naphthoquinones and Stomach-Neoplasms

Napabucasin is a novel oral first-in-class cancer stemness inhibitor. Preclinical and early phase clinical trials showed promising antitumor efficacy signals for napabucasin in a variety of malignancies. In this article, we describe the design and rationale for the now completed BRIGHTER trial, a multicenter, randomized, placebo-controlled, Phase III study designed to determine the efficacy and safety of combining napabucasin with paclitaxel in previously treated patients with advanced gastric and gastroesophageal junction adenocarcinoma (NCT02178956). Patients were randomized in a 1:1 fashion to receive weekly paclitaxel with either napabucasin or placebo. The study failed to achieve its primary end point of overall survival in the intention to treat population. Ongoing analysis of the secondary end points includes progression-free survival, objective response rate, disease control rate, the safety of the combination therapy and evaluation of efficacy in the biomarker-positive subpopulation.

Topics: Adenocarcinoma; Antineoplastic Combined Chemotherapy Protocols; Benzofurans; Clinical Protocols; Disease-Free Survival; Double-Blind Method; Drug Administration Schedule; Esophageal Neoplasms; Humans; Immunohistochemistry; Kaplan-Meier Estimate; Naphthoquinones; Paclitaxel; Stomach Neoplasms; Survival Analysis; Treatment Outcome

Signal transducer and activator of transcription 3 (STAT3) is a key regulator of many human cancers and has been widely recognized as a promising target for cancer therapy. A variety of small-molecule inhibitors have been developed for targeting STAT3, and some of them are now undergoing clinical trials. S3I-201, a known STAT3 inhibitor, may block STAT3 function in cancer cells by binding to the STAT3 SH2 domain to disrupt STAT3 protein complex formation. Using S3I-201 as a starting point for drug development, we synthesized a series of new STAT3 inhibitors 9a-x in this study by introducing naphthoquinone unit, a privileged fragment in STAT3 inhibitors. Most of the compounds exhibited strong anti-proliferation activity of gastric cancer cells (MGC803, MKN28, MNK1, and AGS). The representative compound 9n (SIL-14) could effectively inhibit the colony formation and migration of gastric cancer cells MGC803, arrest the cell cycle and induce MGC803 cell apoptosis at low micromolar concentrations in vitro. In addition, SIL-14 can also inhibit the phosphorylation of STAT3 protein and significantly decrease the expression of total STAT3, suggesting that it may exert anticancer effects by blocking the STAT3 signaling pathway. These results support that SIL-14 may be a promising STAT3 inhibitor for the further development of potential anti-gastric cancer candidates.

Topics: Aminosalicylic Acids; Benzenesulfonates; Cell Line, Tumor; Cell Proliferation; Humans; Naphthoquinones; STAT3 Transcription Factor; Stomach Neoplasms

Gastric cancer is the third leading cause of cancer-related death worldwide. To evaluate the anticancer potential and molecular mechanism of biflorin, a prenyl-ortho-naphthoquinone obtained from Capraria biflora L. roots, we used ACP02, a gastric cancer cell line established from a primary diffuse gastric adenocarcinoma. In this study, biflorin was shown to be a potent cytotoxic agent against ACP02 by Alamar Blue and Trypan Blue assays. Morphological analysis indicated cell death with features of necrosis. Furthermore, a decrease in colony formation, migration and invasion of ACP02 cells was observed after treatment with biflorin (1.0, 2.5 and 5.0 μM). Regarding the underlying molecular mechanism of biflorin in ACP02 cells, we observed a decrease in MYC expression and telomere length using FISH. Our findings suggest a novel molecular target of biflorin in ACP02 cells, which may be a significant therapeutic approach for gastric cancer management.

Topics: Antineoplastic Agents, Phytogenic; Cell Line; Cell Movement; Cell Proliferation; Cell Survival; Humans; Naphthoquinones; Proto-Oncogene Proteins c-myc; Stomach Neoplasms

The 1,4-naphthoquinones and their derivatives have garnered great interest due to their antitumor pharmacological properties in various cancers; however, their clinical application is limited by side effects. In this study, to reduce side effects and improve therapeutic efficacy, a novel 1,4-naphthoquinone derivative-2-(4-methoxyphenylthio)-5,8-dimethoxy-1,4-naphthoquinone (MPTDMNQ) was synthesized. We investigated the effects and underlying mechanisms of MPTDMNQ on cell viability, apoptosis, and reactive oxygen species (ROS) generation in human gastric cancer cells. Our results showed that MPTDMNQ decreased cell viability in nine human gastric cancer cell lines. MPTDMNQ significantly induced apoptosis accompanied by the accumulation of ROS in GC cells. However, pre-treatment with the ROS scavenger N-acetyl-L-cysteine (NAC) attenuated the MPTDMNQ-induced apoptosis. Moreover, MPTDMNQ decreased the phosphorylation levels of extracellular signal-regulated kinase (ERK) and signal transducer and activator of transcription 3 (STAT3); and increased the phosphorylation levels of c-Jun N-terminal kinase (JNK) and p38 kinase. However, phosphorylation was inhibited by NAC and a mitogen-activated protein kinase (MAPK) inhibitor. These findings showed that MPTDMNQ induced AGS cell apoptosis via ROS-mediated MAPK and STAT3 signaling pathways. Thus, MPTDMNQ may be a promising candidate for treating gastric cancer.

