Analysis of chemosensitivity of tumor spheroids exposed to two-dimensional gradient of combination drugs in a hydrogel-based diffusion microfluidic platform

Background: Liver cancer stands as a leading cause of cancer-related deaths globally, challenging conventional treatments due to resistance to chemotherapy and targeted therapy. Although frontline medications show initial efficacy, prolonged use often leads to resistance and harm. Current clinical strategies rely on combination therapies, but evaluating their effectiveness remains challenging. Results: To address this, we developed a hydrogel-based diffusion microfluidic platform for assessing chemosensitivity. This platform features a hydrogel-filled diffusion layer linked to liquid wells, allowing the creation of drug gradients. Tumor spheroids, cultured on the non-adhesive hydrogel surface, were exposed to single or combination drug gradients. Analysis revealed that drug efficacy, quantified by IC₅₀ values, could be determined from responses to single drug gradients. A novel method was introduced to assess spheroid circularity as a time-invariant index of drug efficacy. Furthermore, exposing spheroids to 2D combination drug gradients allowed intuitive visualization of their responses via a color map. This analysis identified optimal drug combinations, exhibiting superior efficacy to monotherapy. Significance: The microfluidic platform enables assessment of synergistic effects and replicates in vivo conditions, enhancing the relevance of test results. By offering a streamlined, fast, and efficient drug screening approach, this platform aims to provide insight into tumor spheroid responses to varying drug combinations, facilitating more effective clinical applications.