A specific and adaptable approach to track CD206 macrophages by molecular MRI and fluorescence imaging.

The mannose receptor (CD206, expressed by the gene ) is a surface marker overexpressed by anti-inflammatory and pro-tumoral macrophages. As such, CD206 macrophages play key roles in the immune response to different pathophysiological conditions and represent a promising diagnostic and therapeutic target. However, methods to specifically target these cells remain challenging. In this study, we describe a multi-mannose approach to develop CD206-targeting fluorescent and MRI agents that specifically and sensitively detect and monitor CD206 macrophage immune response in different disease conditions. We designed and synthesized fluorescent agents MR1-cy5 and MR2-cy5, and MRI agents Mann2-DTPA-Gd and MannGdFish. Cellular assays using pro-inflammatory and anti-inflammatory macrophages differentiated from RAW 264.7 cells were performed, and signals were detected by fluorescence microscopy and inductively coupled plasma mass spectrometry (ICP-MS) to validate specificity . specificity and efficacy of the agents were evaluated by MRI in a subcutaneous wound healing model and experimental glioma with without and with D-mannose treatment, , and mice, and in stroke. One-way ANOVA and two-way ANOVA tests were used for data analysis. P < 0.05 was considered statistically different. Both fluorescence imaging with MR2-cy5, ICP-MS with Mann2-DTPA-Gd, and MRI in mice confirmed the specificity of our approach. Mann2-DTPA-Gd MRI can track the changes of CD206 macrophages at different stages of wound healing, correlating well with flow cytometry data using another anti-inflammatory macrophage marker (arginase-1). The specificity and efficacy of Mann2-DPTA-Gd were further validated in experimental glioma, in which Mann2-DTPA-Gd imaging detected CD206 tumor-associated macrophages (TAMs), demonstrated significantly decreased signals in mice and mice, and tracked treatment changes in D-mannose-treated mice. Furthermore, Mann2-DTPA-Gd can report microglia/macrophages and correlate with histology in stroke. The more Gd-stable agent MannGdFish demonstrated similar efficacy as Mann2-DTPA-Gd with favorable biodistribution and pharmacokinetics. We have developed a fluorescent agent (MR2-cy5) and MRI agents (Mann2-DTPA-Gd and MannGdFish) with two mannose moieties that are highly specific to CD206 and can track CD206 macrophages in disease models of wound healing, tumor, and neurological disease. Importantly, MannGdFish, with its high specificity, stability, favorable biodistribution, and pharmacokinetics, is a promising translational candidate to noninvasively monitor CD206 macrophages in repair/regeneration and tumors in patients. In addition, with the specific binding motif to CD206, other imaging modalities and therapeutic agents could also be introduced for theranostic applications.