Widespread Expression of Sonic Hedgehog (Shh) and Nrf2 in Patient Treated with Cisplatin Predicts Outcome in Resected Tumors and Potential Therapeutic Target for Head and Neck Cancer

Background: Sonic hedgehog (Shh) and Nrf2 play an immense role in chemotherapeutic resistance. These genes have been found to be deregulated in head and neck squamous cell carcinoma (HNSCC). The purpose of this study is to analyze the expression, function and clinical prognostic relationship of Shh and Nrf2 in HNSCC in the context of therapeutic resistance and cancer stem cells (CSCs). Methods: We analyzed a cohort of HNSCC patients to identify potential candidate therapeutic biomarkers correlating with overall survival (OS) and disease-free survival (DFS) from our own data and validated the results using data from TCGA. Expression of Shh and Nrf2 was silenced by siRNA and cell growth, sphere growth and chemotherapeutic resistance were evaluated. Findings: Widespread abundant expression of Shh and Nrf2 protein was associated with shorter OS and DFS. The combination of Shh and Nrf2 expression levels was found to be a mostly significant predictor of patient DFS. The tumor stromal index (SI) was correlated with Shh expression and inversely associated with shorter DFS. Inhibition of Shh by siRNA or cyclopamine resulted in the attenuation of resistant CSC self-renewal, invasion, clonogenic growth and re-sensitization to chemotherapeutic agents. Interpretation: Concomitant high expression of Shh and Nrf2 proved to be an independent predictor of poor OS and DFS in HNSCC patients. These findings suggest that Shh and Nrf2 might serve as therapeutic targets as well as promising dual prognostic therapeutic biomarkers for HNSCC. Funding Statement: Grant-in-Aid from Bangladesh Medical Research Council (BMRC), and University Grant Commission (UGC), Ministry of Education of Bangladesh with the assistance of the World Bank under Higher Education Quality Enhancement Project (HEQEP)– Window -4 (Grant Id. CP4023). Declaration of Interests: The authors declare that they have no conflict of interests. Ethics Approval Statement: This study was approved by the ethics committee of the University of Chittagong and Chittagong Medical Hospital, Chittagong, Bangladesh, and was performed in accordance with the ethical standards laid down in the 1964 declaration of Helsinki and all subsequent revisions. Written informed consent was obtained from all patients used in this study.