Fas Ligand-independent, FADD-mediated Activation of the Fas Death Pathway by Anticancer Drugs

Abstract

Trimerization of the Fas receptor (CD95, APO-1), a membrane bound protein, triggers cell death by apoptosis. The main death pathway activated by Fas receptor involves the adaptor protein FADD (for Fas-associated deathdomain) that connects Fas receptor to the caspase cascade. Anticancer drugs have been shown to enhance both Fas receptor and Fas ligand expression on tumor cells. The contribution of Fas ligand-Fas receptor interactions to the cytotoxic activity of these drugs remains controversial. Here, we show that neither the antagonistic anti-Fas antibody ZB4 nor the Fas-IgG molecule inhibit drug-induced apoptosis in three different cell lines. The expression of Fas ligand on the plasma membrane, which is identified in untreated U937 human leukemic cells but remains undetectable in untreated HT29 and HCT116 human colon cancer cell lines, is not modified by exposure to various cytotoxic agents. These drugs induce the clustering of Fas receptor, as observed by confocal laser scanning microscopy, and its interaction with FADD, as demonstrated by co-immunoprecipitation. Overexpression of FADD by stable transfection sensitizes tumor cells to drug-induced cell death and cytotoxicity, whereas down-regulation of FADD by transient transfection of an antisense construct decreases tumor cell sensitivity to drug-induced apoptosis. These results were confirmed by transient transfection of constructs encoding either a FADD dominant negative mutant or MC159 or E8 viral proteins that inhibit the FADD/caspase-8 pathway. These results suggest that drug-induced cell death involves the Fas/FADD pathway in a Fas ligand-independent fashion. Trimerization of the Fas receptor (CD95, APO-1), a membrane bound protein, triggers cell death by apoptosis. The main death pathway activated by Fas receptor involves the adaptor protein FADD (for Fas-associated deathdomain) that connects Fas receptor to the caspase cascade. Anticancer drugs have been shown to enhance both Fas receptor and Fas ligand expression on tumor cells. The contribution of Fas ligand-Fas receptor interactions to the cytotoxic activity of these drugs remains controversial. Here, we show that neither the antagonistic anti-Fas antibody ZB4 nor the Fas-IgG molecule inhibit drug-induced apoptosis in three different cell lines. The expression of Fas ligand on the plasma membrane, which is identified in untreated U937 human leukemic cells but remains undetectable in untreated HT29 and HCT116 human colon cancer cell lines, is not modified by exposure to various cytotoxic agents. These drugs induce the clustering of Fas receptor, as observed by confocal laser scanning microscopy, and its interaction with FADD, as demonstrated by co-immunoprecipitation. Overexpression of FADD by stable transfection sensitizes tumor cells to drug-induced cell death and cytotoxicity, whereas down-regulation of FADD by transient transfection of an antisense construct decreases tumor cell sensitivity to drug-induced apoptosis. These results were confirmed by transient transfection of constructs encoding either a FADD dominant negative mutant or MC159 or E8 viral proteins that inhibit the FADD/caspase-8 pathway. These results suggest that drug-induced cell death involves the Fas/FADD pathway in a Fas ligand-independent