Mechanisms of the apoptotic activity of Cl-F-araA in a human T-ALL cell line, CCRF-CEM.
OBJECTIVE
The purpose of the present study was to characterize the mechanisms of the antitumor activity of 2-chloro-9-(2-deoxy-2-fluoro-beta- D-arabinofuranosyl)adenine (Cl-F-araA) against lymphocytic leukemia. Recent evidence indicates that Cl-F-araA has more potent antitumor activity in vitro against human leukemia cell lines than against human solid tumor cell lines originating from different tissue. We analyzed the mechanism of action of Cl-F-araA using a human T-acute lymphocytic leukemia cell line, CCRF-CEM, in vitro and in vivo.


RESULTS
Cl-F-araA exhibited marked antitumor activity in vitro and in vivo, and this was correlated with its ability to induce apoptosis, particularly in vivo. To analyze the mechanisms of the apoptotic activity of Cl-F-araA, we sought to determine the effects of the drug on the levels of Bcl-2 family proteins (Bcl-2, Bcl-X(L), Mcl-1, Bax, Bak) and cell survival signals via Akt. Western blot analysis revealed that Cl-F-araA induced a dose- and time-dependent downregulation of Bcl-X(L) and Mcl-1 proteins, and a dose- and time-dependent dephosphorylation of Akt and its downstream effectors (Bad, FKHRL1), particularly in vivo. In addition, there was a marked increase in the population of cells in G(1)/S and early S phase. We therefore investigated the changes in the Cdc25A protein to characterize the mechanism involved in the G(1)/S accumulation. Cl-F-araA induced a dose- and time-dependent downregulation of the Cdc25A protein whereas the Cdc25C protein remained unchanged. We further found that in combination with caffeine, Cl-F-araA potentiated apoptosis induction.


CONCLUSIONS
Taken together, our findings suggest that Cl-F-araA may be an effective drug in vivo.
Takahashi
Takeshi
T
Department of Oncology, Pharmaceutical Research Institute, Kyowa Hakko Kogyo Co. Ltd., 1188 Shimotogari, Nagaizumi-cho, Sunto-gun, Shizuoka, 411-8731, Japan.
Shimizu
Makiko
M
Akinaga
Shiro
S
D000227:Adenine Nucleotides; D017209:Apoptosis / Q000187:drug effects*; D001087:Arabinonucleosides / Q000494:pharmacology*; D000077866:Clofarabine; D015536:Down-Regulation; D006801:Humans; D054198:Precursor Cell Lymphoblastic Leukemia-Lymphoma / Q000188:drug therapy* / Q000473:pathology; D014407:Tumor Cells, Cultured; D020687:cdc25 Phosphatases / Q000096:biosynthesis
D000227:Adenine Nucleotides; D001087:Arabinonucleosides; D000077866:Clofarabine; C517004:CDC25A protein, human; D020687:cdc25 Phosphatases