We first describe the development of a scalable, bioinert hydrogel-based inverted colloidal crystal (BhiCC) framework designed for high-efficiency and uniform cancer spheroid formation as Disease models. This system utilizes uniformly sized alginate microgels assembled into a hexagonal-close-packed structure, facilitating the generation of over 1000 spheroids per well in a standard 96-well plate. The resulting framework supports real-time HCHTS through conventional plate readers and fluorescent microscopy, enhanced by a custom-designed AI-assisted automated data processing algorithm for spatial and temporal analysis of drug response in 3D. This platform provides a robust and reproducible model for applications in disease modeling, and drug discovery.
Leveraging the developed HCHTS cancer spheroid model, we further investigate the role of nanocarrier chirality in enhancing transport and therapeutic efficacy within 3D cellular constructs. Graphene quantum dots (GQDs) functionalized with L- or D-cysteine exhibit distinct diffusion profiles, with L-GQDs demonstrating a 1.7-fold higher apparent diffusion coefficient compared to D-GQDs in HepG2-derived spheroids. Additionally, doxorubicin (DOX)-loaded L-GQDs achieve 25% greater penetration and therapeutic efficacy within tumor-like spheroids relative to free DOX, underscoring the importance of chirality in optimizing nanocarrier-mediated drug delivery.
Additionally, we present a novel chiral GQD-based sEV-loading platform designed to enhance drug encapsulation efficiency in sEV-mediated delivery systems. By tuning ligand design and GQD size to achieve optimal chirality matching with the sEV lipid bilayer, the platform demonstrates high drug loading efficiencies of 66.3% for doxorubicin and 64.1% for siRNA. This approach offers a versatile and efficient strategy for the encapsulation and delivery of both hydrophobic and hydrophilic therapeutic agents.
Collectively, these studies demonstrate the potential of integrating bioinspired material engineering strategies to advance cancer therapeutics. The proposed systems, including the BhiCC framework, chiral nanocarriers, and sEV-based platforms, provide a comprehensive foundation for developing next-generation platforms for high-throughput drug screening and targeted delivery.