Effectiveness of regdanvimab on mortality in COVID-19 infected patients on hemodialysis.

BACKGROUND: Although several therapeutic agents have been evaluated for the treatment of coronavirus disease 2019 (COVID-19), there are lack of effective and proven treatments for end-stage renal disease (ESRD). The present study aims to evaluate the effectiveness of regdanvimab on mortality in COVID-19-infected patients on hemodialysis (HD). METHODS: We conducted an observational retrospective study in 230 COVID-19-infected patients on HD, of whom 77 (33.5%) were administered regdanvimab alone or in combination with dexamethasone or remdesivir during hospitalization (regdanvimab group) and 153 patients (66.5%) were not (no regdanvimab group). The primary outcome was in-hospital mortality. We compared mortality rates according to the use of regdanvimab and investigated the factors associated with mortality. RESULTS: Fifty-nine deaths occurred during hospitalization, 49 in the no regdanvimab group (32.0%) and 10 in the regdanvimab group (13.0%), and the mortality rate was significantly higher in the no regdanvimab group than that in the regdanvimab group (p = 0.001). Multivariate Cox regression analysis showed that malignancy (p = 0.001), SPO2 of <95% at admission (p = 0.003), and administration of antibiotics and regdanvimab (p = 0.007 and p = 0.002, respectively) were significantly associated factors with mortality. CONCLUSION: Regdanvimab administration is beneficial in improving prognosis in hospitalized COVID-19 patients on HD. Considering the vulnerability to infection and high mortality of ESRD patients, regdanvimab may be considered as a therapeutic option in COVID-19 patients on HD.

Brain-targeted exosome-mimetic cell membrane nanovesicles with therapeutic oligonucleotides elicit anti-tumor effects in glioblastoma animal models.

The brain-targeted delivery of therapeutic oligonucleotides has been investigated as a new treatment modality for various brain diseases, such as brain tumors. However, delivery efficiency into the brain has been limited due to the blood-brain barrier. In this research, brain-targeted exosome-mimetic cell membrane nanovesicles (CMNVs) were designed to enhance the delivery of therapeutic oligonucleotides into the brain. First, CMNVs were produced by extrusion with isolated C6 cell membrane fragments. Then, CMNVs were decorated with cholesterol-linked T7 peptides as a targeting ligand by hydrophobic interaction, producing T7-CMNV. T7-CMNV was in aqueous solution maintained its nanoparticle size for over 21 days. The targeting and delivery effects of T7-CMNVs were evaluated in an orthotopic glioblastoma animal model. 2'-O-metyl and cholesterol-TEG modified anti-microRNA-21 oligonucleotides (AMO21c) were loaded into T7-CMNVs, and biodistribution experiments indicated that T7-CMNVs delivered AMO21c more efficiently into the brain than CMNVs, scrambled T7-CMNVs, lipofectamine, and naked AMO21c after systemic administration. In addition, AMO21c down-regulated miRNA-21 (miR-21) levels in glioblastoma tissue most efficiently in the T7-CMNVs group. This enhanced suppression of miR-21 resulted in the up-regulation of PDCD4 and PTEN. Eventually, brain tumor size was reduced in the T7-CMNVs group more efficiently than in the other control groups. With stability, low toxicity, and targeting efficiency, T7-CMNVs may be useful to the development of oligonucleotide therapy for brain tumors.

Antisense-oligonucleotide co-micelles with tumor targeting peptides elicit therapeutic effects by inhibiting microRNA-21 in the glioblastoma animal models.

