Kunhong Kim

p53 and microRNA-34 are suppressors of canonical Wnt signaling.

The tumor suppressor p53 activates miRNA-34 to inhibit Wnt signaling and colorectal cancer cell invasiveness. p53 Activates MicroRNA-34 to Inhibit Wnt Signaling The tumor suppressor p53 is missing or nonfunctional in many cancers, whereas the canonical Wnt signaling pathway is frequently activated. Here, Kim et al. show that p53 restrained Wnt signaling during Xenopus development, whereas loss of p53 function led to aberrant activation of the canonical Wnt signaling pathway, with microRNA-34 (miR-34) providing the connection between the two. They found that p53 stimulated production of miR-34, which, in turn, targeted key genes in the Wnt signaling pathway. Analyses of gene expression data sets indicated that loss of p53 or miR-34 function was associated with activation of Wnt signaling in human cancers; moreover, loss of p53 function increased Wnt signaling in colon cancer cells in vitro. In p53-mutant colon cancer cells, miR-34 attenuated Wnt signaling and decreased the invasiveness of these cells in vitro. Thus, the p53–miR-34–Wnt pathway appears to be crucial not only during development but also for p53’s tumor suppressor function. Although loss of p53 function and activation of canonical Wnt signaling cascades are frequently coupled in cancer, the links between these two pathways remain unclear. We report that p53 transactivated microRNA-34 (miR-34), which consequently suppressed the transcriptional activity of β-catenin–T cell factor and lymphoid enhancer factor (TCF/LEF) complexes by targeting the untranslated regions (UTRs) of a set of conserved targets in a network of genes encoding elements of the Wnt pathway. Loss of p53 function increased canonical Wnt signaling by alleviating miR-34–specific interactions with target UTRs, and miR-34 depletion relieved p53-mediated Wnt repression. Gene expression signatures reflecting the status of β-catenin–TCF/LEF transcriptional activity in breast cancer and pediatric neuroblastoma patients were correlated with p53 and miR-34 functional status. Loss of p53 or miR-34 contributed to neoplastic progression by triggering the Wnt-dependent, tissue-invasive activity of colorectal cancer cells. Further, during development, miR-34 interactions with the β-catenin UTR affected Xenopus body axis polarity and the expression of Wnt-dependent patterning genes. These data provide insight into the mechanisms by which a p53–miR-34 network restrains canonical Wnt signaling cascades in developing organisms and human cancer.

Prostate cancer cell death produced by the co-delivery of Bcl-xL shRNA and doxorubicin using an aptamer-conjugated polyplex

We investigated the synergism between shRNAs against Bcl-xL and doxorubicin (DOX) using aptamer-conjugated polyplexes (APs) in combination cancer therapy. Synergistic and selective cancer cell death was achieved by AP-mediated co-delivery of very small amounts of DOX and Bcl-xL-specific shRNA, which simultaneously activated an intrinsic apoptotic pathway. A branched polyethyleneimine (PEI) was grafted to polyethylene glycol (PEI-PEG) to serve as a vehicle for shRNA delivery, and its surface was further conjugated with an anti-PSMA aptamer (APT) for the selective delivery of APs to prostate cancer cells that express prostate-specific membrane antigens (PSMA) on their cell surface. The APs were finally obtained after intercalation of DOX to form shRNA/PEI-PEG-APT/DOX conjugates. Cell viability assays and FACS analysis of GFP expression against PC3 (PSMA deficient) and LNCaP (PSMA overexpressed) cells demonstrated that the synthesized APs inhibited the growth of PSMA-abundant prostate cancer cells with strong cell selectivity. Consequently, IC(50) values of APs loaded with both DOX and shRNA were approximately 17-fold less than those for the simple mixture of shRNA plus drug (shRNA/Lipofectamine + DOX). These results suggest that AP-mediated co-delivery of an anti-cancer drug and shRNA against Bcl-xL may widen the therapeutic window and allow for the selective destruction of cancer cells.

Development of the Korean versions of WHO Quality of Life scale and WHOQOL-BREF

The purpose of this study was to develop the Korean version of World Health Organization Quality of Life study assessment instrument (WHOQOL) and WHOQOL-BREF, an abbreviated version of WHOQOL and to identify contributing factors in the quality of life of Koreans. The WHOQOL and WHOQOL-BREF were translated into colloquial Korean according to instructions of the WHOQOL study group. Then the Korean questionnaire was applied to 538 subjects, composed of 171 medical patients and 367 healthy subjects who volunteered to rate the scale. Finally, 486 subjects completed the rating. Collected data were analyzed statistically. The Korean version of WHOQOL and WHOQOL-BREF domain scores demonstrated good test–retest reliability, internal consistency, criterion validity, content validity and discriminant validity. The physical, psychological, social and environmental domains made a significant contribution to explaining the variance in the quality of life while the independence and spiritual domains made a lesser contribution. The domain scores produced by the WHOQOL-BREF correlated highly with the WHOQOL. The physical health domain contributed most in overall quality of life, while the social domain made the least contribution. These results suggest that the Korean version of WHOQOL and WHOQOL-BREF are valid and reliable in the assessment of quality of life and that physical domain is contributing most and social and spiritual factors are contributing least to the quality of life in Koreans.