fashion. tumor necrosis factor Fas ligand soluble Fas-L cisplatin vinblastine arbitrary unit phosphate-buffered saline etoposide Fas is a 45-kDa membrane protein that belongs to the tumor necrosis factor (TNF)1/nerve growth factor receptor family (1Itoh N. Yonehara S. Ishii A. Yonehara M. Mizushima S. Sameshima M. Hase A. Seto Y. Nagata S. Cell. 1991; 66: 233-243Abstract Full Text PDF PubMed Scopus (2663) Google Scholar). Engagement of Fas by agonistic anti-Fas antibodies triggers programmed cell death in a variety of cell types (2Trauth B.C. Klas C. Peters A.M.J. Matzuku S. Möller P. Falk W. Debatin K.M. Krammer P.H. Science. 1989; 245: 301-305Crossref PubMed Scopus (1655) Google Scholar, 3Yonehara S. Ishii A. Yonehara M.A. J. Exp. Med. 1989; 169: 1747-1756Crossref PubMed Scopus (1422) Google Scholar). Its natural ligand, Fas ligand (Fas-L), belongs to the TNF family (4Suda T. Takahashi T. Golstein P. Nagata S. Cell. 1993; 75: 1169-1178Abstract Full Text PDF PubMed Scopus (2431) Google Scholar) and can be found as a 40-kDa membrane-bound or a 26-kDa soluble cytokine (5Tanaka M. Suda T. Takahashi T. Nagata S. EMBO J. 1995; 14: 1129-1135Crossref PubMed Scopus (604) Google Scholar). Similarly to agonistic anti-Fas antibodies, binding of membrane-bound or soluble Fas-L to Fas receptor can induce apoptosis in Fas-bearing cells (1Itoh N. Yonehara S. Ishii A. Yonehara M. Mizushima S. Sameshima M. Hase A. Seto Y. Nagata S. Cell. 1991; 66: 233-243Abstract Full Text PDF PubMed Scopus (2663) Google Scholar, 4Suda T. Takahashi T. Golstein P. Nagata S. Cell. 1993; 75: 1169-1178Abstract Full Text PDF PubMed Scopus (2431) Google Scholar). The main death pathway initiated from Fas activation involves a series of death-associated molecules (6Cock J.G. DeVries E. Williams G.T. Borst J. Apoptosis. 1998; 3: 17-25Crossref PubMed Scopus (26) Google Scholar), including FADD (Fas-associateddeath domain-containing protein), which is an adaptor protein that is recruited to Fas receptor upon its engagement (7Chinnaiyan A.M. Tepper C.G. Seldin M.F. O'Rourke K. Kischkel F.C. Hellbardt S. Krammer P.H. Peter M.E. Dixit V.M. J. Biol. Chem. 1996; 271: 4961-4965Abstract Full Text Full Text PDF PubMed Scopus (705) Google Scholar, 8Boldin M.P. Goncharov T.M. Goltsev Y.V. Wallach D. Cell. 1996; 85: 803-815Abstract Full Text Full Text PDF PubMed Scopus (2100) Google Scholar, 9Chinnaiyan A.M. O'Rourke K. Tewari M. Dixit V.M. Cell. 1995; 81: 505-512Abstract Full Text PDF PubMed Scopus (2150) Google Scholar). FADD then binds to and activates procaspase-8 (also called FLICE or MACH) (8Boldin M.P. Goncharov T.M. Goltsev Y.V. Wallach D. Cell. 1996; 85: 803-815Abstract Full Text Full Text PDF PubMed Scopus (2100) Google Scholar, 9Chinnaiyan A.M. O'Rourke K. Tewari M. Dixit V.M. Cell. 1995; 81: 505-512Abstract Full Text PDF PubMed Scopus (2150) Google Scholar, 10Muzio M. Chinnaiyan A.M. Kischkel F.C. O'Rourke K. Shevchenko A. Ni J.N. Scaffidi C. Bretz J.D. Zhang M. Gentz R. Mann M. Krammer P.H. Peter M.E. Dixit V.M. Cell. 1996; 85: 817-827Abstract Full Text Full Text PDF PubMed Scopus (2723) Google Scholar), which is believed to be the first step of a proteolytic cascade that triggers activation of other caspases such as caspases-7, -3, and -6 (11Hirata H. Takahashi A. Kobayashi S. Yonehara S. Sawai H. Okazaki T. Yamamoto K. Sasada M. J. Exp. Med. 1998; 187: 587-600Crossref