Effect of Hybrid Compounds of Stilbene and Pentadienone on Inhibition of Tubulin Polymerization.

INTRODUCTION: Tubulin polymerization inhibitors induce cancer cell death; therefore, they can be developed as chemotherapeutic agents. We hypothesized that hybrid compounds, including the trans-stilbene moiety contained in resveratrol and penta-1,4-dien-3-one contained in curcumin, could inhibit tubulin polymerization. METHODS: Twenty-six hybrid stilbene and pentadienone compounds were designed and synthesized. The cytotoxicity of the hybrid compounds against MDA-MB-231 human breast cancer cells was determined using a clonogenic long-term survival assay. The relationship between cytotoxicity and structural properties was evaluated. Biological activities, including inhibition of tubulin polymerization and cell cycle progression, were investigated to select compounds with excellent anticancer properties. The molecular binding mode between the selected compound and the α, ß-tubulin dimers was investigated. RESULTS: Twenty-six hybrid stilbene and pentadienone compounds were designed and synthesized. Among them, compound 13 exhibited the highest inhibitory effect on the clonogenicity of MDA-MB-231 cells. Compound 13 induced the destabilization of tubulins and inhibited cell cycle progression at the G2/M phase. Through in silico molecular docking analysis, compound 13 was predicted to bind to the colchicine binding site of α, ß-tubulin. CONCLUSION: The stilbene and pentadienone hybrid compound 13 has a binding mode similar to that of colchicine. Compound 13 inhibited the clonogenicity of MDA-MB-231 cells through a mechanism that destabilizes tubulin polymerization, leading to cell cycle arrest at the G2/M phase.

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.

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.

Impairment of Glucose Metabolism and Suppression of Stemness in MCF-7/SC Human Breast Cancer Stem Cells by Nootkatone.

Targeting cancer stem cell metabolism has emerged as a promising therapeutic strategy for cancer treatment. Breast cancer stem cells (BCSCs) exert distinct metabolism machinery, which plays a major role in radiation and multidrug resistance. Therefore, exploring the mechanisms involved in energy utilization of BCSCs could improve the effectiveness of therapeutic strategies aimed at their elimination. This study was conducted to clarify the glucose metabolism machinery and the function of nootkatone, a bioactive component of grapefruit, in regulating glucose metabolism and stemness characteristics in human breast carcinoma MCF-7 stem cells (MCF-7SCs). In vivo experiments, transcriptomic analysis, seahorse XF analysis, MTT assay, Western blotting, mammosphere formation, wound healing, invasion assay, flow cytometric analysis, reverse transcription-quantitative polymerase chain reaction, and in silico docking experiments were performed. MCF-7SCs showed a greater tumorigenic capacity and distinct gene profile with enrichment of the genes involved in stemness and glycolysis signaling pathways compared to parental MCF-7 cells, indicating that MCF-7SCs use glycolysis rather than oxidative phosphorylation (OXPHOS) for their energy supply. Nootkatone impaired glucose metabolism through AMPK activation and reduced the stemness characteristics of MCF-7SCs. In silico docking analysis demonstrated that nootkatone efficiently bound to the active site of AMPK. Therefore, this study indicates that regulation of glucose metabolism through AMPK activation could be an attractive target for BCSCs.

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.

Odd-chain fatty acids as novel histone deacetylase 6 (HDAC6) inhibitors.

The dysregulation of histone deacetylases (HDACs) is closely associated with tumorigenesis and has emerged as a promising target for anti-cancer drugs. Some odd-chain fatty acids are present in trace levels in human tissue. Despite limited health benefits, there is increasing experimental evidence of nutritional benefits of odd-chain fatty acids. This study examines the effects of five odd-chain fatty acids (valeric, heptanoic, nonanoic, undecanoic, and pentadecanoic acid) as novel HDAC6 inhibitors. Examination of these fatty acids on the proliferation and clonogenic ability in various cancer cell lines revealed that pentadecanoic and undecanoic acid can strongly inhibit cancer cell proliferation. Heptanoic and nonanoic acid showed moderate anti-proliferative effects, while valeric acid demonstrated weak anti-proliferative effects. HDAC6 inhibitory activities were in the order of pentadecanoic acid (C15:0) > undecanoic acid (C11:0) > nonanoic acid (C9:0) > heptanoic acid (C7:0) > valeric acid (C5:0), consistent with the anti-proliferative assay results. All of these fatty acids promoted the acetylation of α-tubulin in MCF-7 breast and A549 lung cancer cells dose-dependently. In-silico molecular docking analysis showed that increasing the aliphatic carbon chain length facilitates binding to HDAC6 residues, which might be important for the inhibitory potential of HDAC6. This study shows the potential utility of odd-chain fatty acids for epigenetic-based cancer therapy.

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.

Inhibition of EGR-1-dependent MMP1 transcription by ethanol extract of Ageratum houstonianum in HaCaT keratinocytes.

