Development of a bioluminescence reporter mouse model for tracking and quantifying Ly6G+ neutrophils in vivo 9016

Abstract Description Immunotherapy has revolutionized cancer treatment and despite tremendous success, not all patients respond to immunotherapy. Identifying biomarkers predictive of effective treatment response is of utmost importance. To track neutrophils in vivo, we developed a transgenic mouse where firefly luciferase (Luc2), diphtheria toxin receptor (DTR), and enhanced green fluorescence protein (eGFP) genes were expressed under the endogenous Ly6G promoter. The Ly6G promoter enables tracking of Ly6G+ neutrophils using luciferase-based bioluminescence imaging (BLI), depletion of Ly6G+ neutrophils by diphtheria toxin treatment, and the ability to perform flow analyses using the eGFP marker. To confirm that the Luc2-DTR-eGFP (LDG) reporter cassette expression did not affect neutrophil activity and function, we quantified myeloperoxidase (MPO), a marker of neutrophil activation, in the phorbol myristate acetate (PMA)-induced ear edema mouse model. Neutrophil activity was measured using luminol enabled BLI of MPO. Luminol-BLI following PMA-induced ear edema demonstrated similar MPO activity between wildtype and transgenic mice. Further, topical application of PMA to transgenic mouse earlobe demonstrated neutrophil recruitment as seen by an increase in bioluminescence when compared to DMSO treated earlobe. These transgenic mice provide a valuable tool for profiling oncology compounds, including those targeting immune cells, and for understanding the mechanism of action of such agents. Funding Sources Merck & Co., Inc., Rahway, NJ, USA Topic Categories Tumor Immunology: Cellular Responses and Tumor Microevironment (TIME)