Wol Soon Jo

A novel resveratrol derivative, HS1793, overcomes the resistance conferred by Bcl-2 in human leukemic U937 cells

The chemopreventive and chemotherapeutic properties associated with resveratrol offer promise for the design of new chemotherapeutic agents. However, resveratrol is not a potent cytotoxic compound when compared with other chemotherapeutic drugs. Thus, several studies were undertaken to obtain synthetic analogues of resveratrol with potent activity. The present study was undertaken to examine whether four resveratrol analogues (HS-1784, -1792, -1791 and -1793) that we had designed and synthesized show antitumor activity. Here, we observed that all of these resveratrol analogues displayed stronger antitumor effects than resveratrol in most cancer cells tested. We further examined whether HS-1793, showing potent antitumor effects in most cancer cells tested, overcomes the resistance conferred by Bcl-2, since overcoming the resistance conferred by Bcl-2 represents an attractive therapeutic strategy against cancer. Our viability assay showed that HS-1793 overcomes the resistance conferred by Bcl-2 in human leukemic U937 cells. Various apoptosis assessment assays demonstrated that HS-1793 overcomes the resistance conferred by Bcl-2 in human leukemic U937 cells by inducing apoptosis. Noticeably, we elucidated the marked downregulation of 14-3-3 protein by HS-1793, indicating that HS-1793 overcomes the resistance conferred by Bcl-2 in U937 cells via 14-3-3. We also observed that HS-1793 exerts its antitumor activity via Bad. However, overall data obtained from methylation specific PCR, RT-PCR and real-time PCR suggest that HS-1793 plays a role in the downregulation of 14-3-3 at a post-transcriptional level. Further understanding exactly how HS-1793 overcomes the resistance conferred by Bcl-2 via 14-3-3 may guide the development of future anticancer agents.

Mutant p53 (G199V) Gains Antiapoptotic Function through Signal Transducer and Activator of Transcription 3 in Anaplastic Thyroid Cancer Cells

In the present study, we identified a missense mutation (G199V) in KAT-18 cell line established from primary cultures of anaplastic thyroid cancer (ATC). Notably, knockdown of this mutant (mt) p53 reduced cell viability and exerted antitumor activity equivalent to high doses of several chemotherapeutic agents. We showed that p53 knockdown had an antitumor effect via the induction of apoptosis. We further examined the underlying mechanism by which mt p53 (G199V) gains antiapoptotic function in KAT-18 cells. Microarray analysis revealed that p53 knockdown modified the expression of numerous apoptosis-related genes. Importantly, p53 knockdown led to downregulation of signal transducer and activator of transcription-3 (STAT3) gene expression. We further observed that p53 knockdown induced the downregulation of STAT3 protein. We also observed that a STAT3 inhibitor augmented the reduction of cell viability induced by p53 knockdown, whereas interleukin-6 treatment alleviated this effect. In addition, overexpression of STAT3 protected ATC cells against cell death induced by p53 knockdown. Taken together, these data show that mt p53 (G199V) gains antiapoptotic function mediated by STAT3 in ATC cells. Inhibition of the function of mt p53 (G199V) could be a novel and useful therapeutic strategy for decreasing the extent and severity of toxicity due to chemotherapeutic agents. (Mol Cancer Res 2009;7(10):1645–54)

Interferon gamma induced by resveratrol analog, HS-1793, reverses the properties of tumor associated macrophages

Macrophages are capable of both inhibiting and promoting the growth and spread of cancers, depending on their activation state. Tumor-associated macrophages (TAM) are a kind of alternatively activated M2 macrophage, which may contribute to tumor progression. Following our previous study to evaluate the anti-tumor effect of a synthetic resveratrol analog HS-1793, the current study demonstrated that HS-1793 treatment significantly increased IFN-γ secreting cells in splenocytes and decreased CD206+ macrophage infiltration compared to CD68+ cells in the tumor site with a higher expression of IFN-γ. As these results suggested that IFN-γ increased locally at the tumor sites could modulate the status of TAM, we designed an in vitro model to study macrophage morphology and functions in relation to the tumor microenvironment. Human monocytic cell line THP-1 cells stimulated with phorbol-12-myristate-13-acetate (PMA) differentiated to macrophages with M2-like phenotypes. TAM-like properties of CD206(high), CD204(high), IL-10(high), TGF-β(high), IL-6(low), IL-12(low), VEGF(high), and MMP-9(high) and promotion of tumor cell invasion were more pronounced in M-2-polarized THP-1 macrophages generated by differentiating THP-1 cells with PMA and subsequently polarizing them with Th2 cytokines (IL-4/IL-13). Upon IFN-γ exposure, THP-1-derived TAM changed their phenotypes to the M-1-like morphology and intracellular granular pattern with an expression of an increased level of proinflammatory and immunostimulatory cytokines and a reduced level of immunosuppressive and tumor progressive mediators. These results explain the underlying mechanism of the anti-tumor activity of HS-1793. The elevated level of IFN-γ production after HS-1793 treatment evoked reprogramming of M-2 phenotype TAM, which efficiently countered the immunosuppressive and tumor progressive influences of TAM.

