• HDACi
• cancer
• histone acetyltransferases
• histone deacetylases
• lymphoma

• Humans
• Neoplasms/drug therapy
• Neoplasms/metabolism
• Neoplasms/enzymology
• Neoplasms/genetics
• Neoplasms/pathology
• Neoplasms/etiology
• Histone Deacetylase Inhibitors/therapeutic use
• Histone Deacetylase Inhibitors/pharmacology
• Histone Deacetylases/metabolism
• Histone Deacetylases/genetics
• Epigenesis, Genetic/drug effects
• Animals
• Tumor Microenvironment/drug effects
• Antineoplastic Agents/therapeutic use
• Antineoplastic Agents/pharmacology

• TSA
• cell cycle synchronization
• ovine adult fibroblast
• ovine foetal fibroblast
• roscovitine

• Animals
• Sheep
• Roscovitine/pharmacology
• Roscovitine/metabolism
• Purines/pharmacology
• Purines/metabolism
• Cell Cycle
• Sheep, Domestic
• Fibroblasts

• Türkiye Bilimsel ve Teknolojik Araştırma Kurumu

Compared to pancreatic adenocarcinoma (PDAC), pancreatic neuroendocrine tumors (PanNET) represent a rare and heterogeneous tumor entity. In addition to surgical resection, several therapeutic approaches, including biotherapy, targeted therapy or chemotherapy are applicable. However, primary or secondary resistance to current therapies is still challenging. Recent genome-wide sequencing efforts in PanNET identified a large number of mutations in pathways involved in epigenetic modulation, including acetylation. Therefore, targeting epigenetic modulators in neuroendocrine cells could represent a new therapeutic avenue. Detailed information on functional effects and affected signaling pathways upon epigenetic targeting in PanNETs, however, is missing. The primary human PanNET cells NT-3 and NT-18 as well as the murine insulinoma cell lines beta-TC-6 (mouse) and RIN-T3 (rat) were treated with the non-selective histone-deacetylase (HDAC) inhibitor panobinostat (PB) and analyzed for functional effects and affected signaling pathways by performing Western blot, FACS and qPCR analyses. Additionally, NanoString analysis of more than 500 potentially affected targets was performed. In vivo immunohistochemistry (IHC) analyses on tumor samples from xenografts and the transgenic neuroendocrine Rip1Tag2-mouse model were investigated. PB dose dependently induced cell cycle arrest and apoptosis in neuroendocrine cells in human and murine species. HDAC inhibition stimulated redifferentiation of human primary PanNET cells by increasing mRNA-expression of somatostatin receptors (SSTRs) and insulin production. In addition to hyperacetylation of known targets, PB mediated pleitropic effects via targeting genes involved in the cell cycle and modulation of the JAK2/STAT3 axis. The HDAC subtypes are expressed ubiquitously in the existing cell models and in human samples of metastatic PanNET. Our results uncover epigenetic HDAC modulation using PB as a promising new therapeutic avenue in PanNET, linking cell-cycle modulation and pathways such as JAK2/STAT3 to epigenetic targeting. Based on our data demonstrating a significant impact of HDAC inhibition in clinical relevant in vitro models, further validation in vivo is warranted.

• HDAC
• PanNET
• neuroendocrine
• pancreas
• panobinostat

In human head and neck squamous cell carcinoma (HNSCC), the invasion and metastatic properties of cancer cells are promoted by junctional adhesion molecule-A (JAM-A) and claudin-1; these are epithelial tight junction molecules regulated by histone deacetylases (HDACs) and transcription factor p63. HDAC expression is reportedly upregulated in HNSCC, and HDAC inhibitors suppress cancer cell proliferation by initiating proliferative arrest or apoptosis. However, little is known of the anti-cancer mechanisms of HDAC inhibitors in HNSCC. Thus, in the present study, the HNSCC Detroit 562 cell line and primary cultured HNSCC cells were treated with HDAC inhibitors to investigate their effects in HNSCC. Higher expression of p63, HDAC1, JAM-A and claudin-1 was observed in HNSCC tissues compared with the adjacent dysplastic regions. In Detroit 562 cells, treatment with trichostatin A (TSA), an inhibitor of HDAC1 and 6, downregulated the expression of p63, JAM-A and claudin-1, and upregulated that of acetylated tubulin; conversely, p63 knockdown resulted in the downregulation of JAM-A and claudin-1. Collectively, inhibiting HDAC suppressed the migration and invasiveness of cancer cells. In addition, treatment with TSA suppressed cancer cell proliferation via G₂/M arrest, as well as upregulating p21 and downregulating cyclin D1 expression. TSA also downregulated the expression of epidermal growth factor receptor (EGFR) and phospho-ERK1/2. p63 knockdown and treatment with an EGFR inhibitor induced G₁ arrest and downregulated EGFR and phospho-ERK1/2 levels, respectively. HDAC inhibition also suppressed the migration and invasiveness of primary cultured HNSCC cells. Collectively, the results of the present study indicate that HDAC inhibitors suppress the proliferation, migration and invasiveness of HNSCC by downregulating the p63-mediated tight junction molecules JAM-A and claudin-1, and inducing p63 or p21-mediated growth arrest.

