An in vivo platform to jointly monitor cellular and metabolic responses to chemotherapy.

Drug responses in tumors are shaped by local interactions among cancer cells, immune cells and metabolites, but these relationships are difficult to measure in intact tissue. We developed an in vivo spatial pharmacology platform that combines localized intratumoral drug delivery with paired cyclic immunofluorescence and MALDI imaging mass spectrometry. Applied to MMTV-PyMT mammary tumors, the platform profiled 1.48 million cells and 230,812 metabolic imaging spots across 27 tumor sections and multiple treatment conditions. Metabolic profiles captured histological structure and predicted immune cellular neighborhoods, with strongest performance for myeloid-enriched regions. Integrated analysis revealed two opposing immune-associated metabolic states: a PUFA-rich, CSF1R-high macrophage niche and a purine-catabolic, MPO-high inflammatory niche associated with tumor cell death. Local therapeutic perturbation shifted the balance between these states, distinguishing regions of immune recruitment from regions of drug-induced tumor cell death. This framework enables causal, spatially resolved measurement of drug-induced immunometabolic remodeling in intact tumors.