Cellular response and molecular mechanism of antitumor activity by leinamycin in MiaPaCa human pancreatic cancer cells.
Previous in vitro biochemical studies have revealed that the antitumor drug leinamycin causes oxidative DNA damage and DNA alkylation. However, it is still not clear whether the same mechanism(s) of action operate in cultured human tumor cells. Here, we evaluated the effects of leinamycin in the human pancreatic carcinoma cell line MiaPaCa. Leinamycin was highly toxic to MiaPaCa cells in vitro, with an IC50 value of 50 nM, and extensive DNA fragmentation was observed in leinamycin-treated MiaPaCa cells. Flow cytometric experiments showed that leinamycin was able to disrupt normal cell cycle progression, resulting in an initial arrest of the cells in S phase. With increased time or at higher concentrations of leinamycin, the population of cells in the sub-G1 phase gradually increased, indicative of apoptotic cell death due to DNA damage. Mammalian Chk2, but not Chk1 kinase, was found to be activated in MiaPaCa cells treated with leinamycin, indicating that cellular responses to leinamycin could be attributed to DNA strand break formation rather than DNA adduct formation. Like other DNA-damaging anticancer drugs, the downregulation of telomerase activity was also observed in MiaPaCa cells at cytotoxic concentrations. However, leinamycin failed to induce DNA ligase I expression in MiaPaCa cells, unlike other DNA-damaging agents, which are known to inhibit DNA replication by arresting DNA replication forks. Taken together, the results from our study indicate that the DNA strand breakage caused by the oxidative DNA-damaging property of leinamycin is directly related to the cellular responses of this drug in MiaPaCa cells over the DNA alkylation property in a dose-responsive manner.
Department of Pharmacology and Toxicology, University of Arizona, Tucson 85724, USA.
Stephen
S
Bassett
Rheanna
R
Urrabaz
Daekyu
D
Sun
D000903:Antibiotics, Antineoplastic / Q000494:pharmacology*; D015153:Blotting, Western; D002455:Cell Division / Q000187:drug effects; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D064447:Checkpoint Kinase 2; D004249:DNA Damage; D004273:DNA, Neoplasm / Q000187:drug effects*; D015536:Down-Regulation; D004789:Enzyme Activation / Q000187:drug effects; D005434:Flow Cytometry; D006801:Humans; D007769:Lactams; D018942:Macrolides / Q000494:pharmacology*; D010190:Pancreatic Neoplasms / Q000201:enzymology / Q000473:pathology; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D019098:Telomerase / Q000096:biosynthesis; D013844:Thiazoles / Q000494:pharmacology*; D013871:Thiones / Q000494:pharmacology*
D000903:Antibiotics, Antineoplastic; D004273:DNA, Neoplasm; D007769:Lactams; D018942:Macrolides; D013844:Thiazoles; D013871:Thiones; C059000:leinamycin; D064447:Checkpoint Kinase 2; C578634:CHEK2 protein, human; D017346:Protein Serine-Threonine Kinases; D019098:Telomerase