Evaluation and molecular docking study of two flavonoids from Oroxylum indicum (L.) Kurz and their semi-synthetic derivatives as histone deacetylase inhibitors.

Chrysin (5,7-dihydroxyflavone, 6) and galangin 3-methyl ether (5,7-dihydroxy-3-methoxy flavone, 7) were obtained from the leaves of Oroxylum indicum (L.) Kurz in 4% and 6% yields, respectively. Both compounds could act as pan-histone deacetylase (HDAC) inhibitors. Structural modification of these lead compounds provided thirty-eight derivatives which were further tested as HDAC inhibitors. Compounds 6b, 6c, and 6q were the most potent derivatives with the IC50 values of 97.29 ± 0.63 µM, 91.71 ± 0.27 µM, and 96.87 ± 0.45 µM, respectively. Molecular docking study indicated the selectivity of these three compounds toward HDAC8 and the test against HDAC8 showed IC50 values in the same micromolar range. All three compounds were further evaluated for the anti-proliferative activity against HeLa and A549 cell lines. Compound 6q exhibited the best activity against HeLa cell line with the IC50 value of 13.91 ± 0.34 µM. Moreover, 6q was able to increase the acetylation level of histone H3. These promising HDAC inhibitors deserve investigation as chemotherapeutic agents for treating cancer.

Molecular mechanism and therapeutic potential of HDAC9 in intervertebral disc degeneration.

BACKGROUND: Intervertebral disc degeneration (IVDD) is the major cause of low-back pain. Histone deacetylase 9 (HDAC9) was dramatically decreased in the degenerative nucleus pulposus (NP) samples of patients with intervertebral disc degeneration (IVDD) according to bioinformatics analysis of Gene Expression Omnibus (GEO) GSE56081 dataset. This study aims to investigate the role of HDAC9 in IVDD progression. METHODS: The contribution of HDAC9 to the progression of IVDD was assessed using HDAC9 knockout (HDAC9KO) mice and NP-targeted HDAC9-overexpressing mice by IVD injection of adenovirus-mediated HDAC9 under a Col2a1 promoter. Magnetic resonance imaging (MRI) and histological analysis were used to examine the degeneration of IVD. NP cells were isolated from mice to investigate the effects of HDAC9 on apoptosis and viability. mRNA-seq and coimmunoprecipitation/mass spectrometry (co-IP/MS) analysis were used to analyze the HDAC9-regulated factors in the primary cultured NP cells. RESULTS: HDAC9 was statistically decreased in the NP tissues in aged mice. HDAC9KO mice spontaneously developed age-related IVDD compared with wild-type (HDAC9WT) mice. In addition, overexpression of HDAC9 in NP cells alleviated IVDD symptoms in a surgically-induced IVDD mouse model. In an in vitro assay, knockdown of HDAC9 inhibited cell viability and promoted cell apoptosis of NP cells, and HDAC9 overexpression had the opposite effects in NP cells isolated from HDAC9KO mice. Results of mRNA-seq and co-IP/MS analysis revealed the possible proteins and signaling pathways regulated by HDAC9 in NP cells. RUNX family transcription factor 3 (RUNX3) was screened out for further study, and RUNX3 was found to be deacetylated and stabilized by HDAC9. Knockdown of RUNX3 restored the effects of HDAC9 silencing on NP cells by inhibiting apoptosis and increasing viability. CONCLUSION: Our results suggest that HDAC9 plays an important role in the development and progression of IVDD. It might be required to protect NP cells against the loss of cell viability and apoptosis by inhibiting RUNX3 acetylation and expression during IVDD. Together, our findings suggest that HDAC9 may be a potential therapeutic target in IVDD.

NFYB increases chemosensitivity in glioblastoma by promoting HDAC5-mediated transcriptional inhibition of SHMT2.

