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cells (CSC) from human epithelial tissues and culturing them within an ECM gel .
Under optimized conditions, these cells self-assemble into 3D organoid microstructures,

which recapitulate key architectural and functional elements of the human TME 12–14 .
The integration of organoid culture with microfluidic technology has created a

transformative 'organoid-on-a-chip' platform that significantly advances in vitro

modeling 15 . By enabling precise spatiotemporal control over the cellular

microenvironment through dynamic perfusion and regulation of oxygen, nutrients, and

mechanical forces, this combined approach overcomes limitations of traditional 3D

cultures while better replicating in vivo complexity 16,17 . Microfluidic systems facilitate

high-throughput parallel culturing of organoids with automated monitoring,

revolutionizing drug screening and personalized medicine applications through patient-

derived models. The technology allows reconstruction of sophisticated tissue interfaces

by incorporating vascular networks, immune cells, and stromal components, thereby

faithfully mimicking TMEs and tissue barriers 18–20 . With capabilities for gradient-

based drug testing and enhanced physiological relevance, this next-generation platform

shows tremendous potential for cancer research, regenerative medicine, and toxicology.
Future developments in 3D bioprinting, vascularization, and AI-assisted analysis

promise to further refine these models, potentially reducing animal testing while

accelerating translational research with unprecedented precision. (2) Organ-on-a-

Chip: Micro-engineered systems replicating human organ function and disease states

under controlled conditions. This platform enables precise cellular microenvironment

control, faithfully mimicking in vivo tissue architectures and dynamic physiological

processes 21,22 . Compared to traditional animal models, organ-on-a-chip platforms

provide superior experimental simplicity and cost-efficiency—requiring significantly

less infrastructure, reducing ethical constraints, and enabling parallelized

experimentation—while achieving unprecedented fidelity in recapitulating human

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TME dynamics . These micro-engineered systems facilitate precise modeling of
multistep carcinogenesis: from initial tumor-stroma interactions and angiogenesis to

metastatic intravasation/extravasation, all under physiologically relevant mechanical

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