PubMed Scopus (398) Google Scholar). Although other cell death pathways could be initiated from Fas activation (12Yang X. Khosravi-Far R. Chang H.Y. Baltimore D. Cell. 1997; 89: 1067-1076Abstract Full Text Full Text PDF PubMed Scopus (823) Google Scholar, 13Sillence D.J. Allan D. Biochem. J. 1997; 324: 29-32Crossref PubMed Scopus (54) Google Scholar), analysis of lymphocytes from FADD −/− mice has recently demonstrated the prominent role of the FADD/procaspase-8 pathway in Fas-mediated cell death (14Zhang J.K. Cado D. Chen A. Kabra N.H. Winoto A. Nature. 1998; 392: 296-300Crossref PubMed Scopus (635) Google Scholar).Cytotoxic drugs commonly used in cancer therapy can induce tumor cell death by apoptosis (15Bose R. Verheij M. Haimovitz-Friedman A. Scotto K. Fuks Z. Kolesnick R. Cell. 1995; 82: 405-414Abstract Full Text PDF PubMed Scopus (782) Google Scholar, 16Barry M.A. Behnke C.A. Eastman A. Biochem. Pharmacol. 1990; 40: 2353-2362Crossref PubMed Scopus (909) Google Scholar, 17Hannun Y.A. Blood. 1997; 89: 1845-1853Crossref PubMed Google Scholar). These drugs were shown to enhance the expression of Fas (18Micheau O. Solary E. Hammann A. Martin F. Dimanche-Boitrel M.T. J. Natl. Cancer Inst. 1997; 89: 783-789Crossref PubMed Scopus (271) Google Scholar) and Fas-L (19Friesen C. Herr I. Krammer P.H. Debatin K.M. Nat. Med. 1996; 2: 574-577Crossref PubMed Scopus (951) Google Scholar, 20Herr I. Wilhelm D. Böhler T. Angel P. Debatin K.M. EMBO J. 1997; 16: 6200-6208Crossref PubMed Scopus (277) Google Scholar) on the surface of certain malignant cells. It has been proposed that the molecular process leading from specific cellular damage induced by the drugs to apoptosis might involve an interaction between Fas ligand and Fas (19Friesen C. Herr I. Krammer P.H. Debatin K.M. Nat. Med. 1996; 2: 574-577Crossref PubMed Scopus (951) Google Scholar). However, this issue remains controversial because antagonistic anti-Fas antibodies that block Fas-L-mediated apoptosis do not always inhibit drug-induced cell death (21Eischen C.M. Kottke T.J. Martins L.M. Basi G.S. Tung J.S. Earnshaw W.C. Leibson P.J. Kaufmann S.H. Blood. 1997; 90: 935-943Crossref PubMed Google Scholar, 22McGahon A.J. Costa Pereira A.P. Daly L. Cotter T.G. Br. J. Haematol. 1998; 101: 539-547Crossref PubMed Scopus (81) Google Scholar). In the present study, we further addressed the role played by the Fas/FADD pathway in drug-induced cell death. We show that anticancer drugs can induce Fas receptor clustering and FADD recruitment to Fas receptor in a Fas ligand-independent fashion. By modulating FADD expression, we also demonstrate the role of this adaptor molecule in drug-induced apoptosis.DISCUSSIONThe molecular mechanisms involved during the course of apoptosis upon exposure to anticancer drugs are still poorly understood but are of major importance for the understanding of tumor cell killing. Induction of apoptosis occurs through pathways and on the We have shown that anticancer could Fas receptor the tumor cell surface by Fas The in Fas receptor and tumor cells were to Fas-mediated apoptosis (18Micheau O. Solary E. Hammann A. Martin F. Dimanche-Boitrel M.T. J. Natl. Cancer Inst. 1997; 89: 783-789Crossref PubMed Scopus (271) Google Scholar). interaction of Fas receptor with its ligand a role in anticancer remains a controversial issue (21Eischen C.M. Kottke T.J. Martins L.M. Basi G.S. Tung J.S. Earnshaw W.C. Leibson P.J. Kaufmann S.H. Blood. 