• HDAC inhibitors
• head and neck squamous cell carcinoma
• JAM‑A
• claudin‑1
• p63
• EGFR
• p21

• Apoptosis
• DNA methyltransferases
• Genistein
• Hepatocellular carcinoma
• Trichostatin A

• Costus pictus methanolic extract
• cytotoxicity
• histone deacetylase
• liver hepatocellular carcinoma cells

• Pterocladiella capillacea
• apoptosis
• cell cycle arrest
• cellular effectors
• colon cancer
• mertensene

• Adenocarcinoma/drug therapy
• Adenocarcinoma/metabolism
• Apoptosis/drug effects
• Caspases/metabolism
• Cell Cycle Checkpoints/drug effects
• Cell Cycle Proteins/metabolism
• Cell Line, Tumor
• Colonic Neoplasms/drug therapy
• Colonic Neoplasms/metabolism
• Cyclin-Dependent Kinases/metabolism
• G2 Phase Cell Cycle Checkpoints/drug effects
• HT29 Cells
• Humans
• MAP Kinase Signaling System/drug effects
• Monoterpenes/pharmacology
• NF-kappa B/metabolism
• Poly(ADP-ribose) Polymerases/metabolism
• Proto-Oncogene Proteins c-akt/metabolism
• Signal Transduction/drug effects

The formation of new lymphatic vessels provides an additional route for tumour cells to metastasize. Therefore, inhibiting lymphangiogenesis represents an interesting target in cancer therapy. First evidence suggests that histone deacetylase inhibitors (HDACi) may mediate part of their antitumor effects by interfering with lymphangiogenesis. However, the underlying mechanisms of HDACi induced anti-lymphangiogenic properties are not fully investigated so far and in part remain unknown.

Human lymphatic endothelial cells (LEC) were cultured in vitro and treated with or without HDACi. Effects of HDACi on proliferation and cell cycle progress were analysed by BrdU assay and flow cytometry. Apoptosis was measured by quantifying mono- and oligonucleosomes in the cytoplasmic fraction of cell lysates. In vitro lymphangiogenesis was investigated using the Matrigel short term lymphangiogenesis assay. The effects of TSA on cell cycle regulatory proteins and apoptosis-related proteins were examined by western blotting, immunofluorescence staining and semi-quantitative RT-PCR. Protein- and mRNA half-life of p21 were analysed by western blotting and quantitative RT-PCR. The activity of the p21 promoter was determined using a dual luciferase assay and DNA-binding activity of Sp1/3 was investigated using EMSA. Furthermore, siRNA assays were performed to analyse the role of p21 and p53 on TSA-mediated anti-lymphangiogenic effects.

We found that HDACi inhibited cell proliferation and that the pan-HDACi TSA induced G0/G1 arrest in LEC. Cell cycle arrest was accompanied by up-regulation of p21, p27 and p53. Additionally, we observed that p21 protein accumulated in cellular nuclei after treatment with TSA. Moreover, we found that p21 mRNA was significantly up-regulated by TSA, while the protein and mRNA half-life remained largely unaffected. The promoter activity of p21 was enhanced by TSA indicating a transcriptional mechanism. Subsequent EMSA analyses showed increased constitutive Sp1/3-dependent DNA binding in response to HDACi. We demonstrated that p53 was not required for TSA induced p21 expression and growth inhibition of LECs. Interestingly, siRNA-mediated p21 depletion almost completely reversed the anti-proliferative effects of TSA in LEC. In addition, TSA induced apoptosis by cytochrome c release contributed to activating caspases-9, −7 and −3 and downregulating the anti-apoptotic proteins cIAP-1 and −2.

In conclusion, we demonstrate that TSA - a pan-HDACi - has distinct anti-lymphangiogenic effects in primary human lymphatic endothelial cells by activating intrinsic apoptotic pathway and cell cycle arrest via p21-dependent pathways.

