Nigericin Boosts Anti-Tumor Immune Response via Inducing Pyroptosis in Triple-Negative Breast Cancer.

Although immune checkpoint inhibitors improved the clinical outcomes of advanced triple negative breast cancer (TBNC) patients, the response rate remains relatively low. Nigericin is an antibiotic derived from Streptomyces hydrophobicus. We found that nigericin caused cell death in TNBC cell lines MDA-MB-231 and 4T1 by inducing concurrent pyroptosis and apoptosis. As nigericin facilitated cellular potassium efflux, we discovered that it caused mitochondrial dysfunction, leading to mitochondrial ROS production, as well as activation of Caspase-1/GSDMD-mediated pyroptosis and Caspase-3-mediated apoptosis in TNBC cells. Notably, nigericin-induced pyroptosis could amplify the anti-tumor immune response by enhancing the infiltration and anti-tumor effect of CD4+ and CD8+ T cells. Moreover, nigericin showed a synergistic therapeutic effect when combined with anti-PD-1 antibody in TNBC treatment. Our study reveals that nigericin may be a promising anti-tumor agent, especially in combination with immune checkpoint inhibitors for advanced TNBC treatment.

Nigericin Induces Apoptosis in Primary Effusion Lymphoma Cells by Mitochondrial Membrane Hyperpolarization and ß-Catenin Destabilization.

BACKGROUND/AIM: Primary effusion lymphoma (PEL) is classified as a rare non-Hodgkin's B-cell lymphoma that is caused by Kaposi's sarcoma-associated herpesvirus (KSHV); PEL cells are latently infected with KSHV. PEL is frequently resistant to conventional chemotherapies. Therefore, the development of novel therapeutic agents is urgently required. Nigericin, a H+ and K+ ionophore, possesses unique pharmacological effects. However, the effects of nigericin on PEL cells remain unknown. MATERIALS AND METHODS: We examined the cytotoxic effects of the K+ ionophores, nigericin, nonactin, and valinomycin, on various B-lymphoma cells including PEL. We also evaluated ionophore-induced changes in signaling pathways involved in KSHV-induced oncogenesis. Moreover, the effects of nigericin on mitochondrial membrane potential and viral reactivation in PEL were analyzed. RESULTS: Although the three tested ionophores inhibited the proliferation of several B-lymphoma cell lines, nigericin inhibited the proliferation of PEL cells compared to KSHV-negative cells. In PEL cells, nigericin disrupted the mitochondrial membrane potential and caused the release of cytochrome c, which triggered caspase-9-mediated apoptosis. Nigericin also induced both an increase in phosphorylated p38 MAPK and proteasomal degradation of ß-catenin. Combination treatment of nigericin with the p38 MAPK inhibitor SB203580 potentiated the cytotoxic effects towards PEL cells, compared to either compound alone. Meanwhile, nigericin did not influence viral replication in PEL cells. CONCLUSION: Nigericin induces apoptosis in PEL cells by mitochondrial dysfunction and down-regulation of Wnt/ß-catenin signaling. Thus, nigericin is a novel drug candidate for treating PEL without the risk of de novo KSHV infection.

Sevoflurane preconditioning improves neuroinflammation in cerebral ischemia/reperfusion induced rats through ROS-NLRP3 pathway.

AIM: We aimed to study the influence of sevoflurane on the nucleotide-binding domain and Leucine-rich repeat protein 3 (NLRP3) pathways in rats with cerebral ischemia/reperfusion (I/R) injury. METHODS: Sixty Sprague-Dawley rats were equally divided into five groups randomly: sham-operated, cerebral I/R, sevoflurane (Sevo), NLRP3 inhibitor-treated (MCC950), and sevoflurane and NLRP3 inducer-treated groups. Rats' neurological functions were assessed using Longa scoring after 24 h of reperfusion, after which they were sacrificed, and cerebral infarction area was determined by triphenyl tetrazolium chloride staining. Pathological changes in damaged portions were assessed using hematoxylin-eosin and Nissl staining, and cell apoptosis was detected by terminal-deoxynucleotidyl transferase-mediated nick end labeling staining. Interleukin 1 beta (IL-1ß), tumor necrosis factor α (TNF-α), interleukin-6 (IL-6), interleukin-18 (IL-18), malondialdehyde (MDA), and superoxide dismutase (SOD) levels in brain tissues were determined using enzyme-linked immunosorbent assay. Reactive oxygen species (ROS) levels were analyzed using a ROS assay kit. Protein levels of NLRP3, caspase-1, and IL-1ß were determined by western blot. RESULTS: Neurological function scores, cerebral infarction areas, and neuronal apoptosis index were decreased in the Sevo and MCC950 groups than in the I/R group. IL-1ß, TNF-α, IL-6, IL-18, NLRP3, caspase-1, and IL-1ß levels decreased in the Sevo and MCC950 groups (p < 0.05). ROS and MDA levels increased, but SOD levels increased in the Sevo and MCC950 groups than in the I/R group. NLPR3-inducer nigericin eliminated the protective effects of sevoflurane on cerebral I/R injury in rats. CONCLUSION: Sevoflurane could alleviate cerebral I/R-induced brain damage by inhibiting the ROS-NLRP3 pathway.