1997; 90: 935-943Crossref PubMed Google Scholar). Here, we show that the antagonistic anti-Fas antibody ZB4 and the Fas-IgG used a that Fas-mediated and apoptosis C. P. W. K. F. E. M. 1998; PubMed Scopus Google Scholar), have on anticancer drug-induced apoptosis in three different cell lines. we to of Fas-L expression on the plasma membrane of tumor cells to cytotoxic C. P. W. K. F. E. M. 1998; PubMed Scopus Google Scholar, D. S. D. J. 1998; Google Scholar). In with in other cell (21Eischen C.M. Kottke T.J. Martins L.M. Basi G.S. Tung J.S. Earnshaw W.C. Leibson P.J. Kaufmann S.H. Blood. 1997; 90: 935-943Crossref PubMed Google Scholar, 22McGahon A.J. Costa Pereira A.P. Daly L. Cotter T.G. Br. J. Haematol. 1998; 101: 539-547Crossref PubMed Scopus (81) Google Scholar, A. A. M. S. R. R. Cancer 1997; Google Scholar), these results a role for a Fas interaction in drug-induced cell pathway has been shown to be by Fas in the of Fas-L in cells to Y. D. D. T. J. Biol. 1998; PubMed Scopus Google Scholar, A. C.A. Chinnaiyan A.M. Dixit V.M. J. Biol. Chem. 1997; Full Text Full Text PDF PubMed Scopus Google Scholar). In this the adaptor protein FADD is recruited to Fas receptor through its death (7Chinnaiyan A.M. Tepper C.G. Seldin M.F. O'Rourke K. Kischkel F.C. Hellbardt S. Krammer P.H. Peter M.E. Dixit V.M. J. Biol. Chem. 1996; 271: 4961-4965Abstract Full Text Full Text PDF PubMed Scopus (705) Google Scholar, 8Boldin M.P. Goncharov T.M. Goltsev Y.V. Wallach D. Cell. 1996; 85: 803-815Abstract Full Text Full Text PDF PubMed Scopus (2100) Google Scholar, 9Chinnaiyan A.M. O'Rourke K. Tewari M. Dixit V.M. Cell. 1995; 81: 505-512Abstract Full Text PDF PubMed Scopus (2150) Google Scholar, 10Muzio M. Chinnaiyan A.M. Kischkel F.C. O'Rourke K. Shevchenko A. Ni J.N. Scaffidi C. Bretz J.D. Zhang M. Gentz R. Mann M. Krammer P.H. Peter M.E. Dixit V.M. Cell. 1996; 85: 817-827Abstract Full Text Full Text PDF PubMed Scopus (2723) Google Scholar), by an interaction of the death receptor of FADD with the of procaspase-8 leading to the activation of the proteolytic cascade of caspases (11Hirata H. Takahashi A. Kobayashi S. Yonehara S. Sawai H. Okazaki T. Yamamoto K. Sasada M. J. Exp. Med. 1998; 187: 587-600Crossref PubMed Scopus (398) Google Scholar). laser scanning and that anticancer drugs could induce Fas receptor and FADD to of lymphocytes from FADD −/− mice have recently demonstrated the prominent role of the FADD/procaspase-8 pathway in cell death by either Fas-L or agonistic anti-Fas antibodies (14Zhang J.K. Cado D. Chen A. Kabra N.H. Winoto A. Nature. 1998; 392: 296-300Crossref PubMed Scopus (635) Google Scholar). The of FADD on the cell on the cell and the FADD a activity in cells A.M. O'Rourke K. Tewari M. Dixit V.M. Cell. 1995; 81: 505-512Abstract Full Text PDF PubMed Scopus (2150) Google Scholar), whereas its the growth of cells K. A. EMBO J. 1998; PubMed Scopus Google Scholar). We show that FADD a role in drug-induced apoptosis. We HT29 cell that FADD, that a in FADD protein expression not to apoptosis in these cells. These FADD were to drug-induced apoptosis and cells. role for FADD in drug-induced cell death further demonstrated by the of cytotoxic in HT29 cells in which FADD expression by transient expression of an antisense results were by the FADD/procaspase-8 pathway different transient