High‐Throughput Digital Decoding of Vascular Heterogeneity in Patient‐Specific Tumor Microenvironments

Quantitative characterization of vascular heterogeneity in complex microphysiological systems (MPS), particularly within patient‐derived tumor microenvironments, remains a major challenge for scalable disease modeling and therapeutic evaluation. Existing analysis approaches primarily rely on vessel abundance‐based morphometrics and often fail to resolve network connectivity and spatial remodeling in heterogeneous vascular systems. Here, we present the iMAP platform, a high‐throughput digital vascular profiling framework that integrates an injection‐molded microfluidic chip with an interactive image analysis tool (iMAP Analyzer). This platform enables topology‐resolved and region‐aware quantification of vascular architecture, capturing vessel morphology, branching complexity, connectivity, and spatial variation between tumor‐proximal and distal regions. Applied to 3D co‐cultures of patient‐derived gastric cancer tumor spheroids with either donor‐matched iPSC‐derived endothelial cells (iPSC‐ECs) or primary HUVECs, iMAP identified differences in network organization and spatial stability under standardized conditions, with iPSC‐derived networks exhibiting increased fragmentation and peripheral instability. These findings demonstrate that connectivity‐normalized and region‐resolved analysis provides critical insight beyond conventional whole‐image metrics. The iMAP framework offers a scalable and standardized approach for quantitative vascular phenotyping in complex MPS, supporting high‐content analysis and advancing the development of vascularized in vitro disease models.