FRA1:c-JUN:HDAC1 complex down-regulates filaggrin expression upon TNFα and IFNγ stimulation in keratinocytes.

Filaggrin (FLG), an essential structural protein for skin barrier function, is down-regulated under chronic inflammatory conditions, leading to disruption of the skin barrier. However, the detailed molecular mechanisms of how FLG changes in the context of chronic inflammation are poorly understood. Here, we identified the molecular mechanisms by which inflammatory cytokines inhibit FLG expression in the skin. We found that the AP1 response element within the -343/+25 of the FLG promoter was necessary for TNFα + IFNγ-induced down-regulation of FLG promoter activity. Using DNA affinity precipitation assay, we observed that AP1 subunit composition binding to the FLG promoter was altered from c-FOS:c-JUN (at the early time) to FRA1:c-JUN (at the late time) in response to TNFα + IFNγ stimulation. Knockdown of FRA1 or c-JUN abrogated TNFα + IFNγ-induced FLG suppression. Histone deacetylase (HDAC) 1 interacted with FRA1:c-JUN under TNFα + IFNγ stimulation. Knockdown of HDAC1 abrogated the inhibitory effect of TNFα + IFNγ on FLG expression. The altered expression of FLG, FRA1, c-JUN, and HDAC1 was confirmed in mouse models of 2,4-dinitrochlorobenzene-induced atopic dermatitis and imiquimod-induced psoriasis. Thus, the current study demonstrates that TNFα + IFNγ stimulation suppresses FLG expression by promoting the FRA1:c-JUN:HDAC1 complex. This study provides insight into future therapeutic strategies targeting the FRA1:c-JUN:HDAC1 complex to restore impaired FLG expression in chronic skin inflammation.

Design, synthesis, and biological evaluation of 2-(2-oxoindolin-3-ylidene)hydrazinecarbothioamides as a potential EGR-1 inhibitor for targeted therapy of atopic dermatitis.

Atopic dermatitis is a chronic inflammatory skin disease characterized by intense itching and frequent skin barrier dysfunctions. EGR-1 is a transcription factor that aggravates the pathogenesis of atopic dermatitis by promoting the production of various inflammatory cytokines. Three 2-(2-oxoindolin-3-ylidene)hydrazinecarbothioamides (IT21, IT23, and IT25) were identified as novel inhibitors of EGR-1 DNA-binding activity. In silico docking experiments were performed to elucidate the binding conditions of the EGR-1 zinc-finger (ZnF) DNA-binding domain. Electrophoretic mobility shift assays confirmed the targeted binding effect on the EGR-1 ZnF DNA-binding domain, leading to dose-dependent dissociation of the EGR-1-DNA complex. At the functional cellular level, IT21, IT23, and IT25 effectively reduced mRNA expression of TNFα-induced EGR-1-regulated inflammatory genes, particularly in HaCaT keratinocytes inflamed by TNFα. In the in vivo efficacy study, IT21, IT23, and IT25 demonstrated the potential to alleviate atopic dermatitis-like skin lesions in the ear skin of BALB/c mice. These findings suggest that targeting the EGR-1 ZnF DNA-binding domain with 2-(2-oxoindolin-3-ylidene)hydrazinecarbothioamide derivatives (IT21, IT23, and IT25) could serve as lead compounds for the development of potential therapeutic agents against inflammatory skin disorders, including atopic dermatitis.

Homeobox Protein PROX1 Expression is Negatively Regulated by Histone Deacetylase 1 and c-JUN Complex in MDA-MB-231 Human Breast Cancer Cells.

