P20
Generation and analysis of cardiac progenitors expressing the DTPA-R PET-reporter gene for in vivo tracking of the cells in regenerative cell therapies
S Seyfi(1,2) T Dorn(2) C M Poch(2) V Fricke(2) V Morath(1) K Fritschle(1) A Skerra(4) M Schwaiger(1) W A Weber(1) C Kupatt(2,3) K L Laugwitz(2,3) A Moretti(2,3)
1:Department of Nuclear Medicine, Klinikum rechts der Isar, School of Medicine, Technical University of Munich, 81675 Munich, Germany; 2:Department of Internal Medicine, Klinikum rechts der Isar, School of Medicine, Technical University of Munich, 81675 Munich, Germany; 3:Center for Cardiovascular Research (DZHK), Munich Heart Alliance (MHA), Partner Site Munich, 81377 Munich, Germany; 4:Lehrstuhl für Biologische Chemie, School of Life Sciences, Technical University of Munich, 85354 Freising, Germany
Human heart regeneration is one of the most critical unmet clinical needs at a global level. The development of cell-based strategies for heart regeneration is an intense area of research. Recently, we demonstrated that the transplantation of human pluripotent stem cell (hPSC)-derived ventricular progenitors (HVPs) in injured pig hearts promotes remuscularization and prevents heart failure progression. Here, we introduce a novel reporter gene system (DTPA-R) to human pluripotent stem cell (hPSC) with the potential to quantitatively track the number and viability of transplanted cells in vivo. Tracking the grafted HVPs in vivo can provide useful dynamic information about their migration, viability, and functional effects within the heart. Importantly, the combination of PET and magnetic resonance imaging (MRI) would allow parallel assessment of heart function and its improvement upon HVP treatment.
The engineered hiPSCs were created by nucleofection and antibiotic selection. Correct gene knock-in, the absence of off-site mutations, and the absence of chromosomal abnormalities in the cells were determined. The DTPA-R reporter was expressed at the cell surface at levels of about 2 million copies for hiPSCs. Expression of the receptor was confirmed in HVPs and CMs at later time points. Uptake of the reporter probe [¹⁸F]F-DTPA•ⁿᵃᵗTb of induced cells was measured in 2D cell culture and by ex vivo PET imaging of HVPs applied on native 3D pig heart slices. HVPs survival, differentiation, and ability of specific and efficient binding of the radioligand to the receptor was proven and the bound activity correlated with the number of applied cells. Non-specific binding to native pig heart tissue was negligible. Subsequent immunofluorescent analysis of the tissues overlayed with the obtained PET signals.
Using the [¹⁸F]F-DTPA•ⁿᵃᵗTb system, life monitoring of cellular location and viability of transplanted HVPs and their regenerative effect after myocardial infarction, as well as potential off-target enrichments of cells outside the heart is possible. Further investigations in animal models are essential to get a better understanding of the dynamic behavior of the cells in vivo.
This multimodal imaging could help improve preclinical research as well as clinical studies of regenerative cell therapies.