Inhibiting Glucosylceramide Synthase Sensitizes CML T315I Mutant to Bcr-Abl Inhibitor and Cooperatively Induces Glycogen Synthase Kinase-3-regulated Apoptosis

Inhibiting Glucosylceramide Synthase Sensitizes CML T315I Mutant to

Bcr-Abl Inhibitor and Cooperatively Induces Glycogen Synthase

Kinase-3-regulated Apoptosis

Wei-Ching Huang

, Cheng-Chieh Tsai

, Chia-Ling Chen

, Tsai-Yun Chen

, Ya-Ping

Chen

, Yee-Shin Lin

, Pei-Jung Lu

, Chiung-Wen Tsao

, Chi-Yun Wang

, Yi-Lin

Cheng

, Chia-Yuan Hsieh

, and Chiou-Feng Lin

1

Institute of Clinical Medicine,

Institute of Basic Medical Sciences,

Department of

Microbiology and Immunology,

Internal Medicine, and

Department of Medical Laboratory

Science and Biotechnology, College of Medicine, National Cheng Kung University, Tainan

701, Taiwan;

Department of Nursing, Chung Hwa University of Medical Technology, Tainan

717, Taiwan

Inactivation of glycogen synthase kinase (GSK)-3 has been implicated in cancer progression. We previously showed abundance of inactive GSK-3 in human chronic myeloid leukemia (CML) cell line. CML is a hematopoietic malignancy caused by an oncogenic Bcr-Abl tyrosine kinase. In Bcr-Abl signaling, the role of GSK-3 is not well defined. Here we report that enforced expression of constitutively active GSK-3 reduced proliferation and increased Bcr-Abl-inhibition-induced apoptosis by nearly 1-fold. Bcr-Abl inhibition activated GSK-3 and GSK-3-dependent apoptosis. Inactivation of GSK-3 by Bcr-Abl activity is therefore confirmed. To re-activate GSK-3, we inhibited glucosylceramide synthase (GCS) to accumulate endogenous ceramide, a tumor-suppressor sphingolipid and a potent GSK-3 activator. We found silence of GCS and GCS inhibitor increased Bcr-Abl-inhibition-induced apoptosis by 4-fold and 10% respectively. Furthermore, GCS inhibitor sensitized the most clinical problematic drug-resistant CML T315I mutants to Bcr-Abl inhibitor GNF-2- or imatinib-induced apoptosis by more than 5-fold. Combining GCS and Bcr-Abl inhibitors eliminated transplated-CML-T315I-mutants in vivo and dose-dependently sensitized primary cells from CML T315I patients to Bcr-Abl inhibitor-induced proliferation inhibition and apoptosis. The synergistic efficacy was Bcr-Abl-restricted and acted through GSK-3-mediated apoptosis. This study suggests a feasible novel anti-CML strategy by accumulating endogenous ceramide to re-activate GSK-3 and abrogate drug resistance.

* This work was supported by grant NHRI-EX99-9917NC from the National Health Research Institutes and NSC 96-2320-B-006-018-MY3 from the National Science Council, Taiwan, and by the Landmark Project C020 of National Cheng Kung University.