O2139: Evaluation of Indocyanine Green-Labeled Dinutuximab as a Novel Intraoperative Molecular Imaging Agent for Neuroblastoma: An Orthotopic Murine Model

Background: Neuroblastoma (NB) is the most common extracranial solid tumor in children, accounting for 15% of all pediatric cancer deaths. Gross total resection (>95% tumor removal) of high-risk neuroblastoma (NB) is associated with improved overall survival, but neoadjuvant therapy and the presence of metastatic disease can make intraoperative tumor detection challenging. As GD2 is a cell surface marker uniformly expressed on NB tumor cells, we developed and evaluated the utility of an indocyanine green (ICG)-labeled anti-GD2 antibody, dinutuximab (ICG-D), as a novel intraoperative imaging agent to detect NB in an orthotopic murine model.

Methods: Dinutuximab was conjugated to amine-reactive ICG using an ICG labeling assay. Antibody protein-ICG ratio and ICG-D conjugate concentration were determined via ultraviolet-visible spectrophotometry. Using the human NB cell line SH-SY5Y, orthotopic xenografts were established in NSG mice via subcapsular renal injection. After four weeks, ICG-D, ICG, dinutuximab, or phosphate-buffered saline were administered intraperitoneally at varying time points prior to sacrifice. Preoperative injection timepoints of 2hr, 6hr, 12hr, and 24hr were evaluated. In vivo fluorescence detection of the primary tumor was performed (Stryker SPY-PHI handheld imager) and ex vivo tumor fluorescence was quantified (IVIS Spectrum system). ICG-D tumor fluorescence was compared to ICG alone using the Mann-Whitney U test.

Results: Qualitative evaluation of ICG-D fluorescence demonstrated increased tumor-specific fluorescence compared to ICG alone, and no fluorescence was visualized for the negative controls (dinutuximab, saline). The 24hr preoperative injection timepoint exhibited the greatest fluorescence compared to earlier timepoints. Ex vivo quantification of tumor fluorescence revealed greater fluorescence in the ICG-D group compared to the ICG alone group when injected preoperatively at 12hr (1.72x10^9 ± 6.21x10^6 vs 6.94x10^8 ± 5.91x10^6 [p/s]/[µW/cm2]; p=0.005) and 24hr (2.34x10^10 ± 2.94x10^8 vs 1.48x10^9 ± 1.24x10^7 [p/s]/[µW/cm2]; p=0.005), but not at 2hr (p=0.121) and 6hr (p=0.304). No ICG-D-related toxicity or adverse events were observed.

Conclusion: ICG-D specifically targets and fluoresces in vivo neuroblastoma xenograft tumors. Future studies aim to optimize the antibody protein-ICG ratio and evaluate the utility of ICG-D for recurrent metastatic disease.