Abstract LB-038: Preclinical development and clinical validation of a whole blood pharmacodynamic marker assay for the BET bromodomain inhibitor ZEN-3694 in metastatic castration-resistant prostate cancer (mCRPC) patients

Detection of drug activity in patients is essential to confirm its mechanism of action, as well as to ensure proper target engagement at the selected dose to elicit optimal clinical activity. Pharmacodynamic (PD) markers are often developed to detect pharmacological responses and optimize drug dosing. Whole blood is an easily attainable and minimally invasive source of biological material to measure clinical activity of drugs. We designed, developed, and validated a whole blood PD marker assay to detect the activity of ZEN-3694, an orally available inhibitor of the bromodomain and extra-terminal (BET) domain family of proteins currently in phase I clinical trials in mCRPC (NCT02705469 and NCT02711956). Potential BET-specific PD markers were first identified via comparative microarray analysis using a PI3K inhibitor, a BET inhibitor, and a dual PI3K/BET inhibitor in an MV4-11 acute myeloid leukemia (AML) cell line. Further microarray analysis of subsequent in house data and published data of BET inhibitors from different chemical scaffolds in hematologic cell lines allowed us to develop a short list of ~20 candidate genes. Further testing was done by measuring the modulation of these PD markers by various Zenith BET inhibitors from different chemical scaffolds in a number of human cell lines derived from hematological cancers and solid tumors, as well as cryopreserved human peripheral blood mononuclear cells (PBMCs). In vivo validation was also done in whole blood obtained from xenograft mice, and cynomolgus monkeys that were dosed orally with ZEN-3694, as well as ex-vivo treated human blood derived from normal donors or patients diagnosed with either AML or diffuse large B cell lymphoma. There was also robust target engagement in tumors of mouse AML xenografts, making them suitable tumor PD markers. A quantitative real-time PCR assay was developed for human whole blood matrix with parameters defined based on the multiplex efficiency (85-115%), coefficient of correlation of the standard curve (R2>0.98), and dynamic copy number range (10-106). Assay validation testing demonstrated an inter-assay variability (operator/day/machine) of Citation Format: Laura Tsujikawa, Karen Norek, Cyrus Calosing, Sarah Attwell, Dean Gilham, Nimisha Sharma, Jennifer Tobin, Michelle Haager, Ravi Jahagirdar, Sanjay Lakhotia, Henrik C. Hansen, Eric Campeau. Preclinical development and clinical validation of a whole blood pharmacodynamic marker assay for the BET bromodomain inhibitor ZEN-3694 in metastatic castration-resistant prostate cancer (mCRPC) patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr LB-038. doi:10.1158/1538-7445.AM2017-LB-038