Nigericin Abrogates Maternal and Embryonic Oxidative Stress in the Streptozotocin-Induced Diabetic Pregnant Rats.

Hyperglycemic exposure in diabetic pregnancy can lead to many developmental changes, such as delayed development, fetal malformations, and fetal/embryo death. These detrimental complications are collectively known as diabetic embryopathy or teratogenesis. The current study focuses to discover the therapeutic properties of the nigericin against the STZ-stimulated diabetic embryopathy via alleviation of maternal and embryonic oxidative stress. The male and female rats at a 1:1 ratio were permitted to mate overnight to establish the course of pregnancy. The pregnant rats were distributed into four groups control, diabetic pregnant (via administering 40 mg/kg of STZ), and diabetic + 10 and 20 mg/kg of nigericin-administered (via oral gavage from days 5 to 12) groups, respectively. The glucose level, urine output, diet intake, and body weight were determined carefully. The embryo and placenta weight and implantation rates were examined, and data were tabulated. The total protein and lipid profiles were assessed using respective kits. The oxidative stress markers and antioxidant enzymes were examined using respective assay kits. The 10 and 20 mg/kg of nigericin treatment decreased the glucose level and urine output and improved the diet intake and body weight gain in diabetic pregnant rats. The nigericin also decreased the total protein, cholesterol, triglycerides, and very-low-density lipoprotein (VLDL) and improved the high-density lipoprotein (HDL) in the serum of pregnant rats. The levels of malondialdehyde (MDA), reactive oxygen species (ROS), and protein carbonyls were decreased by the nigericin in both liver and embryos of the pregnant rats. The levels of glutathione (GSH), total thiols, and activities of catalase (CAT), glutathione reductase (GR), superoxide dismutase (SOD), glutathione peroxidase (GPX), and glutathione S-transferase (GST) were improved by the nigericin in the diabetic pregnant rats. Altogether, these results provide evidence that nigericin treatment remarkably attenuates the diabetes-stimulated embryopathy in rats. The nigericin effectively decreased embryo lethality, reduced glucose and dyslipidemia, and relieves oxidative stress via upregulating the antioxidant enzyme activities. Hence, it can be a talented therapeutic agent to treat diabetic pregnancy-associated complications.

Characterization of Pathway-Specific Regulator NigR for High Yield Production of Nigericin in Streptomyces malaysiensis F913.

Nigericin is a polyether antibiotic with potent antibacterial, antifungal, antimalarial and anticancer activity. NigR, the only regulator in the nigericin biosynthetic gene cluster in Streptomyces malaysiensis F913, was identified as a SARP family regulator. Disruption of nigR abolished nigericin biosynthesis, while complementation of nigR restored nigericin production, suggesting that NigR is an essential positive regulator for nigericin biosynthesis. Overexpression of nigR in Streptomyces malaysiensis led to significant increase in nigericin production compared to the wild-type strain. Nigericin production in the overexpression strain was found to reach 0.56 g/L, which may be the highest nigericin titer reported to date. Transcriptional analysis suggested that nigR is required for the transcription of structural genes in the nig gene cluster; quantitative RT-PCR analysis revealed that the expression of structural genes was upregulated in the nigR overexpression strain. Our study suggested that NigR acts in a positive manner to modulate nigericin production by activating transcription of structural genes and provides an effective strategy for scaling up nigericin production.

Senkyunolide A inhibits the progression of osteoarthritis by inhibiting the NLRP3 signalling pathway.

