Bright Visualization of Primary and Metastatic Medullary Thyroid Cancer in an Orthotopic Nude Mouse Model Using a Fluorescent Humanized Anti-CEA Antibody for More Accurate Staging and Improved Resection.

Abstract

The standard treatment for medullary thyroid cancer (MTC) is total thyroidectomy with cervical lymph node dissection. Cure depends on complete surgical resection; however, intraoperative identification of tumor margins and involved lymph nodes remains challenging. The CEA-expressing MTC cell line TT was used to establish orthotopic thyroid tumor models (n = 7), whereas the non-CEA-expressing papillary thyroid cancer (PTC) cell line TPC-1 was used to establish subcutaneous negative control tumor models (n = 3). Orthotopic MTC models were established by injecting TT cells (1x10⁷) into the thyroid gland of athymic nude mice, whereas subcutaneous models were established by implanting TT and TPC-1 tumors into the flanks of nude mice. The humanized anti-CEA antibody M5A was conjugated to the near infrared fluorophore IRDye800CW to generate M5A-IR800. Mice received 50 µg of M5A-IR800 via tail vein injection and were imaged under bright light and fluorescence using a LI-COR Pearl Imaging System. Tumor-to-background ratios (TBR) were calculated as the ratio of mean tumor fluorescence intensity to surrounding tissue. Tumor specimens were analyzed by histology and immunohistochemistry. The mean TBR of orthotopic MTC tumors treated with M5A-IR800 was 6.85 (±3.75) at 96 hours post injection. Mice bearing subcutaneous TT tumors also achieved high TBRs of 4.23, 3.59, and 3.3 at 96 hours. In contrast, non-CEA-expressing TPC-1 tumors demonstrated no detectable fluorescence. Hematoxylin and eosin staining and immunohistochemistry confirmed accurate tumor targeting by M5A-IR800 and strong CEA expression in MTC tumors. M5A-IR800 specifically and brightly targets MTC in preclinical mouse models and represents a promising approach to enhance intraoperative visualization of MTC.