expression of viral proteins J. S. Martin Y. S. T.G. Natl. S. A. 1997; PubMed Scopus Google Scholar) and dominant negative construct J. C.M. J. 1998; Google Scholar). that anticancer drugs can induce Fas receptor clustering in a and FADD to Fas receptor in the cytotoxic process leading to apoptosis. Fas is a 45-kDa membrane protein that belongs to the tumor necrosis factor (TNF)1/nerve growth factor receptor family (1Itoh N. Yonehara S. Ishii A. Yonehara M. Mizushima S. Sameshima M. Hase A. Seto Y. Nagata S. Cell. 1991; 66: 233-243Abstract Full Text PDF PubMed Scopus (2663) Google Scholar). Engagement of Fas by agonistic anti-Fas antibodies triggers programmed cell death in a variety of cell types (2Trauth B.C. Klas C. Peters A.M.J. Matzuku S. Möller P. Falk W. Debatin K.M. Krammer P.H. Science. 1989; 245: 301-305Crossref PubMed Scopus (1655) Google Scholar, 3Yonehara S. Ishii A. Yonehara M.A. J. Exp. Med. 1989; 169: 1747-1756Crossref PubMed Scopus (1422) Google Scholar). Its natural ligand, Fas ligand (Fas-L), belongs to the TNF family (4Suda T. Takahashi T. Golstein P. Nagata S. Cell. 1993; 75: 1169-1178Abstract Full Text PDF PubMed Scopus (2431) Google Scholar) and can be found as a 40-kDa membrane-bound or a 26-kDa soluble cytokine (5Tanaka M. Suda T. Takahashi T. Nagata S. EMBO J. 1995; 14: 1129-1135Crossref PubMed Scopus (604) Google Scholar). Similarly to agonistic anti-Fas antibodies, binding of membrane-bound or soluble Fas-L to Fas receptor can induce apoptosis in Fas-bearing cells (1Itoh N. Yonehara S. Ishii A. Yonehara M. Mizushima S. Sameshima M. Hase A. Seto Y. Nagata S. Cell. 1991; 66: 233-243Abstract Full Text PDF PubMed Scopus (2663) Google Scholar, 4Suda T. Takahashi T. Golstein P. Nagata S. Cell. 1993; 75: 1169-1178Abstract Full Text PDF PubMed Scopus (2431) Google Scholar). The main death pathway initiated from Fas activation involves a series of death-associated molecules (6Cock J.G. DeVries E. Williams G.T. Borst J. Apoptosis. 1998; 3: 17-25Crossref PubMed Scopus (26) Google Scholar), including FADD (Fas-associateddeath domain-containing protein), which is an adaptor protein that is recruited to Fas receptor upon its engagement (7Chinnaiyan A.M. Tepper C.G. Seldin M.F. O'Rourke K. Kischkel F.C. Hellbardt S. Krammer P.H. Peter M.E. Dixit V.M. J. Biol. Chem. 1996; 271: 4961-4965Abstract Full Text Full Text PDF PubMed Scopus (705) Google Scholar, 8Boldin M.P. Goncharov T.M. Goltsev Y.V. Wallach D. Cell. 1996; 85: 803-815Abstract Full Text Full Text PDF PubMed Scopus (2100) Google Scholar, 9Chinnaiyan A.M. O'Rourke K. Tewari M. Dixit V.M. Cell. 1995; 81: 505-512Abstract Full Text PDF PubMed Scopus (2150) Google Scholar). FADD then binds to and activates procaspase-8 (also called FLICE or MACH) (8Boldin M.P. Goncharov T.M. Goltsev Y.V. Wallach D. Cell. 1996; 85: 803-815Abstract Full Text Full Text PDF PubMed Scopus (2100) Google Scholar, 9Chinnaiyan A.M. O'Rourke K. Tewari M. Dixit V.M. Cell. 1995; 81: 505-512Abstract Full Text PDF PubMed Scopus (2150) Google Scholar, 10Muzio M. Chinnaiyan A.M. Kischkel F.C. O'Rourke K. Shevchenko A. Ni J.N. Scaffidi C. Bretz J.D. Zhang M. Gentz R. Mann M. Krammer P.H. Peter M.E. Dixit V.M. Cell. 1996; 85: 817-827Abstract Full Text Full Text PDF PubMed Scopus (2723) Google Scholar), which is believed to be the first step of a proteolytic cascade that triggers activation of other