A 3D-Bioprinted Hydrogel Platform with Tunable Matrix Stiffness Reveals Mechanical Adaptation and Doxorubicin Resistance in Triple-Negative Breast Cancer

Abstract

Triple-negative breast cancer (TNBC) progression and therapeutic response are strongly influenced by biomechanical cues within the tumor microenvironment. However, conventional animal models and two-dimensional (2D) cultures do not faithfully recapitulate the mechanical diversity of potential metastatic tissues. Here, we developed a digital light processing (DLP)-based 3D-bioprinted hydrogel platform using gelatin methacryloyl (GelMA) or its blend with poly(ethylene glycol) dimethacrylate (PEGDM) based bioinks to generate cell-laden constructs with compressive moduli representative of soft (6.7 ± 2.5 kPa) or stiff (43.8 ± 18.4 kPa) tissues. The printed hydrogels exhibited good shape fidelity and supported molecular diffusion, as well as long-term cell viability and proliferation. MDA-MB-231 cells displayed stiffness-dependent phenotypes, adopting a spindle-like morphology and showing a 3.8-fold increase in metabolic activity in soft hydrogels by Day 14. In contrast, cells in stiff hydrogels remained predominantly spherical with minimal metabolic change over the same period. Notably, cells recovered from 3D scaffolds exhibited higher elastic moduli than cells cultured in 2D, indicating matrix-dependent mechanical adaptation. When treated with 1 μM Doxorubicin for 48 h at Day 14, cells in soft hydrogels showed 45% cell death, whereas those in stiff hydrogels showed only 7% cell death. These findings demonstrate that matrix stiffness and culture duration jointly regulate TNBC cell state, mechanics, and chemotherapeutic response. Overall, this 3D-bioprinted platform provides a biomaterials-based in vitro model for investigating TNBC mechanobiology and for developing more predictive drug-screening systems.