In Vivo Detection of Lymph Nodes Metastasis of ESCC Using CXCR4-Targeted Tracer [64Cu]Cu-NOTA-CP01

Abstract Purpose Esophageal squamous cell carcinoma (ESCC) frequently exhibits skip metastasis to lymph nodes. While current efforts have primarily focused on diagnosing the primary tumor, there is a deficiency of noninvasive imaging techniques capable of directly detecting metastatic lymph nodes (MLN) in ESCC. Building on our previous demonstration of the specificity of [⁶⁴Cu]NOTA-CP01 in targeting CXCR4 in ESCC, our aim in this study was to detect MLN in ESCC. Procedures The intralymphatic tumor metastasis model was established by injecting EC109/Luc cells into the left paw pads of mice, and monitored using bioluminescence imaging. Radiolabeling of [⁶⁴Cu]NOTA-CP01 and PET/CT imaging were performed, as in previous studies conducted by our group. Further quantitative analysis was performed. To verify metastatic tumor cells, lymph nodes were collected and stained with H&E. Immunohistochemistry was used to determine the expression levels of CXCR4 in metastatic lymph nodes. Results PET/CT imaging clearly demonstrated the uptake of radioactive signals in the left inguinal lymph nodes following injection of [⁶⁴Cu]Cu-NOTA-CP01, compared to the blocked group. Further quantitative analysis revealed that the SUV of the left inguinal lymph nodes (SUV = 1.55 ± 0.50, n = 5) was significantly higher than that in the blocked group (SUV = 0.50 ± 0.05, n = 3) (p < 0.05). Histological examination using H&E staining confirmed the presence of metastatic tumor cells in the left inguinal lymph nodes, and immunohistochemistry staining confirmed positive expression of CXCR4 in the left inguinal lymph nodes. Conclusions This study investigated a noninvasive approach to detect MLN in a preclinical model using the PET tracer [⁶⁴Cu]Cu-NOTA-CP01. Both in vitro and in vivo biological experiments suggested that CXCR4 could serve as a promising target for visualizing MLN in ESCC. Improving our comprehension of lymphatic metastasis is crucial for devising personalized treatment strategies for ESCC.