A tyrosine kinase inhibitor, beta-hydroxyisovalerylshikonin, induced apoptosis in human lung cancer DMS114 cells through reduction of dUTP nucleotidohydrolase activity.
Apoptotic cell death was induced in human lung cancer DMS114 cells by treatment with beta-hydroxyisovalerylshikonin (beta-HIVS), an ATP-noncompetitive inhibitor of protein tyrosine kinases. Changes in phosphoprotein profiles were analyzed by two-dimensional-polyacrylamide gel electrophoresis (2D-PAGE) after the cells were treated with beta-HIVS. One spot on the 2D gel showed a marked decrease in intensity and the corresponding protein was identified by mass spectrometry as dUTP nucleotidohydrolase (dUTPase). The beta-HIVS-induced decrease of dUTPase in the phosphoprotein fraction of DMS114 cells was confirmed using immunoblotting. Treatment of the cells with beta-HIVS-induced rapid reduction of dUTPase activity. An antioxidant N-acetyl-cysteine inhibited both the reduction of phosphorylated dUTPase and the induction of apoptosis by beta-HIVS treatment of DMS114 cells. Introduction of siRNA directed against dUTPase mRNA into DMS114 cells enhanced the susceptibility of beta-HIVS-induced apoptosis. Treatment of DMS114 cells with beta-HIVS and 5-fluorouracil, a specific inhibitor of thymidylate synthase used as a chemotherapeutic drug, revealed the synergistic effects of these drugs on the inhibition of cell growth. These results suggest that dUTPase activity is one of the crucial factors involved in apoptotic cell death in lung cancer cells.
Kajimoto
Sachiko
S
The Laboratory of Biological Chemistry, School of Pharmaceutical Sciences, Showa University, Tokyo, Japan.
Horie
Masayo
M
Manabe
Hitoshi
H
Masuda
Yutaka
Y
Shibayama-Imazu
Toshiko
T
Nakajo
Shigeo
S
Gong
Xiang Feng
XF
Obama
Takashi
T
Itabe
Hiroyuki
H
Nakaya
Kazuyasu
K
D000595:Amino Acid Sequence; D000970:Antineoplastic Agents / Q000494:pharmacology; D000975:Antioxidants / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D003856:Deoxyuracil Nucleotides / Q000235:genetics / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D005472:Fluorouracil / Q000494:pharmacology; D006801:Humans; D008175:Lung Neoplasms / Q000201:enzymology* / Q000473:pathology; D008969:Molecular Sequence Data; D009285:Naphthoquinones / Q000494:pharmacology*; D010750:Phosphoproteins / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D011755:Pyrophosphatases / Q000737:chemistry / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics
D000970:Antineoplastic Agents; D000975:Antioxidants; D003856:Deoxyuracil Nucleotides; D009285:Naphthoquinones; D010750:Phosphoproteins; D047428:Protein Kinase Inhibitors; D034741:RNA, Small Interfering; C118460:beta-hydroxyisovalerylshikonin; C027078:deoxyuridine triphosphate; D011505:Protein-Tyrosine Kinases; D011755:Pyrophosphatases; D005472:Fluorouracil