Temozolomide (TMZ) is a commonly used chemotherapeutic agent for glioblastoma (GBM), but acquired drug resistance prevents its therapeutic efficacy. We investigated potential mechanisms underlying TMZ resistance and glycolysis in GBM cells through regulation by nuclear transcription factor Y subunit ß (NFYB) of the oncogene serine hydroxymethyltransferase 2 (SHMT2). GBM U251 cells were transfected with NFYB-, SHMT2-, and the potential NFYB target histone deacetylase 5 (HDAC5)-related vectors. Glucose uptake and lactate production were measured with detection kits. CCK-8/colony formation, scratch, Transwell, and flow cytometry assays were performed to detect cell proliferation, migration, invasion, and apoptosis, respectively. The binding of NFYB to the HDAC5 promoter and the regulation of NFYB on HDAC5 promoter activity were detected with chromatin immunoprecipitation and dual-luciferase reporter assays, respectively. NFYB and HDAC5 were poorly expressed and SHMT2 was expressed at high levels in GBM U251 cells. NFYB overexpression or SHMT2 knockdown decreased glucose uptake, lactate production, proliferation, migration, and invasion and increased apoptosis and TMZ sensitivity of the cells. NFYB activated HDAC5 to inhibit SHMT2 expression. SHMT2 overexpression nullified the inhibitory effects of NFYB overexpression on glycolysis and TMZ resistance. Thus, NFYB may reduce tumorigenicity and TMZ resistance of GBM through effects on the HDAC5/SHMT2 axis.

Harnessing pyrimidine as a building block for histone deacetylase inhibitors.

Histone deacetylase (HDAC) inhibitors are well-established multifaceted bioactive agents against tumors and neurodegenerative disorders. Pyrimidine and its fused and substituted derivatives were employed as a surface recognition moiety of HDAC inhibitors. De facto, the literature was loaded with different success stories of pyrimidine-based HDAC inhibitors that garnered much interest. Provoked by our continuous interest in HDAC inhibitors, we summarized and elaborated on the successful harnessing of the pyrimidine scaffold in this regard. Furthermore, we dissect our perspective that may guide medicinal chemists for an effective future design of more active chemotherapeutic agents with potential clinical applications.

Pharmacological inhibition of histone deacetylases ameliorates cognitive impairment after intracerebral hemorrhage with epigenetic alteration in the hippocampus.

OBJECTIVES: Post-stroke cognitive impairment (PSCI) interferes with neurorehabilitation in patients with stroke. Epigenetic regulation of the hippocampus has been targeted to ameliorate cognitive function. In particular, the acetylation level of histones is modulated by exercise, a potent therapy for patients with stroke. MATERIALS AND METHODS: We examined the effects of exercise and pharmacological inhibition of histone deacetylase (HDAC) using sodium butyrate (NaB) on cognitive function and epigenetic factors in the hippocampus after intracerebral hemorrhage (ICH) to seek beneficial neuronal conditioning against PSCI. Forty rats were randomly assigned to five groups: sham, control, NaB, exercise, and NaB plus exercise groups (n=8 in each group). Except for those in the sham group, all rats underwent stereotaxic ICH surgery with a microinjection of collagenase solution. Intraperitoneal administration of NaB (300 mg/kg) and treadmill exercise (11 m/min for 30 min) were conducted for approximately 4 weeks starting 3 days post-surgery. RESULTS: ICH reduced cognitive function, as detected by the object location test, accompanied by enhanced activity of HDACs. Although exercise did not modulate HDAC activity or cognitive function, repetitive NaB administration increased HDAC activity and ameliorated cognitive impairment induced by ICH. CONCLUSIONS: This study suggests that pharmacological treatment with an HDAC inhibitor could potentially present an enriched epigenetic platform in the hippocampus and ameliorate PSCI for neurorehabilitation following ICH.

Melatonin-vorinostat hybrid ligands show higher histone deacetylase and cancer cell growth inhibition than vorinostat.

Anticancer drug conjugates are an emerging approach for future cancer treatment. Here, we report a series of hybrid ligands merging the neurohormone melatonin with the approved histone deacetylase (HDAC) inhibitor vorinostat, using melatonin's amide side chain (3a-e), its indolic nitrogen (5a-d), and its ether oxygen (7a-d) as attachment points. Several hybrid ligands showed higher potency thanvorinostat in both HDAC inhibition and cellular assays on different cultured cancer cell lines. In the most potent HDAC1 and HDAC6 inhibitors, 3e, 5c, and 7c, the hydroxamic acid moiety of vorinostat is linked to melatonin through a hexamethylene spacer. Hybrid ligands 5c and 7c were also found to be potent growth inhibitors of MCF-7, PC-3M-Luc, and HL-60 cancer cell lines. As these compounds showed only weak agonist activity at melatonin MT1 receptors, the findings indicate that their anticancer actions are driven by HDAC inhibition.

