CombiCTx: Screening diffusion gradients of anti-cancer drug combinations

Abstract

The reduced effectiveness of chemotherapy in many patients highlights the need for novel drug combinations that target drug resistance mechanisms contributing to tumor survival. Dynamic conditions within the tumor microenvironment influence the response to anti-cancer drugs. Accordingly, identifying effective drug concentrations and interactions (additive, synergistic, or antagonistic) in relevant tumor tissue models will inform new treatment combinations. To address this need for combinatorial chemotherapeutic (CTx) screening assays, we have developed a new assay called CombiCTx, which uses a device with three reservoirs containing gels loaded with anti-cancer drugs.The drug-loaded device is inverted and placed in a standard culture dish above cancer cells, and both are then enclosed in gel. Drugs diffuse from the reservoirs and expose cancer cells to overlapping dynamic drug gradients. We imaged diffusion of the anti-cancer drug doxorubicin in the assay using timelapse microscopy, and established an imaging protocol for quantifying MDA-MB-231 breast cancer cell survival responses along drug gradients. Finally, evaluating combination effects of navitoclax and gemcitabine in the CombiCTx revealed potent, but localized effects of navitoclax, attributed to limited diffusion, while gemcitabine seemed to diffuse readily throughout the assay and revealed a mild synergy in navitoclax regions. These data demonstrate the capacity of CombiCTx to evaluate the cytotoxic effect of anti-cancer drug combinations while accounting for drug diffusion differences, which is relevant in the context of the 3D tumor environment and may thereby help inform clinical treatment strategies.