FADD Gene Therapy for Malignant GliomasIn VitroandIn Vivo

Abstract

Fas/APO-1 (CD95), a cell surface cytokine receptor, triggers apoptotic cell death by specific agonist antibody, suggesting that Fas/APO-1 may be a promising target for treatment of tumors. In this study, we show that treatment with anti-Fas antibody effectively induced apoptosis in malignant glioma cell lines with high expression of Fas/APO-1 (n = 3). Malignant glioma cells with low or undetectable expression of Fas/APO-1 (n = 6), however, were resistant to Fas/APO-1-dependent cytotoxicity. The purpose of this study, therefore, was to determine whether resistant tumors could be made susceptible to apoptosis. FADD/MORT1 constitutes a novel protein that associates specifically with the cytoplasmic death domain of Fas/APO-1 and induces apoptosis. We investigated whether overexpression of FADD would induce apoptosis in malignant glioma cells without activating Fas/APO-1. Results indicated that about 85% of malignant glioma cells, regardless of Fas/APO-1 expression levels, underwent apoptosis after transient transfection with FADD expression vector. To further improve gene transfer of FADD into malignant glioma cells, we constructed a retroviral vector containing the FADD gene. The retroviral transfer of FADD gene significantly enhanced the transduction efficiency and effectively inhibited both in vitro and in vivo survival of malignant glioma cells through induction of apoptosis. These findings suggest that the FADD gene is a novel and useful tool for the treatment of malignant gliomas.

Targeting proliferating tumor cells via the transcriptional control of therapeutic genes

“…For first strand cDNA synthesis1 mg of total RNA was used. cDNA was synthesized using random hexanucleotide primers (Invitrogen Life Technologies) and oligo-dT [12][13][14][15][16][17] primers, in the presence of superscript II reverse transcriptase (Invitrogen Life Technologies). The expression level of FasL, FADD and the 18S rRNA was quantitated using QuantiTecht SYBR Green PCR kit (Qiagen).…”

“…8 Interestingly, it has been reported that overexpression of FADD is able to induce apoptosis in FasL-insensitive cells. [9][10][11][12] Since cancerous cells such as gliomas are known to be highly heterogeneous, it is likely that different clonal populations within a tumor mass might respond differently to different therapeutic agents. Thus, from a therapeutic point of view, the administration of multiple therapeutic agents would be of great benefit.…”

The death effector domain protein family

“…This has prompted gene therapy studies that use FADD as a 'suicide gene' when targeted to tumor cells, resulting in the apoptosis of FADDpositive cells and regression of tumor volume in mice (Fan et al, 1999;Koga et al, 2001;Komata et al, 2001). Another potential strategy is to sensitize tumors cells with low or undetectable levels of CD95 that are otherwise resistant to CD95-mediated apoptosis, increasing the effectiveness of the apoptotic signal when FADD is overexpressed (Kondo et al, 1998).…”

Novel gene therapy for rheumatoid arthritis by FADD gene transfer: induction of apoptosis of rheumatoid synoviocytes but not chondrocytes

“…In this respect, previous studies have reported that overexpression of FADD can induce apoptosis of various cells. 12,13,18,19 Interestingly, deletion mutants of FADD, which contain DED but not DD, can also induce apoptosis, suggesting that overexpression of FADD can induce apoptosis without involving the DD of Fas antigen. 18,19 We found that overexpression of FADD induced apoptotic cell death of cultured RA synoviocytes accompanied with DNA fragmentation and nuclear condensation.…”

“…11 Recent studies have shown that FADD gene transfer by adenoviral or retroviral vectors potently induced apoptosis of human malignant glioma cells. 12,13 In these studies, Fas-associated death domain protein (FADD), binds to intracellular death domain of Fas and promotes signaling pathways of Fas-mediated apoptosis. In this regard, we have recently reported that FADD is a key signaling molecule for Fas-mediated apoptosis of synoviocytes in patients with RA, 14 suggesting that FADD gene transfer could also induce apoptosis of RA synoviocytes.…”