Abstract 2623: Induction of Bmf and Bim in Vorinostat-induced apoptosis in mantle cell lymphoma

Abstract

Abstract OBJECTIVES: Mantle cell lymphoma (MCL) is an incurable B-cell neoplasm harboring the t(11;14)(q13;q32) which leads to the overexpression of cyclin D1, with the consequent cell cycle deregulation. Usually, MCL is characterized by bad prognosis and an aggressive course of the disease. Current therapies have shown limited efficacy. Recently, Histone Deacetylase Inhibitors (HDACis) have been successfully introduced for the treatment of several cancers, including hematological malignancies. Accordingly, tumor suppressor genes are frequently epigenetically silenced in these entities, due to histone deacetylation in their promoters. Therefore, our purposes were to evaluate the antitumoral properties of the HDACi suberoylanilide hydroxamic acid (Vorinostat; Merck & Co) in MCL, and to describe the molecular mechanisms involved in HDACi signaling in this disorder. METHODS: MCL cell lines and primary MCL cells were used. HDAC activity was measured using a colorimetric HDAC Assay Kit. Flow cytometry was used to determine sensitivity to Vorinostat through measurement of AnnexinV binding, and to analyze apoptosis features. Vorinostat anti-tumoral signaling was evaluated by RQ-PCR determination of gene transcription, western blot analysis and acetyl Histone H4 ChIP assays. RESULTS: Vorinostat exhibited a heterogeneous cytotoxic effect among MCL cell lines, with a median LD50 of 6.6 μM after 24-hour incubation. Nevertheless, cytotoxicity increased notably after 48h of exposure to the drug with LD50 ranging from 0.4 to 5.3 μM. Interestingly, 7 out of the 10 MCL primary samples tested were extremely sensitive to the compound (with a median LD50 of 2.2 μM after 24-hour incubation). Vorinostat increases the acetylation of H3 and H4 histones, as well as inhibits global HDAC activity in just 1 hour of incubation. The drug notably decreases cyclin D1 protein levels while induces upregulation of the proapoptotic BH3-only proteins Bmf and Bim, triggering the mitochondria-dependent cell death and activation of the caspases cascade. Acetyl Histone H4 ChIP assays showed that Vorinostat increases acetylation of BMF and BIM gene promoters, consequently up regulating Bim and Bmf mRNA levels. CONCLUSIONS: This study suggests that Vorinostat could define a new and attractive therapeutic approach for the treatment of MCL. We identify BMF and BIM as possible crucial target genes of HDAC inhibitors in MCL cells, promoting the induction of mitochondria-mediated apoptosis. GRANTS: This work is supported by Ministerio de Ciencia e Innovación SAF 06-8850; S. X-T is a fellow from Ministerio de Ciencia e Innovación (FPU predoctoral fellowship). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2623.