Teratoma formation by human embryonic stem cells: Evaluation of essential parameters for future safety studies

Abstract

Transplantation of human embryonic stem cells (hESC) into immune-deficient mice leads to the formation of differentiated tumors comprising all three germ layers, resembling spontaneous human teratomas. Teratoma assays are considered the gold standard for demonstrating differentiation potential of pluripotent hESC and hold promise as a standard for assessing safety among hESC-derived cell populations intended for therapeutic applications. We tested the potency of teratoma formation in seven anatomical transplantation locations (kidney capsule, muscle, subcutaneous space, peritoneal cavity, testis, liver, epididymal fat pad) in SCID mice with and without addition of Matrigel, and found that intramuscular teratoma formation was the most experimentally convenient, reproducible, and quantifiable. In the same experimental setting, we compared undifferentiated hESC and differentiated populations enriched for either beating cardiomyocytes or definitive endoderm derivatives (insulin-secreting beta cells), and showed that all cell preparations rapidly formed teratomas with varying percentages of mesoderm, ectoderm, and endoderm. In limiting dilution experiments, we found that as little as two hESC colonies spiked into feeder fibroblasts produced a teratoma, while a more rigorous single-cell titration achieved a detection limit of 1/4000. In summary, we established core parameters essential for facilitating safety profiling of hESC-derived products for future therapeutic applications.

Derivation of naïve human embryonic stem cells

“…Studies by others with primed hESC-generated teratomas have noted the disproportionate amount of mesoderm in the tumor mass (amounting to between two-thirds and three-fourths of the tumor) (17,18). As shown in Fig.…”

Mouse Embryonic Stem Cells Are Deficient in Type I Interferon Expression in Response to Viral Infections and Double-stranded RNA

“…Embryonic stem cells (ESCs) 3 have attracted enormous attention in recent years with the expectation that they will be used as a source for cell-based therapy. Although the benefit of this research in medical applications is exciting, currently there is limited understanding of the basic physiology of ESCs and their derived cells.…”

“…However, it is increasingly clear that generating clinically usable cells is a task that faces many biological and technical challenges. For instance, it is well recognized that if ESCs are not fully differentiated and transplanted to the patient, they can grow into tumor-like structures known as teratomas (3). However, there are many other concerns that have not been fully recognized.…”

Induced pluripotent stem cells and severe combined immunodeficiency: merely disease modeling or potentially a novel cure?

“…Because most of the pluripotency genes have been associated with tumorigenesis, incomplete silencing or reactivation of the inserted proviruses can cause tumors (49)(50)(51)(52). As a consequence, iPSCs used for in vivo disease modeling should preferentially be generated by different methods, and when future therapeutic applications are considered, nonintegrating delivery systems are required.…”