In vivo Bioluminescence Imaging of Labile Iron in Xenograft Models and Liver Using FeAl-1, an Iron-Activatable Form of D-Luciferin

Abstract

Dysregulation of cellular iron homeostasis is associated with diverse disease and pathological cell states, from cancer and ischemia-reperfusion injury to epithelial–mesenchymal transition and drug-tolerant “persister” cancer cells. Here we introduce Ferrous Iron Activatable Luciferin-1 (FeAL-1), a small molecule probe for bioluminescent imaging of the labile iron pool (LIP) in luciferase-expressing cells and experimental animals. We find that FeAL-1 reliably detects LIP fluctuations in cells resulting from iron supplementation, depletion, or treatment with hepcidin, the master regulator of systemic iron in mammalian physiology. Utilizing FeAL-1 and a dual-luciferase reporter system, we quantify LIP in mouse liver and three different orthotopic pancreatic ductal adenocarcinoma tumors. Remarkably, we observed up to ten-fold elevation of LIP in tumors as compared to liver, the major organ of iron storage in mammals. Treating mice with hepcidin or doxorubicin further elevated LIP in both liver and tumors. These studies reveal a therapeutic index between tumoral and hepatic LIP and suggest an approach to sensitize tumors toward LIP-activated therapeutics.

Funding Information: This work was funded in part by postdoctoral support (to RLG) from Merck & Co. Inc., and by NIH Grants R01CA260860 (to EAC and ARR) and R01AI105106 (to ARR).

Declaration of Interests: EAC and ARR are co-founders of Tatara Therapeutics.

Ethics Approval Statement: All experiments were conducted in the AAALAC accredited University of California, San Francisco in accordance with all applicable local requirements, including approval by the IACUC. The animal research protocol was reviewed and approved by the Merck Institutional Animal Care and Use Committee (IACUC). The mice were cared for in accordance with the Guide for the Care and Use of Laboratory Animals, 8th Edition.

Keywords: bioluminescence imaging, chemical probes, pancreatic ductal adenocarcinoma, ferroptosis