caspases such as caspases-7, -3, and -6 (11Hirata H. Takahashi A. Kobayashi S. Yonehara S. Sawai H. Okazaki T. Yamamoto K. Sasada M. J. Exp. Med. 1998; 187: 587-600Crossref PubMed Scopus (398) Google Scholar). Although other cell death pathways could be initiated from Fas activation (12Yang X. Khosravi-Far R. Chang H.Y. Baltimore D. Cell. 1997; 89: 1067-1076Abstract Full Text Full Text PDF PubMed Scopus (823) Google Scholar, 13Sillence D.J. Allan D. Biochem. J. 1997; 324: 29-32Crossref PubMed Scopus (54) Google Scholar), analysis of lymphocytes from FADD −/− mice has recently demonstrated the prominent role of the FADD/procaspase-8 pathway in Fas-mediated cell death (14Zhang J.K. Cado D. Chen A. Kabra N.H. Winoto A. Nature. 1998; 392: 296-300Crossref PubMed Scopus (635) Google Scholar). drugs commonly used in cancer therapy can induce tumor cell death by apoptosis (15Bose R. Verheij M. Haimovitz-Friedman A. Scotto K. Fuks Z. Kolesnick R. Cell. 1995; 82: 405-414Abstract Full Text PDF PubMed Scopus (782) Google Scholar, 16Barry M.A. Behnke C.A. Eastman A. Biochem. Pharmacol. 1990; 40: 2353-2362Crossref PubMed Scopus (909) Google Scholar, 17Hannun Y.A. Blood. 1997; 89: 1845-1853Crossref PubMed Google Scholar). These drugs were shown to enhance the expression of Fas (18Micheau O. Solary E. Hammann A. Martin F. Dimanche-Boitrel M.T. J. Natl. Cancer Inst. 1997; 89: 783-789Crossref PubMed Scopus (271) Google Scholar) and Fas-L (19Friesen C. Herr I. Krammer P.H. Debatin K.M. Nat. Med. 1996; 2: 574-577Crossref PubMed Scopus (951) Google Scholar, 20Herr I. Wilhelm D. Böhler T. Angel P. Debatin K.M. EMBO J. 1997; 16: 6200-6208Crossref PubMed Scopus (277) Google Scholar) on the surface of certain malignant cells. It has been proposed that the molecular process leading from specific cellular damage induced by the drugs to apoptosis might involve an interaction between Fas ligand and Fas (19Friesen C. Herr I. Krammer P.H. Debatin K.M. Nat. Med. 1996; 2: 574-577Crossref PubMed Scopus (951) Google Scholar). However, this issue remains controversial because antagonistic anti-Fas antibodies that block Fas-L-mediated apoptosis do not always inhibit drug-induced cell death (21Eischen C.M. Kottke T.J. Martins L.M. Basi G.S. Tung J.S. Earnshaw W.C. Leibson P.J. Kaufmann S.H. Blood. 1997; 90: 935-943Crossref PubMed Google Scholar, 22McGahon A.J. Costa Pereira A.P. Daly L. Cotter T.G. Br. J. Haematol. 1998; 101: 539-547Crossref PubMed Scopus (81) Google Scholar). In the present study, we further addressed the role played by the Fas/FADD pathway in drug-induced cell death. We show that anticancer drugs can induce Fas receptor clustering and FADD recruitment to Fas receptor in a Fas ligand-independent fashion. By modulating FADD expression, we also demonstrate the role of this adaptor molecule in drug-induced apoptosis. molecular mechanisms involved during the course of apoptosis upon exposure to anticancer drugs are still poorly understood but are of major importance for the understanding of tumor cell killing. Induction of apoptosis occurs through pathways and on the We have shown that anticancer could Fas receptor the tumor cell surface by Fas The in Fas receptor and tumor cells were to Fas-mediated apoptosis (18Micheau O. Solary E. Hammann A. Martin F. Dimanche-Boitrel M.T. J. Natl. Cancer Inst. 1997; 89: 783-789Crossref PubMed Scopus (271) Google Scholar). interaction of Fas receptor with its ligand a role in anticancer remains a controversial issue (21Eischen C.M. Kottke T.J. Martins L.M. Basi G.S. Tung J.S. Earnshaw W.C. Leibson P.J. Kaufmann S.H. Blood. 