Stereospecific effects of ginsenoside 20-Rg3 inhibits TGF-β1-induced epithelial–mesenchymal transition and suppresses lung cancer migration, invasion and anoikis resistance

The epithelial-mesenchymal transition (EMT) is a pivotal cellular process during which epithelial polarized cells become motile mesenchymal-appearing cells, which, in turn, promotes the metastatic potential of cancer. Ginseng is a perennial plant belonging to the genus Panax that exhibits a wide range of pharmacological and physiological activities. Ginsenosides 20-Rg3, which is the active component of ginseng, has various medical effects, such as anti-tumorigenic, anti-angiogenesis, and anti-fatiguing activities. In addition, ginsenosides 20(S)-Rg3 and 20(R)-Rg3 are epimers, and this epimerization is produced by steaming. However, the possible role of 20(S)-Rg3 and 20(R)-Rg3 in the EMT is unclear. We investigated the effect of 20(S)-Rg3 and 20(R)-Rg3 on the EMT. Transforming growth factor-beta 1 (TGF-β1) induces the EMT to promote lung adenocarcinoma migration, invasion, and anoikis resistance. To understand the repressive role of 20(S)-Rg3 and 20(R)-Rg3 in lung cancer migration, invasion, and anoikis resistance, we investigated the potential use of 20(S)-Rg3 and 20(R)-Rg3 as inhibitors of TGF-β1-induced EMT development in A549 lung cancer cells in vitro. Here, we show that 20(R)-Rg3, but not 20(S)-Rg3, markedly increased expression of the epithelial marker E-cadherin and repressed Snail upregulation and expression of the mesenchymal marker vimentin during initiation of the TGF-β1-induced EMT. 20(R)-Rg3 also inhibited the TGF-β1-induced increase in cell migration, invasion, and anoikis resistance of A549 lung cancer cells. Additionally, 20(R)-Rg3 markedly inhibited TGF-β1-regulated matrix metalloproteinase-2 and activation of Smad2 and p38 mitogen activated protein kinase. Taken together, our findings provide new evidence that 20(R)-Rg3 suppresses lung cancer migration, invasion, and anoikis resistance in vitro by inhibiting the TGF-β1-induced EMT.

Protein Kinase Casein Kinase 2–Mediated Upregulation of N-Cadherin Confers Anoikis Resistance on Esophageal Carcinoma Cells

Previously, we reported that high PKCK2 activity could protect cancer cells from death receptor–mediated apoptosis through phosphorylation of procaspase-2. Because anoikis is another form of apoptosis, we asked whether PKCK2 could similarly confer resistance to anoikis on cancer cells. Human esophageal squamous cancer cell lines with high PKCK2 activity (HCE4 and HCE7) were anoikis-resistant, whereas cell lines with low PKCK2 activity (TE2 and TE3) were anoikis-sensitive. Because the cells showed different sensitivity to anoikis, we compared the expression of cell adhesion molecules between anoikis-sensitive TE2 and anoikis-resistant HCE4 cells using cDNA microarray. We found that E-cadherin is expressed only in TE2 cells; whereas N-cadherin is expressed instead of E-cadherin in HCE4 cells. To examine whether PKCK2 activity could determine the type of cadherin expressed, we first increased intracellular PKCK2 activity in TE2 cells by overexpressing the PKCK2α catalytic subunit using lentivirus and found that high PKCK2 activity could switch cadherin expression from type E to N and confer anoikis resistance. Conversely, a decrease in PKCK2 activity in HCE4 cells by knockdown of PKCK2α catalytic subunit using shRNA induced N- to E-cadherin switching and the anoikis-resistant cells became sensitive. In addition, N-cadherin expression correlated with PKB/Akt activation and increased invasiveness. We conclude that high intracellular PKCK2 activity confers anoikis resistance on esophageal cancer cells by inducing E- to N-cadherin switching. Mol Cancer Res; 10(8); 1032–8. ©2012 AACR.

The functional relationship between co-repressor N-CoR and SMRT in mediating transcriptional repression by thyroid hormone receptor α

in BCL3, Spot14, FAS and ADRB2 genes. Our data clearly show that SMRT and N-CoR are independently recruited to various TR target genes. We also present evidence that overexpression of N-CoR can restore repression of endogenous genes after knocking down SMRT. Finally, unliganded, co-repressor-free TR is defective in repression and interacts with a co-activator, p300. Collectively, theseresults suggestthatbothSMRTandN-CoRare limited in cells and that knocking down either of them results in co-repressor-free TRandconsequently de-repressionofTRtarget genes.