Retinoic acid and evernyl-based menadione-triazole hybrid cooperate to induce differentiation of neuroblastoma cells.

Neuroblastoma arises when immature neural precursor cells do not mature into specialized cells. Although retinoic acid (RA), a pro-differentiation agent, improves the survival of low-grade neuroblastoma, resistance to retinoic acid is found in high-grade neuroblastoma patients. Histone deacetylases (HDAC) inhibitors induce differentiation and arrest the growth of cancer cells; however, HDAC inhibitors are FDA-approved mostly for liquid tumors. Therefore, combining histone deacetylase (HDAC) inhibitors and retinoic acid can be explored as a strategy to trigger the differentiation of neuroblastoma cells and to overcome resistance to retinoic acid. Based on this rationale, in this study, we linked evernyl group and menadione-triazole motifs to synthesize evernyl-based menadione-triazole hybrids and asked if the hybrids cooperate with retinoic acid to trigger the differentiation of neuroblastoma cells. To answer this question, we treated neuroblastoma cells using evernyl-based menadione-triazole hybrids (6a-6i) or RA or both and examined the differentiation of neuroblastoma cells. Among the hybrids, we found that compound 6b inhibits class-I HDAC activity, induces differentiation, and RA co-treatments increase 6b-induced differentiation of neuroblastoma cells. In addition, 6b reduces cell proliferation, induces expression of differentiation-specific microRNAs leading to N-Myc downregulation, and RA co-treatments enhance the 6b-induced effects. We observed that 6b and RA trigger a switch from glycolysis to oxidative phosphorylation, maintain mitochondrial polarization, and increase oxygen consumption rate. We conclude that in evernyl-based menadione-triazole hybrid, 6b cooperates with RA to induce differentiation of neuroblastoma cells. Based on our results, we suggest that combining RA and 6b can be pursued as therapy for neuroblastoma. Schematic representation of RA and 6b in inducing differentiation of neuroblastoma cells.

Astragaloside IV protects against oxidized low-density lipoprotein-induced injury in human umbilical vein endothelial cells via the histone deacetylase 9 (HDAC9)/NF-κB axis.

BACKGROUND: Atherosclerosis is a main cause of multiple cardiovascular diseases, and cell damage of human umbilical vein endothelial cells (HUVECs) was reported to participate in the development of atherosclerosis. In this study, we aimed to study the action of Astragaloside IV (ASV) on AS development using in vitro AS cell model. METHODS: MTT assay, EdU staining assay, and flow cytometry were utilized for detection of cell proliferation and apoptosis, respectively. The protein expression of histone deacetylase 9 (HDAC9), Bax, Bcl-2, p-P65, P65, p-IκBα, and IκBα was gaged using western blot. The angiogenesis was evaluated by tube formation assay. The inflammatory response was evaluated by ELISA kits. SOD activity and MDA level were detected using the matched commercial kits. RT-qPCR was used for HDAC9 mRNA expression measurement. RESULTS: Oxidized low-density lipoprotein (ox-LDL) significantly repressed cell proliferation, angiogenesis, and enhanced apoptosis, inflammation, and oxidative stress in HUVECs. ASV addition could alleviate ox-LDL-caused cell damage in HUVECs. Moreover, HDAC9 was overexpressed in AS patients and AS cell model. Functionally, HDAC9 knockdown also exhibited the protective role in ox-LDL-treated HUVECs. In addition, ASV treatment protected against ox-LDL-induced damage in HUVECs via targeting HDAC9. ASV could inactivate the NF-κB pathway via regulating HDAC9 in AS cell model. CONCLUSION: ASV exerted the protective effects on ox-LDL-induced damage in HUVECs through the HDAC9/NF-κB axis.

Ginsenoside Rg3 inhibits renal cell carcinoma cell migration, invasion, colony formation, and tube formation and enhances apoptosis through promoting the DNA demethylation and histone acetylation.

