The intratumoral accumulation of nanomedicine has been considered a passive process, referred to as the enhanced permeability and retention effect. Recent studies have suggested that the tumor uptake of nanomedicines follows an energy-dependent pathway rather than being a passive process. Herein, to explore the factor candidates that are associated with nanomedicine tumor uptake, a molecular marker identification platform is developed by integrating microscopic fluorescence images of a nanomedicine distribution with spatial transcriptomics information. When this approach is applied to PEGylated liposomes, molecular markers related to hypoxia, glycolysis, and apoptosis can be identified as being related to the intratumoral distribution of the nanomedicine. It is expected that the method can be applied to explain the distribution of a wide range of nanomedicines and that the data obtained from this analysis can enhance the precise utilization of nanomedicines.