Transfection of thymidine phosphorylase cDNA to human hepatocellular carcinoma cells enhances sensitivity to fluoropyrimidine but augments endothelial cell migration.
OBJECTIVE
To investigate the effects on sensitivity to fluoropyrimidine and endothelial cell (EC) migration by transfection with thymidine phosphorylase (TP) cDNA to a hepatocellular carcinoma (HCC) cell line SMMC-7721.


METHODS
SMMC-7721 was transfected with pcDNA3.1/zeo (+) with human TP cDNA. TP mRNA expression was determined by RT-PCR. Sensitivity to fluoropyrimidine was determined by 3-[4,5-dimethylthiazol-2-yl]-2, 5-diphenyl-tetrazolium bromide (MTT) assay. Induction of EC migration was detected by Boyden chamber assay.


RESULTS
The construction of pcDNA3.1/zeo(+)-TP was verified by digestion with restriction endonuclease Apa1. When comparison was made between SMMC-7721 cell clone transfected with pcDNA3.1/zeo(+)-TP (S-TP) and control clone transfected with pcDNA3.1/zeo(+) (S-vector), we found that TP mRNA expression level was much higher in S-TP, being 2.09+/-0.16 vs 0.48+/-0.06 in S-vector (P < 0.01), sensitivity to 5'-deoxy-5-fluorouridine (5'-dFUrd, a prodrug of 5-fluorouracil) in S-TP was significantly enhanced compared with that in S-vector (IC(50); 56.81+/-9.85 micromol/l vs 162.25+/-11.03 micromol/l, P < 0.01), and the culture medium of S-TP possessed more potential to induce EC migration than that of S-vector (the number of ECs appearing on the outer surfaces of the membrane was 275+/-29 vs 122+/-35 per field, P < 0.01).


CONCLUSIONS
Sensitivity to 5'-dFUrd could be enhanced by transfection with TP cDNA for SMMC-7721 cells. However, EC migration was also promoted at the same time. Therefore, transfection with TP alone might have no potential to enhance anti-tumoral effects of fluoropyrimidine in HCC.
Zhou
Jian
J
Liver Cancer Institute and Zhongshan Hospital, Fudan University, Shanghai, 200032, People's Republic of China.
Xiao
Yong-Sheng
YS
Tang
Zhao-You
ZY
Fan
Jia
J
Wu
Zhi-Quan
ZQ
Zhao
Yan
Y
Xue
Qiong
Q
Shen
Zao-Zhuo
ZZ
Liu
Yin-Kun
YK
Ye
Sheng-Long
SL
D000951:Antigens, Neoplasm / Q000032:analysis; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology*; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy* / Q000201:enzymology / Q000473:pathology; D002465:Cell Movement / Q000187:drug effects; D004250:DNA Topoisomerases, Type II / Q000032:analysis; D018076:DNA, Complementary / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000032:analysis; D018572:Disease-Free Survival; D005467:Floxuridine / Q000494:pharmacology*; D006801:Humans; D007150:Immunohistochemistry; D017404:In Situ Hybridization, Fluorescence; D008113:Liver Neoplasms / Q000188:drug therapy* / Q000201:enzymology / Q000473:pathology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016019:Survival Analysis; D013939:Thymidine Phosphorylase / Q000235:genetics*; D014162:Transfection
D000951:Antigens, Neoplasm; D000964:Antimetabolites, Antineoplastic; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D005467:Floxuridine; D013939:Thymidine Phosphorylase; D004250:DNA Topoisomerases, Type II; C025034:doxifluridine