Assessing novel nano-photosensitizer hepatocellular carcinoma (HCC) tumor uptake in vivo as a candidate platform to enhance Y-90-radioembolization

Cerenkov light emitted from Y-90 can activate light-sensitive nano-photosensitizers producing tumoricidal reactive oxygen species (ROS) and increased hepatocellular carcinoma (HCC) cell death in vitro {1}. A dual nano-photosensitizer consisting of titanium dioxide (TiO₂) nanoparticles and titanocene (Tc) labelled with transferrin (TiO₂-Tf-Tc) holds potential to enhance the efficacy of Y-90 radioembolization. The purpose of this study was to assess the in vivo tumor uptake and penetration of a fluorescently labelled derivative of this dual nano-photosensitizer in an orthotopic rodent HCC model.

Materials and Methods: Orthotopic N1S1 HCC tumors were generated in 8 Sprague-Dawley rats weighing at least 300 gm by inoculating 2-3x10⁶ N1S1 cells into the subcapsular liver either through a laparotomy or percutaneous ultrasound guided approach. Tumor growth was monitored over 2 weeks through T2-weighted magnetic resonance (MR) imaging. After 2-4 weeks, or when tumors were at least 1 cm in diameter, animals were administered TiO₂ nanoparticles fluorescently labelled with Alexa-680-Tf (TiO₂-Alexa-Tf, 1 mg) via tail or portal vein administration. At 24 and 96 hours post administration, selected animals were sacrificed and TiO₂-Alexa-Tf uptake as assessed using ex vivo IVIS fluorescence (Perkin Elmer) imaging at 675nm excitation/720nm emission. Tumors and background livers were harvested fluorescent and light microscopy to assess nano-photosensitizer uptake.

Results:

Both systemic tail vein and portal vein administration resulted in high tumor uptake of TiO₂-Alexa-Tf nano-photosensitizer compared to background liver. Tumor to normal (T/N) ratios ranged from 0.8-2.0 24-hours and increased to 3.5-3.9 96 hours after administration, likely due to nano-photosensitizer elimination from normal liver and retention in tumor. There was minimal uptake in other organs such as kidney and spleen. High uptake was seen in lung metastases that developed in 2 animals.  Fluorescence microscopy using 675nm excitation/720nm emission and corresponding H&E staining demonstrated a clear uptake of TiO₂-Alexa-Tf within the tumor compared to adjacent background normal liver, including central portions of tumor. TiO₂-Alexa-Tf localized to tumor cells as indicated by DAPI staining.

Conclusion: TiO₂-Alexa-Tf demonstrated preferential tumor uptake in an orthotopic HCC animal model, which was retained for at least 4 days. This indicates a favorable retention of nano-photosensitizer in liver tumors, which is ideal to maximize the length of its interaction through Y-90’s radionuclide decay.