A pair of congenic mice for imaging of transplants by positron emission tomography using anti-transferrin receptor nanobodies

Two anti-transferrin receptor (TfR) nanobodies, V_HH123 specific for mouse TfR and V_HH188 specific for human TfR (huTfR) were used to track transplants non-invasively by PET/CT in mouse models, without the need for genetic modification of the transferred cells. We provide a comparison of the specificity and kinetics of the PET signals acquired when using nanobodies radiolabeled with⁸⁹Zr,⁶⁴Cu and¹⁸F, and find that the chelation of the⁸⁹Zr and⁶⁴Cu radioisotopes to anti-TfR nanobodies results in radioisotope release upon endocytosis of the radiolabeled nanobodies. We used a knock-in mouse that expresses a TfR with a human ectodomain (huTfR^+/+) as a source of bone marrow for transplants into C57BL/6 recipients and show that V_HH188 detects such transplants by PET/CT. Conversely, C57BL/6 bone marrow and B16.F10 melanoma cell-line transplanted into huTfR^+/+recipients can be imaged with V_HH123. In C57BL/6 mice impregnated by huTfR^+/+males we saw an intense V_HH188 signal in the placenta, showing that TfR-specific V_HHs accumulate at the placental barrier but do not enter the fetal tissue. We were unable to observe accumulation of the anti-TfR radiotracers in the central nervous system (CNS) by PET/CT but show evidence of CNS accumulation by radiospectrometry. The model presented here can be used to track many transplanted cell types by PET/CT, provided cells express TfR, as is typically the case for proliferating cells such as tumor lines.