The modulation of radiation-induced cell death by genistein in K562 cells: activation of thymidine kinase 1.

ABSTRACTIonizing radiation is one of the most effective tools in cancer therapy. In a previous study, we reported that protein tyrosine kinase (PTK) inhibitors modulate the radiation responses in the human chronic myelogenous leukemia (CML) cell line K562. The receptor tyrosine kinase inhibitor, genistein, delayed radiation-induced cell death, while non-recepter tyrosine kinase inhibitor, herbimycin A (HMA) enhances radiation-induced apoptosis. In this study, we focused on the modulation of radiation-induced cell death by genistein and performed PCR-select suppression subtractive hybridization (SSH) to understand its molecular mechanism. We identified human thymidine kinase 1 (TK1), which is cell cycle regulatory gene and confirmed expression of TK1 mRNA by Northern blot analysis. Expression of TK1 mRNA and TK 1 enzymatic activity were parallel in their increase and decrease. TK1 is involved in G1-S phase transition of cell cycle progression. In cell cycle analysis, we showed that radiation induced G2 arrest in K562 cells but it was not able to sustain. However, the addition of genistein to irradiated cells sustained a prolonged G2 arrest up to 120 h. In addition, the expression of cell cycle-related proteins, cyclin A and cyclin B1, provided the evidences of G1/S progression and G2-arrest, and their relationship with TK1 in cells treated with radiation and genistein. These results suggest that the activation of TK1 may be critical to modulate the radiation-induced cell death and cell cycle progression in irradiated K562 cells.

Resveratrol analog, HS-1793 enhance anti-tumor immunity by reducing the CD4+CD25 + regulatory T cells in FM3A tumor bearing mice

Natural agents with the immunomodulating property have been gaining traction to be employed in the complementary therapy of cancer because the ineffectiveness of numerous therapeutic strategies may be related in part to the tumor-induced immunosuppressive phenotypes, especially regulatory T (Treg) cells found in the tumor microenvironment. The present study was undertaken to examine whether HS-1793, synthetic resvertrol analog free from the restriction of metabolic instability and high dose requirement of resveratrol, induces an in vivo anti-tumor effect in FM3A tumor bearing mice through the suppression of Treg cells, which contribute to an increase in tumor specific cytotoxic T cell responses. Intraperitoneal injections of HS-1793 showed not only therapeutic benefits on established tumors, but also preventive anti-tumor effects. Treg cells (CD4+CD25+Foxp3+ cells) were significantly reduced in the total splenocytes as well as tumor tissues from HS-1793-administered mice, and the production of TGF-β inducing Treg showed a similar pattern. On the contrary, the administration of HS-1793 increased IFN-γ-expressing CD8+ T cells, upregulated IFN-γ production, and enhanced the cytotoxicity of splenocytes against FM3A tumor cells both in therapeutic and preventive experimental animals. These results demonstrated the suppressive role of HS-1793 on the function of Treg cells contributing to tumor specific cytotoxic T lymphocyte responses in tumor-bearing mice, which explained the underlying mechanism of the anti-tumor immunity of HS-1793.

The Anti-inflammatory Effects of Water Extract from Cordyceps militaris in Murine Macrophage

Abstract The aim of this study was to determine the in vitro anti-inflammatory effect of hot water extract from Cordyceps militaris fruiting bodies (CMWE) on lipopolysaccharide (LPS)-stimulated nitric oxide (NO) production, tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) release in RAW 264.7 cells. The treatment of macrophages with various concentrations of hot CMWE significantly reduced LPS-induced production as well as NO, TNF-α and IL-6 secretion in a concentration-dependent manner. These results suggest that CMWE have potent inhibitory effects on the production of these inflammatory mediators.

Protective activity of a novel resveratrol analogue, HS-1793, against DNA damage in 137Cs-irradiated CHO-K1 cells

Resveratrol has received considerable attention as a polyphenol with anti-oxidant, anti-carcinogenic, and anti-inflammatory effects. Radiation is an important component of therapy for a wide range of malignant conditions. However, it causes damage to normal cells and, hence, can result in adverse side effects. This study was conducted to examine whether HS-1793, a novel resveratrol analogue free from the restriction of metabolic instability and the high dose requirement of resveratrol, induces a protective effect against radiation-induced DNA damage. HS-1793 effectively scavenged free radicals and inhibited radiation-induced plasmid DNA strand breaks in an in vitro assay. HS-1793 significantly decreased reactive oxygen species and cellular DNA damage in 2 Gy-irradiated Chinese hamster ovary (CHO)-K1 cells. In addition, HS-1793 dose-dependently reduced the levels of phosphorylated H2AX in irradiated CHO-K1 cells. These results indicate that HS-1793 has chemical radioprotective activity. Glutathione levels and superoxide dismutase activity in irradiated CHO-K1 cells increased significantly following HS-1793 treatment. The enhanced biological anti-oxidant activity and chemical radioprotective activity of HS-1793 maintained survival of irradiated CHO-K1 cells in a clonogenic assay. Therefore, HS-1793 may be of value as a radioprotector to protect healthy tissue surrounding tumor cells during radiotherapy to obtain better tumor control with a higher dose.

