A novel radio-pharmaceutical to image activation of the innate immune system in vivo

Abstract Dysregulation of innate immunity contributes to the pathophysiology of many diseases, including obesity, rheumatoid arthritis and diabetes. Dysregulated production of reactive oxygen (ROS) and nitrogen species (RNS) by the innate immune system act both malevolently and benevolently through tissue damage and T-cell suppression. Optical reporters such as DCFDA and luminol/L-012 are routinely utilized to monitor ROS/RNS. 4-chloro-1-napthol (4CN) has been used to identify innate immune cells, and 1-napthol (N) covalently binds to activated human neutrophils through Nox2 and MPO-dependent mechanisms. However, these compounds are not suitable for deep tissue imaging in vivo. Under-standing this complex interplay in humans may benefit from deep tissue imaging of ROS/RNS by positron emission tomography (PET). Herein we introduce [18F]4FN, a bioisostere of N and 4CN, as a novel PET imaging agent, and validate binding both in vitro and in vivo. In all-trans-retinoic acid-differentiated HL-60 cells, [18F]4FN showed a robust 4.7 +/− 0.5 fold increase in binding to PMA-activated cells vs control cells (SEM n = 4, p = 0.01). Binding was > 99% inhibited by treatment with the peroxidase inhibitor 4-ABAH and the Nox2 inhibitor DPI (p < 0.01, each). [18F]4FN was selectively trapped in inflamed mouse tissues in vivo upon activation by PMA or LPS, robust to injection route (ip vs iv) and strain (C57Bl6/N and Balbc/N). These data quantitatively co-correlated with a gold standard bioluminescent reporter, L-012 (r = 0.94, p = 0.0004), and immune infiltration was cross validated by H&E staining ex vivo. [18F]4FN showed strong promise in pre-clinical models and translational steps toward clinical PET imaging is underway.