Topics: Apoptosis; Cell Line, Tumor; Cell Proliferation; Cell Survival; Humans; Mitogen-Activated Protein Kinases; Naphthoquinones; Reactive Oxygen Species; Signal Transduction; STAT3 Transcription Factor; Stomach Neoplasms

1,4‑Naphthoquinone derivatives have superior anticancer effects, but their use has been severely limited in clinical practice due to adverse side effects. To reduce the side effects and extend the anticancer effects of 1,4‑naphthoquinone derivatives, 2‑(butane‑1‑sulfinyl)‑1,4‑naphthoquinone (BQ) and 2‑(octane‑1‑sulfinyl)‑1,4‑naphthoquinone (OQ) were synthesized, and their anticancer activities were investigated. The anti‑proliferation effects, determined by MTT assays, showed that BQ and OQ significantly inhibited the viability of gastric cancer cells and had no significant cytotoxic effect on normal cell lines. The apoptotic effect was determined by flow cytometry, and the results showed that BQ and OQ induced cell apoptosis by regulating the mitochondrial pathway and cell cycle arrest at the G2/M phase via inhibition of the Akt signaling pathway in AGS cells. Furthermore, BQ and OQ significantly increased the levels of reactive oxygen species (ROS) and this effect was blocked by the ROS scavenger NAC in AGS cells. BQ and OQ induced apoptosis by upregulating the protein expression of p38 and JNK and downregulating the levels of ERK and STAT3. Furthermore, expression levels of these proteins were also blocked after NAC treatment. These results demonstrated that BQ and OQ induced apoptosis and cell cycle arrest at the G2/M phase in AGS cells by stimulating ROS generation, which caused subsequent activation of MAPK, Akt and STAT3 signaling pathways. Thus, BQ and OQ may serve as potential therapeutic agents for the treatment of human gastric cancer.

Topics: Antineoplastic Agents; Apoptosis; Cell Cycle Checkpoints; Cell Line, Tumor; Humans; MAP Kinase Signaling System; Naphthoquinones; Proto-Oncogene Proteins c-akt; Reactive Oxygen Species; Signal Transduction; STAT3 Transcription Factor; Stomach Neoplasms

Objective To investigate the effect and molecular mechanism of β-lapachone combined with NVP-BEZ235 on the proliferation and migration of BGC-823 human gastric cancer cells. Methods BGC-823 cells were randomly divided into four groups: control group, 1 μmol/L β-lapachone group, 50 nmol/L NVP-BEZ235 group and 1 μmol/L β-lapachone combined with 50 nmol/L NVP-BEZ235 group. The proliferation of cells was determined using the MTT assay and colony formation assay. The expression levels of proliferation-related proteins phosphorylated AKT (p-AKT), phosphorylated NF-κB (p-NF-κB), phosphorylated extracellular signal-regulated kinase (p-ERK) and cyclin D1 were detected by Western blotting. The migration of cells was measured by wound healing assay and Transwell

Topics: Antineoplastic Agents; Apoptosis; beta Catenin; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D1; Humans; Imidazoles; Naphthoquinones; NF-kappa B; Phosphatidylinositol 3-Kinases; Phosphorylation; Proto-Oncogene Proteins c-akt; Quinolines; Signal Transduction; Stomach Neoplasms; TOR Serine-Threonine Kinases

The naphthopyranones paepalantine and 5-methoxy-3,4-dehydroxanthomegnin isolated from Paepalanthus sp, showed in previous studies antioxidant, anti-inflammatory, antitumour and antimicrobial potential, such as anti-Helicobacter pylori activity. H. pylori infection is one of the main causes of gastric cancer, causing an excessive inflammatory response through the neutrophils and macrophages infiltration, increasing the release of reactive species and thus inducing the production of pro-inflammatory mediators. In the present study, immunomodulatory activity of naphthopyranones in LPS-stimulated macrophages and cytotoxic action in gastric adenocarcinoma cell lines was evaluated. The potential of interaction of these substances in the iNOS binding site was investigated by molecular docking. Cytotoxic activity in gastric adenocarcinoma cells (AGS) was evaluated by the MTT assay. The results evidenced immunomodulatory potential by inhibiting the pro-inflammatory cytokines and nitric oxide produced by LPS-stimulated macrophages. Cytotoxic activity in AGS cell line was also reported. The results indicated that the studied naphthopyranones are viable alternatives in the treatment and prevention of H. pylori infection as well as the diseases related to this infection, especially gastric cancer.

Topics: Adenocarcinoma; Animals; Antineoplastic Agents, Phytogenic; Cell Line, Tumor; Computer Simulation; Cytokines; Eriocaulaceae; Helicobacter Infections; Helicobacter pylori; Humans; Immunologic Factors; Isocoumarins; Lipopolysaccharides; Macrophages; Mice; Molecular Docking Simulation; Naphthoquinones; Nitric Oxide; Nitric Oxide Synthase Type II; RAW 264.7 Cells; Stomach Neoplasms

Shift metabolism profile from mitochondrial oxidative phosphorylation to aerobic glycolysis (Warburg effect) is a key for tumor cell growth and metastasis. Therefore, suppressing the tumor aerobic glycolysis shows a great promise in anti-tumor therapy. In the present study, we study the role of shikonin, a naphthoquinone isolated from the traditional Chinese medicine Lithospermum, in inhibiting tumor aerobic glycolysis and thus tumor growth. We found that shikonin dose-dependently inhibited glucose uptake and lactate production in Lewis lung carcinoma (LLC) and B16 melanoma cells, confirming the inhibitory effect of shikonin on tumor aerobic glycolysis. Treatment of shikonin also decreased tumor cell ATP production. Furthermore, pyruvate kinase M2 (PKM2) inhibitor or activator respectively altered the effect of shikonin on tumor cell aerobic glycolysis, suggesting that suppression of cell aerobic glycolysis by shikonin is through decreasing PKM2 activity. Western blot analysis confirmed that shikonin treatment reduced tumor cell PKM2 phosphorylation though did not reduce total cellular PKM2 level. In vitro assay also showed that shikonin treatment significantly promoted tumor cell apoptosis compared to untreated control cells. Finally, when mice implanted with B16 cells were administered with shikonin or control vehicle, only shikonin treatment significantly decreased B16 tumor cell growth. In conclusion, this study demonstrates that shikonin inhibits tumor growth in mice by suppressing PKM2-mediated aerobic glycolysis.