OBJECTIVES: This study explored the effect and mechanism of Rg3 on renal cell carcinoma (RCC) progression. METHODS: RCC cells were treated with different concentrations of Rg3, 5-Aza-dc (a methyltransferase inhibitor) or TSA (a deacetylase inhibitor). Rg3-induced cytotoxicity, migration, invasion, colony formation, tube formation and apoptosis of RCC cells were evaluated by CCK-8, wound healing, Transwell, colony formation, tube formation and flow cytometry assays, respectively. Methylation and expressions of p53, p21 and p16, and expressions of methylation-related genes and histone deacetylases and histone acetylation-related genes (H3 (acetyl K14), H3 (acetyl K9), H4 (acetyl K12), H4 (acetyl K5) and H4 (acetyl K16)) were analysed by qRT-PCR and western blot. KEY FINDINGS: Rg3 dose-dependently decreased the viability, inhibited migration, invasion, colony formation and tube formation, and enhanced apoptosis of RCC cells. Rg3 enhanced the demethylation levels and expressions of p53, p21 and p16 as well as the expressions of histone acetylation-related genes, but repressed the expressions of methylation-related genes and histone deacetylases. Rg3 had the same effect as 5-Aza-dc and TSA did on the above-mentioned cellular changes. CONCLUSION: Rg3 restrains RCC cell migration, invasion, colony formation and tube formation, yet enhances apoptosis through promoting demethylation of p53, p21 and p16, and histone acetylation.

Histone-deacetylase-inhibitory effects of periodontopathic-bacterial metabolites induce human gingival epithelial Ca9-22 cell death.

Dental plaque bacteria produce high concentrations of short-chain fatty acids (SCFAs), as bacterial metabolites. SCFA-treated gingival epithelial cells undergo cell death. Our previous reports demonstrated that butyrate-induced cell death depends on autophagy and reactive oxygen species (ROS). However, the precise mechanisms underlying SCFA-induced gingival epithelial cell death is poorly understood. Butyrate is a strong histone deacetylase (HDAC) inhibitor. Therefore, we determined the involvement of HDAC inhibitory activity in SCFA-induced gingival epithelial cells. Ca9-22 cells were used as an in vitro counterpart of gingival epithelial cells. Ca9-22 cells were treated with HDAC inhibitors in the presence or absence of C646, a P300 histone acetyltransferase (HAT) inhibitor, and compared the number of dead cells, which are measured using SYTOX Green dye. Acetylation levels of histone H3 were examined using western blotting. Changes in transcriptomes during the butyrate and C646 treatment were examined using RNA sequencing analysis. The butyrate or propionate-treatment of Ca9-22 cells induced acetylation of histone H3, while the C646 treatment strongly reduced the elevated acetylation levels. Accordingly, butyrate or propionate-induced cell death was inhibited by the C646 treatment. Similar results were obtained when other HDAC inhibitors were used. Whole transcriptome analysis revealed that the expression of numerous genes was altered by butyrate-induced histone acetylation. Moreover, some autophagy and ROS-related genes found in the altered genes might induce cell death. This study suggests the need for HDAC-inhibitory activity of bacterial metabolites to induce cell death, and the effects might enhance autophagy and ROS production.

miR-3,178 contributes to the therapeutic action of baicalein against hepatocellular carcinoma cells via modulating HDAC10.

Hepatocellular carcinoma (HCC) is the most common type of hepatic malignancies with high mortality and poor prognosis. Baicalein, one of the major and bioactive flavonoids isolated from Scutellaria baicalensis Georgi, which is reported to have anti-proliferation effect in varying cancers, including HCC, whose underlying molecular mechanism is still largely unknown. In this study, we found that baicalein significantly inhibited proliferation and colony formation, blocked cell cycle, and promoted apoptosis in HCC cells MHCC-97H and SMMC-7721 in vitro and reduced tumor volume and weight in vivo. Increased microRNA (miR)-3,178 levels and decreased histone deacetylase 10 (HDAC10) expression were found in cells treated with baicalein and in patients' HCC tissues. HDAC10 was identified as a target gene of miR-3,178 by luciferase activity and western blot. Both baicalein treatment and overexpression of miR-3,178 could downregulate HDAC10 protein expression and inactivated AKT, MDM2/p53/Bcl2/Bax and FoxO3α/p27/CDK2/Cyclin E1 signal pathways. Not only that, knockdown of miR-3,178 could partly abolish the effects of baicalein and the restoration of HDAC10 could abated miR-3,178-mediated role in HCC cells. Collectively, baicalein inhibits cell viability, blocks cell cycle, and induces apoptosis in HCC cells by regulating the miR-3,178/HDAC10 pathway. This finding indicated that baicalein might be promising for treatment of HCC.

HDAC inhibitor Vorinostat and BET inhibitor Plx51107 epigenetic agents' combined treatments exert a therapeutic approach upon acute myeloid leukemia cell model.

The process of cancer initiation and development is regulated via the transcriptional expression of cells going under genomic and epigenetic changes. Targeting epigenetic "readers", i.e., bromodomains (BRD) and post-translational modifications of nucleosomal histone proteins regulate gene expression in both cancerous and healthy cells. In this study, the new epigenetic agent BRD inhibitor PLX51107 and histone deacetylase (HDAC) inhibitor SAHA' s (Vorinostat) single/combined applications' reflections were analyzed in case of cell proliferation, cytotoxicity, apoptosis, cell cycle arrest, and finally target gene expression regulation upon both AML and healthy B-lymphocyte cells; HL60 and NCIBL2171, respectively; in vitro. Since mono treatments of either Vorinostat or Plx51107 regulated cellular responses such as growth, proliferation, apoptosis, and cell cycle arrest of tumor cells; their combination treatments exerted accelerated results. We detected that combined treatment of Plx51107 and Vorinostat strengthened effects detected upon leukemic cells for gaining more sensitization to the agents, decreasing cell proliferation, dramatically inducing apoptosis, and cell cycle arrest; thus regulating target gene expressions. We have shown for the first time that the newly analyzed BRD inhibitor Plx51107 could be a promising therapeutic approach for hematological malignancies and its mono or combined usage might support a rapid transition to clinical trials.

Semaglutide ameliorates lipopolysaccharide-induced acute lung injury through inhibiting HDAC5-mediated activation of NF-κB signaling pathway.

BACKGROUND: As a life-threatening respiratory syndrome, acute lung injury (ALI) is characterized by uncontrollable inflammatory activities. Semaglutide (SEM) has been identified as an effective anti-inflammatory drug in a variety of diseases. This study intended to explore the functional effect and potential mechanisms of SEM in ALI. METHODS: Lipopolysaccharide (LPS) was used to construct an in vivo ALI model based on Sprague-Dawley (SD) rats and an in vitro ALI model based on human pulmonary artery endothelial cells (HPAECs). Hematoxylin & eosin (H&E) staining and ELISA were applied to evaluate the histopathological changes in pulmonary tissues and detect TNF-α and IL-6 levels. RT-qPCR and Western blotting were used to measure gene and protein expressions in pulmonary tissues and cells. HPAEC viability and apoptosis were evaluated by CCK-8 method and flow cytometry methods. RESULTS: Semaglutide pretreatment significantly mitigated pulmonary injury, reduced TNF-α and IL-6 production, and led to a decrease in cleaved caspase-3 level and an increase in Bcl-2 level, suggesting SEM could ameliorate LPS-induced ALI in rats. In vitro, SEM increased the proliferative capability and mitigated inflammation and apoptosis in LPS-stimulated HPAECs. In addition, SEM inhibited HDAC5-mediated NF-κB signaling pathway in HPAECs. HDAC5 overexpression or NF-κB signaling activation could partly impair SEM-mediated protective effects against LPS-induced damage to HPAECs. CONCLUSION: Semaglutide restrains LPS-induced ALI by inhibiting HDAC5/NF-κB signaling pathway.

T-17, a novel cyclin-dependent kinases/histone deacetylases dual inhibitor, induces cancer cells death through cell cycle arrest and apoptosis.

Combination of cyclin-dependent kinases (CDKs) and histone deacetylases (HDACs) inhibitors may have statistical synergy in suppressing cancer cell proliferation. Herein, a novel CDKs/HDACs dual inhibitor T-17 was rationally designed, synthesized, and evaluated. Our results demonstrated that T-17 concurrently exhibited potent and balanced inhibitory activity against CDKs (IC50 = 18.0 nM) and HDACs (IC50 = 6.6 nM) and also displayed good cell viability inhibitory effect on four cancer cell lines. Meanwhile, T-17 blocked the MDA-MB-231 and A549 cell cycle at G1 phase and S phase, respectively. In addition, T-17 induced MDA-MB-231 cells apoptosis and inhibited the HDACs and CDKs mediated signaling pathways. Finally, we also found that T-17 had good antitumor activity in vivo. In summary, these results indicated that T-17 would be a promising lead compound which deserves further research.

Dual HDAC/BRD4 Inhibitors Relieves Neuropathic Pain by Attenuating Inflammatory Response in Microglia After Spared Nerve Injury.

Despite the effort on developing new treatments, therapy for neuropathic pain is still a clinical challenge and combination therapy regimes of two or more drugs are often needed to improve efficacy. Accumulating evidence shows an altered expression and activity of histone acetylation enzymes in chronic pain conditions and restoration of these aberrant epigenetic modifications promotes pain-relieving activity. Recent studies showed a synergistic activity in neuropathic pain models by combination of histone deacetylases (HDACs) and bromodomain and extra-terminal domain (BET) inhibitors. On these premises, the present study investigated the pharmacological profile of new dual HDAC/BRD4 inhibitors, named SUM52 and SUM35, in the spared nerve injury (SNI) model in mice as innovative strategy to simultaneously inhibit HDACs and BETs. Intranasal administration of SUM52 and SUM35 attenuated thermal and mechanical hypersensitivity in the absence of locomotor side effects. Both dual inhibitors showed a preferential interaction with BRD4-BD2 domain, and SUM52 resulted the most active compound. SUM52 reduced microglia-mediated spinal neuroinflammation in spinal cord sections of SNI mice as showed by reduction of IBA1 immunostaining, inducible nitric oxide synthase (iNOS) expression, p65 nuclear factor-κB (NF-κB) and p38 MAPK over-phosphorylation. A robust decrease of the spinal proinflammatory cytokines content (IL-6, IL-1ß) was also observed after SUM52 treatment. Present results, showing the pain-relieving activity of HDAC/BRD4 dual inhibitors, indicate that the simultaneous modulation of BET and HDAC activity by a single molecule acting as multi-target agent might represent a promise for neuropathic pain relief.

Class I HDAC overexpression promotes temozolomide resistance in glioma cells by regulating RAD18 expression.

Overexpression of histone deacetylases (HDACs) in cancer commonly causes resistance to genotoxic-based therapies. Here, we report on the novel mechanism whereby overexpressed class I HDACs increase the resistance of glioblastoma cells to the SN1 methylating agent temozolomide (TMZ). The chemotherapeutic TMZ triggers the activation of the DNA damage response (DDR) in resistant glioma cells, leading to DNA lesion bypass and cellular survival. Mass spectrometry analysis revealed that the catalytic activity of class I HDACs stimulates the expression of the E3 ubiquitin ligase RAD18. Furthermore, the data showed that RAD18 is part of the O6-methylguanine-induced DDR as TMZ induces the formation of RAD18 foci at sites of DNA damage. Downregulation of RAD18 by HDAC inhibition prevented glioma cells from activating the DDR upon TMZ exposure. Lastly, RAD18 or O6-methylguanine-DNA methyltransferase (MGMT) overexpression abolished the sensitization effect of HDAC inhibition on TMZ-exposed glioma cells. Our study describes a mechanism whereby class I HDAC overexpression in glioma cells causes resistance to TMZ treatment. HDACs accomplish this by promoting the bypass of O6-methylguanine DNA lesions via enhancing RAD18 expression. It also provides a treatment option with HDAC inhibition to undermine this mechanism.

Effects of tubastatin A on adrenocorticotropic hormone synthesis and proliferation of AtT-20 corticotroph tumor cells.

Cushing's disease is an endocrine disorder characterized by hypercortisolism, mainly caused by autonomous production of ACTH from pituitary adenomas. Autonomous ACTH secretion results in excess cortisol production from the adrenal glands, and corticotroph adenoma cells disrupt the normal cortisol feedback mechanism. Pan-histone deacetylase (HDAC) inhibitors inhibit cell proliferation and ACTH production in AtT-20 corticotroph tumor cells. A selective HDAC6 inhibitor has been known to exert antitumor effects and reduce adverse effects related to the inhibition of other HDACs. The current study demonstrated that the potent and selective HDAC6 inhibitor tubastatin A has inhibitory effects on proopiomelanocortin (Pomc) and pituitary tumor-transforming gene 1 (Pttg1) mRNA expression, involved in cell proliferation. The phosphorylated Akt/Akt protein levels were increased after treatment with tubastatin A. Therefore, the proliferation of corticotroph cells may be regulated through the Akt-Pttg1 pathway. Dexamethasone treatment also decreased the Pomc mRNA level. Combined tubastatin A and dexamethasone treatment showed additive effects on the Pomc mRNA level. Thus, tubastatin A may have applications in the treatment of Cushing's disease.