Neobavaisoflavone sensitizes apoptosis via the inhibition of metastasis in TRAIL-resistant human glioma U373MG cells

AIMS Neobavaisoflavone (NBIF), an isoflavone isolated from Psoralea corylifolia (Leguminosae), has striking anti-inflammatory and anti-cancer effects. NBIF inhibits the proliferation of prostate cancer in vitro and in vivo. MAIN METHODS Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a key endogenous molecule that selectively induces apoptosis in cancer cells with little or no toxicity in normal cells. However, some cancer cells, including U373MG cells, are resistant to TRAIL-mediated apoptosis. We demonstrated that the cell viability, migration and invasion assay were used in U373MG glioma cells. KEY FINDINGS In this study, we found that NBIF sensitizes human U373MG glioma cells to TRAIL-mediated apoptosis. Co-treatment of TRAIL and NBIF effectively induced Bid cleavage and activated caspases 3, 8, and 9. Importantly, DR5 expression was upregulated by NBIF. We also observed that the combination NBIF and TRAIL increased expression of BAX. We further demonstrate that NBIF induced TRAIL-mediated apoptosis in human glioma cells by suppressing migration and invasion, and by inhibiting anoikis resistance. SIGNIFICANCE Taken together, our results suggest that NBIF reduces the resistance of cancer cells to TRAIL and that the combination of NBIF and TRAIL may be a new therapeutic strategy for treating TRAIL-resistant glioma cells.

Nuclear hormone receptor corepressor promotes esophageal cancer cell invasion by transcriptional repression of interferon-γ–inducible protein 10 in a casein kinase 2–dependent manner

Casein kinase 2 (CK2) phosphorylates the nuclear hormone receptor corepressor (NCoR) to stabilize NCoR against the ubiquitin-dependent proteasomal degradation pathway. The CK2-NCoR signaling network suppresses the transcription of IP-10 to promote invasive growth of human esophageal cancer cells.

Role of Promyelocytic Leukemia Zinc Finger (PLZF) in Cell Proliferation and Cyclin-dependent Kinase Inhibitor 1A (p21WAF/CDKN1A) Gene Repression

Background: Promyelocytic leukemia zinc finger (PLZF) is transcription repressor that recruits nuclear co-repressors at the target promoters. PLZF is overexpressed in various human solid tumors, such as clear cell renal carcinoma, glioblastoma, and seminoma. Results: PLZF represses transcription of CDKN1A by inhibition of p53 acetylation, Sp1 binding. Conclusion: PLZF causes cellular transformation and increases cell proliferation by repressing transcription of CDKN1A. Significance: PLZF can act as a proto-oncogene depending on the cell types. Promyelocytic leukemia zinc finger (PLZF) is a transcription repressor that was initially isolated as a fusion protein with retinoic acid receptor α. PLZF is aberrantly overexpressed in various human solid tumors, such as clear cell renal carcinoma, glioblastoma, and seminoma. PLZF causes cellular transformation of NIH3T3 cells and increases cell proliferation in several cell types. PLZF also increases tumor growth in the mouse xenograft tumor model. PLZF may stimulate cell proliferation by controlling expression of the genes of the p53 pathway (ARF, TP53, and CDKN1A). We found that PLZF can directly repress transcription of CDKN1A encoding p21, a negative regulator of cell cycle progression. PLZF binds to the proximal Sp1-binding GC-box 5/6 and the distal p53-responsive elements of the CDKN1A promoter to repress transcription. Interestingly, PLZF interacts with Sp1 or p53 and competes with Sp1 or p53. PLZF interacts with corepressors, such as mSin3A, NCoR, and SMRT, thereby deacetylates Ac-H3 and Ac-H4 histones at the CDKN1A promoter, which indicated the involvement of the corepressor·HDACs complex in transcription repression by PLZF. Also, PLZF represses transcription of TP53 and also decreases p53 protein stability by ubiquitination. PLZF may act as a potential proto-oncoprotein in various cell types.

ZBTB2 increases PDK4 expression by transcriptional repression of RelA/p65

The NF-κB is found in almost all animal cell types and is involved in a myriad of cellular responses. Aberrant expression of NF-κB has been linked to cancer, inflammatory diseases and improper development. Little is known about transcriptional regulation of the NF-κB family member gene RelA/p65. Sp1 plays a key role in the expression of the RelA/p65 gene. ZBTB2 represses transcription of the gene by inhibiting Sp1 binding to a Sp1-binding GC-box in the RelA/p65 proximal promoter (bp, −31 to −21). Moreover, recent studies revealed that RelA/p65 directly binds to the peroxisome proliferator-activated receptor-γ coactivator1α (PGC1α) to decrease transcriptional activation of the PGC1α target gene PDK4, whose gene product inhibits pyruvate dehydrogenase (PDH), a key regulator of TCA cycle flux. Accordingly, we observed that RelA/p65 repression by ZBTB2 indirectly results in increased PDK4 expression, which inhibits PDH. Consequently, in cells with ectopic ZBTB2, the concentrations of pyruvate and lactate were higher than those in normal cells, indicating changes in glucose metabolism flux favoring glycolysis over the TCA cycle. Knockdown of ZBTB2 in mouse xenografts decreased tumor growth. ZBTB2 may increase cell proliferation by reprogramming glucose metabolic pathways to favor glycolysis by upregulating PDK4 expression via repression of RelA/p65 expression.