Topics: Adenosine Triphosphate; Aerobiosis; Animals; Antineoplastic Agents, Phytogenic; Apoptosis; Cell Line, Tumor; Drug Screening Assays, Antitumor; Drugs, Chinese Herbal; Enzyme Inhibitors; Glycolysis; Male; Melanoma, Experimental; Mice; Mice, Nude; Mice, SCID; Molecular Targeted Therapy; Naphthoquinones; Neoplasm Proteins; Phosphorylation; Protein Processing, Post-Translational; Pyruvate Kinase; Stomach Neoplasms

Gastric cancer displays marked molecular heterogeneity with aggressive behavior and treatment resistance. Therefore, good in vitro models that encompass unique subtypes are urgently needed for precision medicine development. Here, we have established a primary gastric cancer organoid (GCO) biobank that comprises normal, dysplastic, cancer, and lymph node metastases (n = 63) from 34 patients, including detailed whole-exome and transcriptome analysis. The cohort encompasses most known molecular subtypes (including EBV, MSI, intestinal/CIN, and diffuse/GS, with CLDN18-ARHGAP6 or CTNND1-ARHGAP26 fusions or RHOA mutations), capturing regional heterogeneity and subclonal architecture, while their morphology, transcriptome, and genomic profiles remain closely similar to in vivo tumors, even after long-term culture. Large-scale drug screening revealed sensitivity to unexpected drugs that were recently approved or in clinical trials, including Napabucasin, Abemaciclib, and the ATR inhibitor VE-822. Overall, this new GCO biobank, with linked genomic data, provides a useful resource for studying both cancer cell biology and precision cancer therapy.

Topics: Aminopyridines; Antineoplastic Agents; Benzimidazoles; Benzofurans; Biological Specimen Banks; Cell Proliferation; Drug Screening Assays, Antitumor; Female; Humans; Isoxazoles; Male; Naphthoquinones; Organoids; Precision Medicine; Pyrazines; Stomach Neoplasms

Gastroenteropancreatic neuroendocrine neoplasms (GEP-NEN) represent a rare and heterogenous tumor entity. Importantly, the highly proliferative subgroup of neuroendocrine carcinoma (GEP-NEC) is characterized by high resistance to conventional chemotherapy. Consequently, there is an urgent need to identify novel therapeutic targets, especially for GEP-NEC. Thus, we focused on Inhibitor of apoptosis protein (IAP) family members survivin and XIAP that orchestrate inhibition of apoptosis, induce resistance against chemotherapeutics and facilitate tumor metastasis. Copy number gains (CNGs) could be detected by microarray comparative genomic hybridization for survivin and XIAP in 60 % and 26.7 % of all GEP-NENs, respectively. Immunohistochemical staining of tissue specimens from 77 consecutive patients with GEP-NEN demonstrated increased survivin protein expression levels in tissue specimens of highly proliferative GEP-NEC or GEP-NEN located in the stomach and colon. In contrast, XIAP overexpression was associated with advanced tumor stages. Knockdown of survivin and XIAP markedly reduced cell proliferation and tumor growth. In vitro, YM155 induced apoptotic cell death accompanied by a reduction in cell proliferation and inhibited GEP-NEC xenograft growth. Taken together, our data provide evidence for a biological relevance of these IAPs in GEP-NEN and support a potential role of survivin as therapeutic target especially in the subgroup of aggressive GEP-NEC.

Topics: Animals; Antineoplastic Agents; Apoptosis; Biomarkers, Tumor; Carcinoma, Neuroendocrine; Cell Line, Tumor; Cell Proliferation; Comparative Genomic Hybridization; DNA Copy Number Variations; Dose-Response Relationship, Drug; Female; Gene Dosage; Gene Expression Regulation, Neoplastic; Humans; Imidazoles; Immunohistochemistry; Inhibitor of Apoptosis Proteins; Intestinal Neoplasms; Male; Masoprocol; Mice, Inbred NOD; Mice, SCID; Molecular Targeted Therapy; Naphthoquinones; Oligonucleotide Array Sequence Analysis; Pancreatic Neoplasms; Retrospective Studies; RNA Interference; Signal Transduction; Stomach Neoplasms; Survivin; Time Factors; Transfection; Tumor Burden; X-Linked Inhibitor of Apoptosis Protein; Xenograft Model Antitumor Assays

