Barnase as a New Therapeutic Agent Triggering Apoptosis in Human Cancer Cells

被引:67
作者
Edelweiss, Evelina [1 ]
Balandin, Taras G. [1 ]
Ivanova, Julia L. [2 ]
Lutsenko, Gennady V. [1 ]
Leonova, Olga G. [2 ]
Popenko, Vladimir I. [2 ]
Sapozhnikov, Alexander M. [1 ]
Deyev, Sergey M. [1 ,3 ]
机构
[1] Russian Acad Sci, Shemyakin & Ovchinnikov Inst Bioorgan Chem, Moscow, Russia
[2] Russian Acad Sci, Engelhardt Inst Mol Biol, Moscow, Russia
[3] Russian Acad Sci, Inst Gene Biol, Moscow, Russia
来源
PLOS ONE | 2008年 / 3卷 / 06期
基金
俄罗斯基础研究基金会;
关键词
D O I
10.1371/journal.pone.0002434
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
Background: RNases are currently studied as non-mutagenic alternatives to the harmful DNA-damaging anticancer drugs commonly used in clinical practice. Many mammalian RNases are not potent toxins due to the strong inhibition by ribonuclease inhibitor (RI) presented in the cytoplasm of mammalian cells. Methodology/Principal Findings: In search of new effective anticancer RNases we studied the effects of barnase, a ribonuclease from Bacillus amyloliquefaciens, on human cancer cells. We found that barnase is resistant to RI. In MTT cell viability assay, barnase was cytotoxic to human carcinoma cell lines with half-inhibitory concentrations (IC50) ranging from 0.2 to 13 mu M and to leukemia cell lines with IC50 values ranging from 2.4 to 82 mu M. Also, we characterized the cytotoxic effects of barnase-based immunoRNase scFv 4D5-dibarnase, which consists of two barnase molecules serially fused to the single-chain variable fragment (scFv) of humanized antibody 4D5 that recognizes the extracellular domain of cancer marker HER2. The scFv 4D5-dibarnase specifically bound to HER2-positive cells and was internalized via receptor-mediated endocytosis. The intracellular localization of internalized scFv 4D5-dibarnase was determined by electronic microscopy. The cytotoxic effect of scFv 4D5-dibarnase on HER2-positive human ovarian carcinoma SKOV-3 cells (IC50 = 1.8 nM) was three orders of magnitude greater than that of barnase alone. Both barnase and scFv 4D5-dibarnase induced apoptosis in SKOV-3 cells accompanied by internucleosomal chromatin fragmentation, membrane blebbing, the appearance of phosphatidylserine on the outer leaflet of the plasma membrane, and the activation of caspase-3. Conclusions/Significance: These results demonstrate that barnase is a potent toxic agent for targeting to cancer cells.
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页数:13
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