An Integrated Approach Identifies Mediators of Local Recurrence in Head and Neck Squamous Carcinoma

PURPOSE: Head and Neck Squamous Cell Carcinomas (HNSCC) cause more than 300,000 deaths worldwide each year. Locoregional and distant recurrences represent worse prognostic events and accepted surrogate markers of patients’ overall survival. No valid biomarker and salvage therapy exist to identify and treat patients at high-risk of recurrence. We aimed to verify if selected microRNAs (miRs) could be used as biomarkers of recurrence in HNSCC. EXPERIMENTAL DESIGN: A Nanostring array was used to identify miRs associated with locoregional recurrence in 44 HNSCC patients. Bioinformatic approaches validated the signature and identified potential miR targets. Validation experiments were performed using an independent cohort of primary HNSCC samples and a panel of HNSCC cell lines. In vivo experiments validated the in vitro results. RESULTS: Our data identified a four-miR signature that classified HNSCC patients at high- or low-risk of recurrence. These miRs collectively impinge on the epithelial-mesenchymal transition process. In silico and wet lab approaches showed that miR-9, expressed at high levels in recurrent HNSCC, targets SASH1 and KRT13, while miR-1, miR-133 and miR-150, expressed at low levels in recurrent HNSCC, collectively target SP1 and TGFβ pathways. A six-genes signature comprising these targets identified patients at high risk of recurrences, as well. Combined pharmacological inhibition of SP1 and TGFβ pathways induced HNSCC cell death and, when timely administered, prevented recurrence formation in a preclinical model of HNSCC recurrence. CONCLUSIONS: By integrating different experimental approaches and competences, we identified critical mediators of recurrence formation in HNSCC that may merit to be considered for future clinical development. TRANSLATIONAL RELEVANCE. Most of HNSCC patients are diagnosed with a locally advanced disease and are treated with the combination of surgery, radiotherapy, and chemotherapy. This highly toxic approach is curative in about half of the cases but recurrent patients do not have effective salvage therapies. Therefore there is the urgency to identify and validate solid biomarkers able to classify patients at high risk that may benefit for specific targeted approaches. Our work tackled these two unmet clinical needs and identified a microRNA signature of locoregional recurrence in HNSCC patients. Starting from this signature, we identified two druggable pathways (i.e. SP1 and TGFβ) that when timely and concomitantly targeted efficiently prevented recurrence formation in a preclinical model. Both SP1 and TGFβ inhibitors have been already used to treat human patients, thus our work is of potential immediate translational relevance.