Tunable 3D Collagen-Based Scaffolds for Biophysical Tumour Microenvironment Studies

The complexity of studying a neoplastic disease relies on understanding specialized cell types within the tumor tissue, which can be recruited and ‘corrupted’ to create a dynamic pro-tumorigenic network called the ‘tumor microenvironment’ (TME). Although the TME is of critical importance during initiation and spread of cancer, relatively little is known about its biophysical evolution during tumor development and progression. In this study a 3D collagen type I–based scaffold model cross-linked with 1,4-butanediol diglycidyl ether (BDDGE) was employed to mimic mechanical changes occurring in normal tissue (2 kPa - soft, So) and advanced cancer tissue (12 kPa - stiff, St) and monitor how these biophysical cues affect the stromal tumor compartment. Viability assays, migration patterns and matrix remodeling together with RNA sequencing investigated cancer-associated fibroblasts (CAFs) response to TME stiffness. In the model, CAFs fail to remodel St scaffolds, showing lower migration and increased cell circularity compared to cells grown on So scaffolds. This behavior is reflected in gene expression profiles, showing an upregulation of DNA replication, DNA repair and chromosome organization gene clusters, with a concommitant loss of their ability to remodel and deposit extracellular matrix after culture on St scaffolds. Soft scaffolds can reproduce biophysically-meaningful microenvironements for tumour early stages investigations, while St scaffolds can better mimic mechanical cues occurring in advanced cancer stages. These results not only establish the need for tunable and affordable 3D scaffolds as effective platforms for cancer research but also reveal the contribution of microenvironment biomechanics in regulating gene expression changes in the stromal tumor tissue compartment.