Therapeutic targeting of c-FLIP with antisense phosphorothioate oligonucleotides
Year: 2008
Session type: Poster / e-Poster / Silent Theatre session
Queen's University, Belfast, UK
Abstract
c-FLIP is a key inhibitor of apoptosis mediated by death receptors Fas, DR4 and DR5 and is expressed as long (c-FLIPL) and short (c-FLIPS) splice forms.
Background
The aim of this study was to design and characterise c-FLIP targeted (FT) phosphorothioate antisense oligonucleotides (ASO) targeting both c-FLIP slice forms.
Method and results
c-FLIP knockdown was assessed using Western Blot analysis. Induction of apoptosis by FT ASO was assessed by Flow Cytometry analysis and evaluating the percentage of apoptotic sub-GO/G1 cells. For in vivo effects of FT ASO, Balb/c nude mice were subcutaneously inoculated with H460 cells using matrigel. FT ASO (20mg/kg) was delivered by intra-peritoneal injection.
FT ASO targeting both splice forms induced apoptosis in H460 non-small cell lung cancer (NSCLC) cells. Flow analysis showed that FT ASO sensitized H460 cells to TRAIL and chemotherapy (taxol and cisplatin). Further in vitro characterisation of FT ASO was carried out in A549 NSCLC, HCT116 colorectal cancer and PC3 prostate cancer cell lines. In all cell lines, AS PTO-mediated down-regulation of c-FLIP induced apoptosis and increased sensitivity to TRAIL. Importantly intra-peritoneal administration of FT ASO induced apoptosis and inhibited the growth of H460 xenografts in BALB/c nude mice. In addition, mice injected with FT ASO displayed enhanced growth inhibitory effects of cisplatin in H460 xenografts.
Conclusion
These results indicate that c-FLIP-targeted antisense phosphorothioates may have therapeutic potential for the treatment of several types of solid cancer.
c-FLIP is a key inhibitor of apoptosis mediated by death receptors Fas, DR4 and DR5 and is expressed as long (c-FLIPL) and short (c-FLIPS) splice forms.
Background
The aim of this study was to design and characterise c-FLIP targeted (FT) phosphorothioate antisense oligonucleotides (ASO) targeting both c-FLIP slice forms.
Method and results
c-FLIP knockdown was assessed using Western Blot analysis. Induction of apoptosis by FT ASO was assessed by Flow Cytometry analysis and evaluating the percentage of apoptotic sub-GO/G1 cells. For in vivo effects of FT ASO, Balb/c nude mice were subcutaneously inoculated with H460 cells using matrigel. FT ASO (20mg/kg) was delivered by intra-peritoneal injection.
FT ASO targeting both splice forms induced apoptosis in H460 non-small cell lung cancer (NSCLC) cells. Flow analysis showed that FT ASO sensitized H460 cells to TRAIL and chemotherapy (taxol and cisplatin). Further in vitro characterisation of FT ASO was carried out in A549 NSCLC, HCT116 colorectal cancer and PC3 prostate cancer cell lines. In all cell lines, AS PTO-mediated down-regulation of c-FLIP induced apoptosis and increased sensitivity to TRAIL. Importantly intra-peritoneal administration of FT ASO induced apoptosis and inhibited the growth of H460 xenografts in BALB/c nude mice. In addition, mice injected with FT ASO displayed enhanced growth inhibitory effects of cisplatin in H460 xenografts.
Conclusion
These results indicate that c-FLIP-targeted antisense phosphorothioates may have therapeutic potential for the treatment of several types of solid cancer.