Prospero homeobox 1 (PROX1) is a member of the homeobox transcription factor family that plays a critical role in the development of multiple tissues and specification of cell fate. PROX1 expression is differentially regulated based on the cellular context and plays an antagonistic role as a tumour promoter or suppressor in different tumour types. In human breast cancer, PROX1 expression is suppress-ed; however, the molecular mechanism by which it is down-regulated remains poorly understood. Here, we show that ectopic expression of PROX1 reduces the motility and invasiveness of MDA-MB-231 human breast cancer cells, suggesting that PROX1 functions as a negative regulator of tumour invasion in MDA-MB-231 cells. Treatment with histone deacetylase (HDAC) inhibitors up-regulates PROX1 mRNA and protein expression levels. Knockdown of HDAC1 using short hairpin RNA also up-regulates PROX1 mRNA and protein expression levels. We found that HDAC1 interacted with c-JUN at the activator protein (AP)-1-binding site located at -734 to -710 in the PROX1 promoter region to suppress PROX1 expression. In addition, c-JUN N-terminal kinase-mediated c-JUN phosphorylation was found to be crucial for silencing PROX1 expression. In conclusion, PROX1 expression can be silenced by the epigenetic mechanism involved in the complex formation of HDAC1 and c-JUN at the AP-1 site in the PROX1 promoter region in MDA-MB-231 human breast cancer cells. Therefore, this study revealed the epigenetic regulatory mechanism involved in the suppression of PROX1 expression in breast cancer cells.

EGR1 Regulation of Vasculogenic Mimicry in the MDA-MB-231 Triple-Negative Breast Cancer Cell Line through the Upregulation of KLF4 Expression.

Vasculogenic mimicry (VM) is an intriguing phenomenon observed in tumor masses, in which cancer cells organize themselves into capillary-like channels that closely resemble the structure and function of blood vessels. Although VM is believed to contribute to alternative tumor vascularization, the detailed regulatory mechanisms controlling these cellular processes remain poorly understood. Our study aimed to investigate the role of Early Growth Response 1 (EGR1) in regulating VM in aggressive cancer cells, specifically MDA-MB-231 triple-negative breast cancer cells. Our study revealed that EGR1 promotes the formation of capillary-like tubes by MDA-MB-231 cells in a 3-dimensional Matrigel matrix. EGR1 was observed to upregulate Kruppel-like factor 4 (KLF4) expression, which regulates the formation of the capillary-like tube structure. Additionally, our findings highlight the involvement of the ERK1/2 and p38 mitogen-activated protein kinase pathways in mediating the expression of EGR1 and KLF4, underscoring their crucial role in VM in MDA-MB-231 cells. Understanding these regulatory mechanisms will provide valuable insights into potential therapeutic targets for preventing VM during the treatment of triple-negative breast cancer.

Design, synthesis, and biological activities of 3-((4,6-diphenylpyrimidin-2-ylamino)methylene)-2,3-dihydrochromen-4-ones.

Novel (Z)-3-((4,6-diphenylpyrimidin-2-ylamino)methylene)-2,3-dihydrochromen-4-one derivatives were designed and synthesized to find chemotherapeutic agents. Derivative 9 was selected based on its clonogenicity against cancer cells and synthetic yield for further biological experiments. It showed decreases in aurora kinase A, B, and C phosphorylation from western blot analysis. Derivative 9 upregulated the expression of G1 cell cycle inhibitory proteins including p21 and p27, and G1 progressive cyclin D1, and downregulated G1-to-S progressive cyclins, resulting in cell cycle arrest at the G1/S boundary. It stimulated the cleavage of caspase-9, -3, -7, and poly (ADP-ribose) polymerase, resulting in triggering apoptosis through a caspase-dependent pathway. In addition, derivative 9 inhibited in vivo tumor growth in a syngeneic tumor implantation mouse model. The findings of this study suggest that derivative 9 can be considered as a lead compound for chemotherapeutic agents.

Saikosaponin A and Saikosaponin C Reduce TNF-α-Induced TSLP Expression through Inhibition of MAPK-Mediated EGR1 Expression in HaCaT Keratinocytes.