Context • Gastric cancer (GC) is the fourth most common cancer and the second leading cause of cancer-related deaths in the world. The current treatments include surgery and chemotherapy, either alone or in combination with radiotherapy, but the prognosis for patients with GC is usually poor. A safe and effective chemopreventive treatment for this malignant disease is urgently needed. Objective • The study intended to investigate the effects and underlying mechanisms of plumbagin, a quinonoid constituent that is derived from the roots of the medicinal plant Plumbago zeylanica, which exhibits potent anticancer properties against a number of cancers. Design • The in vitro study used the human GC cell line SGC-7901. Setting • All experiments were conducted at the Hubei University of Chinese Medicine and Tongji Medical College, Huazhong University of Science and Technology (Wuhan, China). Intervention • SGC-7901 cells were cultured in 30-mm dishes and treated with plumbagin at concentrations of 0, 5, 10, to 20 μmol/L. The cells were incubated with 10 μmol/L plumbagin for different amounts of time (0, 2, 4, 8, 12, and 24 h) in contact with the cancer cells. Outcome Measures • The cell viability was examined using a cell counting kit-8 viability assay, and the cell proliferation rate was determined using a 5-ethynyl-2'-deoxyuridine incorporation assay. The cell cycle distribution was assessed by flow cytometry using propidium iodide staining, and Western blotting was used to assess the expression of BAX, BCL-2, and caspase-3 and to identify any downregulation in the activation of transcription 3 (STAT3), protein kinase B (Akt), and extracellular signal-regulated kinase (ERK1/2). Results • The plumbagin concentrations of 5-20 mmol/L reduced the viability of the GC cells in a dependent manner. Plumbagin suppressed the expression of BAX, BCL-2, pro-caspase-3, and cleaved-caspase-3. It also restrained the expression and phosphorylation of STAT3 and decreased the phosphorylation of Akt1 but did not change the total protein or phosphorylation levels of ERK1/2. Conclusions • Plumbagin inhibits cell apoptosis in human GC cells, and that effect may be related with its ability to suppress phosphorylation of STAT3 and Akt. Given those 2 effects, plumbagin may be a promising agent in the treatment of gastric cancer.

Topics: Antineoplastic Agents, Phytogenic; Apoptosis; Caspase 3; Cell Line, Tumor; Cell Proliferation; Humans; Naphthoquinones; Phytotherapy; Plumbaginaceae; Signal Transduction; STAT3 Transcription Factor; Stomach Neoplasms

Survivin overexpression is associated with poor prognosis of human gastric cancer, and is a target for gastric cancer therapy. YM155 is originally identified as a specific inhibitor of survivin. In this study, we investigated the antitumor effect of YM155 on human gastric cancer. Our results showed that YM155 treatment significantly inhibited cell proliferation, reduced colony formation and induced apoptosis of gastric cancer cells in a dose-dependent manner. Accordingly, YM155 treatment significantly decreased survivin expression without affecting XIAP expression and increased the cleavage of apoptosis-associated proteins caspase 3, 7, 8, 9. YM155 significantly inhibited sphere formation of gastric cancer cells, suppressed expansion and growth of the formed spheres (cancer stem cell-like cells, CSCs) and downregulated the protein levels of β-catenin, c-Myc, Cyclin D1 and CD44 in gastric cancer cells. YM155 infusion at 5 mg/kg/day for 7 days markedly inhibited growth of gastric cancer xenograft in a nude mouse model. Immunohistochemistry staining and Western Blot showed that YM155 treatment inhibited expression of survivin and CD44, induced apoptosis and reduced CD44+ CSCs in xenograft tumor tissues in vivo. No obvious pathological changes were observed in organs (e.g. heart, liver, lung and kidney) in YM155-treated mice. Our results demonstrated that YM155 inhibits cell proliferation, induces cell apoptosis, reduces cancer stem cell expansion, and inhibits xenograft tumor growth in gastric cancer cells. Our results elucidate a new mechanism by which YM155 inhibits gastric cancer growth by inhibition of CSCs. YM155 may be a promising agent for gastric cancer treatment.

Topics: Animals; Apoptosis; Blotting, Western; Cell Proliferation; Female; Gastric Mucosa; Humans; Imidazoles; Immunoenzyme Techniques; Inhibitor of Apoptosis Proteins; Mice; Mice, Inbred BALB C; Mice, Nude; Naphthoquinones; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Stomach; Stomach Neoplasms; Survivin; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

The prognosis of gastric cancer remains poor due to clinical drug resistance. Novel drugs are urgently needed. Shikonin (SHK), a natural naphthoquinone, has been reported to trigger cell death and overcome drug resistance in anti-tumour therapy. In this study, we investigated the effectiveness and molecular mechanisms of SHK in treatment with gastric cancer. In vitro, SHK suppresses proliferation and triggers cell death of gastric cancer cells but leads minor damage to gastric epithelial cells. SHK induces the generation of intracellular reactive oxygen species (ROS), depolarizes the mitochondrial membrane potential (MMP) and ultimately triggers mitochondria-mediated apoptosis. We confirmed that SHK induces apoptosis of gastric cancer cells not only in a caspase-dependent manner which releases Cytochrome C and triggers the caspase cascade, but also in a caspase-independent manner which mediates the nuclear translocation of apoptosis-inducing factor and Endonuclease G. Furthermore, we demonstrated that SHK enhanced the chemotherapeutic sensitivity of 5-fluorouracil and oxaliplatin in vitro and in vivo. Taken together, our data show that SHK may be a novel therapeutic agent in the clinical treatment of gastric cancer.

Topics: Apoptosis; Cell Line, Tumor; Fluorouracil; Humans; Mitochondria; Naphthoquinones; Organoplatinum Compounds; Pyridines; Reactive Oxygen Species; Stomach Neoplasms

A recent study reported that plumbagin downregulated the activity of Janus kinase 2 (JAK2)-signal transducer and activator of transcription 3 (STAT3) pathway to show various antitumor effects in multiple myeloma cells. We aimed in this in vitro study to demonstrate the inhibition of JAK2/STAT3 pathway by plumbagin through inducing SH2-containing protein tyrosine phosphatase 1 (SHP1) expression in the MKN-28 gastric cancer cell line. We performed western blot analysis to measure SHP1, phosphor-JAK2/STAT3 level, and observed that plumbagin induced SHP1 expression and simultaneously downregulated phosphor-JAK2/STAT3 in MKN-28 cells, with negative SHP1 expression. This effect was consistent when JAK2/STAT3 signaling was activated by interleukin-6 (IL-6), and ameliorated when cells were treated with prevanadate, a protein tyrosin phosphatase inhibitor. Furthermore, plumbagin significantly reduced gene expression of cyclin D1, vascular endothelial growth factor (VEGF)-1, Bcl-xL, survivin and matrix metalloproteinase-9 (MMP-9), known target products of STAT3 activation in gastric carcinogenesis by reverse transcription-polymerase chain reaction (RT-PCR). Several functional studies such as water soluble tetrazolium salt-1 (WST-1) assay, wound closure assay, Matrigel invasion assay and Annexin V assay were also performed, and we validated the functional effect of plumbagin for inhibition of cell proliferation, migration and invasion, and induction of apoptosis. Collectively, our findings suggest that plumbagin is a potential regulator of cellular growth, migration, invasion and apoptosis by inhibiting both constitutive and inducible STAT3 activity through induction of SHP1 in gastric cancer cells.

