Three-dimensional cell culture has revolutionized cellular biology research and opened the door to novel discoveries in terms of cellular behavior and response to microenvironment stimuli. Different types of 3D culture exist today, including hydrogel scaffold-based models, which possess a complex structure mimicking the extracellular matrix. These hydrogels can be made of polymers (natural or synthetic) or low-molecular weight gelators that, via the supramolecular assembly of molecules, allow the production of a reproducible hydrogel with tunable mechanical properties. When cancer cells are grown in this type of hydrogel, they develop into multicellular tumor spheroids (MCTS). Three-dimensional (3D) cancer culture combined with a complex microenvironment that consists of a platform to study tumor development and also to assess the toxicity of physico-chemical entities such as ions, molecules or particles. With the emergence of nanoparticles of different origins and natures, implementing a reproducible in vitro model that consists of a bio-indicator for nano-toxicity assays is inevitable. However, the maneuver process of such a bio-indicator requires the implementation of a repeatable system that undergoes an exhaustive follow-up. Hence, the biggest challenge in this matter is the reproducibility of the MCTS and the associated full-scale characterization of this system’s components.

Biocompatible materials are of paramount importance in numerous fields. Unlike chemically-bridge polymer-based hydrogels, low molecular weight gelators can form a reversible hydrogel as their structure rely on non-covalent interaction. Although many applications with this type of hydrogels can be envisioned, we still lack their understanding due to the complexity of their self-assembly process and the difficulty predicting their behaviors (transition temperature, gelation kinetic, impact of solvent…). In this study, we extend the investigations of a series of nucleoside-derivatives gelators which only differ by subtle chemical modifications. Using a multi-technique approach, we determined their thermodynamic and kinetic features at various scale (molecular to macro) in different conditions. Monitored at supramolecular level by circular dichroism as well as macroscopic scales by rheology and turbidimetry, we found out that sol-gel and gel-sol transition are greatly depending on the concentration and on the mechanisms that are probed. Self-assembly kinetic depends on hydrogel molecules and is modulated by temperature and solvent. This fundamental study provides insight on the impact of some parameters on the gelation process, such as concentration, cooling rate and nature of the solvent.