Developing Preclinical Mouse Models That Employ Fluorophore-Nanoluc Bret Reporters (Lumifluor) For Monitoring Tumorigenesis And Response To Therapy

Fluorescent proteins are widely used to study molecular and cellular events, yet this traditionally relies on delivery of excitation light, which can trigger autofluorescence, photoxicity, and photobleaching, impairing their use in vivo. Accordingly, chemiluminescent light sources such as those generated by luciferases have emerged, as they do not require excitation light. However, current luciferase reporters lack the brightness needed to visualize events in deep tissues. We recently reported the creation of chimeric eGFP-NanoLuc (GpNLuc) and LSSmOrange-NanoLuc (OgNLuc) fusion reporter proteins coined LumiFluors, which combine the benefits of eGFP or LSSmOrange fluorescent proteins with the bright, glow-type bioluminescent light generated by an enhanced small luciferase subunit (NanoLuc) of the deep sea shrimp Oplophorus gracilirostris. The intramolecular bioluminescence resonance energy transfer (BRET) that occurs between NanoLuc and the fused fluorophore generates the brightest bioluminescent signal known to date, enabling greatly improved spatio-temporal monitoring of very small numbers of tumor cells via in vivo optical imaging while also allowing isolation and analysis of single cells by flow cytometry. Here we report the generation of an inducible and conditional knockin reporter mouse (Rosa26-LSL-GpNLuc) and the application of this mouse to creating preclinical mouse models suitable for the noninvasive evaluation of therapeutic interventions. To model NSCLC, Rosa26-LSL-GpNLuc were crossed to LSL-KRasG12D and initiated tumorigenesis by intra-nasal administration of adenovirus expressing Cre-recombinase. Longitudinal monitoring of disease progression using bioluminescent imaging detected extremely early lesions corresponding to points in time characteristic of atypical adenomatous hyperplastic lesions. The ability to identify pre-neoplastic lesions at an early stage will enable the development of more precise preclinical models suitable for evaluating responses to drug treatment. Ongoing studies are aimed at modeling HPV-induced HNSCC by crossing Rosa26-LSL-GpNLuc to KRT14-CreERT2;LSL-E6/E7 for monitoring oral tumorigenesis and response to novel treatment strategies. Thus, LumiFluor reporters are inexpensive, robust, non-invasive tools that allow for markedly improved in vivo optical imaging of tumorigenic processes and response to therapy. Note: This abstract was not presented at the meeting. Citation Format: Miranda Carper, Scott Troutman, Franz Schaub, Adele Musicant, Weimin Li, Erin Henry, Joseph Kissil, John Cleveland, Antonio L. Amelio. Developing preclinical mouse models that employ fluorophore-nanoLuc BRET reporters (LumiFluor) for monitoring tumorigenesis and response to therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr LB-137. doi:10.1158/1538-7445.AM2017-LB-137