Topics: Antineoplastic Agents, Phytogenic; Cell Line, Tumor; Gene Expression Regulation, Neoplastic; Humans; Janus Kinase 2; Naphthoquinones; Phosphorylation; Protein Tyrosine Phosphatase, Non-Receptor Type 6; Signal Transduction; STAT3 Transcription Factor; Stomach Neoplasms

Shikonin is an active naphthoquinone pigment isolated from the root of Lithospermum erythrorhizon. This study was designed to explore the inhibition of Shikonin on cell viability, adhesion, migration and invasion ability of gastric cancer (GC) and its possible mechanism.. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was performed for cell viability and adhesion ability of MGC-803 cells. Cell scratch repair experiments were conducted for the determination of migration ability while transwell assay for cell invasion ability. Western blot analysis and real-time polymerase chain reaction assay were used for the detection of protein and mRNA expressions.. Fifty per cent inhibitory concentration of Shikonin on MGC-803 cells was 1.854 μm. Shikonin (1 μm) inhibited significantly the adhesion, invasion and migratory ability of MGC-803 cells. Interestingly, Shikonin in the presence or absence of anti-Toll-like receptor 2 (TLR2) antibody (2 μg) and nuclear factor-kappa B (NF-κB) inhibitor MG-132 (10 μm) could decrease these ability of MGC-803 cells markedly, as well as the expression levels of matrix metalloproteinases (MMP)-2, MMP-7, TLR2 and p65 NF-κB. In addition, the co-incubation of Shikonin and anti-TLR2/MG-132 has a significant stronger activity than anti-TLR2 or MG-132 alone.. The results indicated that Shikonin could suppress the cell viability, adhesion, invasion and migratory ability of MGC-803 cells through TLR2- or NF-κB-mediated pathway. Our findings provide novel information for the treatment of Shikonin on GC.

Topics: Antineoplastic Agents; Cell Adhesion; Cell Line, Tumor; Cell Survival; Humans; Inhibitory Concentration 50; Lithospermum; Matrix Metalloproteinase 2; Matrix Metalloproteinase 7; Naphthoquinones; NF-kappa B; Real-Time Polymerase Chain Reaction; Stomach Neoplasms; Toll-Like Receptor 2

ThyX is an essential thymidylate synthase that is mechanistically and structurally unrelated to the functionally analogous human enzyme, thus providing means for selective inhibition of bacterial growth. To identify novel compounds with anti-bacterial activity against the human pathogenic bacterium Helicobacter pylori, based on our earlier biochemical and structural analyses, we designed a series of eighteen 2-hydroxy-1,4-naphthoquinones (2-OH-1,4-NQs) that target HpThyX. Our lead-like molecules markedly inhibited the NADPH oxidation and 2'-deoxythymidine-5'-monophosphate-forming activities of HpThyX enzyme in vitro, with inhibitory constants in the low nanomolar range. The identification of non-cytotoxic and non-mitotoxic 2-OH-1,4-NQ inhibitors permitted testing their in vivo efficacy in a mouse model for H. pylori infections. Despite the widely assumed toxicity of naphthoquinones (NQs), we identified tight-binding ThyX inhibitors that were tolerated in mice and can be associated with a modest effect in reducing the number of colonizing bacteria. Our results thus provide proof-of-concept that targeting ThyX enzymes is a highly feasible strategy for the development of therapies against H. pylori and a high number of other ThyX-dependent pathogenic bacteria. We also demonstrate that chemical reactivity of NQs does not prevent their exploitation as anti-microbial compounds, particularly when mitotoxicity screening is used to prioritize these compounds for further experimentation.

Topics: Animals; Apoptosis; Blotting, Western; Cell Proliferation; Enzyme Inhibitors; Flow Cytometry; Helicobacter Infections; Helicobacter pylori; Humans; Mice; Mitosis; Naphthoquinones; Stomach Neoplasms; Thymidylate Synthase; Tumor Cells, Cultured

Lithospermum erythrorhizon, a naphthoquinone compound derived from a shikonin, has long been used as traditional Chinese medicine for treatment of various diseases, including cancer. To evaluate the cytotoxic effects of shikonin on AGS gastric cancer cells via induction of cell cycle arrest.. We observed the effects of 12.5-100 ng/mL dosage of shikonin treatment on AGS cancer cell line with the incubation time of 6h. Cytotoxic effects were assessed by measuring the changes in the intracellular ROS, appearance of senescence phenotype, cell cycle progression, CDK and cyclins expression levels upon shikonin treatment. We also examined upon the activation of Egr1-mediated p21 expression, by siRNA transfection, Luciferase assay, and ChIP assay.. In this study, we found that shikonin inhibits cell proliferation by arresting cell cycle progression at the G2/M phase via modulation of p21 in AGS cells. Also, our results revealed that the p21 gene was transactivated by early growth response1 (Egr1) in response to the shikonin treatment. Transient Egr1 expression enhanced shikonin-induced p21 promoter activity, whereas the suppression of Egr1 expression by small interfering RNA attenuated the ability of shikonin to induce p21 promoter activity.. Our results suggested that the anti-proliferative activity of shikonin was due to its ability to induce cell cycle arrest via Egr1-p21 signaling pathway. Thus, the work stated here validates the traditional use of shikonin in the treatment of cancer.

