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

Abstract

We developed a novel approach to bioluminescent T cell imaging (BLI) 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), and membrane-anchored RLuc (extRLuc). In vivo, mouse extGLuc+ T cells exhibited 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 demonstrate 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 (FFLuc). This sensitive imaging technology has application to many in vivo cell based studies in a wide array of mouse models.

The TAK1-NF-κB axis as therapeutic target for AML

“…ExtGLuc was retrieved from the rPLSI1180-ExtGLuc-IRES-hr-green fluorescent protein (GFP) vector (kindly provided by the Brentjens Laboratory 29 ) by EcoRI/StuI (blunt), and ligated into EcoRI/SalI (blunt) sites of the third generation lentivector CD711B_1_pCDH_MSCV. Lentiviral particles were produced by transient transfection in 293T cells using pVSV, pREV, and pMDL-PRRE helper plasmids.…”

Natural Expression of the CD19 Antigen Impacts the Long-Term Engraftment but Not Antitumor Activity of CD19-Specific Engineered T Cells

“…More recently, mouse models demonstrated the efficacy of gene-modified T cells targeting B cell malignancies through chimeric Ag receptors (CARs). Murine and human T cells expressing CAR consisting of an Ab-type recognition domain (scFv) giving CD19 specificity linked to the CD3z signaling molecule have been shown to eradicate established human CD19 + lymphoma when given in conjunction with lymphodepleting conditioning regimens or to immunodeficient mice (3)(4)(5)(6)(7). These receptors are now in earlystage phase I clinical trials at several sites worldwide.…”

In Vivo Cell Tracking with Bioluminescence Imaging

“…However, the secreting nature of the native GLuc enzyme significantly attenuates the in vivo bioluminescent signal from the cell expressing GLuc [44]. Elmer et al [45] reported that this limitation could be overcome by modified Gluc by addition of a CD8 transmembrane domain, and also they produced the membraneanchored form of GLuc-positive cells from both mouse and human primary T cells and demonstrated that the cells have a markedly superior in vivo BLI signal when compared with native GLuc positive T cells.…”