Real‐Time In Vivo Visualization of Tumor‐Associated Macrophage Reprogramming Using a Nitric Oxide‐Activatable NIR‐II Nanoinducer

ABSTRACT Tumor‐associated macrophages (TAMs) are key regulators of the tumor microenvironment (TME). They typically adopt an M2‐like phenotype that promotes tumor progression by providing survival signals, suppressing anti‐tumor immunity, and facilitating pre‐metastatic niche formation. Reprogramming TAMs toward an anti‐tumor phenotype has emerged as a promising therapeutic strategy, with the repolarization of M2‐like TAMs into an M1‐like phenotype being central to this approach. Here, a nitric oxide (NO)‐activatable near‐infrared‐II (NIR‐II) fluorescence/photoacoustic nanoinducer (I/E@M2pep) that selectively targets M2‐like TAMs and reprograms them toward an M1‐like phenotype, thereby enhancing anti‐tumor efficacy is reported. In this construct, the M2pep peptide enables M2‐like TAM targeting, IPI549 reprograms them toward an M1‐like phenotype while inducing NO production, and the NO‐activatable NIR‐II probe (ETNO) allows for in vivo visualization of macrophage repolarization via NIR‐II fluorescence/photoacoustic imaging. In a mouse breast cancer model, intravenous administration of I/E@M2pep produced a ratiometric NIR‐II photoacoustic signal change that correlated with M2‐to‐M1 repolarization. Furthermore, combining this nanoinducer with a CD47 monoclonal antibody markedly enhanced anti‐tumor immunity through M1 macrophage‐mediated tumor killing and TME remodeling. This work presents an effective in vivo strategy that simultaneously facilitates and visualizes TAM repolarization, holding promise for broader applications in studying tumor initiation, metastasis, and treatment response.