Topics: Antineoplastic Agents; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Proliferation; Cyclin-Dependent Kinase Inhibitor p21; Early Growth Response Protein 1; Gene Expression Regulation, Neoplastic; Humans; Naphthoquinones; RNA, Small Interfering; Stomach Neoplasms

The effects of shikonin on gastric cancer cells were investigated in this study. Exposure to shikonin reduced the viability of gastric cancer cells in a time- and dose-dependent manner. However, apoptosis was not observed in gastric cancer cell treatment with different concentrations of shikonin for 6h. By contrast, treatment with shikonin for 24h significantly induced apoptosis, as evidenced by the results of TUNEL assay and flow cytometry analysis in proportion to the concentration. Disruption of the mitochondrial membrane potential was observed in gastric cancer cells that were treated with shikonin for 6 and 24h. Pretreatment with necrostatin-1 recovered cell death and mitochondrial membrane potential in the 6h shikonin treatment, but not in the 24h shikonin treatment. Western blot results reveal enhanced p38 phosphorylation, downregulated AKT phosphorylation, and increased caspase3 and PARP cleavage in cells that were treated with shikonin for 24h, but not in cells treated for 6h. Shikonin also triggered reactive oxygen species (ROS) generation both in the 6 and 24h treatments. Pretreatment with N-acetylcysteine blocked shikonin-induced cell death. In summary, our findings suggest that shikonin, which may function as a promising agent in the treatment of gastric cancers, sequentially triggered necrosis or apoptosis through ROS generation in gastric cancer cells.

Topics: Acetylcysteine; Antineoplastic Agents; Apoptosis; Blotting, Western; Cell Line, Tumor; Cell Survival; Flow Cytometry; Free Radical Scavengers; Humans; Membrane Potential, Mitochondrial; Mitochondria; Naphthoquinones; Necrosis; Phosphorylation; Reactive Oxygen Species; Stomach Neoplasms; Time Factors

Gastric cancer is one of the most common malignant tumors and the second cause of cancer-related deaths worldwide. Naphthoquinones such as juglone and plumbagin are compounds used extensively to overcome resistance to chemotherapeutic agents in cancers due to their cytotoxic role. This study is the first to investigate the anti-cancer effect of naphthazarin (Naph), one of the naphthaquinones, in human gastric cancer AGS cells. We showed that Naph exhibited effective preferential cell growth inhibition via G2/M phase arrest and apoptosis, which was associated with reduced levels of Cdc2 and Cdc25C expression. Naph also increased cleaved caspase-3 and Poly ADR(adenosine diphosphate ribose) Polymerase expression, γ-H2AX expression (an indicator of DNA double strand breaks) and DNA fragmentation. We also found the generation of reactive oxygen species is a critical mediator in Naph-induced cell growth inhibition and apoptosis. The non-protein antioxidant, glutathione significantly abolished Naph-mediated inhibition of cell growth and apoptosis. Taken together, our findings showed that Naph not only inhibited cell growth, but also induced apoptosis of AGS cells, suggesting that Naph may be a potential candidate for cancer therapy against gastric cancers.

Topics: Antineoplastic Agents; Apoptosis; Cell Cycle; Cell Line, Tumor; Glutathione; Humans; Naphthoquinones; Oxidative Stress; Reactive Oxygen Species; Stomach Neoplasms

To investigate the effects and underlying mechanisms of plumbagin, a naphthoquinone derived from medicinal plant Plumbago zeylanica, on human gastric cancer (GC) cells.. Human gastric cancer cell lines SGC-7901, MKN-28, and AGS were used. The cell viability was examined using CCK-8 viability assay. Cell proliferation rate was determined using both clonogenic assay and EdU incorporation assay. Apoptosis was detected via Annexin V/propidium iodide double-labeled flow cytometry. Western blotting was used to assess the expression of both NF-κB-regulated gene products and TNF-α-induced activation of p65, IκBα, and IKK. The intracellular location of NF-κB p65 was detected using confocal microscopy.. Plumbagin (2.5-40 μmol/L) concentration-dependently reduced the viability of the GC cells. The IC(50) value of plumbagin in SGC-7901, MKN-28, and AGS cells was 19.12, 13.64, and 10.12 μmol/L, respectively. The compound (5-20 μmol/L) concentration-dependently induced apoptosis of SGC-7901 cells, and potentiated the sensitivity of SGC-7901 cells to chemotherapeutic agents TNF-αand cisplatin. The compound (10 μmol/L) downregulated the expression of NF-κB-regulated gene products, including IAP1, XIAP, Bcl-2, Bcl-xL, tumor factor (TF), and VEGF. In addition to inhibition of NF-κB p65 nuclear translocation, the compound also suppressed TNF-α-induced phosphorylation of p65 and IKK, and the degradation of IκBα.. Plumbagin inhibits cell growth and potentiates apoptosis in human GC cells through the NF-κB pathway.

