Abstract
Understanding the pharmacokinetic/pharmacodynamic (PK/PD)-relationship of a drug candidate is key to determine effective, yet safe treatment regimens for patients. However, current testing strategies are inefficient in characterizing in vivo responses to fluctuating drug concentrations during multi-day treatment cycles. Methods based on animal models are resource-intensive and require time, while traditional in vitro cell-culturing methods usually do not provide temporally-resolved information on the effects of in vivo–like drug exposure scenarios. To address this issue, we developed a microfluidic system to 1) culture arrays of three-dimensional spheroids in vitro, to 2) apply specific dynamic drug exposure profiles, and to 3) in-situ analyze spheroid growth and the invoked drug effects in 3D by means of 2-photon microscopy at tissue and single-cell level. Spheroids of fluorescently-labeled T-47D breast cancer cells were monitored under perfusion-culture conditions at short time intervals over three days and exposed to either three 24 h-PK-cycles or a dose-matched constant concentration of the phosphatidylinositol 3-kinase inhibitor BYL719. While the overall efficacy of the two treatment regimens was similar, spheroids exposed to the PK profile displayed cycle-dependent oscillations between regression and regrowth. Spheroids treated with a constant BYL719 concentration regressed at a steady, albeit slower rate. At a single-cell level, the cell density in BYL719-treated spheroids oscillated in a concentration-dependent manner. Our system represents a versatile tool for in-depth preclinical characterization of PK/PD parameters, as it enables an evaluation of drug efficacy and/or toxicity under realistic exposure conditions.
Highlights
Understanding the relationship between drug exposure and the biological response is a key element in pharmaceutical drug discovery (Tuntland et al, 2014)
This was achieved by modifying the microfluidic chip by computer numerical controlled (CNC) micro milling and by developing a metal frame that enabled connection of the chip liquid ports to pressure pumps
We developed a microfluidic microtissue culturing system that enables the exposure of arrays of spheroids to pharmacokinetic compound-concentration profiles, while monitoring them optically in 3D at high resolution
Summary
Understanding the relationship between drug exposure (pharmacokinetics; PK) and the biological response (pharmacodynamics; PD) is a key element in pharmaceutical drug discovery (Tuntland et al, 2014). Despite this thorough preclinical characterization, 9 out of 10 potential drug candidates fail in clinical trials, 75% of which are due to PK/PD-related toxicity or a lack of efficacy (Khalil et al, 2020). These numbers illustrate the need for more predictive preclinical methods that allow for translation of in vitro exposure-response-relationships to drug-dosing regimens in patients (Lave et al, 2016). Due to the systemic insights that animal models provide, animal-based, preclinical characterization of PK/PD properties of a drug is and will remain a prerequisite to identify effective/safe dosing regimens and to start clinical studies on patients (Prantil-Baun et al, 2018). A major shortcoming of such in vivo experiments, are, the readout methods, which cannot continuously provide information on the effects of dynamic PK concentration changes in target tissues
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