Panobinostat potentiates temozolomide effects and reverses epithelial-mesenchymal transition in glioblastoma cells

Glioblastoma is the most common form of glioma, as well as the most aggressive. Patients suffering from this disease have a very poor prognosis. Surgery, radiotherapy, and temozolomide are the only approved treatments nowadays. Panobinostat is a pan-inhibitor of histone deacetylases (HDACs) that has been shown to break some pathways which play an important role in cancer development. A global intention of using panobinostat as a therapeutic agent against glioblastoma is beginning to be a reality. We have treated the LN405 glioblastoma cell line with temozolomide, panobinostat, and combined treatment, in order to test apoptosis, colony formation, and a possible molecular reversion of the mesenchymal phenotype of the cells to an epithelial one. Our results show that panobinostat decreased N-cadherin levels in the LN405 glioblastoma cell line while it increased the expression of E-cadherin, which might be associated with a mesenchymal–epithelial transition in glioblastoma cells. Colony formation was reduced, and apoptosis was increased with treatments. Our research highlights the importance of panobinostat as a potential adjuvant therapy to be used with temozolomide to treat glioblastoma and the advantages of the combined treatment versus temozolomide alone, which is currently the first-line treatment used to treat this tumo

Panobinostat potentiates temozolomide effects and reverses epithelial-mesenchymal transition in glioblastoma cells

Glioblastoma is the most common form of glioma, as well as the most aggressive. Patients suffering from this disease have a very poor prognosis. Surgery, radiotherapy, and temozolomide are the only approved treatments nowadays. Panobinostat is a pan-inhibitor of histone deacetylases (HDACs) that has been shown to break some pathways which play an important role in cancer development. A global intention of using panobinostat as a therapeutic agent against glioblastoma is beginning to be a reality. We have treated the LN405 glioblastoma cell line with temozolomide, panobinostat, and combined treatment, in order to test apoptosis, colony formation, and a possible molecular reversion of the mesenchymal phenotype of the cells to an epithelial one. Our results show that panobinostat decreased N-cadherin levels in the LN405 glioblastoma cell line while it increased the expression of E-cadherin, which might be associated with a mesenchymal–epithelial transition in glioblastoma cells. Colony formation was reduced, and apoptosis was increased with treatments. Our research highlights the importance of panobinostat as a potential adjuvant therapy to be used with temozolomide to treat glioblastoma and the advantages of the combined treatment versus temozolomide alone, which is currently the first-line treatment used to treat this tumo

Epigenetic regulation of human hedgehog interacting protein in glioma cell lines and primary tumor samples

Glioma constitutes one of the most common groups of brain tumors, and its prognosis is influenced by different genetic and epigenetic modulations. In this study, we demonstrated low or no expression of hedgehog interacting protein (HHIP) in most of the cell lines and primary glioma tumor samples. We further proceeded to promoter methylation study of this gene in the same cell lines and primary tumor samples and found 87 % (7/8) HHIP methylation in glioblastoma cell lines and 75 % (33/44) in primary tumor samples. These methylation pattern correlates with low or unexpressed HHIP in both cell lines and primary tumor samples. Our results suggest the possibility of epigenetic regulation of this gene in glioma, similarly to medulloblastoma, gastric, hepatic, and pancreatic cancers. Also, HHIP might be a diagnostic or prognostic marker in glioma and help to the detection of these tumors in early stages of disease

Tubastatin A, an inhibitor of HDAC6, enhances temozolomide‐induced apoptosis and reverses the malignant phenotype of glioblastoma cells

Glioblastoma or grade IV astrocytoma is the most common and lethal form of glioma. Current glioblastoma treatment strategies use surgery followed by chemotherapy with temozolomide. Despite this, numerous glioblastoma cases develop resistance to temozolomide treatments, resulting in a poor prognosis for the patients. Novel approaches are being investigated, including the inhibition of histone deacetylase 6 (HDAC6), an enzyme that deacetylates α-tubulin, and whose overexpression in glioblastoma is associated with the loss of primary cilia. The aim of the present study was to treat glioblastoma cells with a selective HDAC6 inhibitor, tubastatin A, to determine if the malignant phenotype may be reverted. The results demonstrated a notable increase in acetylated α-tubulin levels in treated cells, which associated with downregulation of the sonic hedgehog pathway, and may hypothetically promote ciliogenesis in those cells. Treatment with tubastatin A also reduced glioblastoma clonogenicity and migration capacities, and accelerated temozolomide-induced apoptosis. Finally, HDAC6 inhibition decreased the expression of mesenchymal markers, contributing to reverse epithelial-mesenchymal transition in glioblastoma cells

