Gyoonhee Han

Histone deacetylase inhibitor KBH-A42 inhibits cytokine production in RAW 264.7 macrophage cells and in vivo endotoxemia model

In light of the anti-inflammatory properties of histone deacetylase (HDAC) inhibitors, such as suberoylanilide hydroxamic acid (SAHA) and trichostatin A (TSA), we examined a new HDAC inhibitor KBH-A42 for its anti-inflammatory activities. KBH-A42 showed noteworthy anti-inflammatory properties in vitro via suppression of the production of TNF-α, a proinflammatory cytokine, and nitric oxide (NO), a proinflammatory effector molecule, in LPS-stimulated RAW264.7 cells and peritoneal macrophages. It also inhibited TNF-α production in vivo as demonstrated in a LPS-induced mouse endotoxemia model. The levels of TNF-α, IL-1β, IL-6 and iNOS mRNAs determined by RT-PCR propose that the inhibition of these pro-inflammatory mediators by KBH-A42 resulted from inhibiting expression of these genes. However, the EMSA study to see the effect of KBH-A42 on the binding of NF-κB, a transcription factor, to a specific DNA sequence showed that the binding of NF-κB to DNA was not changed regardless of increasing the concentration of KBH-A42 in the presence and absence of LPS stimulation. Interestingly, DNA binding of another transcription factor AP-1 dose-dependently increased by KBH-A42. KBH-A42 differentially regulated the phosphorylation of MAP kinases. While the phosphprylation of ERK1/2 and SAPK/JNK was not affected by KBH-A42, the phosphorylation of p38 decreased by KBH-A42. These results showed that KBH-A42 inhibits production of proinflammatory cytokines in macrophages by decreasing their mRNA levels, and p38 kinase is involved in the KBH-A42-mediated inhibition.

RhoB induces apoptosis via direct interaction with TNFAIP1 in HeLa cells.

RhoB, a tumor suppressor, has emerged as an interesting cancer target, and extensive studies aimed at understanding its role in apoptosis have been performed. In our study, we investigated the involvement of RhoB‐interacting molecules in apoptosis. To identify RhoB‐interacting proteins, we performed yeast‐two hybrid screening assays using RhoB as a bait and isolated TNFAIP1, a TNFα‐induced protein containing the BTB/POZ domain. The interaction between RhoB and TNFAIP1 was demonstrated in vivo through coimmunoprecipitation studies and in vitro binding assays. RFP‐TNFAIP1 was found to be partially colocalized with EGFP‐RhoB. The partial colocalization of RhoB and TNFAIP1 in endosomes suggests that RhoB‐TNFAIP1 interactions may have a functional role in apoptosis. TNFAIP1 elicited proapoptotic activity, while simultaneous expression of RhoB and TNFAIP1 resulted in a dramatic increase in apoptosis in HeLa cells. Furthermore, knockdown of RhoB using siRNA clearly rescued cells from apoptosis induced by TNFAIP1. This finding suggests that interactions between RhoB and TNFAIP1 are crucial for induction of apoptosis in HeLa cells. The observation of increased SAPK/JNK phosphorylation in apoptotic cells and the finding that a JNK inhibitor suppressed apoptosis indicates that SAPK/JNK signaling may be involved in apoptosis induced by RhoB‐TNFAIP1 interactions. In conclusion, we found that RhoB interacts with TNFAIP1 to regulate apoptosis via a SAPK/JNK‐mediated signal transduction mechanism.

Inhibition of human ovarian tumor growth by cytokine-induced killer cells.

Despite the recent improvement in the treatment of ovarian cancer, this disease is still leading cause of cancer death in women. In this study, the anti-tumor activity of cytokine-induced killer (CIK) cells against human ovarian cancer was evaluatedin vitro andin vivo. Although CD3+CD56+ cells were rare in fresh human peripheral blood mononuclear cells, they could expand more than 1,000-fold on day 14 in the presence of anti-CD3 antibody plus IL-2. At an effector-target cell ratio of 30:1, CIK cells destroyed 45% of SK-OV-3 human ovarian cancer cells, which was determined by the51Cr-release assay. In addition, CIK cells at a dose of 23 million cells per mouse inhibited 73% of SK-OV-3 tumor growth in nude mouse xenograft assay. This study suggests that CIK cells may be used as an adoptive immunotherapy for patients with ovarian cancer.

Study of the stability of carbocations by chlorosulfonyl isocyanate reaction with ethers

Abstract The stability order of various alkyl, allyl, and benzyl carbocations was investigated using the novel technique for comparing the stability of carbocations in solution developed by using a simple CSI reaction with various ethers. The p -methoxycinnamyl carbocation was the most stable in our reaction system and the next stable carbocation was the p -methoxybenzyl carbocation. The stability of the other carbocations decreased with methacryl, t -butyl, cinnamyl, acryl, benzyl, 2° and allyl carbocations in that order.

