Abstract 5185: Targeting inhibitors of apoptosis in combination with radiation for the treatment of head and neck squamous cell carcinoma

Abstract

Abstract Background and purpose: Evasion of apoptosis contributes to radioresistance of head and neck squamous cell carcinoma (HNSCC), calling for novel strategies to overcome apoptotic resistance. Second mitochondria-derived activator of caspase (SMAC) - mimetics are a class of targeted drugs that antagonize selected inhibitor of apoptosis protein (IAP) to block pro-survial signalling, and eventually induce cell apoptosis. The present study was designed to investigate the radiosensitizing effect of one SMAC mimetic, LCL161, in HNSCC and the underlying mechanisms for radiosensitization. Material and methods: We analyzed the correlation between apoptotic molecules and HPV status in HNSCC via mRNA expression using The Cancer Genome Atlas (TCGA) database, as well as protein expression in 6 HNSCC cell lines by immunoblotting. We also examined cIAP1 expression on a tissue microarray (TMA) of HNSCC tumors, and assessed its correlation with HPV status and patient outcome. Clonogenic survival and WST-1 assays were carried out to explore the potential of LCL161 as a radiosensitizer in HNSCC cell lines. Cell cycle analysis, Annexin-V assays and immunoblotting were performed to investigate effects of LCL161 on radiation induced cell apoptosis. Human tumor xenografts were generated to explore the radiosensitization effect of LCL161 on HPV[-] tumors in vivo. Results: TCGA database analysis and immunoblotting from HNSCC cells revealed that expression of cIAP1, Survivin, DIABLO, p65, IK-b, TNF-α, BCL-XL, Caspase-7, and Caspase-8 were elevated in HPV[-] compared to HPV[+] HNSCC tumors. IHC staining showed cIAP1 expression was associated with HPV[-] status (p=0.0239). In addition, cIAP1 expression was significantly higher in HNSCC compared with adjacent normal tissue (p=0.0003). In univariate analysis, cIAP1 was significantly associated with poor overall survival (p=0.0359). LCL161 degraded cIAP1 at nanomolar concentrations but has minimal single-agent cytotoxicity with IC50 range of 32 μM – 95 μM. Interestingly, we found that LCL161 could induce radiosensitization only in HPV[-], but not in HPV[+] HNSCC cells. Mechanistic studies showed LCL161 mediated radiosensitisation was associated with increased cell apoptosis, with enhanced activation of caspases-3, -7, -8, -9, and PARP. Finally, in vivo data showed that LCL161 degraded cIAP1, activated caspases, and profoundly radiosensitized two human HPV[-] HNSCC xenograft models. Conclusion: Taken together, our results demonstrate for the first time that cIAP1 is a potential prognostic and therapeutic biomarker for HNSCC patients. Through targeting cIAP1 and other IAP family members, LCL161, radiosensitizes HNSCC tumors, particularly HPV [-] tumors, via caspase activation and apoptosis induction. Taken together, LCL161 holds promise for future clinical development as a novel radiosensitizer in the treatment of HPV[-] HNSCC tumors. Citation Format: Linlin Yang, Bhavna Kumar, Tianyun Li, Thenodoros N. Teknos, Arnab Chakravarti, Terence M. Williams. Targeting inhibitors of apoptosis in combination with radiation for the treatment of head and neck squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5185. doi:10.1158/1538-7445.AM2017-5185