Imatinib enhances human melanoma cell susceptibility to TRAIL-induced cell death: Relationship to Bcl-2 family and caspase activation.
In order to define genetic determinants of primary and metastatic melanoma cell susceptibility to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), we have applied oligonucleotide microarrays to TRAIL-sensitive primary T1 cells and TRAIL-resistant metastatic G1 cells treated or not with TRAIL. T1 and G1 cells are isogenic melanoma cell subclones. We examined 22 000 spots, 4.2% of which displayed differential expression in G1 and T1 cells. Cell susceptibility to TRAIL-mediated apoptosis was found to be correlated with gene expression signatures in this model. Some of the differentially expressed genes were identified as involved in ATP-binding and signaling pathways, based on previously published data. Further analysis provided evidences that c-kit was overexpressed in G1 cells while it was absent in T1 cells. The c-kit inhibitor, imatinib, did not restore TRAIL sensitivity, excluding a role for c-kit in TRAIL resistance in G1 cells. Surprisingly, imatinib inhibited cell proliferation and TRAIL-mediated apoptosis in melanoma cells. We investigated the possible involvement of several molecules, including c-ABL, platelet-derived growth factor receptor (PDGFR), cellular FADD-like interleukin-1 alpha-converting enzyme-like inhibitory protein (c-FLIP)(L/S), Fas-associated DD kinase, p53, p21(WAF1), proteins of B-cell leukemia/lymphoma 2 (Bcl-2) family and cytochrome c. Imatinib did not modulate the expression or activation of its own targets, such as c-ABL, PDGFRalpha and PDGFRbeta, but it did affect the expression of c-FLIP(L), BCL2-associated X protein (Bax) and Bcl-2. Moreover, c-FLIP(L) knockdown sensitized T1 cells to TRAIL-mediated apoptosis, with a sensitivity similar to that of cells previously treated with imatinib. More notably, we found that the resistance to TRAIL in G1 cells was correlated with constitutive c-FLIP(L) recruitment to the DISC and the inhibition of caspase 8, 3 and 9 processing. Moreover, c-FLIP(L) knockdown partly restored TRAIL sensitivity in G1 cells, indicating that the expression level of c-FLIP(L) and its interaction with TRAIL receptor2 play a crucial role in determining TRAIL resistance in metastatic melanoma cells. Our results also show that imatinib enhances TRAIL-induced cell death independently of BH3-interacting domain death agonist translocation, in a process involving the Bax:Bcl-X(L) ratio, Bax:Bcl-X(L)/Bcl-2 translocation, cytochrome c release and caspase activation. Our data indicate that imatinib sensitizes T1 cells by directly downregulating c-FLIP(L), with the use of an alternative pathway for antitumor activity, because PDGFRalpha is not activated in T1 cells and these cells do not express c-kit, c-ABL or PDGFRbeta. Caspase cascade activation and mitochondria also play a key role in the imatinib-mediated sensitization of melanoma cells to the proapoptotic action of TRAIL.
Hamaï
A
A
INSERM, U753, Laboratoire d'Immunologie des Tumeurs Humaines: Interaction effecteurs cytotoxiques-système tumoral, Institut Gustave Roussy PR1 and IFR 54, Villejuif, France.
Richon
C
C
Meslin
F
F
Faure
F
F
Kauffmann
A
A
Lecluse
Y
Y
Jalil
A
A
Larue
L
L
Avril
M F
MF
Chouaib
S
S
Mehrpour
M
M
D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D001549:Benzamides; D020169:Caspases / Q000378:metabolism*; D018929:Cell Culture Techniques; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D004789:Enzyme Activation; D020869:Gene Expression Profiling; D006801:Humans; D000068877:Imatinib Mesylate; D008545:Melanoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D009362:Neoplasm Metastasis / Q000235:genetics / Q000473:pathology; D020411:Oligonucleotide Array Sequence Analysis; D010879:Piperazines / Q000494:pharmacology*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism*; D019009:Proto-Oncogene Proteins c-kit / Q000378:metabolism; D011743:Pyrimidines / Q000494:pharmacology*; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand / Q000378:metabolism; D011994:Recombinant Proteins / Q000494:pharmacology; D053221:TNF-Related Apoptosis-Inducing Ligand / Q000494:pharmacology*
D000970:Antineoplastic Agents; D001549:Benzamides; D010879:Piperazines; D019253:Proto-Oncogene Proteins c-bcl-2; D011743:Pyrimidines; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand; D011994:Recombinant Proteins; D053221:TNF-Related Apoptosis-Inducing Ligand; C505653:TNFSF10 protein, human; D000068877:Imatinib Mesylate; D019009:Proto-Oncogene Proteins c-kit; D020169:Caspases