NIR-II fluorescence imaging-guided colorectal cancer surgery targeting CEACAM5 by a nanobody

BACKGROUND: Surgery is the cornerstone of colorectal cancer (CRC) treatment, yet complete removal of the tumour remains a challenge. The second near-infrared window (NIR-II, 1000–1700 nm) fluorescent molecular imaging is a novel technique, which has broad application prospects in tumour surgical navigation. We aimed to evaluate the ability of CEACAM5-targeted probe for CRC recognition and the value of NIR-II imaging-guided CRC resection. METHODS: We constructed the probe 2D5-IRDye800CW by conjugated anti-CEACAM5 nanobody (2D5) with near-infrared fluorescent dye IRDye800CW. The performance and benefits of 2D5-IRDye800CW at NIR-II were confirmed by imaging experiments in mouse vascular and capillary phantom. Then mouse colorectal cancer subcutaneous tumour model (n = 15), orthotopic model (n = 15), and peritoneal metastasis model (n = 10) were constructed to investigate biodistribution of probe and imaging differences between NIR-I and NIR-II in vivo, and then tumour resection was guided by NIR-II fluorescence. Fresh human colorectal cancer specimens were incubated with 2D5-IRDye800CW to verify its specific targeting ability. FINDINGS: 2D5-IRDye800CW had an NIR-II fluorescence signal extending to 1600 nm and bound specifically to CEACAM5 with an affinity of 2.29 nM. In vivo imaging, 2D5-IRDye800CW accumulated rapidly in tumour (15 min) and could specifically identify orthotopic colorectal cancer and peritoneal metastases. All tumours were resected under NIR-II fluorescence guidance, even smaller than 2 mm tumours were detected, and NIR-II had a higher tumour-to-background ratio than NIR-I (2.55 ± 0.38, 1.94 ± 0.20, respectively). 2D5-IRDye800CW could precisely identify CEACAM5-positive human colorectal cancer tissue. INTERPRETATION: 2D5-IRDye800CW combined with NIR-II fluorescence has translational potential as an aid to improve R0 surgery of colorectal cancer. FUNDINGS: This study was supported by Beijing Natural Science Foundation (JQ19027), the 10.13039/501100012166National Key Research and Development Program of China (2017YFA0205200), 10.13039/501100001809National Natural Science Foundation of China (NSFC) (61971442, 62027901, 81930053, 92059207, 81227901, 82102236), 10.13039/501100004826Beijing Natural Science Foundation (L222054), CAS Youth Interdisciplinary Team (JCTD-2021-08), the Strategic Priority Research Program of the 10.13039/501100002367Chinese Academy of Sciences (XDA16021200), the Zhuhai High-level Health Personnel Team Project (Zhuhai HLHPTP201703), the 10.13039/501100012226Fundamental Research Funds for the Central Universities (JKF-YG-22-B005) and Capital Clinical Characteristic Application Research (Z181100001718178). The authors would like to acknowledge the instrumental and technical support of the multi-modal biomedical imaging experimental platform, Institute of Automation, Chinese Academy of Sciences.