Reversine Increases the Plasticity of Lineage-committed Cells toward Neuroectodermal Lineage

Functional dedifferentiation of lineage-committed cells toward pluripotency may have a great potential in regenerative medicine. Reversine has been shown to induce dedifferentiation of multiple terminally differentiated mesodermal origin cells, which are capable of being directed to differentiate into other cell types within mesodermal lineages. However, the possibilities of these cells to give rise to other lineages have not been examined. Here we show that large scale gene expression profiling of reversine-treated C2C12 myoblasts identifies a subset of up-regulated genes involved in specification of neuroectodermal as well as mesodermal lineages. Reversine treatment leads to up-regulation of priming genes of neuroectodermal lineages, such as Ngn2, Nts, Irx3, Pax7, Hes1, and Hes6, through active histone modifications in the promoter regions of these genes. Additionally, reversine increases the expression of markers for other cell types of mesodermal lineages, Ogn and apoE, via inducing active histone modifications, while down-regulating the myogenic basic helix-loop-helix factor, MyoD, via repressive histone modifications. Consistent with up-regulation of these genes, reversine-treated C2C12 myoblasts redifferentiate into neural as well as mesodermal lineages, under appropriate stimuli. Taken together, these results indicate that reversine induces a multipotency of C2C12 myoblasts via inducing a specific combination of active histone modifications. Collectively, our findings provide a mechanistic rationale for the application of reversine to dedifferentiation of somatic cells.

Antiinflammatory activity of methanol extract isolated from stem bark of Magnolia kobus

The present study reports the antiinflammatory activity of a methanol extract isolated from the stem bark of Magnolia kobus (MK). MK potently inhibited lipopolysaccharide (LPS)‐induced production of nitric oxide and interleukin‐1β (IL‐1β) in RAW 264.7 cells, a murine macrophage‐like cell line. The secretion of tumor necrosis factor‐α (TNF‐α) was also suppressed in LPS‐stimulated RAW 264.7 cells although the magnitude of inhibition was weaker than that of nitric oxide and IL‐1β. The mRNA expressions of inducible nitric oxide synthase (iNOS), IL‐1β and TNF‐α were also suppressed by MK in LPS‐stimulated RAW 264.7 cells. Further study demonstrated that LPS‐induced DNA binding of AP‐1 and phosphorylation of c‐jun N‐terminal kinase (JNK) were inhibited by MK treatment in RAW 264.7 cells, whereas phosphorylation of p38 mitogen‐activated protein kinase was unaffected. Moreover, topical application of MK suppressed ear swelling in 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA)‐induced skin inflammation model. Collectively, these results suggest that MK exerts antiinflammatory effects in vitro and in vivo and this might be mediated, at least in part, by blocking AP‐1 and JNK activation. Copyright

Chemical inhibition of prometastatic lysyl-tRNA synthetase–laminin receptor interaction

Lysyl-tRNA synthetase (KRS), a protein synthesis enzyme in the cytosol, relocates to the plasma membrane after a laminin signal and stabilizes a 67-kDa laminin receptor (67LR) that is implicated in cancer metastasis; however, its potential as an antimetastatic therapeutic target has not been explored. We found that the small compound BC-K-YH16899, which binds KRS, impinged on the interaction of KRS with 67LR and suppressed metastasis in three different mouse models. The compound inhibited the KRS-67LR interaction in two ways. First, it directly blocked the association between KRS and 67LR. Second, it suppressed the dynamic movement of the N-terminal extension of KRS and reduced membrane localization of KRS. However, it did not affect the catalytic activity of KRS. Our results suggest that specific modulation of a cancer-related KRS-67LR interaction may offer a way to control metastasis while avoiding the toxicities associated with inhibition of the normal functions of KRS.

Deprotection of benzyl and p-methoxybenzyl ethers by chlorosulfonyl isocyanate-sodium hydroxide

Abstract CSI–NaOH procedure provided a new and mild methodology for the deprotection of benzyl and p-methoxybenzyl ethers without affecting the other functional groups under similar reaction conditions.

Structure and property based design, synthesis and biological evaluation of γ-lactam based HDAC inhibitors.

Histone deacetylases (HDACs) are involved in post-translational modification and gene expression. Cancer cells recruited amounts of HDACs for their survival by epi-genetic down regulation of tumor suppressor genes. HDACs have been the promising targets for treatment of cancer, and many HDAC inhibitors have been investigated nowadays. In previous study, we synthesized δ-lactam core HDAC inhibitors which showed potent HDAC inhibitory activities as well as cancer cell growth inhibitory activities. Through QSAR study of the δ-lactam based inhibitors, the smaller core is suggested as more active than larger one because it fits better in narrow hydrophobic tunnel of the active pocket of HDAC enzyme. The smaller γ-lactam core HDAC inhibitors were designed and synthesized for biological and property optimization. Phenyl, naphthyl and thiophenyl groups were introduced as the cap groups. Hydrophobic and bulky cap groups increase potency of HDAC inhibition because of hydrophobic interaction between HDAC and inhibitors. In overall, γ-lactam based HDAC inhibitors showed more potent than δ-lactam analogues.

A convenient synthetic method for amide oxidation

Abstract Diazotization of o -aminobenzamides in methanol in the presence of a catalytic amount of CuCl affords α-methoxybenzamides in good yields.