INTRODUCTION: miRNA-21 (miR-21) is highly expressed in glioblastoma, facilitating tumor growth by blocking the expression of apoptosis-related genes. Therefore, an antisense microRNA oligonucleotide (AMO) against miR-21 was suggested as a therapeutic nucleic acid for glioblastoma. OBJECTIVES: AMO21 co-micelles were developed with tumor-targeting T7 peptides as an AMO21 delivery system by intranasal administration. METHODS: Cholesterol-conjugated AMO21 (AMO21c) was mixed with cholesterol-conjugated T7 peptides (T7c) to produce tumor-targeted co-micelles. Physical characterization was performed by dynamic light scattering, gel retardation assay, scanning electron microscope and heparin competition assay. In vitro transfection efficiency to C6 glioblastoma cells was measured by flow cytometry. The AMO21c/T7c co-micelles were administered by intranasal instillation into the brain of intracranial glioblastoma rat models. Scrambled T7 (scrT7) and scrambled AMO21c (scrAMO21c) were used as a negative control. The therapeutic effects of the AMO21c/T7c co-micelles were evaluated by real time RT-PCR, immunohistochemistry, TUNEL assay, and Nissl staining. RESULTS: The formation of the AMO21c/T7c co-micelles was confirmed in gel retardation and heparin competition assays. The highest delivery efficiency in vitro was achieved at a 1:10 wt ratio of AMO21c/T7c. The AMO21c/T7c co-micelles had higher delivery efficiency into C6 glioblastoma cells than naked AMO21c or AMO21c/lipofectamine complexes. After intranasal administration into the intracranial glioblastoma models, the delivery efficiency of the co-micelles into the brain was also higher than those of naked AMO21c and AMO21c/scrambled T7c. Thanks to their enhanced delivery efficiency, the AMO21c/T7c co-micelles downregulated miR-21, inducing the production of the pro-apoptotic phosphatase and tensin homolog (PTEN) and programmed cell death 4 (PDCD4) proteins in the tumor tissues. The tumor size was reduced by the AMO21c/T7c co-micelles more effectively than naked AMO21c, AMO21c/lipofectamine, or scrAMO21c/T7c treatment. CONCLUSION: The results suggest that the co-micelles of AMO21c and T7c may be an efficient delivery system into a brain tumor through intranasal administration.

Dual-Functional Dendrimer Micelles with Glycyrrhizic Acid for Anti-Inflammatory Therapy of Acute Lung Injury.

Dendrimer micelles with glycyrrhizic acid (GA) were developed for anti-inflammatory therapy of acute lung injury (ALI). Cholesterol was conjugated to histidine- and arginine-grafted polyamidoamine (PamHR) for micelle formation. The cholesterol-conjugated PamHR (PamHRchol) was mixed with amphiphilic GA to produce PamHRchol/GA mixed micelles. The GA integrated into the micelles had two functions: it acted as an anti-inflammatory drug and facilitated intracellular gene delivery. The PamHRchol/GA micelles formed stable complexes with plasmid DNA. Integrating GA into the micelles increased their transfection efficiency. Confocal microscopy and flow-cytometry studies confirmed that the PamHRchol/GA micelles improved cellular uptake compared with PamHRchol. A competition assay with free GA suggested that the enhanced transfection efficiency of the micelles might be due to the interaction between GA and its receptor. In addition, GA has a membrane-destabilizing effect, and a chloroquine pretreatment assay confirmed that GA increased endosomal escape. Furthermore, the PamHRchol/GA micelles reduced tumor necrosis factor-α in lipopolysaccharide-activated Raw264.7 cells, suggesting a mechanism for its anti-inflammatory effects. To evaluate the therapeutic potential of the PamHRchol/GA micelles, the heme oxygenase-1 (HO-1) gene was delivered into the lungs of mice with ALI. The PamHRchol/GA micelles had higher gene delivery efficiency into the lungs than polyethylenimine (25 kDa, PEI25k) and the PamHRchol micelles. The combined effects of the HO-1 gene and GA produced effective anti-inflammation response in the lungs of the ALI animals. Therefore, the dual-function PamHRchol/GA micelles, which acted as an anti-inflammatory drug and a gene carrier, could be a useful therapy for inflammatory lung diseases.