Topics: Antineoplastic Agents, Phytogenic; Apoptosis; Cell Line, Tumor; Cell Proliferation; Humans; I-kappa B Kinase; Naphthoquinones; NF-kappa B; Plumbaginaceae; Signal Transduction; Stomach Neoplasms; Tumor Necrosis Factor-alpha

β,β-Dimethylacrylshikonin (DA) is a major component of Radix Lithospermum erythrorhizon and has various biological activities. We have investigated the inhibitory effect of DA on the growth of hepatocellular carcinoma in vitro and in vivo. Notch signaling plays a critical role in maintaining the balance between cell proliferation, differentiation and apoptosis. Hence, perturbed Notch signaling may contribute to tumorigenesis. In the present study, we evaluated whether DA could be an effective inhibitor on cell growth in human gastric cancer cell line, and also the molecular mechanisms. Using multiple cellular and molecular approaches such as MTT assay, colony formation assay, DAPI staining, flow cytometry, real-time PCR and Western blot analysis, we found that DA inhibited cell growth in a dose- and time-dependent manner. Biochemical analysis revealed the involvement of cell cycle regulated proteins in DA-mediated of G₀-G₁ arrest of SGC-7901 cells. Furthermore, DA treatment led to reduced Notch-1 activation, expression of Jagged-1 and its downstream target Hes-1 in vitro and in vivo. Our data demonstrated that DA is a potent inhibitor of progression of gastric cancer cells, which could be due to attenuation of Notch-1. We also suggest that DA could be further developed as a potential therapeutic agent for the treatment of gastric cancer.

Topics: Animals; Antineoplastic Agents, Phytogenic; Apoptosis; Cell Line, Tumor; Cell Proliferation; Down-Regulation; Drugs, Chinese Herbal; Female; G1 Phase; Humans; Mice; Mice, Inbred BALB C; Naphthoquinones; Receptor, Notch1; Resting Phase, Cell Cycle; Signal Transduction; Stomach Neoplasms; Transplantation, Heterologous

β,β-Dimethylacrylshikonin, one of the active components in the root extracts of Lithospermum erythrorhizon, posses antitumor activity. In this study, we discussed the molecular mechanisms of β,β-dimethylacrylshikonin in the apoptosis of SGC-7901 cells. β,β-Dimethylacrylshikonin reduced the cell viability of SGC-7901 cells in a dose- and time-dependent manner and induced cell apoptosis. β,β-Dimethylacrylshikonin treatment in SGC-7901 cells down-regulated the expression of XIAP, cIAP-2, and Bcl-2 and up-regulated the expression of Bak and Bax and caused the loss of mitochondrial membrane potential and release of cytochrome c. Additionally, β,β-dimethylacrylshikonin treatment led to activation of caspases-9, 8 and 3, and cleavage of poly (ADP-ribose) polymerase (PARP), which was abolished by pretreatment with the pan-caspase inhibitor Z-VAD-FMK. β,β-Dimethylacrylshikonin induced phosphorylation of extracellular signal-regulated kinase (ERK) in SGC-7901 cells. U0126, a specific MEK inhibitor, blocked the ERK activation by β,β-dimethylacrylshikonin and abrogated β,β-dimethylacrylshikonin -induced apoptosis. Our results demonstrated that β,β-dimethylacrylshikonin inhibited growth of gastric cancer SGC-7901 cells by inducing ERK signaling pathway, and provided a clue for preclinical and clinical evaluation of β,β-dimethylacrylshikonin for gastric cancer therapy.

Topics: Antineoplastic Agents; Apoptosis; Cell Line, Tumor; Cell Survival; Enzyme Activation; Extracellular Signal-Regulated MAP Kinases; Humans; Mitochondria; Naphthoquinones; Signal Transduction; Stomach Neoplasms

This study is to investigate the effect of (2-methyl-n-butyl) shikonin (MBS) on inducing apoptosis of human gastric cancer cell line SGC-7901 and the role of ERK1/2 signal pathway in the apoptosis. MTT assay was used to detect SGC-7901 cell proliferation. DNA condensation was measured by DAPI stain. Cell apoptosis was analyzed by flow cytometry. Mitochondrial membrane potential (MMP) was analyzed by JC-1 staining. The protein expressions of Bcl-2, Bax, Survivin, cleaved caspase-9, cleaved caspase-3, cleaved PARP, p-ERK1/2, ERK1/2, p-JNK, JNK, p-p38 and p38 were detected by Western blotting. The results showed that MBS reduced the cell viability of SGC-7901 cells in a dose- and time-dependent manner. The IC50 at 24 h and 48 h for SGC-7901 cells was 10.113 and 4.196 micromolL(-1), respectively. After being treated with MBS, the typical nuclear condensation was observed in SGC-7901 cells by DAPI stain. Apoptosis in SGC-7901 cells was induced by MBS in a dose dependent manner. The protein expression of Bcl-2 was down-regulated, while the protein expressions of cleaved caspase-9, cleaved caspase-3, cleaved PARP, p-ERK1/2 and p-JNK were up-regulated after MBS treatment. U0126, a specific MAP kinase (MEK1/2) inhibitor, blocked the ERK1/2 activation by MBS. MMP was decreased by MBS treatment. It can be concluded that MBS could inhibit SGC-7901 cell proliferation and induce apoptosis. Mitochondrial apoptosis pathway, ERK1/2 signal pathway and JNK signal pathway might be involved in this process.

Topics: Antineoplastic Agents, Phytogenic; Apoptosis; Caspase 3; Cell Line, Tumor; Cell Proliferation; Dose-Response Relationship, Drug; Drugs, Chinese Herbal; Humans; Inhibitory Concentration 50; Lithospermum; MAP Kinase Signaling System; Membrane Potential, Mitochondrial; Molecular Structure; Naphthoquinones; Plant Roots; Plants, Medicinal; Proto-Oncogene Proteins c-bcl-2; Stomach Neoplasms

