Imaging of genetically engineered T cells by PET using gold nanoparticles complexed to Copper-64

Adoptive transfer of primary T cells genetically modified to have desired specificity can exert an anti-tumor response in some patients. To improve our understanding of their therapeutic potential we have developed a clinically-appealing approach to reveal their in vivo biodistribution using nanoparticles that serve as a radiotracer for imaging by positron emission tomography (PET). T cells electroporated with DNA plasmids from the Sleeping Beauty transposon-transposase system to co-express a chimeric antigen receptor (CAR) specific for CD19 and Firefly luciferase (ffLuc) were propagated on CD19⁺ K562-derived artificial antigen presenting cells. The approach to generating our clinical-grade CAR⁺ T cells was adapted for electro-transfer of gold nanoparticles (GNPs) functionalized with ⁶⁴Cu²⁺ using the macrocyclic chelator (1,4,7,10- tetraazacyclododecane-1,4,7,10-tetraacetic acid, DOTA) and polyethyleneglycol (GNP-⁶⁴Cu/PEG2000). MicroPET/CT was used to visualize CAR ⁺EGFPffLucHyTK⁺GNP-⁶⁴Cu/PEG2000⁺ T cells and correlated with bioluminescence imaging. These data demonstrate that GNPs conjugated with ⁶⁴Cu²⁺ can be prepared as a radiotracer for PET and used to image T cells using an approach that has translational implications.