[Effect of nanosize delivery system for ASODN against hTERT on the expression of telomerase in the esophageal cancer EC9706 cells].
OBJECTIVE
To investigate the inhibitory effect of nanoparticle-mediated antisense oligodeoxynucleotide (ASODN) of human telomerase reverse transcriptase (hTERT) on telomerase in the esophageal cancer EC9706 cells.


METHODS
Line-polyethylenimine (L-PEI) was used to condense ASODN into nanoparticle and to couple NGR peptides into targeting nanoparticle, and the prepared L-PEI/ASODN complexes were transfected into the EC9706 cells. Cellular uptake of L-PEI/ASODN complexes was detected by laser confocal scanning microscopy. MTT assay was used to detect the inhibitory rate of EC9706 cell growth. The level of hTERT mRNA and its protein expression were measured by RT-PCR and immunohistochemistry, respectively. Annexin V FITC/PI double labeling was used to detect cell apoptosis. The distribution of drug in nude mice was observed by laser confocal scanning microscopy, and the growth and morphology of the tumor was examined.


RESULTS
The L-PEI-mediated ASODN uptake was enhanced. After transfection, the inhibitory rate of EC9706 cells was time-dependant and there was a significant difference between control cell group and L-PEI/ASODN group (P < 0.05). At 48 h after transfection, the level of hTERT mRNA was decreased significantly compared with that of control cell group (P < 0.05), and the expression of hTERT protein was negative. There was apparent apoptosis in EC9706 cells after transfection with L-PEI/ASODN complexes. For the two NGR/L-PEI/ASODN groups, fluorescence was observed in the liver, kidney, lung and tumor tissues of nude mice, and their uptake intensity was time-dependent. The mean volume of tumors in the two NGR/L-PEI/ASODN groups was significantly smaller than those in blank control group and SODN group (P < 0.05). Apoptotic bodies were detected in the tumors of L-PEI/ASODN group.


CONCLUSIONS
The NGR/L-PEI/ASODN nanoparticles can effectively reach into the human esophageal cancer xenograft and inhibit the tumor growth in nude mice, and this may provide a theoretical and experimental basis for gene therapy for human esophageal squamous cell carcinoma.
Wang
Jin
J
Department of Pathology, The First Affiliated Hospital, Zhengzhou University, Henan Key Laboratory for Tumor Pathology, Zhengzhou 450052, China.
Zhang
Zhen-zhong
ZZ
Zhou
Tian-yang
TY
Liu
Yu-qiong
YQ
Li
Hui-xiang
HX
D000818:Animals; D017209:Apoptosis; D045744:Cell Line, Tumor; D004938:Esophageal Neoplasms / Q000378:metabolism / Q000473:pathology; D005260:Female; D005822:Genetic Vectors; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D053758:Nanoparticles; D009368:Neoplasm Transplantation; D020319:Oligodeoxyribonucleotides, Antisense / Q000235:genetics / Q000494:pharmacology*; D009842:Oligopeptides / Q000737:chemistry / Q000493:pharmacokinetics; D011094:Polyethyleneimine / Q000737:chemistry / Q000493:pharmacokinetics; D012333:RNA, Messenger / Q000378:metabolism; D019098:Telomerase / Q000235:genetics / Q000378:metabolism*; D014018:Tissue Distribution; D014162:Transfection; D047368:Tumor Burden
C526428:NGR peptide; D020319:Oligodeoxyribonucleotides, Antisense; D009842:Oligopeptides; D012333:RNA, Messenger; D011094:Polyethyleneimine; C509186:TERT protein, human; D019098:Telomerase