a Novel Indocyanine Green-Conjugated Cetuximab for Targeted Photoimmunotherapy in Colorectal Cancer
Indocyanine green (ICG) labeled antibodies have shown a promising effect on intraoperative cancer imaging. However, conventional ICG-conjugated antibodies suffer from heterogeneous labeling-due to random conjugation sites, or suboptimal fluorescence quantum yield, thereby limiting their clinical translatability. In this study, we synthesized a chemically modified ICG derivative bearing disulfonate and maleimide groups (ICG-disulfo-Mal), and engineered a modified Cetuximab-ICG conjugate (mCet-ICG) via Michael addition by linking the maleimide group of ICG-disulfo-Mal to free thiol groups of reduced Cetuximab. We demonstrate that mCet-ICG enables high-contrast fluorescent detection of colorectal cancer (CRC) in in vivo models, achieving an optimal tumor-to-background ratio (TBR) of 8-10 that remains stable for up to 5 days. Moreover, compared with free ICG, mCet-ICG exhibits significantly superior performance in detecting lymph node metastasis, with an improved sensitivity (100% vs. 63.2%), specificity (88.5% vs. 55.4%), and overall accuracy (92.7% vs. 56.5%), as confirmed by histopathological assessment. Consistent with these findings, the tissue enrichment specificity of mCet-ICG in human CRC is significantly higher than ICG (0.82 vs 0.33, P<0.01), as demonstrated in clinical CRC specimens. Notably, we revealed that mCet-ICG elicited a robust immune-activating effect characterized by enhanced T-cell activation and intratumoral infiltration upon irradiation with an 808nm laser, leading to pronounced immunogenic cell death (ICD) and significant tumor regression. In conclusion, the mCet-ICG holds dual potential as a precise intraoperative tumor tracer for real-time surgical guidance and as a potent photosensitizer for photoimmunotherapy.