⁶⁸ Ga-NODAGA-SNA009: A first-in-class single-domain antibody probe for rapid PET imaging of GPA33 in colorectal cancer.

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Background: GPA33 is a cell-surface glycoprotein highly expressed in normal intestinal epithelium and over 95% of colorectal cancers (CRC), with minimal expression in other tissues, making it an ideal target for radiopharmaceutical therapy. While prior studies utilized full-length antibodies labeled with ¹²⁴ I, their clinical utility is hampered by prolonged imaging windows and isotope availability. This study developed ⁶⁸ Ga-NODAGA-SNA009, a GPA33-targeted single-domain antibody probe, to enable rapid and efficient clinical imaging. Methods: GPA33 expression in colon cancer cell lines was determined by Western blot and flow cytometry. The binding affinity of SNA009 was evaluated via flow cytometry using the GPA33-positive LS174T cell line. Biodistribution, pharmacokinetics, and PET/CT imaging were assessed in xenograft models. A first-in-human study was conducted in 15 CRC patients to evaluate ⁶⁸ Ga-NODAGA-SNA009 PET/CT. The correlation between SUVmax and GPA33 expression was analyzed and imaging features were compared with standard ¹⁸ F-FDG PET. Results: SNA009 demonstrated high affinity and specificity for GPA33 with no cytotoxicity. Preclinical studies showed significant tumor and renal uptake with sustained retention at 2 h post-injection. In patients, clear tumor visualization was achieved within 1 h. A preliminary linear correlation was observed between SUVmax and GPA33 expression. Compared to ¹⁸ F-FDG, ⁶⁸ Ga-NODAGA-SNA009 exhibited similar tumor-to-background ratios for extra-intestinal lesions but demonstrated superior specificity with low uptake in inflammatory or benign lesions. However, high physiological intestinal uptake—attributable to rapid clearance and early imaging kinetics—limited the assessment of adjacent lesions, which could not be mitigated by cold antibody blockade. Conclusions: ⁶⁸ Ga-NODAGA-SNA009 is a safe and tumor-specific probe enabling rapid, non-invasive assessment of GPA33 expression. It serves as a valuable complement to ¹⁸ F-FDG PET, particularly for differentiating extra-intestinal tumors from inflammation. Although high intestinal uptake poses a challenge for local lesion evaluation, this probe lays the foundation for future GPA33-directed theranostic applications. Strategies such as pre-targeting will be explored to overcome on-target/off-tumor physiological uptake.