Microfluidic bioreactor for 3D Cell Culture and Drug Screening of Hepatocellular CarcinomaTumoroids

Hepatocellular carcinoma (HCC) represents the second largest contributor to cancer mortality worldwide. Currently, there is no curative treatment for advanced stage HCC, so development of new therapies is urgently required. However, testing new drugs in standard 2D cell culture does not reflect the complex structure of a 3D tumor. Therefore, the objective of this project is to develop a novel microfluidic bioreactor capable of cultivating 3D tumoroids, as in vitro models, for drug screening research. We aim to mimic the in vivo situation of tumor treatment with biocompatible materials and continuous perfusion of culture medium and drugs.

Microsystem technology was used for the microfabrication of the microbioreactor with technologies such as photolithography, Deep Reactive Ion Etching, Self-Assembled Monolayer, Replica Molding, and Oxygen plasma bonding.
We designed a capture system for the tumoroids within the microfluidic bioreactor consisting of micro pillars which also allows continuous supply with medium and drugs. Computational FluidDynamics was used to understand the effect of parameters such as flow velocity and stress. During the experiments, we observed that micropillars size, shape and configuration, are important factors for their stability and function when increasing microflow pressure and the design was optimized accordingly. In order to avoid cell attachment to the bottom surface, we cultured HepG2, Hep3B and Huh-7 cell lines on polydimethylsiloxane. We confirmed biocompatibility and HepG2 cells were successfully grown as spherical tumoroids, which has remained healthy and viable for more than one week in the microbioreactor.

Furthermore, our design enables in situ viability assays and continuous monitoring for days by using calcein staining followed by fluorescent microscopy.
Thereby, we have developed a novel microfluidic bioreactor for HCC tumoroids culture and drug experimentation which could collaborate with the fight against cancer.