Hypoxia-Targeting Fluorescent Nanobodies for Optical Molecular Imaging of Pre-Invasive Breast Cancer.

PURPOSE: The aim of this work was to develop a CAIX-specific nanobody conjugated to IRDye800CW for molecular imaging of pre-invasive breast cancer. PROCEDURES: CAIX-specific nanobodies were selected using a modified phage display technology, conjugated site-specifically to IRDye800CW and evaluated in a xenograft breast cancer mouse model using ductal carcinoma in situ cells (DCIS). RESULTS: Specific anti-CAIX nanobodies were obtained. Administration of a CAIX-specific nanobody into mice with DCIS xenografts overexpressing CAIX showed after 2 h a mean tumor-to-normal tissue ratio (TNR) of 4.3 ± 0.6, compared to a TNR of 1.4 ± 0.2 in mice injected with the negative control nanobody R2-IR. In DCIS mice, a TNR of 1.8 ± 0.1 was obtained. Biodistribution studies demonstrated an uptake of 14.0 ± 1.1 %I.D./g in DCIS + CAIX tumors, 4.6 ± 0.8 %I.D./g in DCIS tumors, while 2.0 ± 0.2 %I.D./g was obtained with R2-IR. CONCLUSIONS: These results demonstrate the successful generation of a CAIX-specific nanobody-IRDye800CW conjugate that can be used for rapid imaging of (pre-)invasive breast cancer.