BackgroundThree-dimensional (3D) tumor microdevices are promising platform for biomimetic antitumor prediction and high-throughput chemotherapeutic screening and play crucial roles in the exploration of cancer-associated pharmaceutics and therapeutics. Traditional cell manipulation tools (e.g., non-adhesive surfaces and hanging drops) and recent microengineered systems (e.g., microfluidic chips and micropatterned array chips) have progressed in terms of microscale control, substantial tumor production, programmable drug combinations, and throughput analysis. However, establishing a facile 3D tumor microdevice to construct heterotypic tumor-microenvironmental profiles and for throughput, implementable, multi-instrument-compatible analysis of chemotherapies to advance consumer-grade tumor modelling tools is still being explored. ResultsIn this study, we present a facilely operated tumor-on-a-chip platform for massive production of heterotypic 3D tumors and diverse investigations of combinatorial chemotherapy screening. Large quantity of heterotypic tumor generation with high geometric controllability (size difference: 19.6 μm) and operational repeatability (n = 10) was achieved using simple-to-fabricate micropatterned chips. Multiple characteristics of solid tumors, including phenotypic gradients (viability and proliferation) and heterogeneous cellular compositions (multi-cell participation and stroma composition), were reproduced in heterotypic tumors, being more biomimetic than homotypic tumors. We completed the user-friendly analytical evaluation of individual and combinatorial drug therapies, and demonstrated the high applicability of the platform in biomimetic tumor-related large-scale manipulation and on-chip analysis, as well as its high compatibility for off-chip detection. The entire operative process during tumor production and chemotherapy only requires the routine and easy-to-master pipetting manipulation. SignificanceThe establishment of a biomimetic and easy-to-use 3D tumor platform and the large-scale screening-like evaluation of combinatorial chemotherapies based on the usage of the micropatterned chip was achieved in a user-friendly manner. This advancement has significant application potential in the fields of oncology, drug discovery, and tissue engineering, and is expected to be valuable for developing accessible and generalizable tumor-on-a-chip microsystems for exploring cancer therapies.