Matrix metalloproteinase 1 (MMP-1) initiates the breakdown of matrix networks by cleaving fibrillar collagen during the pathophysiological progression of skin aging. Ageratum houstonianum ethanol extract (AHE) has been used as a traditional herbal medicine to treat external wounds and skin diseases. However, the mechanism of action underlying A. houstonianum-mediated modulation of skin aging has not been investigated. In this study, we evaluated the effect of AHE on MMP-1 expression in HaCaT keratinocytes. Gene expression was analyzed by Reverse transcription-PCR (RT-PCR), Quantitative real-time PCR (Q-PCR), gene promoter-reporter assay, and immunoblotting. We found that AHE abrogated TNFα-induced MMP1 expression at the transcriptional level via the suppression of ERK1/2 mitogen-activated protein kinase (MAPK)-mediated Early Growth Response 1 (EGR1) expression. We also demonstrated that ß-caryophyllene, a cannabinoid receptor 2 (CB2) agonist, is a functional component of the AHE that inhibits TNFα-induced EGR-1 and MMP1 expression. AHE exerts inhibitory activity on TNFα-induced MMP1 expression at the transcription level through EGR-1 downregulation in keratinocytes. ß-Caryophyllene is a bioactive ingredient of AHE that is responsible for the inhibition of TNFα-induced EGR1 expression. ß-Caryophyllene can be used as a potential agent to prevent inflammation-induced skin aging.

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.

Chrysin Inhibits NF-κB-Dependent CCL5 Transcription by Targeting IκB Kinase in the Atopic Dermatitis-Like Inflammatory Microenvironment.

Chrysin (5,7-dihydroxyflavone) is a natural polyphenolic compound that induces an anti-inflammatory response. In this study, we investigated the molecular mechanism underlying the chrysin-induced suppression of C-C motif chemokine ligand 5 (CCL5) gene expression in atopic dermatitis (AD)-like inflammatory microenvironment. We showed that chrysin inhibited CCL5 expression at the transcriptional level through the suppression of nuclear factor kappa B (NF-κB) in the inflammatory environment. Chrysin could bind to the ATP-binding pocket of the inhibitor of κB (IκB) kinase (IKK) and, subsequently, prevent IκB degradation and NF-κB activation. The clinical efficacy of chrysin in targeting IKK was evaluated in 2,4-dinitrochlorobenzene-induced skin lesions in BALB/c mice. Our results suggested that chrysin prevented CCL5 expression by targeting IKK to reduce the infiltration of mast cells to the inflammatory sites and at least partially attenuate the inflammatory responses. These findings suggested that chrysin might be useful as a platform for the design and synthesis of small-molecule IKK-targeting drugs for the treatment of chronic inflammatory diseases, such as AD.

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.

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].

Design, synthesis, and biological evaluation of polyphenols with 4,6-diphenylpyrimidin-2-amine derivatives for inhibition of Aurora kinase A.

BACKGROUND: Several 4,6-diarylpyrimidin-2-amine derivatives show anticancer properties. However, their mode of action is not fully characterized. To develop potent anticancer chemotherapeutic agents, we designed and synthesized 25 4,6-diphenylpyrimidin-2-amine derivatives containing a guanidine moiety. METHODS: Clonogenic long-term survival assays were performed to screen anticancer compounds. To derive the structural conditions showing good cytotoxicities against cancer cells, quantitative structure-activity relationships (QSAR) were calculated. Biological activities were determined by flow cytometry for cell cycle analysis and by immunoblot analysis for the detection of Aurora kinase A (AURKA) activity. Because 2-(2-Amino-6-(2,4-dimethoxyphenyl)pyrimidin-4-yl) phenol (derivative 12) selectively inhibited AURKA activity from the kinome assay, in silico docking experiments were performed to elucidate the molecular binding mode between derivative 12 and AURKA. RESULTS: The pharmacophores were derived based on the QSAR calculations. Derivative 12 inhibited AURKA activity and reduced phosphorylation of AURKA at Thr283 in HCT116 human colon cancer cells. Derivative 12 caused the accumulation of the G2/M phase of the cell cycle and triggered the cleavages of caspase-3, caspase -7, and poly(ADP-ribose) polymerase. The binding energies of 30 apo-AURKA - derivative 12 complexes obtained from in silico docking ranged from -16.72 to -11.63 kcal/mol. CONCLUSIONS: Derivative 12 is an AURKA inhibitor, which reduces clonogenicity, arrests the cell cycle at the G2/M phase, and induces caspase-mediated apoptotic cell death in HCT116 human colon cancer cells. In silico docking demonstrated that derivative 12 binds to AURKA well. The structure-activity relationship calculations showed hydrophobic substituents and 1-naphthalenyl group at the R2 position increased the activity. The existence of an H-bond acceptor at C-2 of the R1 position increased the activity, too. Graphical abstract Derivative 12 inhibits Aurora kinase A activity and causes the G2/M phase arrest of the cell cycle.

Inhibition of TNFα-induced interleukin-6 gene expression by barley (Hordeum vulgare) ethanol extract in BV-2 microglia.