Effect of Microalgal Extracts of Tetraselmis suecica against UVB-Induced Photoaging in Human Skin Fibroblasts

Exposure of cells to ultraviolet B (UVB) radiation can induce production of free radicals and reactive oxygen species (ROS), which damage cellular components. In addition, these agents can stimulate the expression of matrix metalloproteinase (MMP) and decrease collagen synthesis in human skin cells. In this study, we examined the anti-photoaging effects of extracts of Tetraselmis suecica (W-TS). W-TS showed the strongest scavenging activity against 2,2-difenyl-1-picrylhydrazyl (DPPH) and peroxyl radicals, followed by superoxide anions from the xanthine/xanthine oxidase system. We observed that the levels of both intracellular ROS and lipid peroxidation significantly increased in UVB-irradiated human skin fibroblast cells. Furthermore, the activities of enzymatic antioxidants (e.g., superoxide dismutase) and the levels of non-enzymatic antioxidants (e.g., glutathione) significantly decreased in cells. However, W-TS pretreatment, at the maximum tested concentration, significantly decreased intracellular ROS and malondialdehyde (MDA) levels, and increased superoxide dismutase and glutathione levels in the cells. At this same concentration, W-TS did not show cytotoxicity. Type 1 procollagen and MMP-1 released were quantified using RT-PCR techniques. The results showed that W-TS protected type 1 procollagen against UVBinduced depletion in fibroblast cells in a dose-dependent manner via inhibition of UVB-induced MMP-1. Taken together, the results of the study suggest that W-TS effectively inhibits UVB-induced photoaging in skin fibroblasts by its strong anti-oxidant ability.

Tumor associated macrophages provide the survival resistance of tumor cells to hypoxic microenvironmental condition through IL-6 receptor-mediated signals.

Hypoxia and infiltration of tumor-associated macrophages (TAM) are intrinsic features of the tumor microenvironment. Tumor cells that remain viable in hypoxic conditions often possess an increased survival potential and tend to grow aggressively. TAM also respond to a variety of signals in the hypoxic tumor microenvironment and express a more M2-like phenotype. In this study, the established mouse tumor tissues showed a dense infiltration of CD206+ macrophages at the junctions between the normoxic and hypoxic regions and an increased IL-6 receptor (IL-6R) expression of tumor cells in the areas of CD206+ TAM accumulation, which indicates a role of M2 phenotype TAM in survival adaptation of tumor cells preparing for an impending hypoxic injury before changes in oxygen availability. Cocultured mouse FM3A or human MCF-7 tumor cells with tumor infiltrating macrophages isolated from mouse tumor tissues and M2-polarized macrophages generated from human THP-1 cells, respectively, showed significantly decreased rate of cell death in cultures exposed to hypoxia. The acquisition of survival resistance was attributed to increased IL-6 production by M2 TAM and increased expression of IL-6R in tumor cells in the coculture system. MCF-7 cells cocultured with M2 TAM showed activated JAK1/STAT3 and Raf/MEK/JNK pathways contributing to tyrosine and serine phophorylation of STAT3, respectively. However, only tyrosine phosphorylated STAT3 was detected in the nucleus, which induced upregulation of Bcl-2 and downregulation of Bax and Bak. Finally, knockdown of IL-6R by small interfering RNA significantly counteracted coculture-induced signals and completely abolished the survival resistance to hypoxic injury. Thus, we present evidence for the role of M2 phenotype TAM in IL-6 receptor-mediated signals, particularly tyrosine phosphorylation of STAT3, responsible for the prosurvival adaptation of tumor cells to hypoxia.

Bub1 is required for maintaining cancer stem cells in breast cancer cell lines.

Breast cancer is a leading cause of death among women worldwide due to therapeutic resistance and cancer recurrence. Cancer stem cells are believed to be responsible for resistance and recurrence. Many efforts to overcome resistance and recurrence by regulating cancer stem cells are ongoing. Bub1 (Budding uninhibited by benzimidazoles 1) is a mitotic checkpoint serine/threonine kinase that plays an important role in chromosome segregation. Bub1 expression is correlated with a poor clinical prognosis in patients with breast cancer. We identified that depleting Bub1 using shRNAs reduces cancer stem cell potential of the MDA-MB-231 breast cancer cell line, resulting in inhibited formation of xenografts in immunocompromised mice. These results suggest that Bub1 may be associated with cancer stem cell potential and could be a target for developing anti-breast cancer stem cell therapies.