CONTEXT: Osteoarthritis (OA) is a degenerative disease. Senkyunolide A (SenA) is an important phthalide from Ligusticum chuanxiong Hort (Umbelliferae) with anti-spasmodic and neuroprotective effects. OBJECTIVE: We explored the effect of SenA on IL-1ß-stimulated chondrocytes and OA mice. MATERIALS AND METHODS: Chondrocytes were stimulated by IL-1ß (10 ng/mL) to establish an OA model in vitro. Cells were treated with SenA (20, 40, 80 and 160 µg/mL) for 48 h. The in vivo OA model was established by cutting off the medial meniscus tibial ligament (MMTL) at right knee incision of male C57BL/6 mice. One week after surgery, mice were injected with SenA (intraperitoneally one week) and divided into four groups (n = 6 per group): Sham, OA, OA + SenA 20 mg/kg and OA + SenA 40 mg/kg. The OA progression was examined by haematoxylin and eosin (H&E) staining. RESULTS: SenA treatment increased cell viability (33%), proliferation (71%), inhibited apoptosis (21%), decreased levels of catabolic marker proteins (MMP13, 23%; ADAMTS4, 31%; ADAMTS5, 19%), increased levels of anabolic marker proteins (IGF-1, 57%; aggrecan, 75%; Col2a1, 48%), reduced levels of inflammation cytokines (TNF-α, 31%; IL-6, 19%; IL-18, 20%) and decreased levels of NLRP3 (21%), ASC (20%) and caspase-1 (29%) of chondrocytes. However, NLRP3 agonist nigericin increased levels of MMP13 (55%), ADAMTS4 (70%), ADAMTS5 (53%), decreased levels of IGF-1 (36%), aggrecan (26%), Col2a1 (25%), inhibited proliferation (61%) and promoted apoptosis (76%). DISCUSSION AND CONCLUSIONS: SenA alleviates OA progression by inhibiting NLRP3 signalling pathways. These findings provide an experimental basis for the clinical application of drugs in the treatment of OA.

Nigericin exerts anticancer effects through inhibition of the SRC/STAT3/BCL-2 in osteosarcoma.

The treatment of osteosarcoma has reached a bottleneck period in recent 30 years, there is an urgent need to find new drugs and treatment methods. Nigericin, an antibiotic derived from Streptomyces hygroscopicus, has exerted promising antitumoral effect in various tumors. The anticancer effect of Nigericin in human osteosarcoma has never been reported. In the present study, we explored the anticancer effects of Nigericin in osteosarcoma in vitro and in vivo. Our results showed that nigericin treatment significantly reduced tumor cell proliferation in dose-dependent and time-dependent in human osteosarcoma cells. Nigericin can inhibit cell growth of osteosarcoma cells, in addition to S-phase cycle arrest, the nigericin induces apoptosis. Furthermore, bioinformatics predicted that Nigericin exerts anticancer effects through inhibiting SRC/STAT3 signaling pathway in osteosarcoma. The direct binding between SRC and activator of transcription 3 (STAT3) was confirmed by Western blot. Nigericin can down regulate STAT3 and Bcl-2. In order to further elucidate the inhibitory effect of nigericin on SRC/STAT3/Bcl-2 signal transduction mechanism, we established human osteosarcoma cancer cells stably expressing STAT3. Western blot confirmed that nigericin exerts anticancer effects on human osteosarcoma cancer cells by directly targeting STAT3. In addition, Nigericin can significantly inhibit tumor migration and invasion. Finally, Nigericin inhibits tumor growth in a mouse osteosarcoma model. The nigericin targeting the SRC/STAT3/BCL-2 signaling pathway may provide new insights into the molecular mechanism of nigericin on cancer cells and suggest its possible clinical application in osteosarcoma.

Combination of nigericin with cisplatin enhances the inhibitory effect of cisplatin on epithelial ovarian cancer metastasis by inhibiting slug expression via the Wnt/ß-catenin signalling pathway.

Epithelial ovarian cancer (EOC) is the most lethal cancer among female genital tumours. Standard therapies, including postoperative chemotherapy, exhibit high proportions of recurrence and resistance. Novel therapeutic strategies are combined with chemotherapy. Emerging studies have demonstrated that nigericin, an H+, K+ and Pb2+ ionophore, exhibits promising anticancer activity in various types of malignancy, such as colorectal and epithelial ovarian cancer. Our previous study suggested that nigericin could regulate EOC cell proliferation, migration and invasion, and may be a novel chemotherapy candidate for EOC. However, to the best of our knowledge, the effects of combined therapy with cisplatin, and the associated underlying mechanisms, are not yet fully understood. The present study aimed to clarify the effects of combined chemical therapy with nigericin and cisplatin on EOC cells and to reveal its mechanism. Wound healing, Transwell, cell viability and colony formation assays were used to measure the migration, invasion and proliferation of EOC cells. Western blotting was used to detect protein expression. A slug overexpression lentivirus was used to create a slug overexpression model in SK-OV-3 cells. Small interfering RNA was used to knock down slug expression. Nigericin combined with cisplatin enhanced the inhibitory effects of cisplatin on the migration and colony formation of EOC cells. Nigericin also enhanced the inhibitory effects of cisplatin on the expression levels of MMP7, as well as the inhibitory effects of cisplatin on the expression levels of ß-catenin and GSK-3ß, indicating that nigericin and cisplatin regulated in the Wnt/ß-catenin signalling pathway. When slug was knocked down, the effect of nigericin was weakened. Overexpression of slug could repress the inhibitory effect of nigericin on the Wnt/ß-catenin signalling pathway. Furthermore, nigericin inhibited slug expression by enhancing its modification through small ubiquitin-like modifiers (SUMOs; referred to as SUMOylation). Overall, the present results demonstrated that nigericin combined with cisplatin might serve as a novel therapeutic strategy in patients with metastatic EOC because the combined therapy had higher effectiveness than single drug use. The underlying mechanism of combined therapy maybe the enhanced inhibitory effect of slug through its nigericin-induced SUMOylation.

NLRP3 Triggers Attenuate Lipocalin-2 Expression Independent with Inflammasome Activation.

Lipocalin-2 (LCN2), a small secretory glycoprotein, is upregulated by toll-like receptor (TLR) signaling in various cells and tissues. LCN2 inhibits bacterial growth by iron sequestration and regulates the innate immune system. Inflammasome activates the inflammatory caspases leading to pyroptosis and cytokine maturation. This study examined the effects of inflammasome activation on LCN2 secretion in response to TLR signaling. The triggers of NLRP3 inflammasome activation attenuated LCN2 secretion while it induced interleukin-1ß in mouse macrophages. In mice, NLRP3 inflammasome activation inhibited TLR-mediated LCN2 secretion. The inhibition of NLRP3 triggers on LCN2 secretion was caused by the inhibited transcription and translation of LCN2. At the same time, no changes in the other cytokines and IκBζ, a well-known transcriptional factor of Lcn2 transcription, were observed. Overall, NLRP3 triggers are a regulator of LCN2 expression suggesting a new linkage of inflammasome activation and LCN2 secretion in the innate immunity.

Evidence of nigericin as a potential therapeutic candidate for cancers: A review.

Emerging studies have shown that nigericin, an H+, K+ and Pb2+ ionophore, has exhibited a promising anti-cancer activity in various cancers. However, its anti-cancer mechanisms have not been fully elucidated. In this review, the recent progresses on the use of nigericin in human cancers have been summarized. By exchanging H+ and K+ across cell membranes, nigericin shows promising anti-cancer activities in in vitro and in vivo as a single agent or in combination with other anti-cancer drugs through decreasing intracellular pH (pHi). The underlying mechanisms of nigericin also include the inactivation of Wnt/ß-catenin signals, blockade of Androgen Receptor (AR) signaling, and activation of Stress-Activated Protein Kinase/c-Jun N-terminal Kinase (SAPK/JNK) signaling pathways. In many cancers, nigericin is proved to specifically target putative Cancer Stem Cells (CSCs), and its synergistic effects on photodynamic therapy are also reported. Other mechanisms of nigericin including influencing the mitochondrial membrane potentials, inducing an increase in drug accumulation and autophagy, controlling insulin accumulation in nuclei, and increasing the cytotoxic activity of liposome-entrapped drugs, are also discussed. Notably, the potential adverse effects such as teratogenic effects, insulin resistance and eryptosis shall not be ignored. Taken together, these reports suggest that treatment of cancer cells with nigericin may offer a novel therapeutic strategy and future potential of translation to clinics.

Streptomyces hygroscopicus UFPEDA 3370: A valuable source of the potent cytotoxic agent nigericin and its evaluation against human colorectal cancer cells.