Intranasal delivery of self-assembled nanoparticles of therapeutic peptides and antagomirs elicits anti-tumor effects in an intracranial glioblastoma model.

MicroRNA-21 (miR-21) is involved in the progression of glioblastoma through inhibition of pro-apoptotic genes. Antisense RNA against miR-21 (antagomir-21) has been developed as a potential therapeutic reagent for the treatment of glioblastoma. The receptor for advanced glycation end-products (RAGE) is also involved in the progression of glioblastoma through induction of angiogenic factors. Therefore, RAGE-antagonist peptide (RAP) is proposed to be an anti-tumor reagent. In this study, self-assembled nanoparticles were produced solely with therapeutic agents, antagomir-21 and RAP, with no additional carrier. The therapeutic effects of the nanoparticles by intranasal delivery were evaluated in intracranial glioblastoma animal models. First, physical characterizations such as size/zeta-potential study, scanning electron microscopy, and gel retardation assays showed that antagomir-21 and RAP formed stable nanoparticles without any additional reagents. The ratio between antagomir-21 and RAP was optimized by an in vitro cellular uptake study. The antagomir-21/RAP nanoparticles were administrated intranasally in the intracranial glioblastoma animal models to bypass the blood-brain-barrier. As a result, the nanoparticles reduced the miR-21 levels in tumors. Inhibition of miR-21 by the nanoparticles induced the expression of pro-apoptotic genes, such as PTEN and PDCD4, which enhanced tumor cell apoptosis. In addition, the expression of RAGE was suppressed by the nanoparticles, resulting in decreased levels of vascular endothelial growth factor in the tumor. The reduction of CD31-positive endothelial cells confirmed the anti-angiogenic effects of the nanoparticles. The results indicate that the intranasal delivery of the self-assembled nanoparticles of antagomir-21 and RAP is an efficient treatment of glioblastoma.

Hypoxia-specific anti-RAGE exosomes for nose-to-brain delivery of anti-miR-181a oligonucleotide in an ischemic stroke model.

Ischemic stroke is caused by a reduction in blood flow to the brain due to narrowed cerebral arteries. Thrombolytic agents have been used to induce reperfusion of occluded cerebral arteries. However, brain damage continues to progress after reperfusion and induces ischemia-reperfusion (I/R) injury. The receptor for advanced glycation end-products (RAGE) is overexpressed in hypoxic cells of the ischemic brain. In this study, an exosome linked to RAGE-binding-peptide (RBP-Exo) was developed as a hypoxia-specific carrier for nose-to-brain delivery of anti-microRNA oligonucleotide (AMO). The RBP-Exos were less than 50 nm in size and had negative surface charge. In vitro studies showed that RBP-Exos delivered AMO181a to Neuro2A cells more efficiently than unmodified exosomes (Unmod-Exos). In addition, RAGE was downregulated by RBP-Exos, suggesting that the RBP moiety of the RBP-Exos reduced the RAGE-mediated signal pathway. MicroRNA-181a (miR-181a) is one of the upregulated miRNAs in the ischemic brain and its downregulation can reduce the damage to the ischemic brain. Cholesterol-modified AMO181a (AMO181a-chol) was loaded onto the RBP-Exo by hydrophobic interaction. The AMO181a-chol-loaded RBP-Exo (RBP-Exo/AMO181a-chol) was administered intranasally to a rat middle cerebral artery occlusion (MCAO) model. MiR-181a was knocked down and Bcl-2 was upregulated by intranasal delivery of RBP-Exo/AMO181a-chol. In addition, tumor necrosis factor-α (TNF-α) expression and apoptosis were reduced by RBP-Exo/AMO181a-chol. As a result, RBP-Exo/AMO181a-chol significantly suppressed infarct size compared with the controls. In conclusion, RBP-Exo was a hypoxia-specific carrier for nose-to-brain delivery of AMO181a-chol in an ischemic stroke model. Furthermore, the combined effects of RBP and AMO181a-chol exerted neuroprotective effects in the ischemic brain.

