3D tumor models are emerging as valuable tools for drug screening and nanoparticle based personalized cancer treatments. The main challenges in building microfluidic chip-based 3D tumor models currently include the development of bioinks with high bioactivity and the reproduction of the key tumor extracellular matrix (ECM) with heterogeneous tumor microenvironments. In this study, we designed a triangular multi-chamber tumor-on-a-chip (TM-CTC) platform, which consisted of three circular chambers at the vertices of a triangle connected by three rectangular chambers; it significantly improved the culture efficiency of 3D tumor tissues. MCF-7 tumor cells were cultured in a 3D ECM and then dynamically perfused for 7 days of culture to obtain abundant tumor spheroids with uniform size (100 ± 4.1 μm). The biological features of the 3D tumor tissue including epithelial transformation (EMT), hypoxia and proliferation activities were reproduced in the triangular multi-chamber tumor-on-a-chip (TM-CTC) platform. The permeability results of NPs confirmed that the ECM exhibited a significant barrier effect on the transportation of NPs when compared with free drugs, indicating that the ECM barrier should be considered as one of the key factors of drug delivery carrier development. In addition, this TM-CTC model provided a suitable platform for constructing a complex heterogeneous tumor microenvironment with multiple cells (MCF-7, HUVEC and MRC-5) involved, which was beneficial for exploring the dynamic interaction between tumor cells and other cells in the tumor microenvironment. The above results suggest that this TM-CTC model can simulate the dynamic transportation of NPs around 3D tumor tissues, and thus provide a reliable platform for NP evaluation.