TY - JOUR
T1 - The effect of FK506 on transforming growth factor β signaling and apoptosis in chronic lymphocytic leukemia B cells
AU - Romano, Simona
AU - Mallardo, Maria
AU - Chiurazzi, Federico
AU - Bisogni, Rita
AU - D'Angelillo, Anna
AU - Liuzzi, Raffaele
AU - Compare, Giovanna
AU - Romano, Maria Fiammetta
PY - 2008/7
Y1 - 2008/7
N2 - Background Loss of response to transforming growth factor-beta (TGF-β) is thought to contribute to the progression of chronic lymphocytic leukemia. Recent findings of over-activation of the TGF-β signal in FKBP12-knockout mouse prompted us to investigate whether FK506, the canonical ligand of FKBR can activate the TGF-β signal in chronic lymphocytic leukemia. Design and Methods We studied 62 chronic lymphocytic leukemia samples from patients with Rai/Binet stage 0 to 4 disease. The TGF-β signal was investigated by western blotting and flow eytometry. The levels of Bc12-family members and death-associated-protein kinase were also investigated by western blotting, whereas apoptosis was studied in flow cytometry. Down-modulation of FKBP12 was obtained by gene silencing with short interfering RNA. Results Twenty-two out of 62 chronic lymphocytic leukemia samples were sensitive to TGF-β-induced apoptosis. All but two of the responsive samples underwent apoptosis also when cultured with FK506, but not with cyclosporine. Thirteen samples that were not sensitive to TGF-β were sensitive to FK506. Overall, response to FK506 occurred in 33 samples. FK506 induced Smad2 phosphorylation and nuclear translocation. Accordingly, death-associated-protein kinase, a transcriptional target of Smad, was induced. At the same time, Bcl-2 and Bcl-xL levels decreased whereas the levels of Bim and Bmf increased. A loss of mitochondrial membrane potential preceded caspase activation and cell death. FK506 removed FKBP12 from its binding to the TGF-β-receptor. FKBP12 release activated the receptor-kinase activity as suggested by the enhanced levels of phospho-Smad found in cells depleted of FKBP12. Conclusions Our study shows that most chronic lymphocytic leukemia cells escape the homeostatic control of TGF-β and that FK506 restores the TGF-β signal in a proportion of non-responsive samples. We demonstrated that FK506 activates TGF-β receptor I kinase activity in chronic lymphocytic leukemia, which transduces apoptosis by a mitochondrial-dependent pathway.
AB - Background Loss of response to transforming growth factor-beta (TGF-β) is thought to contribute to the progression of chronic lymphocytic leukemia. Recent findings of over-activation of the TGF-β signal in FKBP12-knockout mouse prompted us to investigate whether FK506, the canonical ligand of FKBR can activate the TGF-β signal in chronic lymphocytic leukemia. Design and Methods We studied 62 chronic lymphocytic leukemia samples from patients with Rai/Binet stage 0 to 4 disease. The TGF-β signal was investigated by western blotting and flow eytometry. The levels of Bc12-family members and death-associated-protein kinase were also investigated by western blotting, whereas apoptosis was studied in flow cytometry. Down-modulation of FKBP12 was obtained by gene silencing with short interfering RNA. Results Twenty-two out of 62 chronic lymphocytic leukemia samples were sensitive to TGF-β-induced apoptosis. All but two of the responsive samples underwent apoptosis also when cultured with FK506, but not with cyclosporine. Thirteen samples that were not sensitive to TGF-β were sensitive to FK506. Overall, response to FK506 occurred in 33 samples. FK506 induced Smad2 phosphorylation and nuclear translocation. Accordingly, death-associated-protein kinase, a transcriptional target of Smad, was induced. At the same time, Bcl-2 and Bcl-xL levels decreased whereas the levels of Bim and Bmf increased. A loss of mitochondrial membrane potential preceded caspase activation and cell death. FK506 removed FKBP12 from its binding to the TGF-β-receptor. FKBP12 release activated the receptor-kinase activity as suggested by the enhanced levels of phospho-Smad found in cells depleted of FKBP12. Conclusions Our study shows that most chronic lymphocytic leukemia cells escape the homeostatic control of TGF-β and that FK506 restores the TGF-β signal in a proportion of non-responsive samples. We demonstrated that FK506 activates TGF-β receptor I kinase activity in chronic lymphocytic leukemia, which transduces apoptosis by a mitochondrial-dependent pathway.
KW - Apoptosis
KW - Chronic lymphocytic leukemia
KW - FK506
KW - TGF-β
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U2 - 10.3324/haematol.12402
DO - 10.3324/haematol.12402
M3 - Article
C2 - 18492692
AN - SCOPUS:46849115402
VL - 93
SP - 1039
EP - 1048
JO - Haematologica
JF - Haematologica
SN - 0390-6078
IS - 7
ER -