Biomimetic cell membrane-coated DNA nanoparticles for gene delivery to glioblastoma.

Gene therapy has been introduced as an alternative to radiation and chemical therapy for glioblastoma. Biomimetic nanoparticles coated with cell membranes (CM) have advantages such as high biocompatibility and prolong half-life. To apply CM coated nanoparticles to gene delivery, the polyethylenimine (PEI25k)/plasmid DNA (pDNA) complexes were coated with CM from C6 rat glioblastoma cells. With the CM covering, the PEI25k/pDNA complexes formed stable nanoparticles with negative surface charge. The PEI25k/pDNA/CM nanoparticles had high colloidal stability and could be stored for approximately 20 days without aggregation. The transfection efficiency of the PEI25k/pDNA/CM nanoparticles was higher than that of the PEI25k/pDNA complex in serum-containing medium. This suggests that serum does not interfere with transfection efficiency of the nanoparticles. Moreover, the PEI25k/pDNA/CM nanoparticles had lower toxicity than the PEI25k/DNA complex in vitro and in vivo. The PEI25k/pDNA/CM nanoparticles prepared with CMs of different types of cells were transfected into cells. The results showed that the PEI25k/pDNA/CM nanoparticles with the C6 CM had the highest transfection efficiency to C6 cells, suggesting the homotypic targeting effect. The therapeutic effects of the nanoparticles were evaluated in intracranial C6 transplanted glioblastoma animal models. The PEI25k/pDNA/CM nanoparticles were prepared with herpes simplex virus thymidine kinase plasmid (pHSVtk) and injected into the tumor locally. The results showed that the PEI25k/pHSVtk/CM nanoparticles induced higher HSVtk expression compared with the PEI25k/pHSVtk complex. Furthermore, tumor size was reduced more efficiently by the PEI25k/pHSVtk/CM nanoparticles than by the PEI25k/pHSVtk complex. Overall results indicate that PEI25k/pDNA/CM nanoparticles are suitable for pDNA delivery to glioblastoma.

Antitumor Effect of Metformin in Combination with Binimetinib on Melanoma Cells.

Cutaneous melanoma is a fatal disease for patients with distant metastasis. Metformin is the most widely used anti-diabetic drug, and proved to suppress cell proliferation and metastasis in diverse cancers including melanoma. We previously reported that MEK inhibitor trametinib increases the expression of epithelial-mesenchymal transition (EMT) regulators and melanoma cell motility, which are suppressed by addition of metformin in A375 melanoma cells. To confirm our findings further, we first evaluated the effect of metformin in combination with another MEK inhibitor binimetinib on cell viability in G361 melanoma cells. We then investigated whether binimetinib affects the expression of EMT regulators and cell motility. We finally monitored the effect of metformin on binimetinib-induced cell migration. Cell viability assay showed that combination index (CI) value at ED50 is 0.80, suggesting synergy for the combination of metformin with binimetinib. Our results also revealed that binimetinib increased the expression of EMT regulators such as integrin αV, fibronectin and slug, which correlate well with the enhanced cell migration in wound healing assay. Metformin, on the contrary, suppressed the expression of sparc, integrin αV, fibronectin and N-cadherin with the reduced cell motility. The combination treatment showed that metformin counteracts the binimetinib-induced increase of cell motility. Overall, these results suggest that metformin with binimetinib might be useful as a potential therapeutic adjuvant against cell survival and metastatic activity in melanoma patients.

Effect of Metformin in Combination With Trametinib and Paclitaxel on Cell Survival and Metastasis in Melanoma Cells.

