PDE4 inhibitors activate a mitochondrial apoptotic pathway in chronic lymphocytic leukemia cells that is regulated by protein phosphatase 2A.
Chronic lymphocytic leukemia (CLL) cells, but not peripheral blood T cells, undergo apoptosis following treatment with inhibitors of type 4 cyclic nucleotide phosphodiesterase (PDE4), a process that correlates dose dependently with elevation of adenosine 3',5'-cyclic monophosphate (cAMP) in leukemic cells. We show that treatment of CLL cells with rolipram, a prototypic PDE4 inhibitor, and forskolin, an adenylate cyclase activator, induces mitochondrial depolarization, release of cytochrome c into the cytosol, caspase-9 and -3 activation, and cleavage of poly(adenosine diphosphate [ADP]-ribose)polymerase. Inhibitors of caspase-9, but not caspase-8, block rolipram/forskolin-induced CLL apoptosis. In a subset of CLL patients, B-cell lymphoma 2 (Bcl-2)-associated death promoter homolog (Bad), a proapoptotic Bcl-2 family member that when phosphorylated on specific serine residues is sequestered in the cytosol by 14-3-3, was dephosphorylated at Ser112 following rolipram/forskolin treatment of leukemic cells. Rolipram/forskolin treatment also induced Bad to accumulate in CLL heavy-membrane fractions, consistent with Bad translocation to mitochondria. To determine the mechanism for rolipram/forskolin-induced Bad dephosphorylation, we examined CLL phosphatase activity. Rolipram/forskolin treatment augmented protein phosphatase 2A (PP2A) activity, as well as levels of immunoreactive PP2A catalytic subunit. Treatment of CLL cells with a concentration of okadaic acid (5 nM) that selectively inhibits PP2A, reduced both rolipram/forskolin-induced mitochondrial cytochrome c release and mitochondrial depolarization. Okadaic acid restored Bad Ser112 phosphorylation and Bad association with 14-3-3 in rolipram/forskolin-treated CLL cells. These results suggest that PDE4 inhibitors may induce CLL apoptosis by activating PP2A-induced dephosphorylation of proapoptotic BH3-only Bcl-2 family members such as Bad.
Moon
Eun-Yi
EY
Evans Department of Medicine, Section of Hematology and Oncology, Boston Medical Center, Boston, MA, USA.
Lerner
Adam
A
D015105:3',5'-Cyclic-AMP Phosphodiesterases / Q000037:antagonists & inhibitors*; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D002458:Cell Fractionation; D005576:Colforsin / Q000494:pharmacology; D054703:Cyclic Nucleotide Phosphodiesterases, Type 4; D003574:Cytochrome c Group / Q000378:metabolism; D004171:Diploidy; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005434:Flow Cytometry; D006801:Humans; D007425:Intracellular Membranes / Q000187:drug effects / Q000473:pathology / Q000502:physiology; D007700:Kinetics; D015451:Leukemia, Lymphocytic, Chronic, B-Cell / Q000201:enzymology / Q000473:pathology*; D008928:Mitochondria / Q000187:drug effects / Q000473:pathology* / Q000502:physiology; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D054648:Protein Phosphatase 2; D020889:Rolipram / Q000494:pharmacology*; D014407:Tumor Cells, Cultured
D003574:Cytochrome c Group; D004791:Enzyme Inhibitors; D005576:Colforsin; D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2; D015105:3',5'-Cyclic-AMP Phosphodiesterases; D054703:Cyclic Nucleotide Phosphodiesterases, Type 4; C505407:CASP3 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases; D020889:Rolipram