Atopic dermatitis (AD) is one of the most common chronic inflammatory skin diseases worldwide, characterized by intense pruritus and eczematous lesions. Aberrant expression of thymic stromal lymphopoietin (TSLP) in keratinocytes is associated with the pathogenesis of AD and is considered a therapeutic target for the treatment of this disease. Saikosaponin A (SSA) and saikosaponin C (SSC), identified from Radix Bupleuri, exert anti-inflammatory effects. However, the topical effects of SSA and SSC on chronic inflammatory skin diseases are unclear. In this study, we investigated the effects of SSA and SSC on TSLP suppression in an AD-like inflammatory environment. We observed that SSA and SSC suppressed tumor necrosis factor-α-induced TSLP expression by downregulating the expression of the transcription factor early growth response 1 (EGR1) via inhibition of the extracellular signal-regulated kinase 1/2, c-Jun N-terminal kinase 1/2, and p38 mitogen-activated protein kinase pathways. We also confirmed that topical application of SSA or SSC reduced AD-like skin lesions in BALB/c mice challenged with 2,4-dinitrochlorobenzene. Our findings suggest that suppression of EGR1-regulated TSLP expression in keratinocytes might be attributable to the anti-inflammatory effects of SSA and SSC in AD-like skin lesions.

ß-Caryophyllene Ameliorates 2,4-Dinitrochlorobenzene-Induced Atopic Dermatitis through the Downregulation of Mitogen-Activated Protein Kinase/EGR1/TSLP Signaling Axis.

Atopic dermatitis (AD) is one of the most common inflammatory skin diseases accompanied by severe itching. ß-caryophyllene (BCP), which displays anti-inflammatory activity, is a natural agonist of cannabinoid receptor 2. However, the therapeutic effects of BCP on atopic dermatitis (AD) remain poorly understood. The current study aimed to evaluate the topical therapeutic efficacy of BCP in an AD-like mouse model. Thymic Stromal Lymphopoietin (TSLP) is a keratinocyte-derived cytokine that drives AD pathogenesis. This study also investigated the effect of BCP on the interleukin 4 (IL-4)-induced expression of TSLP in HaCaT keratinocytes. We found that the topical application of BCP alleviated AD-like skin inflammation and inhibited the infiltration of proinflammatory cells into skin lesions. Moreover, the topical application of BCP reduced EGR1 (Early Growth Response 1) and TSLP expression in AD-like skin lesions. We also found that BCP inhibited IL-4-induced TSLP expression by downregulating mitogen-activated protein kinase (MAPK)-mediated EGR1 expression in HaCaT keratinocytes. These findings demonstrate that BCP ameliorates DNCB-induced AD-like skin lesions through the downregulation of the MAPK/EGR1/TSLP signaling axis. BCP may be applicable for developing topical therapeutic agents for chronic skin inflammatory diseases, such as AD.

The Natural Janus Kinase Inhibitor Agerarin Downregulates Interleukin-4-Induced PER2 Expression in HaCaT Keratinocytes.

The circadian clock system is closely associated with inflammatory responses. Dysregulation of the circadian clock genes in the skin impairs the skin barrier function and affects the pathophysiology of atopic dermatitis. Interleukin 4 (IL-4) is a proinflammatory cytokine derived from T-helper type 2 cells; it plays a critical role in the pathogenesis of atopic dermatitis. Agerarin (6,7-dimethoxy-2,2-dimethyl-2H-chromene) is a natural JAK1/2/3 inhibitor isolated from Ageratum houstonianum that has a protective effect on the epidermal skin barrier. However, it remains unclear whether agerarin affects the circadian clock system. The aim of this study is to investigate the effect of agerarin on IL-4-induced PER2 gene expression in human keratinocytes through reverse transcription (RT)-PCR, quantitative real-time PCR (qPCR), immunoblotting, immunofluorescence microscopic analysis, and real-time bioluminescence analysis. We found that agerarin reduced IL-4-induced PER2 mRNA expression by suppressing the JAK-STAT3 pathway. In addition, real-time bioluminescence analysis in PER2:luc2p promoter-reporter cells revealed that agerarin restored the oscillatory rhythmicity of PER2 promoter activity altered by IL-4. These findings suggest that agerarin may be useful as a cosmeceutical agent against inflammatory skin conditions associated with disrupted circadian rhythms, such as atopic dermatitis.

Chrysin Inhibits TNFα-Induced TSLP Expression through Downregulation of EGR1 Expression in Keratinocytes.

