Secretory Transactivating Transcription-apoptin fusion protein induces apoptosis in hepatocellular carcinoma HepG2 cells.
OBJECTIVE
To determine whether SP-TAT-apoptin induces apoptosis and also maintains its tumor cell specificity.


METHODS
In this study, we designed a secretory protein by adding a secretory signal peptide (SP) to the N terminus of Transactivating Transcription (TAT)-apoptin (SP-TAT-apoptin), to test the hypothesis that it gains an additive bystander effect as an anti-cancer therapy. We used an artificial human secretory SP whose amino acid sequence and corresponding cDNA sequence were generated by the SP hidden Markov model.


RESULTS
In human liver carcinoma HepG2 cells, SP-TAT-apoptin expression showed a diffuse pattern in the early phase after transfection. After 48 h, however, it translocated into the nuclear compartment and caused massive apoptotic cell death, as determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and annexin-V binding assay. SP-TAT-apoptin did not, however, cause any cell death in non-malignant human umbilical vein endothelial cells (HUVECs). Most importantly, the conditioned medium from Chinese hamster ovary (CHO) cells transfected with SP-TAT-apoptin also induced significant cell death in HepG2 cells, but not in HUVECs.


CONCLUSIONS
The data demonstrated that SP-TAT-apoptin induces apoptosis only in malignant cells, and its secretory property might greatly increase its potency once it is delivered in vivo for cancer therapy.
Han
Su-Xia
SX
College of Life Sciences, Northwest University, Xi'an 710069, Shaanxi Province, China.
Ma
Jin-Lu
JL
Lv
Yi
Y
Huang
Chen
C
Liang
Hai-Hua
HH
Duan
Kang-Min
KM
D000818:Animals; D017209:Apoptosis; D016466:CHO Cells; D036022:Capsid Proteins / Q000235:genetics / Q000378:metabolism*; D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000378:metabolism* / Q000473:pathology / Q000628:therapy; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism / Q000473:pathology; D006224:Cricetinae; D003412:Cricetulus; D017077:Culture Media, Conditioned / Q000378:metabolism; D042783:Endothelial Cells / Q000378:metabolism; D015696:Gene Products, tat / Q000235:genetics / Q000378:metabolism*; D015316:Genetic Therapy / Q000379:methods; D006801:Humans; D008113:Liver Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D021382:Protein Sorting Signals / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013997:Time Factors; D014162:Transfection
D036022:Capsid Proteins; D017077:Culture Media, Conditioned; D015696:Gene Products, tat; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins