Somy Yoon

HDAC and HDAC Inhibitor: From Cancer to Cardiovascular Diseases

Histone deacetylases (HDACs) are epigenetic regulators that regulate the histone tail, chromatin conformation, protein-DNA interaction, and even transcription. HDACs are also post-transcriptional modifiers that regulate the protein acetylation implicated in several pathophysiologic states. HDAC inhibitors have been highlighted as a novel category of anti-cancer drugs. To date, four HDAC inhibitors, Vorinostat, Romidepsin, Panobinostat, and Belinostat, have been approved by the United States Food and Drug Administration. Principally, these HDAC inhibitors are used for hematologic cancers in clinic with less severe side effects. Clinical trials are continuously expanding to address other types of cancer and also nonmalignant diseases. HDAC inhibition also results in beneficial outcomes in various types of neurodegenerative diseases, inflammation disorders, and cardiovascular diseases. In this review, we will briefly discuss 1) the roles of HDACs in the acquisition of a cancers phenotype and the general outcome of the HDAC inhibitors in cancer, 2) the functional relevance of HDACs in cardiovascular diseases and the possible therapeutic implications of HDAC inhibitors in cardiovascular disease.

Proteome response to ochratoxin A-induced apoptotic cell death in mouse hippocampal HT22 cells.

Mycotoxins are commonly encountered natural products, and are capable of poisoning animals or humans that inhale mold particles from mycotoxin-contaminated foods. Ochratoxin A (OTA) is produced by Aspergillu ochracus and Penicillium verrucosum, and is often found in cereals and agricultural products. Although previous studies have focused on the potent nephrotoxicity and renal carcinogenicity of OTA, more recent studies suggest that it accumulates in the brain and causes oxidative stress and DNA damage in various brain regions and neuronal populations. In the present study, we undertook to investigate the potential harm caused by environmental exposure to OTA in terms of its effects on neuronal cell viability and proteome profiles. OTA was found to significantly reduce the viabilities of human neuroblastoma SH-SY5Y and mouse hippocampal HT22 cells, as assessed by lactic dehydrogenase release into culture media. Generation of reactive oxygen species was detected in OTA-treated SH-SY5Y and HT22 cells, however, caspase activation and increase in p53 phosphorylation were only detected in HT22 cells, and the expressions of several proteins were found to be significantly altered after treating HT22 cells with OTA. Valosin containing protein, prolyl 4-hydroxylase, Atp5b protein, nucleophosmin 1, eukaryotic translation elongation factor 1 delta isoform, ornithine aminotransferase, prohibitin, and peroxiredoxin 6, which have been suggested to be implicated in the pathogenesis of neurodegenerative disorders, were up-regulated. Our findings suggest that coordinated regulations of molecular networks are involved in the OTA-induced cytotoxicity and that proteome response can be an indicative for neurodegeneration.

Lichen Secondary Metabolites in Flavocetraria cucullata Exhibit Anti-Cancer Effects on Human Cancer Cells through the Induction of Apoptosis and Suppression of Tumorigenic Potentials

Lichens are symbiotic organisms which produce distinct secondary metabolic products. In the present study, we tested the cytotoxic activity of 17 lichen species against several human cancer cells and further investigated the molecular mechanisms underlying their anti-cancer activity. We found that among 17 lichens species, F. cucullata exhibited the most potent cytotoxicity in several human cancer cells. High performance liquid chromatography analysis revealed that the acetone extract of F. cucullata contains usnic acid, salazinic acid, Squamatic acid, Baeomycesic acid, d-protolichesterinic acid, and lichesterinic acid as subcomponents. MTT assay showed that cancer cell lines were more vulnerable to the cytotoxic effects of the extract than non-cancer cell lines. Furthermore, among the identified subcomponents, usnic acid treatment had a similar cytotoxic effect on cancer cell lines but with lower potency than the extract. At a lethal dose, treatment with the extract or with usnic acid greatly increased the apoptotic cell population and specifically activated the apoptotic signaling pathway; however, using sub-lethal doses, extract and usnic acid treatment decreased cancer cell motility and inhibited in vitro and in vivo tumorigenic potentials. In these cells, we observed significantly reduced levels of epithelial-mesenchymal transition (EMT) markers and phosphor-Akt, while phosphor-c-Jun and phosphor-ERK1/2 levels were only marginally affected. Overall, the anti-cancer activity of the extract is more potent than that of usnic acid alone. Taken together, F. cucullata and its subcomponent, usnic acid together with additional component, exert anti-cancer effects on human cancer cells through the induction of apoptosis and the inhibition of EMT.

