Dose Optimization of ClpP Agonists Using an In Vitro Microfluidic Perfusion Platform and In Silico...

Small molecule activators of the mitochondrial caseinolytic protease P (ClpP agonists) can disrupt tumor metabolism and deprive tumors of their energy needs. The imipridone, ONC201, is a ClpP agonist currently undergoing clinical evaluation across multiple cancer types, while additional analogs with improved potency and selectivity are in preclinical development. Preclinical studies in mice have demonstrated a unique pharmacokinetic-pharmacodynamic (PK-PD) relationship for ONC201 characterized by prolonged pharmacology following a single dose. This motivated the selection of an initial human dosing regimen of every three weeks, and subsequent dose exploration studies in mice led to dose intensification in human patients. However, a systematic analysis of ClpP agonist PK-PD relationships has not been performed, and the optimal exposure profile for ClpP agonists remains undefined. To address this gap, we combined PK-PD modeling with a microfluidic perfusion platform as an animal-alternative approach for translational PK-PD of ClpP agonists. We demonstrate that the anti-proliferative effect on triple negative breast cancer cells correlates with the magnitude and duration of ClpP agonist exposure above a threshold concentration required for ClpP activation. Moreover, we demonstrate that PK-PD model simulations using parameters derived from microfluidic perfusion datasets can successfully predict the anti-tumor efficacy of a ClpP agonist in a mouse tumor xenograft study. These studies support the translational relevance of the animal-alternative in vitro PK-PD platform and its utility to help guide dose optimization of ClpP agonists as cancer therapeutics.