1997; 90: 935-943Crossref PubMed Google Scholar). Here, we show that the antagonistic anti-Fas antibody ZB4 and the Fas-IgG used a that Fas-mediated and apoptosis C. P. W. K. F. E. M. 1998; PubMed Scopus Google Scholar), have on anticancer drug-induced apoptosis in three different cell lines. we to of Fas-L expression on the plasma membrane of tumor cells to cytotoxic C. P. W. K. F. E. M. 1998; PubMed Scopus Google Scholar, D. S. D. J. 1998; Google Scholar). In with in other cell (21Eischen C.M. Kottke T.J. Martins L.M. Basi G.S. Tung J.S. Earnshaw W.C. Leibson P.J. Kaufmann S.H. Blood. 1997; 90: 935-943Crossref PubMed Google Scholar, 22McGahon A.J. Costa Pereira A.P. Daly L. Cotter T.G. Br. J. Haematol. 1998; 101: 539-547Crossref PubMed Scopus (81) Google Scholar, A. A. M. S. R. R. Cancer 1997; Google Scholar), these results a role for a Fas interaction in drug-induced cell pathway has been shown to be by Fas in the of Fas-L in cells to Y. D. D. T. J. Biol. 1998; PubMed Scopus Google Scholar, A. C.A. Chinnaiyan A.M. Dixit V.M. J. Biol. Chem. 1997; Full Text Full Text PDF PubMed Scopus Google Scholar). In this the adaptor protein FADD is recruited to Fas receptor through its death (7Chinnaiyan A.M. Tepper C.G. Seldin M.F. O'Rourke K. Kischkel F.C. Hellbardt S. Krammer P.H. Peter M.E. Dixit V.M. J. Biol. Chem. 1996; 271: 4961-4965Abstract Full Text Full Text PDF PubMed Scopus (705) Google Scholar, 8Boldin M.P. Goncharov T.M. Goltsev Y.V. Wallach D. Cell. 1996; 85: 803-815Abstract Full Text Full Text PDF PubMed Scopus (2100) Google Scholar, 9Chinnaiyan A.M. O'Rourke K. Tewari M. Dixit V.M. Cell. 1995; 81: 505-512Abstract Full Text PDF PubMed Scopus (2150) Google Scholar, 10Muzio M. Chinnaiyan A.M. Kischkel F.C. O'Rourke K. Shevchenko A. Ni J.N. Scaffidi C. Bretz J.D. Zhang M. Gentz R. Mann M. Krammer P.H. Peter M.E. Dixit V.M. Cell. 1996; 85: 817-827Abstract Full Text Full Text PDF PubMed Scopus (2723) Google Scholar), by an interaction of the death receptor of FADD with the of procaspase-8 leading to the activation of the proteolytic cascade of caspases (11Hirata H. Takahashi A. Kobayashi S. Yonehara S. Sawai H. Okazaki T. Yamamoto K. Sasada M. J. Exp. Med. 1998; 187: 587-600Crossref PubMed Scopus (398) Google Scholar). laser scanning and that anticancer drugs could induce Fas receptor and FADD to of lymphocytes from FADD −/− mice have recently demonstrated the prominent role of the FADD/procaspase-8 pathway in cell death by either Fas-L or agonistic anti-Fas antibodies (14Zhang J.K. Cado D. Chen A. Kabra N.H. Winoto A. Nature. 1998; 392: 296-300Crossref PubMed Scopus (635) Google Scholar). The of FADD on the cell on the cell and the FADD a activity in cells A.M. O'Rourke K. Tewari M. Dixit V.M. Cell. 1995; 81: 505-512Abstract Full Text PDF PubMed Scopus (2150) Google Scholar), whereas its the growth of cells K. A. EMBO J. 1998; PubMed Scopus Google Scholar). We show that FADD a role in drug-induced apoptosis. We HT29 cell that FADD, that a in FADD protein expression not to apoptosis in these cells. These FADD were to drug-induced apoptosis and cells. role for FADD in drug-induced cell death further demonstrated by the of