BACKGROUND: Inflammation in the central nervous system is closely associated with pathological neurodegenerative diseases as well as psychiatric disorders. Prolonged activation of microglia can produce many inflammatory mediators, which may result in pathological neurotoxic side effects. Interleukin (IL)-6 serves as a hallmark of the injured brain. OBJECTIVE: Whole grains are known to contain many bioactive components. However, little information is available about anti-neuroinflammatory effects of grains in the CNS. This study aims to investigate the effect of Hordeum vulgare ethanol extract (HVE) on the suppression of IL-6 expression in BV2 microglia. METHODS: Inhibitory effects of HVE on IL-6 expression were analyzed by immunoblot anaysis, immunofluoresce microscopic analysis, reverse transcription-polymerase chain reaction, and luciferase promoter reporter assay. RESULTS: HVE inhibited TNFα-induced phosphorylation of IKKα/ß, IκB, and p65/RelA NF-κB. TNFα-induced IL-6 mRNA expression and promoter activity were reduced by HVE. Point mutation of NF-κB-binding site within the IL-6 gene promoter abolished TNFα-induced reporter activity, whereas exogenous expression of p65 NF-κB enhanced IL-6 promoter activity. CONCLUSION: NF-κB-binding site within the IL-6 promoter region is a HVE target element involved in the inhibition of TNFα-induced IL-6 gene transcription. HVE inhibits TNFα-induced IL-6 expression via suppression of NF-κB signaling in BV2 microglial cells.

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.

Design, synthesis, and biological activities of 1-aryl-(3-(2-styryl)phenyl)prop-2-en-1-ones.

A moderate elevation in reactive oxygen species (ROS) levels can generally be controlled in normal cells, but may lead to death of cancer cells as the ROS level in cancer cells is already elevated. Therefore, a ROS-generating compound can act as a selective chemotherapeutic agent for cancer cells that does not affect normal cells. In our previous study, a compound containing a Michael acceptor was selectively cytotoxic to cancer cells without affecting normal cells; therefore, we designed and synthesized 26 compounds containing a Michael acceptor. Their cytotoxicities against HCT116 human colon cancer cell lines were measured by using a clonogenic long-term survival assay. To derive the structural conditions required to obtain stronger cytotoxicity against cancer cells, the relationships between the half-maximal cell growth inhibitory concentration values of the synthesized compounds and their physicochemical properties were evaluated by Comparative Molecular Field Analysis and Comparative Molecular Similarity Indices Analysis. It was confirmed that the compound with the best half-maximal cell growth inhibitory concentration triggered apoptosis through ROS generation, which then led to stimulation of the caspase pathway.

Gomisin G Suppresses the Growth of Colon Cancer Cells by Attenuation of AKT Phosphorylation and Arrest of Cell Cycle Progression.

Colorectal cancer is one of the leading causes of cancer related death due to a poor prognosis. In this study, we investigated the effect of Gomisin G on colon cancer growth and examined the underlying mechanism of action. We found that Gomisin G significantly suppressed the viability and colony formation of LoVo cells. Gomisin G reduced the phosphorylation level of AKT implying that Gomisin G suppressed the PI3K-AKT signaling pathway. Gomisin G also induced apoptosis shown by Annexin V staining and an increased level of cleaved poly-ADP ribose polymerase (PARP) and Caspase-3 proteins. Furthermore, Gomisin G remarkably triggered the accumulation of cells at the sub-G1 phase which represents apoptotic cells. In addition, the level of cyclin D1 and phosphorylated retinoblastoma tumor suppressor protein (Rb) was also reduced by the treatment with Gomisin G thus curtailing cell cycle progression. These findings show the suppressive effect of Gomisin G by inhibiting proliferation and inducing apoptosis in LoVo cells. Taken together, these results suggest Gomisin G could be developed as a potential therapeutic compound against colon cancer.

Inhibitory Effect of Synthetic Flavone Derivatives on Pan-Aurora Kinases: Induction of G2/M Cell-Cycle Arrest and Apoptosis in HCT116 Human Colon Cancer Cells.

Members of the aurora kinase family are Ser/Thr kinases involved in regulating mitosis. Multiple promising clinical trials to target aurora kinases are in development. To discover flavones showing growth inhibitory effects on cancer cells, 36 flavone derivatives were prepared, and their cytotoxicity was measured using a long-term clonogenic survival assay. Their half-maximal growth inhibitory effects against HCT116 human colon cancer cells were observed at the sub-micromolar level. Pharmacophores were derived based on three-dimensional quantitative structure⁻activity calculations. Because plant-derived flavones inhibit aurora kinase B, we selected 5-methoxy-2-(2-methoxynaphthalen-1-yl)-4H-chromen-4-one (derivative 31), which showed the best half-maximal cell growth inhibitory effect, and tested whether it can inhibit aurora kinases in HCT116 colon cancer cells. We found that derivative 31 inhibited the phosphorylation of aurora kinases A, aurora kinases B and aurora kinases C, suggesting that derivative 31 is a potential pan-aurora kinase inhibitor. The results of our analysis of the binding modes between derivative 31 and aurora A and aurora B kinases using in-silico docking were consistent with the pharmacophores proposed in this study.