Conditional cell suicide using dox-dependent caspase-2 expression.
BACKGROUND
Adoptive immune transfer is used as an efficient treatment modality to achieve a graft-versus-leukemia effect in persisting or relapsing residual leukemic disease. Safety considerations dictate the need for equipping the transferred cells with a conditional suicide mechanism to eliminate donor T cells when graft-versus-host disease occurs. We have examined in a model system using HeLa cells whether doxycycline (dox)-dependent expression of pro-apoptotic proteins could be used as a potential new strategy for conditional cell elimination.


METHODS
Four constructs encoding pro-apoptotic proteins were tested in transient transfections to identify suitable cell death inducers. Murine caspase-2 placed under Tet-control was chosen for stable transfection into cell lines carrying different dox-dependent transregulators. The efficiency of cell death induction and the expression patterns of caspase-2 were analyzed in the respective clones.


RESULTS
Different levels of induced cell death were obtained depending on the properties of the transregulators used to control target gene expression. High expression levels of caspase-2 in the presence of dox were required to achieve efficient induction of cell death, while tight repression in the absence of inducer was not necessary for cell survival. Dox treatment for 48 h resulted in 94% cell death indicating a very efficient conditional suicide mechanism.


CONCLUSIONS
We propose that the principle of using pro-apoptotic cellular proteins placed under appropriate dox-dependent regulation may represent an alternative conditional suicide mechanism to the frequently used herpes simplex virus thymidine kinase (HSV-TK)/ganciclovir-system, which harbors immunological and toxicological risks.
Knott
Anja
A
Lehrstuhl für Mikrobiologie, Friedrich-Alexander-Universität Erlangen-Nürnberg, Staudtstr. 5, 91058 Erlangen, Germany.
Müller
Yvonne
Y
Zunino
Susan J
SJ
Berens
Christian
C
Hillen
Wolfgang
W
D000818:Animals; D017209:Apoptosis / Q000502:physiology; D029681:Arabidopsis Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D053143:Caspase 2; D053148:Caspase 3; D020169:Caspases / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects / Q000502:physiology*; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D004247:DNA / Q000032:analysis; D018076:DNA, Complementary; D004318:Doxycycline / Q000494:pharmacology*; D044943:Fatty Acid Desaturases / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D017346:Protein Serine-Threonine Kinases / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D053475:Receptor-Interacting Protein Serine-Threonine Kinase 2; D053422:Receptor-Interacting Protein Serine-Threonine Kinases; D014162:Transfection
D029681:Arabidopsis Proteins; D018076:DNA, Complementary; D004247:DNA; D044943:Fatty Acid Desaturases; C084083:Fad7 protein, Arabidopsis; D017346:Protein Serine-Threonine Kinases; C506253:RIPK2 protein, human; D053475:Receptor-Interacting Protein Serine-Threonine Kinase 2; D053422:Receptor-Interacting Protein Serine-Threonine Kinases; C506255:Ripk2 protein, mouse; C505407:CASP3 protein, human; C505409:Casp3 protein, mouse; D053143:Caspase 2; D053148:Caspase 3; D020169:Caspases; D004318:Doxycycline