Genipin-Crosslinked, Silane-Anchored 3D Tumor-Stroma Microtissues for High-Content On-Chip Drug Testing.

Area of Science:

• Biomaterials Engineering
• Cancer Research
• Microfluidics

Background:

• Physiologically relevant 3D tumor models are crucial for understanding tumor progression and drug resistance.
• Existing models often face hydrogel instability and extracellular matrix (ECM) deformation in microfluidic systems, hindering long-term studies and imaging.
• Cancer-associated fibroblasts (CAFs) play a significant role in tumor microenvironments, necessitating their inclusion in advanced models.

Purpose of the Study:

• To develop a stable microfluidic co-culture platform for head and neck squamous cell carcinoma (HNSCC) that overcomes limitations of hydrogel contraction and ECM deformation.
• To enable physiologically relevant modeling of tumor-fibroblast interactions for studying drug resistance and progression.
• To provide a scalable tool for preclinical drug chemosensitivity screening and clinical translation.

Main Methods:

• A dual strategy involving APTES-mediated surface silanization to anchor the ECM and Genipin-based crosslinking to reinforce the hydrogel network.
• Utilizing Fourier-transform infrared spectroscopy (FTIR) to confirm collagen crosslinking and preserve reactive groups for bonding.
• Integrating semi-automated segmentation and high-content imaging for quantitative analysis of drug responses at single-cell and multicellular levels.
• Performing drug chemosensitivity assays with patient-derived CAFs to assess chemoprotective effects.

Main Results:

• The developed platform demonstrated enhanced stability and prevented ECM deformation in microfluidic formats.
• Genipin crosslinking modestly increased hydrogel stiffness and progressively reinforced the network without compromising cell viability, confirmed by rheology.
• FTIR analysis confirmed successful collagen crosslinking.
• The system enabled quantitative assessment of drug responses and clinically relevant chemoprotective effects of CAFs.

Conclusions:

• The microfluidic platform provides a reproducible and physiologically relevant model for studying tumor-fibroblast interactions in HNSCC.
• This engineered system overcomes key biomaterial limitations, offering a scalable solution for preclinical drug screening.
• The platform facilitates quantitative evaluation of drug efficacy and potential chemoprotective effects within the tumor microenvironment.