Dye labeling for optical imaging biases drug carriers' biodistribution and tumor uptake.

Biodistribution analyses of nanocarriers are often performed with optical imaging. Though dye tags can interact with transporters, e.g., organic anion transporting polypeptides (OATPs), their influence on biodistribution was hardly studied. Therefore, this study compared tumor cell uptake and biodistribution (in A431 tumor-bearing mice) of four near-infrared fluorescent dyes (AF750, IRDye750, Cy7, DY-750) and dye-labeled poly(N-(2-hydroxypropyl)methacrylamide)-based nanocarriers (dye-pHPMAs). Tumor cell uptake of hydrophobic dyes (Cy7, DY-750) was higher than that of hydrophilic dyes (AF750, IRDye750), and was actively mediated but not related to OATPs. Free dyes' elimination depended on their hydrophobicity, and tumor uptake correlated with blood circulation times. Dye-pHPMAs circulated longer and accumulated stronger in tumors than free dyes. Dye labeling significantly influenced nanocarriers' tumor accumulation and biodistribution. Therefore, low-interference dyes and further exploration of dye tags are required to achieve the most unbiased results possible. In our assessment, AF750 and IRDye750 best qualified for labeling hydrophilic nanocarriers.

CT- and MRI-Aided Fluorescence Tomography Reconstructions for Biodistribution Analysis.

OBJECTIVES: Optical fluorescence imaging can track the biodistribution of fluorophore-labeled drugs, nanoparticles, and antibodies longitudinally. In hybrid computed tomography-fluorescence tomography (CT-FLT), CT provides the anatomical information to generate scattering and absorption maps supporting a 3-dimensional reconstruction from the raw optical data. However, given the CT's limited soft tissue contrast, fluorescence reconstruction and quantification can be inaccurate and not sufficiently detailed. Magnetic resonance imaging (MRI) can overcome these limitations and extend the options for tissue characterization. Thus, we aimed to establish a hybrid CT-MRI-FLT approach for whole-body imaging and compared it with CT-FLT. MATERIALS AND METHODS: The MRI-based hybrid imaging approaches were established first by scanning a water and coconut oil-filled phantom, second by quantifying Cy7 concentrations of inserts in dead mice, and finally by analyzing the biodistribution of AF750-labeled immunoglobulins (IgG, IgA) in living SKH1 mice. Magnetic resonance imaging, acquired with a fat-water-separated mDixon sequence, CT, and FLT were co-registered using markers in the mouse holder frame filled with white petrolatum, which was solid, stable, and visible in both modalities. RESULTS: Computed tomography-MRI fusion was confirmed by comparing the segmentation agreement using Dice scores. Phantom segmentations showed good agreement, after correction for gradient linearity distortion and chemical shift. Organ segmentations in dead and living mice revealed adequate agreement for fusion. Marking the mouse holder frame and the successful CT-MRI fusion enabled MRI-FLT as well as CT-MRI-FLT reconstructions. Fluorescence tomography reconstructions supported by CT, MRI, or CT-MRI were comparable in dead mice with 60 pmol fluorescence inserts at different locations. Although standard CT-FLT reconstruction only considered general values for soft tissue, skin, lung, fat, and bone scattering, MRI's more versatile soft tissue contrast enabled the additional consideration of liver, kidneys, and brain. However, this did not change FLT reconstructions and quantifications significantly, whereas for extending scattering maps, it was important to accurately segment the organs and the entire mouse body. The various FLT reconstructions also provided comparable results for the in vivo biodistribution analyses with fluorescent immunoglobulins. However, MRI additionally enabled the visualization of gallbladder, thyroid, and brain. Furthermore, segmentations of liver, spleen, and kidney were more reliable due to better-defined contours than in CT. Therefore, the improved segmentations enabled better assignment of fluorescence signals and more differentiated conclusions with MRI-FLT. CONCLUSIONS: Whole-body CT-MRI-FLT was implemented as a novel trimodal imaging approach, which allowed to more accurately assign fluorescence signals, thereby significantly improving pharmacokinetic analyses.