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EMBRYONIC STEM CELLS: CHARACTERIZATION SERIES

Ablation of Undifferentiated Human Embryonic Stem Cells: Exploiting Innate Immunity Against the Gal 1-3Galß1-4GlcNAc-R (-Gal) Epitope

Department of Gene Function and Development, Roslin Institute, Roslin, Midlothian, United Kingdom

Key Words. Regenerative medicine • Selective ablation • Human embryonic stem cells • Stem cell therapy Complement-mediated lysis • Gal 1,3-galactosyltransferase

Correspondence: Jim McWhir, Ph.D., Roslin Institute, Roslin, Midlothian, EH25 9PS, United Kingdom. Telephone: +44 131 527 4334; Fax: +44 131440 0434; e-mail: jim.mcwhir{at}bbsrc.ac.uk

Received September 30, 2005; accepted for publication August 29, 2006.
First published online in STEM CELLS EXPRESS   September 7, 2006.



Although undifferentiated human embryonic stem cells (hESCs) are tumorigenic, this capacity is lost after differentiation, and hESCs are being widely investigated for applications in regenerative medicine. To engineer protection against the unintentional transplantation of undifferentiated cells, we generated hESCs carrying a construct in which the 1,3-galactosyltransferase (GalT) open reading frame was transcribed from the hTERT promoter (pmGT). Because the endogenous GalT gene is inactive, GalT expression was limited to undifferentiated cells. A second chimeric construct (pmfGT) differed by replacement of the GalT leader sequence for that of the fucosyltransferase gene. Two subclones containing stable integrations of pmGT and pmfGT (M2 and F11, respectively) were assessed for their response to human serum containing antibodies to the Gal1-3Galß1-4GlcNAc-R (-gal) epitope. The low-variegation line, M2, and to a lesser extent the more variegated line F11, were sensitive to human serum when exposed in the undifferentiated state. However, M2 cells were largely insensitive after differentiation and retained both a normal karyotype and the ability to differentiate into derivatives of the three germ layers in severe combined immunodeficient mice. These data exemplify a method of protection against residual, undifferentiated hESCs prior to engraftment and may provide ongoing immune surveillance after engraftment against dedifferentiation or against de novo tumorigenesis involving hTERT reactivation. Untransfected H9 cells were not sensitive to the human serum used in this study. Hence, in our system, interactions of hESCs with other circulating antibodies, such as anti-Neu5Gc, were not observed.

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