Sensitive in vivo imaging of T cells using a membrane-bound Gaussia princeps luciferase


We developed a new approach to bioluminescent T cell imaging using a membrane-anchored form of the Gaussia luciferase (GLuc) enzyme, termed extGLuc, which we could stably express in both mouse and human primary T cells. In vitro, extGLuc+ cells emitted significantly higher bioluminescent signal when compared to cells expressing GLuc, Renilla luciferase (RLuc) or membrane-anchored RLuc (extRLuc). In vivo, mouse extGLuc+ T cells showed higher bioluminescent signal when compared to GLuc+ and RLuc+ T cells. Application of this imaging approach to human T cells genetically modified to express tumor-specific chimeric antigen receptors (CARs) enabled us to show in vivo CAR-mediated T cell accumulation in tumor, T cell persistence over time and concomitant imaging of T cells and tumor cells modified to express firefly luciferase. This sensitive imaging technology has application to many in vivo cell-based studies in a wide array of mouse models.

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Figure 1: In vitro analyses of GLuc, extGLuc, RLuc and extRLuc bioluminescent signaling.
Figure 2: In vivo comparison of extGLuc+, GLuc+ and RLuc+ T cell bioluminescent signaling in a mouse graft-versus-host disease model.
Figure 3: OVA-specific extGLuc+ DO11.10 T cells specifically traffic to subcutaneous A20(OVA) tumors, as assessed by bioluminescent imaging.
Figure 4: Human extGLuc+ T cells further modified to express the CD19-targeted 19z1 CAR specifically accumulate in subcutaneous CD19+ NALM6 tumors in SCID-beige mice, as assessed by bioluminescent imaging.
Figure 5: Human 1928zIRESextGLuc+ T cells retain targeted cytotoxic potential in vitro when compared to 1928z+ T cells and specifically traffic to and eradicate systemic NALM6 tumors in SCID-beige mice, as assessed by dual bioluminescent imaging of T cells and tumor cells.


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This work was supported by US National Institutes of Health grants CA95152, CA059350, CA08748, CA086438, CA096945, CA094060 and CA083084, The Alliance for Cancer Gene Therapy, the Damon Runyon Clinical Investigator Award (R.J.B.), The Annual Terry Fox Run for Cancer Research organized by the Canada Club of New York, Kate's Team, W.H. Goodwin and A. Goodwin and the Commonwealth Cancer Foundation for Research and the Experimental Therapeutics Center of MSKCC, the Geoffrey Beene Cancer Foundation, the Chronic Lymphocytic Leukemia Foundation, the Ludwig Center for Cancer Immunotherapy and the Bocina Cancer Research Fund.

Author information

E.B.S. contributed to the design of the experiments, supervised and conducted the experiments and generated the manuscript figures; R.Y. contributed to the design of the experiments; J.L. helped conduct experiments for Figure 5 and Supplementary Figures 1 and 2; Y.N. assisted in the conduct experiments from Figures 1, 2, 3; Blesida and Blesserene Punzalan assisted in the conduct of the in vivo experiments; K.L.P. contributed to the conduct and interpretation of histological analyses; S.M.L. contributed to the design of the experiments; M.S. contributed to the design of the experiments and the review of the manuscript and R.J.B. supervised the project, contributed to the design of the experiments and wrote the manuscript.

Correspondence to Renier J Brentjens.

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Santos, E., Yeh, R., Lee, J. et al. Sensitive in vivo imaging of T cells using a membrane-bound Gaussia princeps luciferase. Nat Med 15, 338–344 (2009).

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