Abstract 5223: Valproic acid, by preventing cisplatin/cetuximab-induced EGFR nuclear translocation and increasing cisplatin uptake, potentiates the antitumor effect of the combination treatment in head and neck squamous cell carcinomas
Authors: Iannelli Federica , Zotti Andrea Ilaria , Roca Maria Serena , Grumetti Laura , Moccia Tania , Vitagliano Carlo , Ciardiello Chiara , Bruzzese Francesca , Leone Alessandra , Caponigro Francesco , Ionna Franco , Longo Francesco , Gennaro Elena Di , Budillon Alfredo 
Published: September 2020

Abstract
INTRODUCTION: Recurrent metastatic (R/M) squamous cell carcinoma of the head and neck (HNSCC) is a devastating malignancy with a poor prognosis and the combination of cisplatin (CDDP) plus cetuximab (CX) is one of the gold standard for first-line treatment. However, this therapy is often associated with toxicity and resistance, suggesting that new combinatorial strategies are needed to improve the therapeutic index of this regimen. In our study, we evaluated the synergistic antitumor effect of valproic acid (VPA), an anticonvulsant compound with histone deacetylase inhibitor (HDACi) activity, in combination with CDDP/CX in HNSCC models in vitro and in vivo.
METHODS: Synergistic anti-proliferative effects were assessed on HNSCC Cal27, FaDu and Cal33 cell lines and BJ-hTERT normal fibroblast, by calculating combination index (CI) accordingly to Chou and Talalay method. Apoptosis was measured by flow cytometry analysis and caspase assay. Tumor spheroids were obtained by low attach systems and scored with luminescence 3D-cell viability assay. In vivo experiment was performed on xenograft models in athymic mice.
RESULTS: We demonstrated, in HNSCC cells, but not in normal human fibroblasts, that simultaneous exposure to equitoxic doses of VPA plus CDDP/CX results in a clear synergistic antiproliferative and proapoptotic effect. Interestingly, in order to better recapitulate tumor growth complexity compared to 2D monolayers conditions, the synergistic antitumor effect was confirmed in four different 3D-self-assembled spheroid models, as well as in in vivo Cal27 xenograft model. Mechanistically, VPA induced a dose-dependent down-regulation of EGFR expression/activation affecting its downstream canonical pathway (pAKT and pMAPK), which plays a driver role in HNSCC. Moreover, we demonstrated that VPA was able to prevent the CDDP and/or CX induced EGFR nuclear translocation, a well-known mechanism of resistance to chemotherapy. Indeed, VPA impaired the transcription of cyclin D1 and DNA repair genes, regulated by non-canonical activity of nuclear EGFR, thus increasing DNA damage induced by CDDP/CX combination. Moreover, VPA was able to enhance the sensitivity to CDDP, by upregulating, at transcriptional level, the CDDP influx channel copper transporter 1 (CTR1).
CONCLUSIONS: The introduction of a safe and generic drug such as VPA into the conventional treatment for R/M HNSCC, represents an innovative and feasible antitumor strategy that warrants further clinical evaluation. Indeed, we are currently enrolling patients in a phase-2 clinical trial in order to explore whether the addition of VPA to the standard combination CDDP/CX can increase the response rate in patients with R/M HNSCC.
Citation Format: Federica Iannelli, Andrea Ilaria Zotti, Maria Serena Roca, Laura Grumetti, Tania Moccia, Carlo Vitagliano, Chiara Ciardiello, Francesca Bruzzese, Alessandra Leone, Francesco Caponigro, Franco Ionna, Francesco Longo, Elena Di Gennaro, Alfredo Budillon. Valproic acid, by preventing cisplatin/cetuximab-induced EGFR nuclear translocation and increasing cisplatin uptake, potentiates the antitumor effect of the combination treatment in head and neck squamous cell carcinomas [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5223.