Preclinical study on a newly developed near-infrared fluorescence endoscopy system using molecular targeted fluorescence antibodies - Towards instant detection of (pre)malignant colorectal lesions

Abstract

Introduction and aim: Colonoscopy is considered to be the most sensitive tool to detect colorectal (pre)malignant lesions, including small and flat lesions. Nevertheless, detection of such lesions is still incomplete, giving rise to interval cancers, particularly in patients at high risk for developing colorectal cancer (CRC), like people with Lynch syndrome (LS). To improve the detection of (pre)malignant lesions, near-infrared (NIR) fluorescence endoscopy guided by molecular targeted fluorescent labeled tracers may hold great promise. Therefore, we developed a combined white-light and near-infrared (WLNI) fluorescence endoscopy system, as well as three fluorescent labeled antibodies targeted to vascular endothelial growth factor A (VEGF-A), epithelial growth factor receptor (EGFR) and human epidermal growth factor receptor type 2 (HER-2). These molecular targets are clinically relevant since VEGF-A and EGFR, but not HER-2, are shown to be highly upregulated in (pre)malignant lesions of LS patients. In the present study we validated the newly developed WLNI fluorescence endoscopy system, together with intravenously injected NIR fluorescence antibodies, in human CRC mouse models. Materials and Methods: The prototype WLNI fluorescence endoscopy system consists of a custom made clinically approved WLNI camera, comprising a color camera and an ultra-sensitive camera for concurrent white-light and NIR fluorescence acquisition, attached to either a clinical fiberscope or a multi-modal fiber-optic bundle. The sensitivity of the WLNI fluorescence endoscopy system using bevacizumab-800CW (anti-VEGF-A), cetuximab-800CW (anti-EGFR) and trastuzumab-800CW (anti-HER-2) was evaluated in vivo in CRC subcutaneous (s.c.) and intraperitoneal (i.p.) mouse models of bioluminescent CRC cell lines (HCT116luc, HT29luc2). Results: The WLNI fluorescence endoscopy system allowed simultaneous display of real-time conventional white-light, fluorescence and overlay images at video-rate. It demonstrated excellent instant visualization of the s.c. and i.p. tumors (diameter ≥1 mm), with clear tumor boundaries and low background autofluorescence. The fluorescence was detected despite the intact overlying skin, even providing clear fluorescence images up to 8 cm distance from the s.c. tumors. The 800CW-labeled antibodies showed high TBRs in the HCT116luc s.c. model using IVIS (3.2±0.9 for bevacizumab-800CW, 5.7±3.0 for cetuximab-800CW and 6.9±3.4 for trastuzumab-CW800, n=3). The Odyssey Infrared Imager revealed accumulation of the tracers in the HCT116luc and HT29luc2 s.c. and i.p. tumors and in the liver, with almost no accumulation in muscles and colon. The specificity will be examined in a following study. Conclusion: This WLNI fluorescence endoscopy system offers unique advantages for the detection and characterization of (pre)malignant colorectal lesions in combination with the 800CW-labeled antibodies: it promises high sensitivity and provides dynamic, real-time in vivo, high-quality images at video-rate. At this moment, it is not possible to draw conclusions about the binding specificity of the 800CW-labeled antibodies used. These results support clinical evaluation of WLNI endoscopy, in order to enhance early detection of colorectal (pre)malignancies and improve potentially outcome in high-risk patients.