Cell Fate Control Gene Therapy Based on Engineered Variants of Human Deoxycytidine Kinase

被引:15
作者
Neschadim, Anton [2 ]
Wang, James C. M. [3 ]
Sato, Takeya [4 ]
Fowler, Daniel H. [5 ]
Lavie, Arnon [6 ]
Medin, Jeffrey A. [1 ,2 ,3 ]
机构
[1] Univ Hlth Network, Toronto, ON M5G 2M1, Canada
[2] Univ Toronto, Dept Med Biophys, Toronto, ON, Canada
[3] Univ Toronto, Inst Med Sci, Toronto, ON M5S 1A1, Canada
[4] Tohoku Univ, Sendai, Miyagi 980, Japan
[5] NCI, Expt Transplantat & Immunol Branch, NIH, Bethesda, MD 20892 USA
[6] Univ Illinois, Chicago, IL USA
关键词
DONOR T-CELLS; SUICIDE GENE; THYMIDYLATE SYNTHASE; VARICELLA-ZOSTER; STRUCTURAL BASIS; FUSION PROTEIN; SAFETY SWITCH; HOST-DISEASE; THYMIDINE; (E)-5-(2-BROMOVINYL)-2'-DEOXYURIDINE;
D O I
10.1038/mt.2011.298
中图分类号
Q81 [生物工程学(生物技术)]; Q93 [微生物学];
学科分类号
071005 ; 0836 ; 090102 ; 100705 ;
摘要
The safety of cell therapy applications can be enhanced by the introduction of Cell Fate Control (CFC) elements, which encode pharmacologically controlled cellular suicide switches. CFC Gene Therapy (CFCGT) offers the possibility of establishing control over gene-modified cells (GMCs) with regards to their proliferation, differentiation, or function. However, enzymes commonly employed in these approaches often possess poor kinetics and high immunogenicity. We describe a novel CFCGT system based on engineered variants of human deoxyCytidine Kinase (dCK) that overcomes limitations of current modalities. Mutants of dCK with rationally designed active sites that make them thymidine-activating were stably introduced into cells by recombinant lentiviral vectors (LVs). Transduced cells maintained growth kinetics and function. These dCK mutants efficiently activate bromovinyl-deoxyuridine (BVdU), L-deoxythymidine (LdT), and L-deoxyuridine (LdU), which are otherwise not toxic to wild-type cells. We show that mutant dCK-expressing Jurkat, Molt-4, and U87mg cells could be efficiently eliminated in vitro and in xenogeneic leukemia and tumor models in vivo. We also describe a fusion construct of the thymidine-activating dCK to the cytoplasmic tail-truncated LNGFR molecule and applications to in vivo eradication of primary human T cells. This novel CFCGT system offers unique plasticity with respect to the wide range of prodrugs it can potentiate, and can be used as a reliable safety switch in cell and gene therapy.
引用
收藏
页码:1002 / 1013
页数:12
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