APR-246 combined with 3-deazaneplanocin A, panobinostat or temozolomide reduces clonogenicity and induces apoptosis in glioblastoma cells

Glioblastoma is the most malignant brain tumor and presents high resistance to chemotherapy and radiotherapy. Surgery, radiotherapy and chemotherapy with temozolomide are the only treatments against this tumor. New targeted therapies, including epigenetic modulators such as 3‑deazaneplanocin A (DZ‑Nep; an EZH2 inhibitor) and panobinostat (a histone deacetylase inhibitor) are being tested in vitro, together with temozolomide. The present study combined APR‑246 with DZ‑Nep, panobinostat and teomozolomide in order to explore the possibility of restoring p53 function in mutated cases of glioblastoma. Following the Chou‑Talalay method it was demonstrated that APR‑246 acts in an additive manner together with the other compounds, reducing clonogenicity and inducing apoptosis in glioblastoma cells independently of p53 status

The synergistic effect of DZ‑NEP, panobinostat and temozolomide reduces clonogenicity and induces apoptosis in glioblastoma cells

Current treatment against glioblastoma consists of surgical resection followed by temozolomide, with or without combined radiotherapy. Glioblastoma frequently acquires resistance to chemotherapy and/or radiotherapy. Novel therapeutic approaches are thus required. The inhibition of enhancer of zeste homolog 2 (EZH2; a histone methylase) and histone deacetylases (HDACs) are possible epigenetic treatments. Temozolomide, 3-deazaneplanocin A (DZ-Nep; an EZH2 inhibitor) and panobinostat (an HDAC inhibitor) were tested in regular and temozolomide-resistant glioblastoma cells to confirm whether the compounds could behave in a synergistic, additive or antagonistic manner. A total of six commercial cell lines, two temozolomide-induced resistant cell lines and two primary cultures derived from glioblastoma samples were used. Cell lines were exposed to single treatments of the drugs in addition to all possible two- and three-drug combinations. Colony formation assays, synergistic assays and reverse transcription-quantitative PCR analysis of apoptosis-associated genes were performed. The highest synergistic combination was DZ-Nep + panobinostat. Triple treatment was also synergistic. Reduced clonogenicity and increased apoptosis were both induced. It was concluded that the therapeutic potential of the combination of these three drugs in glioblastoma was evident and should be further explored

Silencing of histone deacetylase 6 decreases cellular malignancy and contributes to primary cilium restoration, epithelial-to-mesenchymal transition reversion, and autophagy inhibition in glioblastoma cell lines

Glioblastoma multiforme, the most common type of malignant brain tumor as well as the most aggressive one, lacks an effective therapy. Glioblastoma presents overexpression of mesenchymal markers Snail, Slug, and N-Cadherin and of the autophagic marker p62. Glioblastoma cell lines also present increased autophagy, overexpression of mesenchymal markers, Shh pathway activation, and lack of primary cilia. In this study, we aimed to evaluate the role of HDAC6 in the pathogenesis of glioblastoma, as HDAC6 is the most overexpressed of all HDACs isoforms in this tumor. We treated glioblastoma cell lines with siHDAC6. HDAC6 silencing inhibited proliferation, migration, and clonogenicity of glioblastoma cell lines. They also reversed the mesenchymal phenotype, decreased autophagy, inhibited Shh pathway, and recovered the expression of primary cilia in glioblastoma cell lines. These results demonstrate that HDAC6 might be a good target for glioblastoma treatment

Epigenetic regulation of human hedgehog interacting protein in glioma cell lines and primary tumor samples

Glioma constitutes one of the most common groups of brain tumors, and its prognosis is influenced by different genetic and epigenetic modulations. In this study, we demonstrated low or no expression of hedgehog interacting protein (HHIP) in most of the cell lines and primary glioma tumor samples. We further proceeded to promoter methylation study of this gene in the same cell lines and primary tumor samples and found 87 % (7/8) HHIP methylation in glioblastoma cell lines and 75 % (33/44) in primary tumor samples. These methylation pattern correlates with low or unexpressed HHIP in both cell lines and primary tumor samples. Our results suggest the possibility of epigenetic regulation of this gene in glioma, similarly to medulloblastoma, gastric, hepatic, and pancreatic cancers. Also, HHIP might be a diagnostic or prognostic marker in glioma and help to the detection of these tumors in early stages of disease