Kenjiro Sawada

CD95 promotes tumour growth

CD95 (also called Fas and APO-1) is a prototypical death receptor that regulates tissue homeostasis mainly in the immune system through the induction of apoptosis. During cancer progression CD95 is frequently downregulated or cells are rendered apoptosis resistant, raising the possibility that loss of CD95 is part of a mechanism for tumour evasion. However, complete loss of CD95 is rarely seen in human cancers and many cancer cells express large quantities of CD95 and are highly sensitive to CD95-mediated apoptosis in vitro. Furthermore, cancer patients frequently have elevated levels of the physiological ligand for CD95, CD95L. These data raise the possibility that CD95 could actually promote the growth of tumours through its non-apoptotic activities. Here we show that cancer cells in general, regardless of their CD95 apoptosis sensitivity, depend on constitutive activity of CD95, stimulated by cancer-produced CD95L, for optimal growth. Consistently, loss of CD95 in mouse models of ovarian cancer and liver cancer reduces cancer incidence as well as the size of the tumours. The tumorigenic activity of CD95 is mediated by a pathway involving JNK and Jun. These results demonstrate that CD95 has a growth-promoting role during tumorigenesis and indicate that efforts to inhibit its activity rather than to enhance it should be considered during cancer therapy.

Loss of E-Cadherin Promotes Ovarian Cancer Metastasis via α5-Integrin, which Is a Therapeutic Target

E-cadherin loss is frequently associated with ovarian cancer metastasis. Given that adhesion to the abdominal peritoneum is the first step in ovarian cancer dissemination, we reasoned that down-regulation of E-cadherin would affect expression of cell matrix adhesion receptors. We show here that inhibition of E-cadherin in ovarian cancer cells causes up-regulation of alpha(5)-integrin protein expression and transcription. When E-cadherin was blocked, RMUG-S ovarian cancer cells were able to attach and invade more efficiently. This greater efficiency could, in turn, be inhibited both in vitro and in vivo with an alpha(5)beta(1)-integrin-blocking antibody. When E-cadherin is silenced, alpha(5)-integrin is up-regulated through activation of an epidermal growth factor receptor/FAK/Erk1-mitogen-activated protein kinase-dependent signaling pathway and not through the canonical E-cadherin/beta-catenin signaling pathway. In SKOV-3ip1 ovarian cancer xenografts, which express high levels of alpha(5)-integrin, i.p. treatment with an alpha(5)beta(1)-integrin antibody significantly reduced tumor burden, ascites, and number of metastasis and increased survival by an average of 12 days when compared with IgG treatment (P < 0.0005). alpha(5)-Integrin expression was detected by immunohistochemistry in 107 advanced stage ovarian cancers using a tissue microarray annotated with disease-specific patient follow-up. Ten of 107 tissues (9%) had alpha(5)-integrin overexpression, and 39% had some level of alpha(5)-integrin expression. The median survival for patients with high alpha(5)-integrin levels was 26 months versus 35 months for those with low integrin expression (P < 0.05). Taken together, we have identified alpha(5)-integrin up-regulation as a molecular mechanism by which E-cadherin loss promotes tumor progression, providing an explanation for how E-cadherin loss increases metastasis. Targeting this integrin could be a promising therapy for a subset of ovarian cancer patients.

Foretinib (GSK1363089), an Orally Available Multikinase Inhibitor of c-Met and VEGFR-2, Blocks Proliferation, Induces Anoikis, and Impairs Ovarian Cancer Metastasis

Purpose: Currently, there are no approved targeted therapies for the treatment of ovarian cancer, despite the fact that it is the most lethal gynecological malignancy. One proposed target is c-Met, which has been shown to be an important prognostic indicator in a number of malignancies, including ovarian cancer. The objective of this study was to determine whether an orally available multikinase inhibitor of c-Met and vascular endothelial growth factor receptor-2 (foretinib, GSK1363089) blocks ovarian cancer growth. Experimental Design: The effect of foretinib was tested in a genetic mouse model of endometrioid ovarian cancer, several ovarian cancer cell lines, and an organotypic 3D model of the human omentum. Results: In the genetic mouse model, treatment with foretinib prevented the progression of primary tumors to invasive adenocarcinoma. Invasion through the basement membrane was completely blocked in treated mice, whereas in control mice, invasive tumors entirely replaced the normal ovary. In 2 xenograft mouse models using human ovarian cancer cell lines, the inhibitor reduced overall tumor burden (86% inhibition, P < 0.0001) and metastasis (67% inhibition, P < 0.0001). The mechanism of inhibition by foretinib involved (a) inhibition of c-Met activation and downstream signaling, (b) reduction of ovarian cancer cell adhesion, (c) a block in migration and invasion, (d) reduced proliferation mediated by a G2–M cell-cycle arrest, and (e) induction of anoikis. Conclusions: This study shows that foretinib blocks tumorigenesis and reduces invasive tumor growth in different models of ovarian cancer by affecting several critical tumor functions. We believe that it provides a rationale for the further clinical development of foretinib for the treatment of ovarian cancer. Clin Cancer Res; 17(12); 4042–51. ©2011 AACR.

Abstract LB-341: CD95/FAS promotes tumorigenesis

Proceedings: AACR 101st Annual Meeting 2010‐‐ Apr 17‐21, 2010; Washington, DCnnFAS/CD95 has been known as a typical death receptor that enhances cell apoptosis mainly in the immune system. Recent studies indicate that CD95 expression is rarely absent or down-regulated in solid tumors and CD95L, the cognate ligand that binds cell membrane CD95, is constantly shown to be elevated in patient serum as well as tumor microenviroment, raising the question whether CD95 acturally enhances tumorigenesis and through what mechanisms. By stably knocking down endogeneous CD95 expression level using lentivirus based shRNA in six different tumor cell lines that represent liver (HepG2), colon (HCT116), renal (CAKI-1), breast (MCF7), ovary (SKOV3ip1 and HeyA8), in all cases the CD95 knockdown tumor cell lines show a clear reduction in proliferation rate comparing with the corresponding vector control cells. Injection CD95 knockdown SKOV3ip1 cells i.p. into nude mice according to method of well established xenograft ovary cancer animal model again shows a reduced tumor load and this is also confirmed by using a early passage human ovary cancer cell line MONTY-1. Human neutralizing anti-CD95L mAb (NOK-1) rather than mouse anti-CD95 mAb (MFL3) injection into these xenografted nude mice significantly reduced the tumor load, indicating that its the CD95L secreted by tumor cells rather than the mice microenviroment that stimulates the tumor proliferation. Knocking down FasL expression from the six tumor cell lines mentioned above reduced cell proliferation even more dramatically than knocking down Fas and some cell lines, like HepG2, CAKI-1, MCF7 and HeyA8, even stop growing with reduced FasL expression. The reduced tumor formation by losing FAS/CD95 was also confirmed in vivo in two animal models: DEN induced liver cancer model by using FAS/CD95 liver specific knockout mice and Ade-Cre induced ovary cancer mouse model by generating Fas knockout mice from a mice stain with background of mutated k-ras and deleted Pten expression (LSK-K-ras(G12D/+)Pten(loxP/loxP)). The tumorigenic activity of CD95 is further identified as mediated through the CD95L-CD95-JNK-c-Jun-Fos and EGR1 pathway. These results demonstrate that CD95 plays a major growth promoting role during tumorigenesis and suggest that efforts to inhibit its activity rather than to enhance its activation should be considered during cancer therapy.nnCitation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr LB-341.