Flavopiridol downregulates the expression of both the inducible NO synthase and p27(kip1) in malignant cells from B-cell chronic lymphocytic leukemia.
Flavopiridol, an inhibitor of cyclin-dependent kinases and other protein kinases, induces in vitro apoptosis of malignant cells from B-cell chronic lymphocytic leukemia (B-CLL). Previously, we reported that nitric oxide (NO), produced by an inducible NO synthase (iNOS), spontaneously expressed by the B-CLL cells, contributed to their deficiency in apoptosis. In the present work, we show that ex vivo treatment of leukemic cells from B-CLL patients with flavopiridol results in the inhibition of iNOS expression, as determined by immunofluorescence and Western blotting, and in a marked inhibition of NO production measured in situ with a specific fluorescent probe (DAF-2 DA). These effects are accompanied by membrane, mitochondrial and nuclear events of apoptosis. Flavopiridol exposure also results in the stimulation of caspase 3 activity and in caspase-dependent cleavage of p27(kip1), a negative regulator of the cell cycle, which is overexpressed in B-CLL. Thus, flavopiridol is capable of downregulating both iNOS and p27(kip1) expression in B-CLL cells. Furthermore, flavopiridol-promoted apoptosis is partly reverted by an NO donor, suggesting that inhibition of the NO pathway could participate in the apoptotic effects of flavopiridol on the leukemic cells.
Billard
C
C
U 365 INSERM, Institut Curie, Paris, France.
Kern
C
C
Tang
R
R
Ajchenbaum-Cymbalista
F
F
Kolb
J-P
JP
D000368:Aged; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D018797:Cell Cycle Proteins / Q000378:metabolism*; D050760:Cyclin-Dependent Kinase Inhibitor p27; D015536:Down-Regulation / Q000187:drug effects; D005260:Female; D005419:Flavonoids / Q000494:pharmacology*; D006801:Humans; D066298:In Vitro Techniques; D015451:Leukemia, Lymphocytic, Chronic, B-Cell / Q000188:drug therapy* / Q000378:metabolism*; D008297:Male; D008875:Middle Aged; D009569:Nitric Oxide / Q000378:metabolism; D020030:Nitric Oxide Donors / Q000494:pharmacology; D019001:Nitric Oxide Synthase / Q000378:metabolism*; D052247:Nitric Oxide Synthase Type II; D010880:Piperidines / Q000494:pharmacology*; D025521:Tumor Suppressor Proteins / Q000378:metabolism*
D000970:Antineoplastic Agents; D018797:Cell Cycle Proteins; D005419:Flavonoids; D020030:Nitric Oxide Donors; D010880:Piperidines; D025521:Tumor Suppressor Proteins; D050760:Cyclin-Dependent Kinase Inhibitor p27; D009569:Nitric Oxide; C077990:alvocidib; C496319:NOS2 protein, human; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II