Engraftment and tumor formation after allogeneic in utero transplantation of primate embryonic stem cells

Background. To achieve human embryonic stem (ES) cell-based transplantation therapies, allogeneic transplantation models of nonhuman primates would be useful. We have prepared cynomolgus ES cells genetically marked with the green fluorescent protein (GFP). The cells were transplanted into the allogeneic fetus, taking advantage of the fact that the fetus is so immunologically immature as not to induce immune responses to transplanted cells and that fetal tissue compartments are rapidly expanding and thus providing space for the engraftment. Methods. Cynomolgus ES cells were genetically modified to express the GFP gene using a simian immunodeficiency viral vector or electroporation. These cells were transplanted in utero with ultrasound guidance into the cynomolgus fetus in the abdominal cavity (n=2) or liver (n=2) at the end of the first trimester. Three fetuses were delivered 1 month after transplantation, and the other, 3 months after transplantation. Fetal tissues were examined for transplanted cell progeny by quantitative polymerase chain reaction and in situ polymerase chain reaction of the GFP sequence. Results. A fluorescent tumor, obviously derived from transplanted ES cells, was found in the thoracic cavity at 3 months after transplantation in one fetus. However, transplanted cell progeny were also detected (∼1%) without teratomas in multiple fetal tissues. The cells were solitary and indistinguishable from surrounding host cells. Conclusions. Transplanted cynomolgus ES cells can be engrafted in allogeneic fetuses. The cells will, however, form a tumor if they leak into an improper space such as the thoracic cavity.