Intracellular triggering of Fas aggregation and recruitment of apoptotic molecules into Fas-enriched rafts in selective tumor cell apoptosis.
We have discovered a new and specific cell-killing mechanism mediated by the selective uptake of the antitumor drug 1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine (ET-18-OCH(3), Edelfosine) into lipid rafts of tumor cells, followed by its coaggregation with Fas death receptor (also known as APO-1 or CD95) and recruitment of apoptotic molecules into Fas-enriched rafts. Drug sensitivity was dependent on drug uptake and Fas expression, regardless of the presence of other major death receptors, such as tumor necrosis factor (TNF) receptor 1 or TNF-related apoptosis-inducing ligand R2/DR5 in the target cell. Drug microinjection experiments in Fas-deficient and Fas-transfected cells unable to incorporate exogenous ET-18-OCH(3) demonstrated that Fas was intracellularly activated. Partial deletion of the Fas intracellular domain prevented apoptosis. Unlike normal lymphocytes, leukemic T cells incorporated ET-18-OCH(3) into rafts coaggregating with Fas and underwent apoptosis. Fas-associated death domain protein, procaspase-8, procaspase-10, c-Jun amino-terminal kinase, and Bid were recruited into rafts, linking Fas and mitochondrial signaling routes. Clustering of rafts was necessary but not sufficient for ET-18-OCH(3)-mediated cell death, with Fas being required as the apoptosis trigger. ET-18-OCH(3)-mediated apoptosis did not require sphingomyelinase activation. Normal cells, including human and rat hepatocytes, did not incorporate ET-18-OCH(3) and were spared. This mechanism represents the first selective activation of Fas in tumor cells. Our data set a framework for the development of more targeted therapies leading to intracellular Fas activation and recruitment of downstream signaling molecules into Fas-enriched rafts.
Centro de Investigación del Cáncer, Instituto de Biología Molecular y Celular del Cáncer, CSIC, Universidad de Salamanca, Campus Miguel de Unamuno, E-37007 Salamanca, Spain.
Consuelo
C
Gajate
Esther
E
Del Canto-Jañez
A Ulises
AU
Acuña
Francisco
F
Amat-Guerri
Emilio
E
Geijo
Antonio M
AM
Santos-Beneit
Robert J
RJ
Veldman
Faustino
F
Mollinedo
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D051017:Apoptosis Regulatory Proteins; D018922:HL-60 Cells; D022781:Hepatocytes / Q000187:drug effects; D006801:Humans; D008562:Membrane Glycoproteins / Q000502:physiology; D021962:Membrane Microdomains / Q000378:metabolism*; D010742:Phospholipid Ethers / Q000494:pharmacology*; D014176:Protein Biosynthesis; D051381:Rats; D013108:Sphingomyelin Phosphodiesterase / Q000378:metabolism; D053221:TNF-Related Apoptosis-Inducing Ligand; D014409:Tumor Necrosis Factor-alpha / Q000502:physiology; D020298:U937 Cells; D019014:fas Receptor / Q000502:physiology*
D000970:Antineoplastic Agents; D051017:Apoptosis Regulatory Proteins; D008562:Membrane Glycoproteins; D010742:Phospholipid Ethers; D053221:TNF-Related Apoptosis-Inducing Ligand; C505653:TNFSF10 protein, human; C505655:Tnfsf10 protein, rat; D014409:Tumor Necrosis Factor-alpha; D019014:fas Receptor; C026659:edelfosine; D013108:Sphingomyelin Phosphodiesterase