Thymic stromal lymphopoietin (TSLP) is an epithelial cell-derived cytokine that acts as a critical mediator in the pathogenesis of atopic dermatitis (AD). Various therapeutic agents that prevent TSLP function can efficiently relieve the clinical symptoms of AD. However, the downregulation of TSLP expression by therapeutic agents remains poorly understood. In this study, we investigated the mode of action of chrysin in TSLP suppression in an AD-like inflammatory environment. We observed that the transcription factor early growth response (EGR1) contributed to the tumor necrosis factor alpha (TNFα)-induced transcription of TSLP. Chrysin attenuated TNFα-induced TSLP expression by downregulating EGR1 expression in HaCaT keratinocytes. We also showed that the oral administration of chrysin improved AD-like skin lesions in the ear and neck of BALB/c mice challenged with 2,4-dinitrochlorobenzene. We also showed that chrysin suppressed the expression of EGR1 and TSLP by inhibiting the extracellular signal-regulated kinase (ERK) 1/2 and c-Jun N-terminal kinase (JNK) 1/2 mitogen-activated protein kinase pathways. Collectively, the findings of this study suggest that chrysin improves AD-like skin lesions, at least in part, through the downregulation of the ERK1/2 or JNK1/2-EGR1-TSLP signaling axis in keratinocytes.

Chrysoeriol Prevents TNFα-Induced CYP19 Gene Expression via EGR-1 Downregulation in MCF7 Breast Cancer Cells.

Estrogen overproduction is closely associated with the development of estrogen receptor-positive breast cancer. Aromatase, encoded by the cytochrome P450 19 (CYP19) gene, regulates estrogen biosynthesis. This study aimed to identify active flavones that inhibit CYP19 expression and to explore the underlying mechanisms. CYP19 expression was evaluated using reverse transcription PCR, quantitative real-time PCR, and immunoblot analysis. The role of transcription factor early growth response gene 1 (EGR-1) in CYP19 expression was assessed using the short-hairpin RNA (shRNA)-mediated knockdown of EGR-1 expression in estrogen receptor-positive MCF-7 breast cancer cells. We screened 39 flavonoids containing 26 flavones and 13 flavanones using the EGR1 promoter reporter activity assay and observed that chrysoeriol exerted the highest inhibitory activity on tumor necrosis factor alpha (TNFα)-induced EGR-1 expression. We further characterized and demonstrated that chrysoeriol inhibits TNFα-induced CYP19 expression through inhibition of extracellular signal-regulated kinase 1/2 (ERK1/2)-mediated EGR-1 expression. Chrysoeriol may be beneficial as a dietary supplement for the prevention of estrogen receptor-positive breast cancer, or as a chemotherapeutic adjuvant in the treatment of this condition.

A Novel Synthetic Compound (E)-5-((4-oxo-4H-chromen-3-yl)methyleneamino)-1-phenyl-1H-pyrazole-4-carbonitrile Inhibits TNFα-Induced MMP9 Expression via EGR-1 Downregulation in MDA-MB-231 Human Breast Cancer Cells.

Breast cancer is a common malignancy among women worldwide. Gelatinases such as matrix metallopeptidase 2 (MMP2) and MMP9 play crucial roles in cancer cell migration, invasion, and metastasis. To develop a novel platform compound, we synthesized a flavonoid derivative, (E)-5-((4-oxo-4H-chromen-3-yl)methyleneamino)-1-phenyl-1H-pyrazole-4-carbonitrile (named DK4023) and characterized its inhibitory effects on the motility and MMP2 and MMP9 expression of highly metastatic MDA-MB-231 breast cancer cells. We found that DK4023 inhibited tumor necrosis factor alpha (TNFα)-induced motility and F-actin formation of MDA-MB-231 cells. DK4023 also suppressed the TNFα-induced mRNA expression of MMP9 through the downregulation of the TNFα-extracellular signal-regulated kinase (ERK)/early growth response 1 (EGR-1) signaling axis. These results suggest that DK4023 could serve as a potential platform compound for the development of novel chemopreventive/chemotherapeutic agents against invasive breast cancer.

Leptin is a direct transcriptional target of EGR1 in human breast cancer cells.

