Noninvasive longitudinal PET/CT imaging of CAR T cells using PSMA reporter gene

PurposeChimeric antigen receptor (CAR) T cell therapy has achieved great success in treating hematologic malignancies. However, it is yet to prove effective in the treatment of solid tumors. Thus, it is necessary to develop appropriate methodology for the long-term, accurate, and quantitative evaluation of the distribution and activities of CAR T cells in solid tumors. In the present study, we engineered TfR Delta PSMA CAR (CAR-Delta PSMA) T cells, which targeted the transferrin receptor (TfR) expressed by tumor cells and could be tracked in vivo via a reporter gene encoding the truncated prostate specific membrane antigen (Delta PSMA). We then quantitatively monitored these CAR T cells in vitro and in vivo using [68Ga]Ga-PSMA-617 positron emission tomography (PET)/computed tomography (CT).MethodsThe CAR-Delta PSMA T cells were genetically engineered by transducing T cells with a lentiviral vector encoding TfR41BB zeta-T2A-Delta PSMA. Firstly, the target expression, activation, and cytotoxicity of CAR-Delta PSMA T cells were validated in vitro. Secondly, the minimum thresholds of CAR-Delta PSMA T cells detection for [68Ga]Ga-PSMA-617 PET/CT were also determined in vitro and in vivo respectively. Lastly, the feasibility of monitoring the biodistribution and infiltration of CAR-Delta PSMA T cells after systematic administration was evaluated in the breast cancer subcutaneous xenograft model.ResultsThe CAR-Delta PSMA T cells retained activation and tumor killing capacity after transduction of the Delta PSMA-encoding reporter gene. Next, the CAR-Delta PSMA T cells could be reliably tracked by [68Ga]Ga-PSMA-617 PET/CT, the detection sensitivity of which was 250 cells/mm3 in vitro and 100 cells/mm3 in vivo. Next, the sequential imaging assays revealed that [68Ga]Ga-PSMA-617 PET/CT could be used to specifically visualize Delta PSMA+ CAR T cells at the tumor site. The increase in the [68Ga]Ga-PSMA-617 signal intensity over time allowed us to effectively detect CAR T cells in vivo.ConclusionOur findings preliminarily confirmed that [68Ga]Ga-PSMA-617 PET/CT could reliably detect CAR-Delta PSMA T cells in vitro and in vivo in solid tumors, laying the foundation for the monitoring CAR T cell therapy in the future.