Streptomyces hygroscopicus UFPEDA 3370 was fermented in submerged cultivation and the biomass extract was partitioned, obtaining a fraction purified named EB1. After purification of EB1 fraction, nigericin free acid was obtained and identified. Nigericin presented cytotoxic activity against several cancer cell lines, being most active against HL-60 (human leukemia) and HCT-116 (human colon carcinoma) cell lines, presenting IC50 and (IS) values: 0.0014 µM, (30.0) and 0.0138 µM (3.0), respectively. On HCT-116, nigericin caused apoptosis and autophagy. In this study, nigericin was also screened both in vitro and in silico against a panel of cancer-related kinases. Nigericin was able to inhibit both JAK3 and GSK-3ß kinases in vitro and its binding affinities were mapped through the intermolecular interactions with each target in silico.

Molecular Screening for Nigericin Treatment in Pancreatic Cancer by High-Throughput RNA Sequencing.

Objectives: Nigericin, an antibiotic derived from Streptomyces hygroscopicus, has been proved to exhibit promising anti-cancer effects on a variety of cancers. Our previous study investigated the potential anti-cancer properties in pancreatic cancer (PC), and demonstrated that nigericin could inhibit the cell viabilities in concentration- and time-dependent manners via differentially expressed circular RNAs (circRNAs). However, the knowledge of nigericin associated with long non-coding RNA (lncRNA) and mRNA in pancreatic cancer (PC) has not been studied. This study is to elucidate the underlying mechanism from the perspective of lncRNA and mRNA. Methods: The continuously varying molecules (lncRNAs and mRNAs) were comprehensively screened by high-throughput RNA sequencing. Results: Our data showed that 76 lncRNAs and 172 mRNAs were common differentially expressed in the nigericin anti-cancer process. Subsequently, the bioinformatics analyses, including Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, coding and non-coding co-expression network, cis- and trans-regulation predictions and protein-protein interaction (PPI) network, were applied to annotate the potential regulatory mechanisms among these coding and non-coding RNAs during the nigericin anti-cancer process. Conclusions: These findings provided new insight into the molecular mechanism of nigericin toward cancer cells, and suggested a possible clinical application in PC.

Maackiain, a compound derived from Sophora flavescens, increases IL-1ß production by amplifying nigericin-mediated inflammasome activation.

Sophora flavescens is used as a traditional herbal medicine to modulate inflammatory responses. However, little is known about the impact of (-)-maackiain, a compound derived from S. flavescens, on the activation of inflammasome/caspase-1, a key factor in interleukin-1ß (IL-1ß) processing. Here, we report that (-)-maackiain potently amplified caspase-1 cleavage in macrophages in response to nigericin (Nig). In macrophages primed with either lipopolysaccharide or monophosphoryl lipid A, Nig-mediated caspase-1 cleavage was also markedly promoted by (-)-maackiain. Notably, (-)-maackiain induced the production of vimentin, an essential mediator for the activation of the NOD-, LRR-, and pyrin domain-containing protein 3 inflammasome, thereby contributing to promotion of the formation of the inflammasome complex to activate caspase-1. Taken together, our data suggest that (-)-maackiain exerts an immunostimulatory effect by promoting IL-1ß production via activation of the inflammasome/caspase-1 pathway. Thus, the potent inflammasome-activating effect of (-)-maackiain may be clinically useful as an acute immune-stimulating agent.

Therapeutic Potential of Pharmacological Targeting NLRP3 Inflammasome Complex in Cancer.