Leptin is a cytokine that regulates energy metabolism. Leptin can promote breast cancer progression in obese women. However, the mechanism of regulation of leptin expression in breast cancer cells is unclear. Tumor necrosis factor-alpha (TNF-α) stimulated the transcription of the leptin gene. Using mutant promoter constructs, we demonstrated that the EGR1-binding motif in the proximal region of the leptin gene is required for leptin transcription by TNF-α. Forced expression of EGR1 stimulated leptin promoter activity, whereas silencing of EGR1 by RNA interference reduced TNF-α-induced leptin protein accumulation. The ERK1/2 pathway contributed to the expression of EGR1 and leptin by TNF-α. Our results suggest that EGR1 targets the leptin gene in response to TNF-α stimulation in breast cancer cells.

p38 mitogen-activated protein kinase contributes to TNFα-induced endothelial tube formation of bone-marrow-derived mesenchymal stem cells by activating the JAK/STAT/TIE2 signaling axis.

Bone marrow-derived mesenchymal stem cells (BM-MSCs) can differentiate into endothelial cells in an inflammatory microenvironment. However, the regulatory mechanisms underlying this process are not entirely understood. Here, we found that TIE2 in BM-MSCs was upregulated at the transcriptional level after stimulation with tumor necrosis factor-alpha (TNFα), a major pro-inflammatory cytokine. Additionally, the STAT-binding sequence within the proximal region of TIE2 was necessary for TNFα-induced TIE2 promoter activation. TIE2 and STAT3 knockdown reduced TNFα-induced endothelial tube formation in BMMSCs. Among the major TNFα-activated MAP kinases (ERK1/2, JNK1/2, and p38 MAPK) in BM-MSCs, only inhibition of the p38 kinase abrogated TNFα-induced TIE2 upregulation by inhibiting the JAK-STAT signaling pathway. These findings suggest that p38 MAP contributes to the endothelial differentiation of BM-MSCs by activating the JAK-STAT-TIE2 signaling axis in the inflammatory microenvironment.

The JNK-EGR1 signaling axis promotes TNF-α-induced endothelial differentiation of human mesenchymal stem cells via VEGFR2 expression.

Mesenchymal stem cells (MSCs) can differentiate into endothelial cells; however, the mechanisms underlying this process in the tumor microenvironment (TME) remain elusive. This study shows that tumor necrosis factor alpha (TNF-α), a key cytokine present in the TME, promotes the endothelial differentiation of MSCs by inducing vascular endothelial growth factor receptor 2 (VEGFR2) gene expression. EGR1 is a member of the zinc-finger transcription factor family induced by TNF-α. Our findings indicate that EGR1 directly binds to the VEGFR2 promoter and transactivates VEGFR2 expression. We also demonstrate that EGR1 forms a complex with c-JUN activated by c-JUN N-terminal kinase (JNK) to promote VEGFR2 transcription and endothelial differentiation in MSCs in response to TNF-α stimulation. The shRNA-mediated silencing of EGR1 or c-JUN abrogates TNF-α-induced VEGFR2 transcription and the endothelial differentiation of MSCs. To further evaluated the role of EGR1 in the endothelial differentiation of BM-MSCs, we used a syngenic tumor implantation model. 4T1 mouse mammary tumor cells were injected subcutaneously into BALB/c mice with primary mBM-MSCs isolated from wild-type (Egr1+/+) or Egr1-null (Egr1-/-) mice. CD31-positive cells were predominantly observed at the border of the tumor in the 4T1 plus wild-type MSC group, while staining less in the 4T1 alone or 4T1 plus Egr1-null MSC group. Collectively, these findings demonstrate that the JNK-EGR1 signaling axis plays a crucial role in the TNF-α-induced endothelial differentiation of MSCs in the TME, which could be a potential therapeutic target for solid tumors vasculatures.

Transcription Factor EGR1 Regulates the Expression of the Clock Gene PER2 under IL-4 Stimulation in Human Keratinocytes.