4-Acyl Pyrrole Capped HDAC Inhibitors: A New Scaffold for Hybrid Inhibitors of BET Proteins and Histone Deacetylases as Antileukemia Drug Leads.

Multitarget drugs are an emerging alternative to combination therapies. In three iterative cycles of design, synthesis, and biological evaluation, we developed a novel type of potent hybrid inhibitors of bromodomain, and extra-terminal (BET) proteins and histone deacetylases (HDACs) based on the BET inhibitor XD14 and well-established HDAC inhibitors. The most promising new hybrids, 49 and 61, displayed submicromolar inhibitory activity against HDAC1-3 and 6, and BRD4(1), and possess potent antileukemia activity. 49 induced apoptosis more effectively than the combination of ricolinostat and birabresib (1:1). The most balanced dual inhibitor, 61, induced significantly more apoptosis than the related control compounds 62 (no BRD4(1) affinity) and 63 (no HDAC inhibition) as well as the 1:1 combination of both. Additionally, 61 was well tolerated in an in vivo zebrafish toxicity model. Overall, our data suggest an advantage of dual HDAC/BET inhibitors over the combination of two single targeted compounds.

The synergistic proapoptotic effect of PARP-1 and HDAC inhibition in cutaneous T-cell lymphoma is mediated via Blimp-1.

The therapy of advanced mycosis fungoides (MF) presents a therapeutic challenge, and the search for new therapeutic targets is ongoing. Poly(ADP-ribose) polymerase 1 was shown to be upregulated in patients with advanced MF and could be druggable by a new class of chemotherapeutic agents, PARP-1 inhibitors, which are already in clinical trials for other malignancies; however, the role of PARP-1 inhibitors in MF has never been established. We examined the efficacy of talazoparib in the murine model of cutaneous T-cell lymphoma. The cytotoxic effect of talazoparib on Moloney MuLV-induced T-cell lymphoma (MBL2) cells was a result of G2/M cell cycle arrest via the upregulation of p53. The in vivo experiments confirmed the clinical impact of talazoparib on MF tumors. When talazoparib was combined with the histone deacetylase (HDAC) inhibitor, romidepsin, the cytotoxic effect was synergized via downregulation of the DNA-repair genes Fanconianemia complementation group A (FANCA), Fanconi anemia complementation group D2 (FANCD2), and DNA topoisomerase II binding protein 1(TOPBP1)and stimulation of apoptosis via Blimp-1 (PRDM1)/Bax axis. Romidepsin increased the expression of IRF8 and Bcl-6, leading to upregulation of Blimp1and Bax; whereas talazoparib upregulated Blimp-1 and Bax via upregulation of interferon regulatory factor 4 (IRF4), leading to cleavage of caspases 6 and 7. Thus, a combination of talazoparib with romidepsin demonstrated the synergistic antilymphoma effect and warranted further investigation in a clinical trial.

Establishment and characterization of NCC-DFSP3-C1: a novel patient-derived dermatofibrosarcoma protuberans cell line.

Dermatofibrosarcoma protuberans (DFSP) is the most common dermal sarcoma; it is characterized by the presence of the COL1A1-PDGFB translocation, which causes the constitutive activation of the platelet-derived growth factor ß (PDGFB) signaling pathway. DFSP frequently exhibits local recurrence and is refractory to conventional chemotherapy. Therefore, a novel therapeutic strategy is required for improving the prognosis of DFSP. Although patient-derived cell lines are important tools for pre-clinical studies, currently, only a few such cell lines are available for DFSP in cell banks. Here, we report the establishment of a novel DFSP cell line. Using a surgically resected metastatic tumor tissue from a patient with DFSP, we established a cell line called NCC-DFSP3-C1. The NCC-DFSP3-C1 cells had a COL1A1-PDGFB translocation and retained the same copy number aberrations as the original tumor tissue. NCC-DFSP3-C1 cells exhibited constant growth, spheroid formation, and invasive ability. By screening a drug library, we identified anti-cancer agents with inhibitory effects on the proliferation of NCC-DFSP3-C1 cells; these anti-cancer agents included proteasomal, histone deacetylase, and kinase inhibitors. We concluded that the NCC-DFSP3-C1 cell line may serve as a useful tool for performing basic and pre-clinical studies on DFSP.