In Living COLOR: Using Bioluminescence Imaging to Quantify Tumor Burden

Aug 5, 2021 2:00:00 PM / by Champions Oncology

Bioluminescence imaging (BLI) in mouse models

Bioluminescence imaging (BLI) is a highly sensitive non-invasive molecular imaging technique that has been used across biological research areas for insights into how tissues and organs function in real-time under normal physiological conditions or different disease states. BLI was developed using transgenic mice that expressed the luciferase enzyme as a reporter in specific cells. These cells can be visualized in the presence of oxidated luciferin substrate, which generates bioluminescent light (photons) that can be detected using microscope-based or portable photon detectors[1]. This method can detect biologically active tumor cells and can monitor changes in the growth or clearance these cells because bioluminescence is an ATP-dependent process in living animals[2], [3]. More recently, functional BLI probes have been developed that use caged luciferin substrates that must be cleaved through a specific biological process for luminescence[4]. These functional probes have been useful in monitoring metabolism and enzyme activity, and such read-outs can be used to understand basic tumor biology or identify mechanisms of action for anti-tumor drugs.

Early BLI methods typically yielded qualitative results, but advances in reagents and visualization methods have led to the development of semi-quantitative BLI methods that are useful to different types of preclinical oncology studies. One application uses BLI to verify injection of tumor cells into mouse models. Some models require the injection of very small quantities of tumor cells, but uncertainty about the volume and site of inoculation can make data interpretation difficult. Using BLI-based tumor cell lines, tumor cell inoculation can be verified quickly, accurately, and non-invasively[5]. BLI can also track longitudinal changes in tumor loads, but it is not appropriate for absolute tumor mass measurements due to limitations of luciferin penetration into tumor tissue and constraints of spatial resolution. Longitudinal studies like this are critical for preclinical evaluations anti-tumor treatments, especially intratumoral-based treatments, and investigators have developed quantitative indexes of bioluminescence using standardized BLI methods to improve the interpretation of this data[5].

Luciferase Reaction diagram

Although BLI methods are noninvasive and not affected by body movements, spatial resolution is limited, and genetically modified cells, tissues, or mouse models that express luciferase must be used. BLI does show a strong direct correlation between signal and tumor mass for subcutaneous focal lesions. Thus, BLI studies cannot be used for direct translation but are well suited to longitudinal studies of orthotopic tumors that are in deep tissues or organs[6]. BLI can also track tumor metastases but the correlation between BLI signal and tumor mass is not as strong. BLI can complement other imaging methods, including ultrasound, PET, and MRI. These other methods remain essential to clinical diagnostics, and MRI is a superior method for absolute tumor mass measurements and delineation of tumor margins[7].

Despite these limitations, BLI is a valuable non-invasive preclinical method that allows real-time monitoring of tumor cell delivery, growth, and metastasis. As such, fewer mice can be used in studies because tumors can be monitored and measured in living mice. Advances in BLI reagents and imaging technology have allowed for this method to be used in larger animal models and opens the door for potential use in human subjects[8].

 

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[1] Yevtodiyenko A, Bazhin A, Khodakivskyi P, et al. Portable bioluminescent platform for in vivo monitoring of biological processes in non-transgenic animals. Nat. Commun. 2021;12(1):2680.

[2] Edinger M, et al. Revealing lymphoma growth and the efficacy of immune cell therapies using in vivo bioluminescence imaging. Blood. 2003;101:640–648.

[3] Sweeney TJ, et al. Visualizing the kinetics of tumor-cell clearance in living animals. Proc. Natl. Acad. Sci. USA. 1999;96:12044–12049.

[4] Mezzanotte L, van ‘t Root M, Karatas H, Goun EA, Lowik C. In Vivo Molecular Bioluminescence Imaging: new Tools and Applications. Trends Biotechnol. 2017;35:640–652. 

[5] Cosette J, Ben Abdelwahed R, Donnou-Triffault S, Sautès-Fridman C, Flaud P, Fisson S. Bioluminescence-Based Tumor Quantification Method for Monitoring Tumor Progression and Treatment Effects in Mouse Lymphoma Models. J. Vis. Exp. 2016 Jul 7;(113):53609.

[6] Klerk CPW, Overmeer RM, Niers TMH, et al. Validity of bioluminescence measurements for noninvasive in vivo imaging of tumor load in small animals. Biotechniques. 2018 May 16; 43:1S.

[7] Ravoori MK, Margalit O, Singh S, Kim SH, Wei W, Menter DG, DuBois RN, Kundra V. Magnetic Resonance Imaging and Bioluminescence Imaging for Evaluating Tumor Burden in Orthotopic Colon Cancer. Sci. Rep. 2019 Apr 15;9(1):6100.

[8] Pirovano G, Roberts S, Kossatz S, Reiner T. Optical Imaging Modalities: Principles and Applications in Preclinical Research and Clinical Settings. J. Nucl. Med. 2020 Oct;61(10):1419-1427.

Tags: Preclinical In Vivo Imaging