Radiosynthesis and preclinical evaluation of [(68)Ga]Ga-NOTA-folate for PET imaging of folate receptor β-positive macrophages

Folate receptor β (FR-β), a marker expressed on macrophages, is a promising target for imaging of inflammation. Here, we report the radiosynthesis and preclinical evaluation of [(68)Ga]Ga-NOTA-folate ((68)Ga-FOL). After determining the affinity of (68)Ga-FOL using cells expressing FR-β, we studied atherosclerotic mice with (68)Ga-FOL and (18)F-FDG PET/CT. In addition, we studied tracer distribution and co-localization with macrophages in aorta cryosections using autoradiography, histology, and immunostaining. The specificity of (68)Ga-FOL was assessed in a blocking study with folate glucosamine. As a final step, human radiation doses were extrapolated from rat PET data. We were able to produce (68)Ga-FOL with high radiochemical purity and moderate molar activity. Cell binding studies revealed that (68)Ga-FOL had 5.1 nM affinity for FR-β. Myocardial uptake of (68)Ga-FOL was 20-fold lower than that of (18)F-FDG. Autoradiography and immunohistochemistry of the aorta revealed that (68)Ga-FOL radioactivity co-localized with Mac-3–positive macrophage-rich atherosclerotic plaques. The plaque-to-healthy vessel wall ratio of (68)Ga-FOL was significantly higher than that of (18)F-FDG. Blocking studies verified that (68)Ga-FOL was specific for FR. Based on estimations from rat data, the human effective dose was 0.0105 mSv/MBq. Together, these findings show that (68)Ga-FOL represents a promising new FR-β–targeted tracer for imaging macrophage-associated inflammation.