cytotoxic in HT29 cells in which FADD expression by transient expression of an antisense results were by the FADD/procaspase-8 pathway different transient expression of viral proteins J. S. Martin Y. S. T.G. Natl. S. A. 1997; PubMed Scopus Google Scholar) and dominant negative construct J. C.M. J. 1998; Google Scholar). that anticancer drugs can induce Fas receptor clustering in a and FADD to Fas receptor in the cytotoxic process leading to apoptosis. The molecular mechanisms involved during the course of apoptosis upon exposure to anticancer drugs are still poorly understood but are of major importance for the understanding of tumor cell killing. Induction of apoptosis occurs through pathways and on the We have shown that anticancer could Fas receptor the tumor cell surface by Fas The in Fas receptor and tumor cells were to Fas-mediated apoptosis (18Micheau O. Solary E. Hammann A. Martin F. Dimanche-Boitrel M.T. J. Natl. Cancer Inst. 1997; 89: 783-789Crossref PubMed Scopus (271) Google Scholar). interaction of Fas receptor with its ligand a role in anticancer remains a controversial issue (21Eischen C.M. Kottke T.J. Martins L.M. Basi G.S. Tung J.S. Earnshaw W.C. Leibson P.J. Kaufmann S.H. Blood. 1997; 90: 935-943Crossref PubMed Google Scholar). Here, we show that the antagonistic anti-Fas antibody ZB4 and the Fas-IgG used a that Fas-mediated and apoptosis C. P. W. K. F. E. M. 1998; PubMed Scopus Google Scholar), have on anticancer drug-induced apoptosis in three different cell lines. we to of Fas-L expression on the plasma membrane of tumor cells to cytotoxic C. P. W. K. F. E. M. 1998; PubMed Scopus Google Scholar, D. S. D. J. 1998; Google Scholar). In with in other cell (21Eischen C.M. Kottke T.J. Martins L.M. Basi G.S. Tung J.S. Earnshaw W.C. Leibson P.J. Kaufmann S.H. Blood. 1997; 90: 935-943Crossref PubMed Google Scholar, 22McGahon A.J. Costa Pereira A.P. Daly L. Cotter T.G. Br. J. Haematol. 1998; 101: 539-547Crossref PubMed Scopus (81) Google Scholar, A. A. M. S. R. R. Cancer 1997; Google Scholar), these results a role for a Fas interaction in drug-induced cell death. Fas pathway has been shown to be by Fas in the of Fas-L in cells to Y. D. D. T. J. Biol. 1998; PubMed Scopus Google Scholar, A. C.A. Chinnaiyan A.M. Dixit V.M. J. Biol. Chem. 1997; Full Text Full Text PDF PubMed Scopus Google Scholar). In this the adaptor protein FADD is recruited to Fas receptor through its death (7Chinnaiyan A.M. Tepper C.G. Seldin M.F. O'Rourke K. Kischkel F.C. Hellbardt S. Krammer P.H. Peter M.E. Dixit V.M. J. Biol. Chem. 1996; 271: 4961-4965Abstract Full Text Full Text PDF PubMed Scopus (705) Google Scholar, 8Boldin M.P. Goncharov T.M. Goltsev Y.V. Wallach D. Cell. 1996; 85: 803-815Abstract Full Text Full Text PDF PubMed Scopus (2100) Google Scholar, 9Chinnaiyan A.M. O'Rourke K. Tewari M. Dixit V.M. Cell. 1995; 81: 505-512Abstract Full Text PDF PubMed Scopus (2150) Google Scholar, 10Muzio M. Chinnaiyan A.M. Kischkel F.C. O'Rourke K. Shevchenko A. Ni J.N. Scaffidi C. Bretz J.D. Zhang M. Gentz R. Mann M. Krammer P.H. Peter M.E. Dixit V.M. Cell. 1996; 85: 817-827Abstract Full Text Full Text PDF PubMed Scopus (2723) Google Scholar), by an interaction of the death receptor of FADD with the of procaspase-8 leading to the activation of the proteolytic cascade of caspases (11Hirata H. Takahashi A. 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