Introduction: Dysregulation of NLRP3 inflammasome complex formation can promote chronic inflammation by increased release of IL-1ß. However, the effect of NLRP3 complex formation on tumor progression remains controversial. Therefore, we sought to determine the effect of NLRP3 modulation on the growth of the different types of cancer cells, derived from lung, breast, and prostate cancers as well as neuroblastoma and glioblastoma in-vitro. Method: The effect of Caspase 1 inhibitor (VX765) and combination of LPS/Nigericin on NLRP3 inflammasome activity was analyzed in A549 (lung cancer), MCF-7 (breast cancer), PC3 (prostate cancer), SH-SY5Y (neuroblastoma), and U138MG (glioblastoma) cells. Human fibroblasts were used as control cells. The effect of VX765 and LPS/Nigericin on NLRP3 expression was analyzed using western blot, while IL-1ß and IL-18 secretion was detected by ELISA. Tumor cell viability and progression were determined using Annexin V, cell proliferation assay, LDH assay, sphere formation assay, transmission electron microscopy, and a multiplex cytokine assay. Also, angiogenesis was investigated by a tube formation assay. VEGF and MMPs secretion were detected by ELISA and a multiplex assay, respectively. Statistical analysis was done using one-way ANOVA with Tukey's analyses and Kruskal-Wallis one-way analysis of variance. Results: LPS/Nigericin increased NRLP3 protein expression as well as IL-1ß and IL-18 secretion in PC3 and U138MG cells compared to A549, MCF7, SH-SY5Y cells, and fibroblasts. In contrast, MIF expression was commonly found upregulated in A549, PC3, SH-SY5Y, and U138MG cells and fibroblasts after Nigericin treatment. Nigericin and a combination of LPS/Nigericin decreased the cell viability and proliferation. Also, LPS/Nigericin significantly increased tumorsphere size in PC3 and U138MG cells. In contrast, the sphere size was reduced in MCF7 and SH-SY5Y cells treated with LPS/Nigericin, while no effect was detected in A549 cells. VX765 increased secretion of CCL24 in A549, MCF7, PC3, and fibroblasts as well as CCL11 and CCL26 in SH-SY5Y cells. Also, VX765 significantly increased the production of VEGF and MMPs and stimulated angiogenesis in all tumor cell lines. Discussion: Our data suggest that NLRP3 activation using Nigericin could be a novel therapeutic approach to control the growth of tumors producing a low level of IL-1ß and IL-18.

Doxorubicin-carboxymethyl xanthan gum capped gold nanoparticles: Microwave synthesis, characterization, and anti-cancer activity.

An ultrafast (e.g. 75 s) synthesis of carboxymethyl xanthan gum (CMXG) capped gold nanoparticles (AuNPs) (CMXG@AuNPs) was developed using microwave irradiation (MWI) method. The synthesis of AuNPs was optimized by varying CMXG amount, gold ion concentration, and MWI time. The CMXG@AuNPs exhibited a spherical shape, high crystallinity, and narrow size distribution (i.e. 8-10 nm). The electrostatic interaction-mediated the loading of doxorubicin (DOX) onto CMXG@AuNPs. The release of DOX, loaded on CMXG@AuNPs was extensive in an acidic condition but negligible at physiological pH value. The in vitro anticancer efficacy of DOX loaded on CMXG@AuNPs (i.e. DOX@CMXG@AuNPs) in the presence of an ionophore (i.e. nigericin) was about 4.6 folds higher than that of free DOX. Flow cytometry revealed that DOX@CMXG@AuNPs exhibited a higher cellular uptake under an acidic condition than free DOX. CMXG@AuNPs showed unique excellence in the pH-responsive DOX-releasing property and the cancer cell-killing capability.

Nigericin Promotes NLRP3-Independent Bacterial Killing in Macrophages.

Altered microbiota has been associated with a number of diseases, including inflammatory bowel diseases, diabetes, and cancer. This dysregulation is thought to relate the host inflammatory response to enteric pathogens. Macrophages play a key role in host response to microbes and are involved in bacterial killing and clearance. This process is partially mediated through the potassium efflux-dependent, cytosolic, PYCARD-containing inflammasome protein complex. Surprisingly, we discovered an alternative mechanism for bacterial killing, independent of the NLRP3 inflammasome/PYCARD. Using the NLRP3 inflammasome-deficient Raw 264.7 and PYCARD-deficient J77 macrophages, which both lack PYCARD, we found that the potassium efflux activator nigericin enhances bacterial killing. Macrophage response to nigericin was examined by RT gene profiling and subsequent qPCR, which demonstrated altered expression of a series of genes involved in the IL-18 bacterial killing pathway. Based on our results we propose a model of bacterial killing, unrelated to NLRP3 inflammasome activation in macrophage cells. Improving understanding of the molecular pathways driving bacterial clearance within macrophage cells will aid in the development of novel immune-targeted therapeutics in a number of diseases.

High-throughput sequencing of circRNAs reveals novel insights into mechanisms of nigericin in pancreatic cancer.

