Feasibility of real-time in vivo Zr-89-DFO-labeled CAR T-cell trafficking using PET imaging

Abstract

Introduction Chimeric antigen receptor (CAR) T-cells have been recently developed and are producing impressive outcomes in patients with hematologic malignancies. However, there is no standardized method for cell trafficking and in vivo CAR T-cell monitoring. We assessed the feasibility of real-time in vivo Zr-89-p-Isothiocyanatobenzyl-desferrioxamine (Df-Bz-NCS, DFO) labeled CAR T-cell trafficking using positron emission tomography (PET). Results The Zr-89-DFO radiolabeling efficiency of Jurkat/CAR and human peripheral blood mononuclear cells (hPBMC)/CAR T-cells was 70%-79%, and cell radiolabeling activity was 98.1-103.6 kBq/10(6) cells. Cell viability after radiolabeling was > 95%. Cell proliferation was not significantly different during the early period after radiolabeling, compared with unlabeled cells; however, the proliferative capacity decreased over time (day 7 after labeling). IL-2 or IFN-. secretion was not significantly different between unlabeled and labeled CAR T-cells. PET/magnetic resonance imaging in the xenograft model showed that most of the (ZrDFO)-Zr-89-labeled Jurkat/CAR T-cells were distributed in the lung (24.4% +/- 3.4%ID) and liver (22.9% +/- 5.6%ID) by one hour after injection. The cells gradually migrated from the lung to the liver and spleen by day 1, and remained stable in these sites until day 7 (on day 7: lung 3.9% +/- 0.3%ID, liver 36.4% +/- 2.7%ID, spleen 1.4% +/- 0.3%ID). No significant accumulation of labeled cells was identified in tumors. A similar pattern was observed in ex vivo biodistributions on day 7 (lung 3.0% +/- 1.0%ID, liver 19.8% +/- 2.2%ID, spleen 2.3% +/- 1.7%ID). Zr-89-DFO-labeled hPBMC/CAR T-cells showed a similar distribution, compared with Jurkat/CAR T-cells, on serial PET images. CAR T cell distribution was cross-confirmed by flow cytometry, Alu polymerase chain reaction, and immunohistochemistry. Conclusion Real-time in vivo cell trafficking is feasible using PET imaging of Zr-89-DFO-labeled CAR T-cells. This can be used to investigate cellular kinetics, initial in vivo biodistribution, and safety profiles in future CAR T-cell development.