A benefit of biomedical application of nanosystems is implementation of a precise effect at the level of an individual cell, and magnetic nanoparticles (MNPs) are some of the best candidates for the development of an intelligent nanosystem with remote control. To develop a nanosystem for precise therapy, a deep understanding of the nanosystem's in vivo behavior is required. Here, we studied penetration and distribution of PEGylated iron oxide MNPs unmodified or modified with the pH low insertion peptide (a ligand for smart targeting of the tumor acidic microenvironment) in vivo in a 4T1 mouse tumor. We revealed that MNPs penetrate into the tumor via both vascular burst and endothelial transcytosis. By implementing an approach based on single-cell high-throughput RNA sequencing, we identified the populations of the cells that took up MNPs in the 4T1 tumor and revealed preferential accumulation of MNPs in regulatory Trem2+ tumor-associated macrophages. © 2024