BACKGROUND/AIM: Despite clinical benefit from treatment with dabrafenib and trametinib in melanoma patients with BRAF mutations, half relapse within months and one-third are unresponsive to treatment. We evaluated the anticancer potential of metformin in combination with trametinib plus paclitaxel, against four melanoma cell lines. MATERIALS AND METHODS: Metformin with trametinib and paclitaxel was tested for effects on cell viability, signaling molecules in MAPK and mTOR pathways, factors involved in epithelial-mesenchymal transition (EMT), and cell motility. RESULTS: The combination of metformin with trametinib and paclitaxel showed differential growth inhibitory effects; synergistic effects were observed in a cell line in which metformin suppresses ERK activity, whereas the combination showed antagonistic effects in a cell line with metformin-induced ERK activation. Trametinib or paclitaxel increased the expression of EMT regulators and melanoma cell motility, which were suppressed by combining metformin with trametinib and paclitaxel. CONCLUSION: The combined treatment of metformin with trametinib and paclitaxel showed divergent effects on melanoma cell viability. Metformin might be useful as a potential adjuvant against cell proliferation and metastatic activity in melanoma patients.

Stepwise achievement of high convection volume in post-dilution hemodiafiltration: A prospective observational study.

High-volume online hemodiafiltration (HDF) has been reported to reduce the patient's mortality. However, achieving a high convection volume is challenging. In this prospective study, we investigated the feasibility of achieving high-volume HDF with ≥21 L substitution volume via modification of blood flow rate (BFR), needle size, and dialysis membrane. In 30 patients undergoing hemodialysis, we followed a stepwise protocol and gradually increased the BFR (280→300→330 ml/min; steps 1, 2, and 3) and needle size (16→15 G; step 4). After changing dialyzer surface area (1.8 m2 →2.5 m2 ), the BFR and needle size were similarly increased stepwise (steps 5, 6, 7, and 8). The mean substitution volume was 18.7 ± 2.2 L at step 1 and it significantly increased to 25.1 ± 2.6 L by step 8. A substitution volume of 21 L was achieved by 13.3% of patients in step 1 and by 96.7% after step 8. The substitution volume was higher for the dialyzer with a large surface area and for the larger needle (15 G). Between steps 1 and 8, the Kt/V and ß2 microglobulin reduction ratios also improved significantly. High-volume HDF is feasible through a stepwise increase in the BFR, needle size, and surface area of the dialysis membrane.

Engineering exosomes for pulmonary delivery of peptides and drugs to inflammatory lung cells by inhalation.

Exosomes have been investigated as delivery vesicles for various drugs. However, exosome-mediated peptide delivery into the lungs has not been studied. In this study, exosomes were engineered for the pulmonary delivery of RAGE-binding peptide (RBP), an anti-inflammatory peptide. To load the peptide into exosomes, RBP was linked to an exosome membrane integral protein, Lamp2b, to produce RBP-linked exosomes (RBP-exo). The anti-inflammatory effects of RBP-exo were confirmed by cytokine assays in lipopolysaccharide (LPS)-activated macrophage cells. To increase anti-inflammatory effects, curcumin was loaded into RBP-exo. Curcumin loaded RBP-exo (RBP-exo/Cur) had higher intracellular curcumin delivery efficiency than curcumin alone or curcumin loaded into unmodified exosomes (unmod-exo/Cur). This suggests that RBP on the surface of RBP-exo may interact with RAGE and increase the intracellular delivery efficiency of curcumin. In addition, RBP-exo/Cur had higher anti-inflammatory effects than curcumin alone, a mixture of RBP and curcumin, and unmod-exo/Cur in vitro. For in vivo evaluation, RBP-exo/Cur was administrated by intratracheal instillation into the lungs of an acute lung injury (ALI) model. The results showed that RBP-exo/Cur reduced pro-inflammatory cytokines more efficiently than curcumin alone, RBP-exo, and unmod-exo/Cur. Hematoxylin and eosin staining confirmed that the inflammation reaction was inhibited in the RBP-exo and RBP-exo/Cur groups. Immunostaining assays showed that RBP-exo was co-localized mostly with type I epithelial cells. In conclusion, RBP was successfully delivered with exosomes into the lungs by inhalation. A combination of RBP and curcumin using exosomes as carriers may be useful as ALI therapy.

