Enhanced caspase-8 recruitment to and activation at the DISC is critical for sensitisation of human hepatocellular carcinoma cells to TRAIL-induced apoptosis by chemotherapeutic drugs.
Tumour necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) exhibits potent antitumour activity upon systemic administration in mice without showing the deleterious side effects observed with other apoptosis-inducing members of the TNF family such as TNF and CD95L. TRAIL may, thus, have great potential in the treatment of human cancer. However, about 60% of tumour cell lines are not sensitive to TRAIL. To evaluate the mechanisms of tumour resistance to TRAIL, we investigated hepatocellular carcinoma (HCC) cell lines that exhibit differential sensitivity to TRAIL. Pretreatment with chemotherapeutic drugs, for example, 5-fluorouracil (5-FU), rendered the TRAIL-resistant HCC cell lines sensitive to TRAIL-induced apoptosis. Analysis of the TRAIL death-inducing signalling complex (DISC) revealed upregulation of TRAIL-R2. Caspase-8 recruitment to and its activation at the DISC were substantially increased after 5-FU sensitisation, while FADD recruitment remained essentially unchanged. 5-FU pretreatment downregulated cellular FLICE-inhibitory protein (cFLIP) and specific cFLIP downregulation by small interfering RNA was sufficient to sensitise TRAIL-resistant HCC cell lines for TRAIL-induced apoptosis. Thus, a potential mechanism for TRAIL sensitisation by 5-FU is the increased effectiveness of caspase-8 recruitment to and activation at the DISC facilitated by the downregulation of cFLIP and the consequent shift in the ratio of caspase-8 to cFLIP at the DISC.
Ganten
T M
TM
Divison of Apoptosis Regulation, German Cancer Research Center (DKFZ), Heidelberg, Germany.
Haas
T L
TL
Sykora
J
J
Stahl
H
H
Sprick
M R
MR
Fas
S C
SC
Krueger
A
A
Weigand
M A
MA
Grosse-Wilde
A
A
Stremmel
W
W
Krammer
P H
PH
Walczak
H
H
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D051017:Apoptosis Regulatory Proteins; D015153:Blotting, Western; D053446:CASP8 and FADD-Like Apoptosis Regulating Protein; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D053148:Caspase 3; D053178:Caspase 6; D053181:Caspase 8; D020169:Caspases / Q000378:metabolism*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D053418:Death Domain Receptor Signaling Adaptor Proteins; D015536:Down-Regulation; D019008:Drug Resistance, Neoplasm / Q000187:drug effects; D004357:Drug Synergism; D004792:Enzyme Precursors / Q000378:metabolism; D053200:Fas-Associated Death Domain Protein; D005434:Flow Cytometry; D005472:Fluorouracil / Q000494:pharmacology; D058851:GPI-Linked Proteins; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D022781:Hepatocytes / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism; D008113:Liver Neoplasms / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D008562:Membrane Glycoproteins / Q000494:pharmacology / Q000502:physiology*; D008856:Microscopy, Fluorescence; D034741:RNA, Small Interfering / Q000235:genetics; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand; D018124:Receptors, Tumor Necrosis Factor / Q000032:analysis / Q000276:immunology / Q000378:metabolism*; D053320:Receptors, Tumor Necrosis Factor, Member 10c; D011993:Recombinant Fusion Proteins; D053221:TNF-Related Apoptosis-Inducing Ligand; D014162:Transfection; D053319:Tumor Necrosis Factor Decoy Receptors; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology / Q000502:physiology*; D016159:Tumor Suppressor Protein p53 / Q000502:physiology; D015854:Up-Regulation; D019014:fas Receptor / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D000911:Antibodies, Monoclonal; D000970:Antineoplastic Agents; D051017:Apoptosis Regulatory Proteins; D053446:CASP8 and FADD-Like Apoptosis Regulating Protein; C506386:CFLAR protein, human; D053418:Death Domain Receptor Signaling Adaptor Proteins; D004792:Enzyme Precursors; C505533:FADD protein, human; D053200:Fas-Associated Death Domain Protein; D058851:GPI-Linked Proteins; D047908:Intracellular Signaling Peptides and Proteins; C448905:LZ-TRAIL protein, recombinant; D008562:Membrane Glycoproteins; D034741:RNA, Small Interfering; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand; D018124:Receptors, Tumor Necrosis Factor; D053320:Receptors, Tumor Necrosis Factor, Member 10c; D011993:Recombinant Fusion Proteins; D053221:TNF-Related Apoptosis-Inducing Ligand; C105190:TNFRSF10A protein, human; C505639:TNFRSF10B protein, human; C108661:TNFRSF10C protein, human; C505653:TNFSF10 protein, human; D053319:Tumor Necrosis Factor Decoy Receptors; D014409:Tumor Necrosis Factor-alpha; D016159:Tumor Suppressor Protein p53; D019014:fas Receptor; C485192:green fluorescent protein, Aequorea victoria; D049452:Green Fluorescent Proteins; C505407:CASP3 protein, human; C505530:CASP8 protein, human; D053148:Caspase 3; D053178:Caspase 6; D053181:Caspase 8; D020169:Caspases; D005472:Fluorouracil