RESEARCH ARTICLE


Eliminating Target Cells by Inducing Apoptosis-Related Factors Using hTERT Promoter



Tomoki Takashinaa, Manabu Nakayama*, a, b
a Laboratory of Pharmacogenomics, Graduate School of Pharmaceutical Sciences, Chiba University, 2-6-7 Kazusa- Kamatari, Kisarazu, Chiba 292-0818, Japan
b Department of Human Genome Research, Kazusa DNA Research Institute, 2-6-7 Kazusa-Kamatari, Kisarazu, Chiba 292-0818, Japan


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© 2010 Takashina and Nakayama

open-access license: This is an open access article distributed under the terms of the Creative Commons Attribution 4.0 International Public License (CC-BY 4.0), a copy of which is available at: (https://creativecommons.org/licenses/by/4.0/legalcode). This license permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

* Address correspondence to this author at the 2-6-7 Kazusa-Kamatari, Kisarazu, Chiba 292-0818, Japan; Tel: +81-438-52-3909; Fax: +81-438-52-3931; E-mail: nmanabu@kazusa.or.jp


Abstract

Inducing the complete apoptosis cascade in target cells is useful for eliminating cancer cells and for producing animal models lacking specific cell types. Neuroblastoma SH-SY5Y cells lack caspase-8 and are thus resistant to the apoptosis-inducing effects of 2DEDplusE, an engineered Fas-associated death domain protein (FADD) containing tandem death effector domains (DEDs) of FADD and lambda phage E protein. To overcome tumor cells' resistance to apoptosisinducing factors, we produced new engineered factors ― 2DEDcasp8, 2DEDcasp8 Δ DEDa, and 2DEDcasp8 Δ DEDab ― by fusing the tandem DEDs of FADD to procaspase-8 or its different segments. Of the three, 2DEDcasp8 Δ DEDa most effectively induced apoptosis in SH-SY5Y cells. Finally, by using the human telomerase reverse transcriptase (hTERT) promoter, a known cancer cell-specific promoter, and adapting it to a mifepristone-inducing system, we expressed engineered factors that induced apoptosis in HeLa and A549 tumor cells. The same system did not induce apoptosis in normal Wi-38 and MRC-5 cells, although driving the same system with a constitutive thymidine kinase (TK) promoter induced apoptosis in both cell types.

Keywords: Apoptosis, FADD, Biotechnology, Engineered protein, Human telomerase reverse transcriptase.