PER2 is a core circadian clock gene that regulates circadian rhythms. IL-4 plays a critical role in the pathogenesis of skin inflammation, including atopic dermatitis. IL-4 enhances PER2 expression, suggesting a relationship between inflammation and the circadian clock. However, little is known about the molecular and cellular mechanisms regulating PER2 expression by inflammatory cytokines. This study showed that transcription factor EGR1 interacted with the PER2 promoter and promoted IL-4‒induced transcriptional activation of the PER2, as revealed by promoter‒reporter assay, electrophoretic mobility shift assay, DNA affinity precipitation assay, and chromatin immunoprecipitation analysis. We also found that IL-4 can use both MAPK and Jak signaling pathways to induce EGR1-mediated PER2 expression, and c-Jun N-terminal kinase 1/2 can augment IL-4‒induced activation of the Jak‒signal transducer and activator of transcription 3 pathway. Consistently, Per2 expression was reduced in dinitrochlorobenzene-induced atopic dermatitis‒like skin lesions in Egr1‒/‒ mice compared with that in Egr1+/+ mice. In addition, using a real-time bioluminescence assay, we observed that EGR1 is required for rhythmic oscillation of PER2 expression under IL-4 exposure. These findings provide further insight into the role of EGR1 in regulating PER2 expression in impaired circadian rhythm in skin inflammation.

Overcoming multidrug resistance by activating unfolded protein response of the endoplasmic reticulum in cisplatin-resistant A2780/CisR ovarian cancer cells.

Cisplatin is a widely used anti-cancer agent. However, the effectiveness of cisplatin has been limited by the commonly developed drug resistance. This study aimed to investigate the potential effects of endoplasmic reticulum (ER) stress to overcome drug resistance using the cisplatin-resistant A2780/CisR ovarian cancer cell model. The synthetic chalcone derivative (E)-3-(3,5-dimethoxyphenyl)-1-(2-methoxyphenyl)prop-2-en-1-one (named DPP23) is an ER stress inducer. We found that DPP23 triggered apoptosis in both parental cisplatinsensitive A2780 and cisplatin-resistant A2780/CisR ovarian cancer cells due to activation of reactive oxygen species (ROS)-mediated unfolded protein response (UPR) pathway in the endoplasmic reticulum. This result suggests that ROSmediated UPR activation is potential in overcoming drug resistance. DPP23 can be used as a target pharmacophore for the development of novel chemotherapeutic agents capable of overcoming drug resistance in cancer cells, particularly ovarian cancer cells. [BMB Reports 2020; 53(2): 88-93].

Transcriptomic analysis of the effect of (E)-3-(3,5-dimethoxyphenyl)-1-(2-methoxyphenyl) prop-2-en-1-one (DPP23) on reactive oxygen species generation in MIA PaCa-2 pancreatic cancer cells.

BACKGROUND: Reactive oxygen species (ROS) generation specifically in cancer cells may be a promising strategy for their selective killing. The synthetic chalcone derivative (E)-3-(3,5-dimethoxyphenyl)-1-(2-methoxyphenyl)prop-2-en-1-one (DPP23) exerts antitumor activity through ROS-mediated apoptosis in cancer cells but not in healthy cells. However, the mechanism underlying ROS generation by DPP23 remains unknown. OBJECTIVE: The current study aims to identify possible DPP23 target genes responsible for ROS generation through the mining of microarray data stored in NCBI's Gene Expression Omnibus (GEO). METHODS: A comprehensive expression profile of genes modulated by DPP23 was examined by gene ontology analysis. DPP23-modulated genes in Mia-PaCa2 pancreatic cells were validated by reverse transcription-PCR. RESULTS: Multiple genes were up and downregulated by DPP23 treatment in MiaPaCa2 pancreatic cancer cells. Genes with absolute fold-change (FC) of > 2 were selected as the cut-off criteria and grouped into 10 clusters to analyze expression patterns systematically. We observed that genes with increased expression at 6 h were significantly affected by ROS increase, unfolded protein response, and cell death. Expression of 13 genes involved in glutathione metabolism, including CHAC1, GCLC, G6PD, GSTO2, GSTA5, GSTM2, GSR, GPX3/6/8, GGT1, PGD, ATF4, and NAT8B, are modulated by DPP23. Of these, CHAC1 was most highly upregulated upon DPP23 treatment. CONCLUSION: DPP23 alters global gene expression associated with multiple cellular responses, including oxidative stress and apoptosis. We found that DPP23 may induce GSH depletion through modulation of gene expression, which is especially involved in glutathione metabolism. Of these, CHAC1 emerged as the most prominent candidate for DPP23 as it was the most responsive to DPP23 treatment.

