PET Imaging of CAR T Cell Proliferation with ⁶⁸Ga-DOTA-Biotin

Objectives: Chimeric antigen receptor (CAR) T cell therapeutics have revolutionized the treatment landscape for hematologic malignancies, however, many challenges exist in quantitatively monitoring the proliferation and toxic characteristic of CART cells in vivo. The non-invasive PET imaging of CAR T cell with ⁶⁸Ga-DOTA-Biotin may address those issues.

Methods: Anti-CD19 CAR T cells were transduced with the streptavidin (SA) and the specific molecular probe ⁶⁸Ga-DOTA-Biotin was synthesized. Cell uptake experiments were performed to test the binding ability of ⁶⁸Ga-DOTA-Biotin with CD19-tSA CAR T cells. Serial microPET imaging were performed in NOD-scid gamma (NSG) mice models bearing stable expressing hCD19 K562 tumors and normal K562 tumors after intravenous injected with 1*10^6 CD19-tSA CAR T cells, 1*10^6 CD19-CAR T cells and without CART cells, respectively.

Results: AntiCD19 CART cells were successfully transduced with streptavidin at the transduction rate of approximately 10%. The radiolabeled yield of ⁶⁸Ga-DOTA-Biotin was high over 90%. Cell uptake results showed that the radioactivity in the CD19-tSA CAR T cells was linear with the number of cells (R²=0.965). Preliminary PET imaging revealed that the tumor uptake (%ID/g) at day 4 of hCD19-K562 animal models administrated with CD19-tSA CAR T cells was 3-fold higher than day 1, which increased from 0.38 to 1.20. While for other models and administration, the tumor uptake was basically consistent with day 1 (approximately 0.20-0.40). The uptakes of lung, liver and spleen were also quantified in hCD19-K562 animal models. From day 1 to day 4, lung uptake (%ID/g) with CD19-tSA CAR T cells was decreased from 1.19 to 0.76, while for the same animal models without CART cells, lung uptake (%ID/g) was as low as 0.26. The liver and spleen uptake remained stable from day 1 to day 4 for hCD19-K562 animal models administrated with CD19-tSA CAR T cells, and 2-3 folds higher than the same animal models without CD19-tSA CAR T cells.

Conclusions: These preliminary data suggest that the ⁶⁸Ga-DOTA-Biotin may be used as a specific probe to monitor CD19-tSA cells proliferation in vivo. This non-invasive method requires further in-depth research and is expected to provide a fast, easy and repeatable tool for cell proliferation monitoring in CAR T cell therapy. Research Support: This work was supported by National Significant New Drugs Creation Program (2017ZX09304021), Jiangsu Provincial Medical Innovation Team (CXTDA2017024). CAR T cells were provided by Shanghai Unicar-Therapy Bio-Medicine Technology Co., Ltd. We acknowledge the contributions of Prof. Liyan Miao from Department of Clinical Pharmacology, the First Affiliated Hospital of Soochow University.