The online version of this article (doi:10.1186/s12885-016-2807-y) contains supplementary material, which is available to authorized users.

• G0/G1 cell cycle arrest
• Histon deacetylase inhibitors (HDACi)
• Lymphangiogenesis
• intrinsic apoptotic pathway
• p21

In previous papers, we presented data on studies on the anticancer activity of the vitamin D₃ analogs, named PRI-2191 and PRI-2205, in different cancer models. In this study, we showed the improved antiproliferative activity of a combination of imatinib mesylate (Gleevec, GV) and cytostatic agents in in vitro studies, when used with a third compound, namely PRI-2191, in an A549 human lung cancer model. Furthermore, we analyzed the influence of both PRI-2191, as well as PRI-2205 on the anticancer activity of GV in mice bearing A549 tumors. The route of PRI-2191 analog administration showed a significant impact on the outcome of GV treatment: subcutaneous injection was more efficient and less toxic than oral gavage. Moreover, both vitamin D compounds increased the anticancer activity of GV; however, they might also potentiate some adverse effects. We also evaluated in tumor tissue the expression of VEGF, PDGF-BB, vitamin D receptor, CYP27B1, CYP24, p53 and Bcl-2, as well as PDGF receptors: α and β. We observed the upregulation of p53 expression and the downregulation of Bcl-2, as well as VEGF in A549 tumors as a result of the tested treatment. However, vitamin D analogs did not significantly influence the expression of these proteins.

• NSCLC
• imatinib
• vitamin D analogs

• Antineoplastic Agents/pharmacology
• Antineoplastic Agents/therapeutic use
• Apoptosis/drug effects
• Cell Line, Tumor
• Cell Proliferation/drug effects
• Endoplasmic Reticulum/drug effects
• Humans
• Lung Neoplasms/drug therapy
• M Phase Cell Cycle Checkpoints/drug effects
• Mitochondria/drug effects
• Quinazolines/pharmacology
• Quinazolines/therapeutic use
• Small Cell Lung Carcinoma/drug therapy

• Anticancer therapy
• Bcl-2 family
• Histone deacetylase
• Proteasome inhibition
• Synergism

• SET1
• Sp1
• Sp3
• p300

In the present study, we evaluated the antitumor effect of two synthetic analogs of vitamin D, namely PRI-2191 [(24R)-1,24-dihydroxyvitamin D₃] and PRI-2205 (5,6-trans calcipotriol), in combined human colon HT-29 cancer treatment with 5-fluorouracil (5-FU). Mice bearing HT-29 tumors transplanted subcutaneously or orthotopically were injected with vitamin D analogs and 5-FU in various schedules. A statistically significant inhibition of subcutaneous or orthotopic tumor growth was observed as a result of combined therapy. In HT-29 tumors and in cells from in vitro culture, we observed increased vitamin D receptor (VDR) expression after treatment with either PRI-2205 or 5-FU alone, or in combination. Moreover, PRI-2205 decreased the percentage of cells from intestinal tumors in G₂/M and S stages and increased sub-G₁. Increased VDR expression was also observed after combined treatment of mice with 5-FU and PRI-2191. Moreover, our docking studies showed that PRI-2205 has stronger affinity for VDR, DBP and CAR/RXR ligand binding domains than PRI-2191. PRI-2191 analog, used with 5-FU, increased the percentage of subcutaneous tumor cells in G₀/G₁ and decreased the percentage in G₂/M, S and sub-G₁ populations as compared to 5-FU alone. In in vitro studies, we observed increased expression of p21 and p-ERK1/2 diminution via use of both analogs as compared to use of 5-FU alone. Simultaneously, PRI-2191 antagonizes some pro-apoptotic activities of 5-FU in vitro. However, in spite of these disadvantageous effects in terms of apoptosis, the therapeutic effect expressed as tumor growth retardation by PRI-2191 is significant. Our results suggest that the mechanism of potentiation of 5-FU antitumor action by both analogs is realized via increased p21 expression and decreased p-ERK1/2 level which may lead to diminution of thymidylate synthase expression. Higher binding affinity for VDR, DBP, but also for CAR\RXR ligand binding domain of PRI-2205 may, in part, explain its very low toxicity with sustained anticancer activity.

Histone deacetylase inhibitors (HDACi) are promising antineoplastic agents, but their precise mechanisms of actions are not well understood. In particular, the relevance of p53 for HDACi-induced effects has not been fully elucidated. We investigated the anticancer effects of four structurally distinct HDACi, vorinostat, entinostat, apicidin and valproic acid, using isogenic HCT-116 colon cancer cell lines differing in p53 status.

