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Clinical‐grade preparation of human natural regulatory T‐cells encoding the thymidine kinase suicide gene as a safety gene
Author(s) -
GuillotDelost Maude,
Cheraï Mustapha,
Hamel Yamina,
Rosenzwajg Michelle,
Baillou Claude,
Simonin Ghislaine,
Leclercq Virginie,
MariottiFerrandiz Maria Encarnita,
Six Adrien,
BonDurand Véronique,
Maury Sébastien,
Salomon Benoit L.,
Cohen José L.,
Klatzmann David,
Lemoine François M.
Publication year - 2008
Publication title -
the journal of gene medicine
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.689
H-Index - 91
eISSN - 1521-2254
pISSN - 1099-498X
DOI - 10.1002/jgm.1220
Subject(s) - foxp3 , il 2 receptor , thymidine kinase , suicide gene , cd90 , ganciclovir , ex vivo , immunology , cd8 , context (archaeology) , immune system , biology , cancer research , medicine , t cell , in vivo , genetic enhancement , microbiology and biotechnology , gene , stem cell , genetics , human cytomegalovirus , paleontology , virus , herpes simplex virus , cd34
Background Human CD4 + CD25 + FOXP3 + natural regulatory T‐cells (nTreg) have a great therapeutic potential for the induction of tolerance in allo‐transplanted patients or for the control of severe auto‐immune diseases. However, clinical‐grade production of nTreg remains difficult to achieve because of the absence of a truly specific surface marker and of their low frequency that implies a need for their ex vivo expansion. Furthermore, safety issues should be taken into consideration due to the risk of either uncontrolled nTreg‐induced immunosuppression or uncontrolled proliferation of autoreactive contaminating T‐cells particularly in an auto‐immune context. Methods We compared different clinical‐grade conditions for immuno‐magnetic selection and ex vivo expansion of nTreg. For safety, expanded cells were genetically modified with retroviral vectors co‐expressing human CD90 and HSV1 thymidine kinase. The CD90 surface marker and thymidine kinase allow for selection and elimination of transduced cells by ganciclovir, respectively. Results We showed that (i) nTreg could be enriched in a one step using CD25 microbeads, were functionally suppressive and mainly FOXP3 + ; (ii) using anti‐CD28‐ and anti‐CD3‐coated beads, interleukin‐2 and rapamycin, nTreg were expanded 150–200‐fold after 3 weeks. Under these clinical‐grade conditions, they remained suppressive, and no major alteration of the TCR repertoire was observed; (iii) after efficient retroviral transduction and CD90 selection, nTreg maintained their suppressive activity; (iv) transduced nTreg could be eliminated by ganciclovir upon activation. Conclusions The efficient procedure reported here for the preparation of nTreg, whose safety has been ensured, is now applicable for further clinical trials. Copyright © 2008 John Wiley & Sons, Ltd.