Abstract 55: A claudin-1 near-infrared fluorescent antibody conjugate for in vivo primary and metastatic colorectal cancer detection

Abstract Background: Claudins are a large family of tight junction (TJ) proteins being investigated as biomarkers and therapeutic targets in cancer. Their availability at the cell surface, tissue-specific expression, and upregulation in some carcinomas make them amenable to monoclonal antibody targeting. In addition, epitope masking within the intact TJs of healthy tissue limits off-target binding. Claudin-1 (CLDN1) is upregulated in colorectal cancer (CRC) and promotes therapy resistance and metastasis. Significance of Problem: CRC is the second deadliest cancer in the United States. While localized CRC can be treated surgically and has a 5-year survival rate of 91.1%, the survival rate of metastatic CRC falls to 15.7%. Due to resistance to current systemic treatments, surgery is the only curative option for CRC. However, tumors recur after ∼30% of curative surgeries for stage II or III CRC. The rate of positive surgical margins for CRC has been reported at 6.83% and is associated with poor prognosis and metastasis. Fluorescence-guided surgery using targeted near-infrared (NIR) dyes allows surgeons to more accurately identify tumor margins. METHODS: scRNAseq was used to characterize CLDN1 expression in CRC tumors. A NIR fluorescent dye, IRDye800 NHS-Ester, was conjugated to an anti-CLDN1 mAb (1C5-D9, CLDN1-IR800) and an IgG isotype control (IgG-IR800). Specificity was evaluated using fluorescent microscopy and Western blot in CLDN1-overexpression and knockout cell lines. The antibody was then tested in vivo using CRC cell line xenograft, patient-derived organoid (PDO) xenograft, and orthotopic syngeneic mouse models. Animals were injected IV and imaged using the LI-COR Pearl Trilogy System at multiple time points post-injection. Finally, an ex vivo panel of PDOs representing different ethnic and mutational backgrounds was used to examine the conjugate’s translation potential. Results: CLDN1 is highly expressed in CRC tumor cells, with low levels in the healthy colon. CLDN1-IR800 showed specific binding and robust fluorescence in CLDN1-expressing CRC cells compared to non-expressing cell lines or IgG-800. In vivo, CLDN1-IR800 showed significantly higher signal in CLDN1-expressing CRC cell line and PDO xenografts when compared to CLDN1 knockout tumors or IgG-IR800-injected mice. Biodistribution upon necropsy showed a significantly higher signal in the tumor compared to other organs. In an orthotopic syngeneic mouse model, both primary and metastatic lesions were detectable. Finally, ex vivo imaging showed significant signal in a panel of PDOs with diverse mutational and racial backgrounds. Conclusions: CLDN1’s selective expression on CRC tumor cells makes it an ideal candidate for in vivo tumor detection. In this study, we demonstrate the translation potential in targeting CLDN1 for tumor detection using multiple in vitro, in vivo, and ex vivo models. Citation Format: Mark Primeaux, Ritika Gupta, Iram Fatima, Sumbal Talib, Aaron Mohs, Michael Bouvet, Punita Dhawan. A claudin-1 near-infrared fluorescent antibody conjugate for in vivo primary and metastatic colorectal cancer detection [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 55.