WNT11 is a direct target of early growth response protein 1.

WNT11 is a member of the non-canonical Wnt family and plays a crucial role in tumor progression. However, the regulatory mechanisms underlying WNT11 expression are unclear. Tumor necrosis factor-alpha (TNFα) is a major inflammatory cytokine produced in the tumor microenvironment and contributes to processes associated with tumor progression, such as tumor invasion and metastasis. By using site-directed mutagenesis and introducing a serial deletion in the 5'-regulatory region of WNT11, we observed that TNFα activates the early growth response 1 (EGR1)-binding sequence (EBS) in the proximal region of WNT11 and that the transcription factor EGR1 is necessary for the TNFα-induced transcription of WNT11. EGR1 bound directly to the EBSs within the proximal 5'-regulatory region of WNT11 and ectopic expression of EGR1 stimulated WNT11 promoter activity, whereas the knockdown of EGR1 expression by RNA interference reduced TNFα-induced WNT11 expression in T47D breast cancer cells. We also observed that mitogen-activated protein kinases (MAPK), extracellular signalregulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 kinase mediated TNFα-induced transcription of WNT11 via EGR1. Our results suggest that EGR1 directly targets WNT11 in response to TNFα stimulation in breast cancer cells. [BMB Reports 2020; 53(12): 628-633].

Leptin stimulates IGF-1 transcription by activating AP-1 in human breast cancer cells.

Leptin, an adipokine regulating energy metabolism, appears to be associated with breast cancer progression. Insulin-like growth factor-1 (IGF-1) mediates the pathogenesis of breast cancer. The regulation of IGF-1 expression by leptin in breast cancer cells is unclear. Here, we found that leptin upregulates IGF-1 expression at the transcriptional level in breast cancer cells. Activating protein-1 (AP-1)-binding element within the proximal region of IGF-1 was necessary for leptin-induced IGF-1 promoter activation. Forced expression of AP-1 components, c-FOS or c-JUN, enhanced leptin-induced IGF-1 expression, while knockdown of c-FOS or c-JUN abrogated leptin responsiveness. All three MAPKs (ERK1/2, JNK1/2, and p38 MAPK) mediated leptin-induced IGF-1 expression. These results suggest that leptin contributes to breast cancer progression through the transcriptional upregulation of leptin via the MAPK pathway. [BMB Reports 2019; 52(6): 385-390].

A synthetic chalcone derivative, 2-hydroxy-3',5,5'-trimethoxychalcone (DK-139), triggers reactive oxygen species-induced apoptosis independently of p53 in A549 lung cancer cells.

2-Hydroxy-3',5,5'-trimethoxychalcone (named DK-139) is a synthetic chalcone derivative that has anti-inflammatory, anti-tumor, and endoplasmic reticulum-mediated apoptosis activities. However, the mode of action of DK-139 on reactive oxygen species (ROS)-induced apoptosis remains unknown. In this study, we found that DK-139 activated DNA damage responses, as was revealed by the accumulation of the tumor suppressor p53 and the phosphorylation of histone H2AX at Ser139 (called γ-H2AX), which are hallmarks of DNA damage responses. The occurrence of DK-139-induced DNA damage was confirmed through single-cell gel electrophoresis (comet tail assay). Interestingly, using p53-null HCT116 cells revealed that p53 was not involved in DK-139-induced apoptosis. Instead, we found that DK-139 increased the production of ROS, which led to the processing of caspase-2, BH3 interacting-domain death agonist (BID), caspase-9, and caspase-7. Pretreatment with the ROS scavenger N-acetyl cysteine reduced the frequency of DK-139-induced γ-H2AX formation, demonstrating that DK-139 triggered DNA damage through ROS production. In addition, NAC pretreatment prevented DK-139-induced processing of caspase-2, BID, caspase-9, caspase-7, and poly(ADP-ribose) polymerase. These results suggest that DK-139 triggers apoptosis through ROS-mediated DNA damage and activation of the caspase-2 cascade in A549 human lung cancer cells.