Off-Peak Near-Infrared-II (NIR-II) Bioimaging of an Immunoconjugate Having Peak Fluorescence Emission in the NIR-I Spectral Region for Improving Tumor Margin Delineation.

The primary treatment for malignant tumors remains to be resection. The strongest predictor of recurrence and postoperative prognosis is whether diseased tissue/cells remain(s) at the surgical margin. Cancer surgery entails surgeons having the capability to visually distinguish between subtle shades of color in attempts of differentiating between diseased tissue and healthy tissue under standard white-light illumination, as such tissue states appear identical at the meso-/macroscopic level. Accordingly, enhancing the capability of surgeons to do so such that they can accurately delineate the tumor margin is of paramount importance. Fluorescence-guided surgery facilitates in enhancing such capability by color-coding the surgical field with overlaid contrasting pseudo-colors from real-time intraoperative fluorescence emission via utilizing fluorescent constructs in tandem. Constructs undergoing clinical trials or that are FDA-approved provide peak fluorescence emission in the visible (405 - 700 nm) or near-infrared-I (NIR-I) spectral region (700-900 nm), whereby differentiation between tissue states progressively improves in sync with using constructs that emit longer wavelengths of light. Here, we repurpose the usage of such fluorescent constructs by establishing feasibility of a tumor-targeting immunoconjugate (cetuximab-IRDye800) having peak fluorescence emission at the NIR-I spectral region to provide improved tumor margin delineation by affording higher tumor-to-background ratios (TBRs) when measuring its off-peak fluorescence emission at the near-infrared-II (NIR-II) spectral region (1000-1700 nm) in in vivo applications. We prepared murine tumor models, administered such immunoconjugate, and imaged such models pre-/post-administration via utilizing imaging systems that separately afforded acquisition of fluorescence emission in the NIR-I or NIR-II spectral region. On doing so, we determined in vivo TBRs, ex vivo TBRs with/-out skin, and ex vivo biodistribution, all via measuring the fluorescence emission of the immunoconjugate at tumor site(s) at both spectral regions. Collectively, we established feasibility of using the immunoconjugate to afford improved tumor margin delineation by providing 2-fold higher TBRs via utilizing the NIR-II spectral region to capture off-peak fluorescence emission from a fluorescent construct having NIR-I peak fluorescence emission.