This study was designed to investigate the effect of juglone on the apoptosis of human gastric cancer SGC-7901 cells. The cytotoxic activity of juglone on SGC-7901 cells was tested by the sulforhodamine B (SRB) assay. The morphological changes in the cells were observed by transmission electron microscopy (TEM). The apoptotic rate, the level of reactive oxygen species (ROS), mitochondrial transmembrane potential and the expression of cytochrome c protein were detected by flow cytometry (FCM). The expression of Bcl-2 and Bax proteins were examined by Western blot. Caspase 3 activity was determined with a microplate reader. Our results were as follows: the GI(50) values for SGC-7901 cells were 36.51 ± 1.05 μmol/L (24h) and 25.37 ± 1.19 μmol/L (48 h). After 24h of exposure to juglone (5, 10, 15 and 20 μmol/L), the cells presented the typical morphological changes of apoptosis, and the rate of apoptosis was found to increase in a dose-dependent manner. After cells were treated with juglone at the same dose for 24h, the level of ROS was significantly higher, the expression of Bcl-2 was significantly down-regulated and the expression of Bax was significantly up-regulated compared to the control. The mitochondrial transmembrane potential was significantly lower, and the expression of the cytochrome c protein was significantly higher relative to the control. Caspase 3 was activated in a concentration-dependent manner. In conclusion, juglone can induce apoptosis in SGC-7901 cells through a mitochondrial pathway that seems to be mediated by the generation of ROS and a reduction in the Bcl-2/Bax ratio.

Topics: Antineoplastic Agents; Apoptosis; bcl-2-Associated X Protein; Blotting, Western; Caspase 3; Cell Line, Tumor; Cytochromes c; Humans; Membrane Potential, Mitochondrial; Microscopy, Electron, Transmission; Mitochondria; Naphthoquinones; Proto-Oncogene Proteins c-bcl-2; Reactive Oxygen Species; Stomach Neoplasms

Increasing evidence indicates that the interaction between the CXC chemokine receptor-4 (CXCR4) and its ligand CXCL12 is critical in the process of metastasis that accounts for more than 90% of cancer-related deaths. Thus, novel agents that can downregulate the CXCR4/CXCL12 axis have therapeutic potential in inhibiting cancer metastasis.. In this report, we investigated the potential of an agent, plumbagin (5-hydroxy-2-methyl-1, 4-naphthoquinone), for its ability to modulate CXCR4 expression and function in various tumor cells using Western blot analysis, DNA binding assay, transient transfection, real time PCR analysis, chromatin immunoprecipitation, and cellular migration and invasion assays.. We found that plumbagin downregulated the expression of CXCR4 in breast cancer cells irrespective of their HER2 status. The decrease in CXCR4 expression induced by plumbagin was not cell type-specific as the inhibition also occurred in gastric, lung, renal, oral, and hepatocellular tumor cell lines. Neither proteasome inhibition nor lysosomal stabilization had any effect on plumbagin-induced decrease in CXCR4 expression. Detailed study of the underlying molecular mechanism(s) revealed that the regulation of the downregulation of CXCR4 was at the transcriptional level, as indicated by downregulation of mRNA expression, inhibition of NF-κB activation, and suppression of chromatin immunoprecipitation activity. In addition, using a virtual, predictive, functional proteomics-based tumor pathway platform, we tested the hypothesis that NF-κB inhibition by plumbagin causes the decrease in CXCR4 and other metastatic genes. Suppression of CXCR4 expression by plumbagin was found to correlate with the inhibition of CXCL12-induced migration and invasion of both breast and gastric cancer cells.. Overall, our results indicate, for the first time, that plumbagin is a novel blocker of CXCR4 expression and thus has the potential to suppress metastasis of cancer.

Topics: Antineoplastic Agents, Phytogenic; Breast Neoplasms; Cell Line, Tumor; Cell Movement; Chemokine CXCL12; Computer Simulation; Down-Regulation; Female; Gene Expression Regulation, Neoplastic; Genes, Reporter; Humans; Luciferases; Models, Biological; Naphthoquinones; Neoplasm Invasiveness; NF-kappa B; Protein Binding; Receptors, CXCR4; Stomach Neoplasms; Transcription, Genetic

Salvicine (a novel diterpenoid quinone compound) exhibited a marked antitumor activity on human solid tumor cell lines and BALB/c-nu human carcinoma xenografts in our earlier studies, and it has been chosen as a candidate anticarcinogenic compound in the preclinical research stage. The present study was undertaken in order to observe whether or not the antitumor effect of salvicine is associated with its ability to induce apoptosis. Our results show that salvicine is capable of inhibiting cell proliferation and inducing characteristic changes of apoptosis in both human leukemia K-562 and gastric carcinoma SGC-7901 cells. These effects are dose and time dependent. The results of this study strongly suggest that the antitumor effect of salvicine is associated with its ability to induce apoptosis. Meanwhile, this study also shows that the activity of salvicine against K-562 and SGC-7901 cells is similar with regards to both growth inhibition and apoptosis induction, further indicating that salvicine causes these particular effects on solid tumor cells.

Topics: Adenocarcinoma; Antineoplastic Agents, Phytogenic; Apoptosis; Cell Division; DNA Fragmentation; Dose-Response Relationship, Drug; Drug Screening Assays, Antitumor; Humans; Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Naphthoquinones; Stomach Neoplasms; Tumor Cells, Cultured

Naphthoquinone pigment-LIII, an extract from Arnebia euchroma, could apparently inhibit the proliferation of stomach cancer cell line and esophagus cancer cell line. At the effective concentration of 5 micrograms/ml, the mitotic index and growth curve declined without showing any damage to human normal cells. At 5-10 micrograms/ml, the colony efficiency of cancer cells became significantly low. The anti-cancer effect of Naphthoquinone pigment-LIII might be related to its role of influencing the amount of RNA and ultrastructure of cancer cells which was discussed in this paper.

Topics: Antineoplastic Agents, Phytogenic; Esophageal Neoplasms; Humans; Naphthoquinones; Stomach Neoplasms; Tumor Cells, Cultured