Measurement of ECM stiffness and liver cancer radiosensitivity using shear- wave-based elasticity in millimeter-sized 3D culture models

Abstract

Abstract Background The stiffness of the tumor microenvironment (TME) directly influences cellular behaviors. Radiotherapy (RT) is a common treatment for solid tumors, but the TME can impact its efficacy. In the case of liver cancer, clinical observations have shown that tumors within a cirrhotic, stiffer background respond less to RT, suggesting that the extracellular matrix (ECM) stiffness plays a critical role in the development of radioresistance. Methods This study explored the effects of ECM stiffness and the inhibition of lysyl oxidase (LOX) isoenzymes on the radiosensitivity of liver cancer in a millimeter-sized three-dimensional (3D) culture. We constructed a cube-shaped ECM-based millimeter-sized hydrogel containing Huh7 human liver cancer cells. By modulating the collagen concentration, we produced two groups of samples with different ECM stiffnesses to mimic the clinical scenarios of normal and cirrhotic livers. We used a single-transducer system for shear-wave-based elasticity measurement, to derive Young’s modulus of the 3D cell culture to investigate how the ECM stiffness affects radiosensitivity. This is the first demonstration of a workflow for assessing radiosensitivity in a millimeter-sized 3D culture. Results A higher ECM stiffness was associated with decreased radiosensitivity. Moreover, sonoporation-aided LOX inhibition with BAPN (β-aminopropionitrile monofumarate) significantly decreased the initial ECM stiffness and increased RT-induced cell death. Inhibiting LOX reduced the ECM stiffness, particularly when the ECM was stiffer. Importantly, combining LOX inhibition with RT sensitized cirrhotic liver cancer cells to irradiation-induced DNA damage, leading to a significant increase in radiosensitivity. Furthermore, LOX inhibition can be combined with sonoporation to overcome stiffness-related radioresistance, potentially leading to better treatment outcomes for patients with liver cancer. Conclusions This study reveals the critical role of ECM stiffness in liver cancer radioresistance. Sonoporation-aided LOX inhibition emerges as a promising strategy to mitigate stiffness-related resistance, offering potential improvements in liver cancer treatment outcomes.