Molecular Engineering of Activatable NIR-II Hemicyanine Reporters for Early Diagnosis and Prognostic Assessment of Inflammatory Bowel Disease

Molecular imaging in the second near-infrared window (NIR-II) provides high-fidelity visualization of biopathological events in deep tissue. However, most NIR-II probes produce "always-on" output and demonstrate poor signal specificity toward biomarkers. Herein, we report a series of hemicyanine reporters (HBCs) with tunable emission to NIR-II window (715-1188 nm) and structurally amenable to constructing activatable probes. Such manipulation of emission wavelengths relies on rational molecular engineering by integrating benz[c,d]indolium, benzo[b]xanthonium, and thiophene moieties to a conventional hemicyanine skeleton. In particular, HBC4 and HBC5 possess bright and record long emission over 1050 nm, enabling improved tissue penetration depth and superior signal to background ratio for intestinal tract mapping than NIR-I fluorophore HC1. An activatable inflammatory reporter (AIR-PE) is further constructed for pH-triggered site-specific release in colon. Due to minimized background interference, oral gavage of AIR-PE allows clear delineation of irritated intestines and assessment of therapeutic responses in a mouse model of inflammatory bowel disease (IBD) through real-time NIRF-II imaging. Benefiting from its high fecal clearance efficiency (>90%), AIR-PE can also detect IBD and evaluate the effectiveness of colitis treatments via in vitro optical fecalysis, which outperforms typical clinical assays including fecal occult blood testing and histological examination. This study thus presents NIR-II molecular scaffolds that are not only applicable to developing versatile activatable probes for early diagnosis and prognostic monitoring of deeply seated diseases but also hold promise for future clinical translations.