Real-time morphometric analysis of targeted therapy for neuroblastoma cells in monolayer and 3D hydrogels using digital holographic microscopy.

High-risk neuroblastoma (HR-NB) patient treatment is currently insufficient and challenging due to its high clinical, morphological, and genetic heterogeneity as well as the scarcity of available samples for research. We used a gelatin- and silk fibroin-based hydrogel system with cross-linked vitronectin (VN) as an artificial biomimetic three-dimensional (3D) environment to mirror aggressive neuroblastoma (NB) tumors and tested long-term cell response to Cilengitide (CLG). Based on our previous studies and others using the integrin inhibitor CLG as a potential mechanotherapy drug, we show that CLG caused cell detachment in monolayer cultures of MYCN-amplified SK-N-BE (2) and ALK-mutated SH-SY5Y human neuroblastoma cell lines. Cell detachment and aggregation were maintained in hydrogel-free monolayer cells whereas cells embedded in hydrogels presented different responses to treatment, suggesting differential anoikis resistance between the two cell lines. This underscores the advantages of testing therapeutic approaches using real-time imaging of tumor cells in 3D biomimetic models and its contribution to precision medicine.