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CD95 ligand induces motility and invasiveness of apoptosis-resistant tumor cells.

ABSTRACT: The apoptosis-inducing death receptor CD95 (APO-1/Fas) controls the homeostasis of many tissues. Despite its apoptotic potential, most human tumors are refractory to the cytotoxic effects of CD95 ligand. We now show that CD95 stimulation of multiple apoptosis-resistant tumor cells by CD95 ligand induces increased motility and invasiveness, a response much less efficiently triggered by TNFalpha or TRAIL. Three signaling pathways resulting in activation of NF-kappaB, Erk1/2 and caspase-8 were found to be important to this novel activity of CD95. Gene chip analyses of a CD95-stimulated tumor cell line identified a number of potential survival genes and genes that are known to regulate increased motility and invasiveness of tumor cells to be induced. Among these genes, urokinase plasminogen activator was found to be required for the CD95 ligand-induced motility and invasiveness. Our data suggest that CD95L, which is found elevated in many human cancer patients, has tumorigenic activities on human cancer cells. This could become highly relevant during chemotherapy, which can cause upregulation of CD95 ligand by both tumor and nontumor cells.

SUBMITTER: Barnhart BC

PROVIDER: S-EPMC514938 | biostudies-literature | 2004 Aug

REPOSITORIES: biostudies-literature

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CD95 ligand induces motility and invasiveness of apoptosis-resistant tumor cells.

Barnhart Bryan C BC Legembre Patrick P Pietras Eric E Bubici Concetta C Franzoso Guido G Peter Marcus E ME

The EMBO journal 20040722 15

The apoptosis-inducing death receptor CD95 (APO-1/Fas) controls the homeostasis of many tissues. Despite its apoptotic potential, most human tumors are refractory to the cytotoxic effects of CD95 ligand. We now show that CD95 stimulation of multiple apoptosis-resistant tumor cells by CD95 ligand induces increased motility and invasiveness, a response much less efficiently triggered by TNFalpha or TRAIL. Three signaling pathways resulting in activation of NF-kappaB, Erk1/2 and caspase-8 were foun ...[more]

PMID: 15272306

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Project description:Abstract BACKGROUND CD95 ligand (TNFSF6/APO-1L/FASLG) is a member of the Tumour Necrosis Factor Super Family (TNF-SF). Binding of the CD95 ligand (CD95L) to CD95 expressed on intrinsically apoptosis-resistant cancer cells like glioblastoma, triggers intracellular signal transduction resulting in increased tumour growth and invasiveness. A phase II clinical trial in patients with recurrent glioblastoma (NCT01071837) treated by Asunercept (a recombinant glycosylated fusion protein, which selectively binds and inhibits CD95L activity) was followed by a genome-wide assessment of DNA methylation. Here, a CpG-site (designated CpG2) within the CD95L-promotor was identified, exhibiting differential methylation between Asunercept responders (PFS>5 months) and non-responders (PFS < 2 months) with a significant survival benefit achieved in Asunercept treated patients with low CpG2 methylation (HR 0.34, p = 0.025). METHODS AND RESULTS We have performed in vitro studies to establish a link between the CpG2 methylation status and cellular characteristics of glioblastoma cell lines. A 3-dimensional spheroid invasion assay showed that highly methylated glioma cells like T98G did not grow and invade the surrounding matrix as aggressively compared to spheroids formed from low methylated glioma cells, e.g. U87-MG. Invasive growth of U87-MG spheroids in these assays was suppressed in the presence of Asunercept. A 1 kb fragment of the T98G CD95L promoter was subsequently cloned into a CpG-free reporter gene plasmid. Luciferase-based reporter gene assays of in vitro methylated and unmethylated plasmids in transfected HEK cells indicate that the CD95L promoter, despite the observed sparseness of CpG sites, is at least partially regulated by its methylation level. Furthermore, mutational disruption of the CpG2 site completely silenced reporter gene activity in vitro, which insinuates that both methylation level and gene promoter sequence are involved in regulation of the CD95L gene. CONCLUSION In essence, the level of CpG2 methylation correlates to aggressiveness of glioma derived cancer spheroids in vitro, and methylation of the CD95L promoter in glioblastoma tissue from patients might warrant use as a potential biomarker predicting response to therapy with Asunercept. We are currently developing a specific and sensitive assay to quantify CpG2 methylation considered as a companion diagnostic for further clinical studies.

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CD95 ligand induces motility and invasiveness of apoptosis-resistant tumor cells.

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CD95 ligand induces motility and invasiveness of apoptosis-resistant tumor cells.

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