BTK antibody, and total signal is detected using the MSD Sector Imager 6000 as relative light units (RLU).

Summary of Results: Final results in RLU are inversely related to free BTK. The assay was developed and successfully validated to interrogate PBMC lysates as the matrix of choice, using the BTK standard as a reference. The assay was used to test PBMC samples from the clinical study DBL1002. PBMC samples collected from patients (N=9) treated at the recommended dose of 560mg were measured prior to and 4 hrs after the first dose of ibrutinib. The median value for Btk occupancy at pre-treatment was 825.357 pg/mL, while 4 hours after the first dose of ibrutinib the median value dropped to 11.25 pg/mL, indicating full occupancy of BTK.

Conclusions: Treatment with ibrutinib at a dose of 560mg in combination with R-CHOP led to complete inhibition of BTK 4 hours after the first dose of ibrutnib. These results were consistent with previous results from single agent trials using the gel-based assay for BTK occupancy, indicating that the additional chemoimmunotherapy did not interfere with ibrutinib binding to its target BTK.

Citation Format: Shalini Chaturvedi, Cuc Davis, Stella Chang, Mint Sirisawad, Jan de Jong, Betty Chang, Anas Younes, Nele Fourneau, Sriram Balasubramanian, Brett Hall. Quantification of BTK engagement by ibrutinib in peripheral blood mononuclear cells in a phase I clinical study. [abstract]. In: Proceedings of the AACR Special Conference on Hematologic Malignancies: Translating Discoveries to Novel Therapies; Sep 20-23, 2014; Philadelphia, PA. Philadelphia (PA): AACR; Clin Cancer Res 2015;21(17 Suppl):Abstract nr B18.