Ritterazine B, a new cytotoxic natural compound, induces apoptosis in cancer cells.
OBJECTIVE
Ritterazine B, one of the ritterazine analogues extracted from Ritterella tokioka, has been shown to be chemically similar to cephalostatin 1, and among the ritterazine derivatives is the most cytotoxic to P388 murine leukemia cells. The objective of this study was to determine the cytotoxicity of ritterazine B to non-small-cell lung cancer (NSCLC) cells in vitro and its effects on the cell cycle and apoptosis.


METHODS
The cytotoxicity of ritterazine B against PC14 NSCLC cells was investigated using a 4-day MTT assay. Morphological changes in cells after exposure to this compound were evaluated by phase-contrast microscopy. The effects on the cell cycle of HL-60 leukemia cells and PC14 cells were elucidated by flow cytometry and an in vitro CDK/cyclin kinase assay. Induction of apoptosis in HL-60 cells was assessed using the TUNEL assay and Hoechst 33342 staining. In addition, molecules involved in apoptosis were evaluated by Western blotting.


RESULTS
Ritterazine B exerted strong cytotoxic effects against PC14 cells with a mean GI(50) of 75.1 n M. Cell cycle analysis showed that ritterazine B caused accumulation of HL-60 and PC14 cells at the G2/M checkpoint. Furthermore, ritterazine B-treated HL-60 cells became multinucleated, and at a concentration of 20 n M this resulted in the onset of apoptosis. Neither cleavage of caspase target molecules nor phosphorylation of bcl-2 were observed in ritterazine B-treated HL-60 cells.


CONCLUSIONS
These results indicate that ritterazine B might be a potent inducer of apoptosis acting via a novel antimitotic mechanism.
Komiya
Takefumi
T
Division of Medical Oncology, Kinki University School of Medicine, 377-2 Ohnohigashi, 589-8511, Osakasayama City, Osaka, Japan. komiyat@mail.nih.gov
Fusetani
Nobuhiro
N
Matsunaga
Shigeki
S
Kubo
Akihito
A
Kaye
Frederic J
FJ
Kelley
Michael J
MJ
Tamura
Kenji
K
Yoshida
Makoto
M
Fukuoka
Masahiro
M
Nakagawa
Kazuhiko
K
D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D002289:Carcinoma, Non-Small-Cell Lung / Q000473:pathology*; D002453:Cell Cycle / Q000187:drug effects*; D018922:HL-60 Cells; D006801:Humans; D008175:Lung Neoplasms / Q000473:pathology*; D010619:Phenazines / Q000494:pharmacology*; D013141:Spiro Compounds / Q000494:pharmacology*; D013256:Steroids; D014407:Tumor Cells, Cultured
D000970:Antineoplastic Agents; D010619:Phenazines; D013141:Spiro Compounds; D013256:Steroids; C473897:ritterazine B