KITENIN-targeting MicroRNA-124 Suppresses Colorectal Cancer Cell Motility and Tumorigenesis

MicroRNAs are increasingly implicated in the modulation of the progression of various cancers. We previously observed that KAI1 C-terminal interacting tetraspanin (KITENIN) is highly expressed in sporadic human colorectal cancer (CRC) tissues and hence the functional KITENIN complex acts to promote progression of CRC. However, it remains unknown that microRNAs target KITENIN and whether KITENIN-targeting microRNAs modulate CRC cell motility and colorectal tumorigenesis. Here, through bioinformatic analyses and functional studies, we showed that miR-124, miR-27a, and miR-30b negatively regulate KITENIN expression and suppress the migration and invasion of several CRC cell lines via modulation of KITENIN expression. Through in vitro and in vivo induction of mature microRNAs using a tetracycline-inducible system, miR-124 was found to effectively inhibit the invasion of CT-26 colon adenocarcinoma cells and tumor growth in a syngeneic mouse xenograft model. Constitutive overexpression of precursor miR-124 in CT-26 cells suppressed in vivo tumorigenicity and resulted in decreased expression of KITENIN as well as that of MYH9 and SOX9, which are targets of miR-124. Thus, our findings identify that KITENIN-targeting miR-124, miR-27a, and miR-30b function as endogenous inhibitors of CRC cell motility and demonstrate that miR-124 among KITENIN-targeting microRNAs plays a suppressor role in colorectal tumorigenesis.

An Unconventional KITENIN/ErbB4-Mediated Downstream Signal of EGF Upregulates c-Jun and the Invasiveness of Colorectal Cancer Cells

Purpose: EGF-stimulated signaling via EGF receptor (EGFR) is important in colorectal tumorigenesis and drug targeting. However, anti-EGFR therapy is not effective in a subset of patients with colorectal cancer, suggesting that unidentified EGF-stimulated pathways might play roles in colorectal cancer. Previously, we identified KAI1 C-terminal interacting tetraspanin (KITENIN) as a metastasis-enhancing gene and found it to be highly expressed in sporadic colorectal cancer tissues. We recently found that EGF further increases KITENIN-induced elevated AP-1 activity. Here we attempted to clarify this novel EGF-stimulated molecular pathway and its roles in colorectal cancer. Experimental Design: We analyzed how EGF modulates the downstream signaling pathway of oncogenic KITENIN in colorectal cancer cells. Biological alterations following EGF treatment were identified in KITENIN-overexpressed colorectal cancer cells with or without alteration of EGFR activity. Results: We identified the KITENIN/ErbB4–Dvl2–c-Jun axis as a novel downstream signal of EGF that is switched on under elevated KITENIN conditions in an EGFR-independent manner. This unconventional EGF signal upregulates c-Jun and enhances invasion and anchorage-independent growth of colorectal cancer cells. In addition, tumor tissues from metastatic patients with colorectal cancer who showed initial poor responses to cetuximab/chemotherapy expressed higher levels of KITENIN than did responders to therapy. Conclusions: Our results highlight the role of an EGFR-independent EGF signal in mediating the invasiveness and tumorigenesis of colorectal cancer cells. This unconventional pathway might be related to the limited clinical efficacy of anti-EGFR agents in a subset of patients with colorectal cancer. Clin Cancer Res; 20(15); 4115–28. ©2014 AACR.

MDM2 E3 ligase-mediated ubiquitination and degradation of HDAC1 in vascular calcification.

Vascular calcification (VC) is often associated with cardiovascular and metabolic diseases. However, the molecular mechanisms linking VC to these diseases have yet to be elucidated. Here we report that MDM2-induced ubiquitination of histone deacetylase 1 (HDAC1) mediates VC. Loss of HDAC1 activity via either chemical inhibitor or genetic ablation enhances VC. HDAC1 protein, but not mRNA, is reduced in cell and animal calcification models and in human calcified coronary artery. Under calcification-inducing conditions, proteasomal degradation of HDAC1 precedes VC and it is mediated by MDM2 E3 ubiquitin ligase that initiates HDAC1 K74 ubiquitination. Overexpression of MDM2 enhances VC, whereas loss of MDM2 blunts it. Decoy peptide spanning HDAC1 K74 and RG 7112, an MDM2 inhibitor, prevent VC in vivo and in vitro. These results uncover a previously unappreciated ubiquitination pathway and suggest MDM2-mediated HDAC1 ubiquitination as a new therapeutic target in VC.

Elevated Coexpression of KITENIN and the ErbB4 CYT-2 Isoform Promotes the Transition from Colon Adenoma to Carcinoma Following APC loss

Purpose and Experimental Design: The molecular events in the malignant progression of colon adenoma after loss of adenomatous polyposis coli (APC) are not fully understood. KITENIN (KAI1 C-terminal interacting tetraspanin) increases the invasiveness of colorectal cancer cells, and we identified a novel EGFR-independent oncogenic signal of EGF that works under coexpressed KITENIN and ErbB4. Here we tested whether elevated KITENIN and ErbB4 contribute to further progression of intestinal adenoma following APC loss. Results: The intestinal tissues of villin-KITENIN transgenic mice in which villin-driven KITENIN expression induces increased c-Jun expression exhibit mild epithelial cell proliferation but no epithelial lineage changes compared with those of nontransgenic mice. Among the four ErbB4 isoforms, JM-a/CYT-2 and JM-b/CYT-2 exhibited the highest AP-1 activity when cells coexpressing KITENIN and each isoform were stimulated by EGF. Interestingly, predominant overexpression of the ErB4-CYT-2 mRNA as well as increased EGFR expression were observed in intestinal adenoma of APCmin/+ mice, which makes the microenvironment of activated EGF signaling. When we crossed villin-KITENIN mice with APCmin/+ mice, intestinal tumor tissues in the crossed mice showed the characteristics of early-stage invading adenocarcinoma. In patients with colorectal cancer, ErbB4-CYT-2 mRNA expression was significantly greater in tumor tissues than in normal adjacent tissues, but no significant differences in tumor tissue expression were found between different colorectal cancer stages. Furthermore, the mRNA expression of KITENIN and that of ErbB4-CYT-2 were positively correlated in human colorectal cancer tissue. Conclusions: Elevated coexpression of KITENIN and ErbB4-CYT-2 promotes the transition of colon adenoma to adenocarcinoma within an APC loss–associated tumor microenvironment. Clin Cancer Res; 22(5); 1284–94. ©2015 AACR.

Glycoprotein 90K Promotes E-Cadherin Degradation in a Cell Density-Dependent Manner via Dissociation of E-Cadherin–p120-Catenin Complex

Glycoprotein 90K (also known as LGALS3BP or Mac-2BP) is a tumor-associated protein, and high 90K levels are associated with poor prognosis in some cancers. To clarify the role of 90K as an indicator for poor prognosis and metastasis in epithelial cancers, the present study investigated the effect of 90K on an adherens junctional protein, E-cadherin, which is frequently absent or downregulated in human epithelial cancers. Treatment of certain cancer cells with 90K significantly reduced E-cadherin levels in a cell-population-dependent manner, and these cells showed decreases in cell adhesion and increases in invasive cell motility. Mechanistically, 90K-induced E-cadherin downregulation occurred via ubiquitination-mediated proteasomal degradation. 90K interacted with the E-cadherin–p120-catenin complex and induced its dissociation, altering the phosphorylation status of p120-catenin, whereas it did not associate with β-catenin. In subconfluent cells, 90K decreased membrane-localized p120-catenin and the membrane fraction of the p120-catenin. Particularly, 90K-induced E-cadherin downregulation was diminished in p120-catenin knocked-down cells. Taken together, 90K upregulation promotes the dissociation of the E-cadherin–p120-catenin complex, leading to E-cadherin proteasomal degradation, and thereby destabilizing adherens junctions in less confluent tumor cells. Our results provide a potential mechanism to explain the poor prognosis of cancer patients with high serum 90K levels.

Novel NTRK1 mutations associated with congenital insensitivity to pain with anhidrosis verified by functional studies: Nam

Congenital insensitivity to pain with anhidrosis (CIPA), also known as hereditary sensory and autonomic neuropathy type IV, features loss of pain sensation, decreased or absent sweating (anhidrosis), recurrent episodes of unexplained fever, self‐mutilating behavior, and variable mental retardation. Mutations in neurotrophic receptor tyrosine kinase 1 (NTRK1) have been reported to be associated with CIPA. We identified four novel NTRK1 mutations in six Korean patients from four unrelated families. Of the four mutations, we demonstrated using a splicing assay that IVS14+3A>T causes aberrant splicing of NTRK1 mRNA, leading to introduction of a premature termination codon. An NTRK1 autophosphorylation assay showed that c.1786G>A (p.Asp596Asn) abolished autophosphorylation of NTRK1. In addition, Western blotting showed that c.704C>G (p.Ser235*) and c.2350_2363del (p.Leu784Serfs*79) blunted NTRK1 expression to undetectable levels. The four novel NTRK1 mutations we report here will expand the repertoire of NTRK1 mutations in CIPA patients, and further our understanding of CIPA pathogenesis.

Usefulness of Age-Stratified N-Terminal Prohormone of Brain Natriuretic Peptide for Diagnosing Kawasaki Disease

N-terminal prohormone of brain natriuretic peptide (NT-proBNP) was recently reported as a biomarker for diagnosing Kawasaki disease (KD). The basal NT-proBNP level, however, gradually decreases with age. We investigated the usefulness of an age-stratified cutoff value of NT-proBNP for diagnosing KD. All the patients enrolled in this study visited Chonnam National University Hospital between December 2007 and March 2016. The KD groups consisted of 214 patients with complete KD and 129 patients with incomplete KD. The control group included 62 children with simple febrile illness but without heart disease. Laboratory data including NT-proBNP level were evaluated. Each group was divided into subgroups according to patient age (<6 months, 6–12 months, 12–24 months, and >24 months), and different cutoff values of NT-proBNP were calculated. The cutoff values of NT-proBNP used to diagnose total KD and incomplete KD were 762 and 762 pg/mL (<6 months), 310 and 310 pg/mL (6–12 months), 326 and 326 pg/mL (12–24 months), and 208 and 137 pg/mL (>24 months), respectively. In conclusion, age-stratified NT-proBNP is a useful biomarker for the differential diagnosis of KD in patients with a simple febrile illness.