BACKGROUND: Our previous study had proved that nigericin could reduce colorectal cancer cell proliferation in dose- and time-dependent manners by targeting Wnt/ß-catenin signaling. To better elucidate its potential anti-cancer mechanism, two pancreatic cancer (PC) cell lines were exposed to increasing concentrations of nigericin for different time periods, and the high-throughput sequencing was performed to explore the circRNA expression profiles after nigericin exposure on pancreatic cancer (PC) cells. RESULTS: In this study, a total of 183 common differentially expressed circRNAs were identified, and the reliability and validity of the sequencing data were verified by the PCR analysis. According to the parental genes of circRNAs, the GO analysis was performed to predict the most enriched terms in the biological process, cellular components and molecular functions. The KEGG analysis and pathway-pathway network exhibited the potential signal pathways and their regulatory relationships. Meanwhile, a potential competing endogenous RNA (ceRNA) mechanism through a circRNA-miRNA-mRNA network was applied to annotate potential functions of these common differentially expressed circRNAs, and these predicted miRNAs or mRNAs might be involved in nigericin damage. CONCLUSIONS: By the bioinformatics method, our data will facilitate the understanding of nigericin in PC cells, and provide new insight into the molecular mechanism of nigericin toward cancer cells. This is the first report that discusses the potential functions of nigericin in cancers through the bioinformatics method. Our data will facilitate the understanding of nigericin-mediated anti-cancer mechanisms in PC.

Ionophores: Potential Use as Anticancer Drugs and Chemosensitizers.

Ion homeostasis is extremely important for the survival of both normal as well as neoplastic cells. The altered ion homeostasis found in cancer cells prompted the investigation of several ionophores as potential anticancer agents. Few ionophores, such as Salinomycin, Nigericin and Obatoclax, have demonstrated potent anticancer activities against cancer stem-like cells that are considered highly resistant to chemotherapy and responsible for tumor relapse. The preclinical success of these compounds in in vitro and in vivo models have not been translated into clinical trials. At present, phase I/II clinical trials demonstrated limited benefit of Obatoclax alone or in combination with other anticancer drugs. However, future development in targeted drug delivery may be useful to improve the efficacy of these compounds. Alternatively, these compounds may be used as leading molecules for the development of less toxic derivatives.

Intracellular pH Measurements in Glioblastoma Cells Using the pH-Sensitive Dye BCECF.

The regulation of pH in glioblastoma (GBM) has received significant attention, because it has been linked to tumor metabolism and the stem cell phenotype. The variability in blood perfusion and oxygen tension within tumors suggests that ambient pH values fluctuate across different tumor territories. This chapter describes a detailed protocol for measuring intracellular pH in patient-derived GBM cells in vitro, using the fluorescent pH sensitive dye BCECF.

Nigericin decreases the viability of multidrug-resistant cancer cells and lung tumorspheres and potentiates the effects of cardiac glycosides.

Multiple factors including tumor heterogeneity and intrinsic or acquired resistance have been associated with drug resistance in lung cancer. Increased stemness and the plasticity of cancer cells have been identified as important mechanisms of resistance; therefore, treatments targeting cancer cells independent of stemness phenotype would be much more effective in treating lung cancer. In this article, we have characterized the anticancer effects of the antibiotic Nigericin in cells displaying varying degrees of stemness and resistance to anticancer drugs, arising from (1) routine culture conditions, (2) prolonged periods of serum starvation. These cells are highly resistant to conventional anticancer drugs such as Paclitaxel, Hydroxyurea, Colchicine, Obatoclax, Wortmannin, and LY294002, and the multidrug-resistant phenotype of cells growing under prolonged periods of serum starvation is likely the result of extensive rewiring of signaling pathways, and (3) lung tumorspheres that are enriched for cancer stem-like cells. We found that Nigericin potently inhibited the viability of cells growing under routine culture conditions, prolonged periods of serum starvation, and lung tumorspheres. In addition, we found that Nigericin downregulated the expression of key proteins in the Wnt canonical signaling pathway such as LRP6, Wnt5a/b, and ß-catenin, but promotes ß-catenin translocation into the nucleus. The antitumor effects of Nigericin were potentiated by the Wnt activator HLY78 and by therapeutic levels of the US Food and Drug Administration-approved drug Digitoxin and its novel synthetic analog MonoD. We believe that Nigericin may be used in a co-therapy model in combination with other novel chemotherapeutic agents in order to achieve potent inhibition of cancers that display varying degrees of stemness, potentially leading to sustained anticancer effects.