PO-083 Suppressing the accumulation of cancer stem cells (CSC) using HDAC inhibitor reverses HNSCC chemoresistance by targeting NFKB signalling

IntroductionEpigenetic mechanisms regulate transcription of CSC, controlling self-renewal and differentiation, providing survival advantages, contributing to tumour progression and chemoresistance. Histone acetylation is associated with transcriptional activation. Changes in histone acetylation are observed in cancer initiation and progression, maintenance of CSC, and tumour resistance. Therapeutic agents like Cisplatin target proliferating cells, but CSC survive and regenerate the tumour. We investigated the ability of HDAC inhibitor Vorinostat in disrupting CSC and reversing chemoresistance in HNSCC.Material and methodsWe explored CSC and histone acetylation levels in HNSCC cell lines and tumour samples. Immunofluorescence, immunoblotting, spheres formation, flow cytometry and proliferation were used to investigate cell lines after treatment with Vorinostat, Cisplatin and both. The efficiency of the combined treatment was analysed in vivo using xenograft model. NFkB signalling was assessed using interference RNA (si) for IKKα, TNF-α and Emetine treatments.Results and discussionsHistone acetylation levels are heterogeneous in HNSCC and decreased in the CSC subpopulation. CSCs are chemoresistant and enriched after Cisplatin treatment. HNSCC cells resistant to cisplatin present greater CSC accumulation than sensitive. Very low doses of Vorinostat diminish CSC 8 hour after administration and abrogate CSC after 5 days. Prolonged treatment using low doses of Vorinostat can be applied to efficiently impair the CSC subpopulation. Previous Vorinostat administration sensitises cells to Cisplatin. Cisplatin alone decreases histone acetylation and increases MBD2c, which is associated with pluripotency and stem cells, and previous administration of Vorinostat reverses this, indicating that Vorinostat chemosensitizes HNSCC reducing CSC via epigenetic regulation. Combined treatment reduces tumour volume in xenograft models, abolishing CSC and increasing histone acetylation without increasing toxicity. Combined treatment decreases nuclear NFkB, which was previously associated with resistance to Cisplatin and diminished histone acetylation in HNSCC. SiIKKα and TNF-α treatment showed that the amount of CSC was directly associated with NFkB levels. Emetine, a FDA-approved drug that decreases NFkB, reduced CSC alone and in combination with TNF-α or Cisplatin.ConclusionOur findings suggest that drugs that decrease NFkB inducing chromatin acetylation result in the destruction