Effects were assessed by MTT assay, flow-cytometric analyses of propidium iodide uptake, mitochondrial depolarisation and cell-cycle distribution, as well as by gene expression profiling.

Vorinostat was equally effective in p53 wild-type and null cells, whereas entinostat was less effective in p53 null cells. Histone deacetylase inhibitors treatment suppressed the expression of MDM2 and increased the abundance of p53. Combination treatments showed that vorinostat enhanced the cytotoxic activity of TRAIL and bortezomib, independent of the cellular p53 status. Investigations into the effects of an inhibitor of the sirtuin class of HDAC, tenovin-1, revealed that tenovin-1-mediated cell death hinged on p53.

These results demonstrate that vorinostat activates p53, but does not require p53 for inducing its anticancer action. Yet they also demonstrate that entinostat-induced cytotoxic effects partially depend on p53, indicating that different HDACi have a different requirement for p53.

The aim of this study was to explore the level of apoptosis and p53 expression in the placental villi of patients with unexplained recurrent spontaneous abortion (URSA). Fifty-three pregnant females with URSA and 32 pregnant females who required an induced abortion were selected as the subjects of this study. Placental villus tissues were collected from June 2010 to June 2012 and quantitative polymerase chain reaction (qPCR) and immunohistochemical analysis were performed to determine the mRNA and protein levels of p53 in the placental villus tissues. The level of apoptosis in the tissues was studied using terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling (TUNEL) assay. The mRNA and protein expression levels of p53 in the URSA group were significantly higher than those in the control group (P<0.05). Furthermore, the levels of apoptosis were increased markedly in the URSA group compared with the control group (P<0.05). In conclusion, the placental villi of patients with URSA express a high level of p53, which may result in cell apoptosis and lead to recurrent spontaneous abortion.

• apoptosis
• p53
• unexplained recurrent spontaneous abortion

• ATP5O
• Acetylation modification
• Gastric cancer
• Mass spectrometry
• PKM2
• Trichostatin A

• Acetylation
• Antineoplastic Agents/pharmacology
• Apoptosis/drug effects
• Cell Cycle Checkpoints/drug effects
• Cell Line, Tumor
• Cell Proliferation/drug effects
• Dose-Response Relationship, Drug
• Histone Deacetylase Inhibitors/pharmacology
• Humans
• Hydroxamic Acids/pharmacology
• Lysine
• Neoplasm Proteins/genetics
• Neoplasm Proteins/metabolism
• Protein Processing, Post-Translational/drug effects
• Signal Transduction/drug effects
• Stomach Neoplasms/genetics
• Stomach Neoplasms/metabolism
• Stomach Neoplasms/pathology
• Time Factors

Small cell lung cancer (SCLC) accounts for nearly 15% of human lung cancers and is one of the most aggressive solid tumors. The SCLC cells are thought to derive from self-renewing pulmonary neuroendocrine cells by oncogenic transformation. However, whether the SCLC cells possess stemness and plasticity for differentiation as normal stem cells has not been well understood thus far. In this study, we investigated the expressions of multilineage stem cell markers in the cancer cells of SCLC cell line (NCI-H446) and analyzed their clonogenicity, tumorigenicity, and plasticity for inducing differentiation. It has been found that most cancer cells of the cell line expressed multilineage stem cell markers under the routine culture conditions and generated single-cell clones in anchorage-dependent or -independent conditions. These cancer cells could form subcutaneous xenograft tumors and orthotopic lung xenograft tumors in BALB/C-nude mice. Most cells in xenograft tumors expressed stem cell markers and proliferation cell nuclear antigen Ki67, suggesting that these cancer cells remained stemness and highly proliferative ability in vivo. Intriguingly, the cancer cells could be induced to differentiate into neurons, adipocytes, and osteocytes, respectively, in vitro. During the processes of cellular phenotype-conversions, autophagy and apoptosis were two main metabolic events. There is cross-talking between autophagy and apoptosis in the differentiated cancer cells. In addition, the effects of the inhibitor and agonist for Sirtuin1/2 on the inducing osteogenic differentiation indicated that Sirtuin1/2 had an important role in this process. Taken together, these results indicate that most cancer cells of NCI-H446 cell line possess stemness and plasticity for multilineage differentiation. These findings have potentially some translational applications in treatments of SCLC with inducing differentiation therapy.