Evaluation of the HET-CAM model for biodistribution studies of nanoparticles using PET and MR imaging

Aim: Nanoparticles (NP) are of particular interest for medical applications due to their high biocompatibility and highly adaptable surface. The biodistribution and biostability of the particles is usually studied in animal experiments with rodents. The Hen's Egg Test-Chorioallantoic Membrane (HET-CAM) in vivo model is a good alternative to animal experiments with regard to the 3R principles of animal welfare and has already been successfully used for peptide studies. The suitability of the model for biodistribution studies of particles is analyzed in this project. Therefore, nanodiamonds (ND), mesoporous silica (MSN) and superparamagnetic iron oxide NP (SPION) were studied with respect to biodistribution in the HET-CAM model.

Methods: ND coated with deferoxamine (DFO) coupled with human serum albumin, SPION modified with PEG (polyethylene glycol), and MSN functionalized with a fragment of CD47 protein and DFO were examined. All particles were radiolabeled using ⁸⁹Zr. The stability of the labeling was monitored in 0.9% NaCl, cell culture medium, and human serum over an observation period of five days. After i.v. injection, PET (Focus 120 Siemens Medical Solutions, Inc., Erlangen, Germany) and/or MRI (BioSpec 117/16, Bruker, Ettlingen, Germany) measurements were performed depending on the particle specificity. All measurements were performed between embryo development day 12 and 16.

Results: ⁸⁹Zr-radiolabeling yields of >76% were achieved for all NP. The stability of the labeling ranged from 77% (SPION) to 99% (MSN). An accumulation of MSN (6.5 ± 1.7) %IA/cc (n=3) and ND (10.0 ± 5.3) %IA/cc (n=4) was observed in the liver by overlaying PET and MRI data. In T1-weighted MR images, SPION caused signal reduction in the liver. The ratios of the corresponding signal intensities of liver-to-chest muscle (n=3) were 2.7 ± 0.2 before and 10.2 ± 1.2 (0h) to 9.6 ± 3.7 (4h) after injection.

Conclusions: The assessment of the biodistribution of various radiolabeled NP using high-resolution MR and PET imaging was successfully demonstrated. The particles accumulated in the liver of the chick embryo, similar to the distribution in mice, suggesting poor blood retention and the need to improve the surface modification. Despite model-related limitations, the HET-CAM model could help to reduce the number of animal experiments required, especially in the early stages of compound development.

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