Antitumor Activity of Combination Therapy with Metformin and Trametinib in Non-Small Cell Lung Cancer Cells.

Metformin has been widely used as an antidiabetic drug, and reported to inhibit cell proliferation in many cancers including non-small cell lung cancer (NSCLC). In NSCLC cells, metformin suppresses PI3K/AKT/mTOR signaling pathway, but effect of metformin on RAS/ RAF/MEK/ERK signaling pathway is controversial; several studies showed the inhibition of ERK activity, while others demonstrated the activation of ERK in response to metformin exposure. Metformin-induced activation of ERK is therapeutically important, since metformin could enhance cell proliferation through RAS/RAF/MEK/ERK pathway and lead to impairment of its anticancer activity suppressing PI3K/AKT/mTOR pathway, requiring blockade of both signaling pathways for more efficient antitumor effect. The present study tested the combination therapy of metformin and trametinib by monitoring the alterations of regulatory effector proteins of cell signaling pathways and the effect of the combination on cell viability in NCI-H2087 NSCLC cells with NRAS and BRAF mutations. We show that metformin alone blocks PI3K/AKT/mTOR signaling pathway but induces the activation and phosphorylation of ERK. The combination therapy synergistically decreased cell viability in treatment with low doses of two drugs, while it gave antagonistic effect with high doses. These findings suggest that the efficacy of metformin and trametinib combination therapy may depend on the alteration of ERK activity induced by metformin and specific cellular context of cancer cells.

Messenger RNA/polymeric carrier nanoparticles for delivery of heme oxygenase-1 gene in the post-ischemic brain.

Ischemic stroke is a cerebrovascular disease caused by narrowed cerebral arteries. Thrombolytic agents such as tissue-plasminogen activators have been used for recanalization of the blood supply into the ischemic region. However, ischemia-reperfusion damage continues to increase the infarction volume. In this study, heme oxygenase-1 (HO1)-mRNA was delivered into the brain, using a non-viral carrier. Various non-viral carriers such as polyethylenimine (25 kDa, PEI25k), lipofectamine, dexamethasone-conjugated PEI2k (Dexa-PEI2k), deoxycholic acid-conjugated PEI2k (DA-PEI2k), and R3V6 peptides were evaluated as carriers of mRNA into the brain. Gene delivery assays showed that DA-PEI2k and lipofectamine had a higher mRNA delivery efficiency than the other carriers in Neuro2A cells in vitro and a rat brain in vivo. Cytotoxicity assays showed that lipofectamine had higher toxicity than DA-PEI2k. Therefore, DA-PEI2k was used for delivery of HO1-mRNA. Unlike plasmid DNA (pDNA), mRNA is expressed in the cytosol without nuclear translocation. This suggests that mRNA may have higher gene expression than pDNA, since the nuclear location of pDNA is an inefficient step. Indeed, in in vitro transfection assays, HO1-mRNA/DA-PEI2k had higher gene expression than HO1-pDNA/DA-PEI2k without induction of a pro-inflammatory cytokine. The therapeutic effects of HO1-mRNA delivery using DA-PEI2k were evaluated in the middle cerebral artery occlusion animal model after local injection. HO1-mRNA delivery had higher gene expression than HO1-pDNA delivery 24 h after the local injection. In addition, HO1-mRNA delivery reduced the infarct size more efficiently than HO1-pDNA delivery. The results suggest that the delivery of mRNA using DA-PEI2k may be useful for gene therapy of ischemic stroke.

Systemic delivery of microRNA-21 antisense oligonucleotides to the brain using T7-peptide decorated exosomes.

Antisense miRNA oligonucleotides against miR-21 (AMO-21) have a therapeutic potential for treatment of glioblastoma. However, glioblastoma-targeted delivery through systemic injection requires development of an efficient targeting carrier. For this purpose, a glioblastoma-targeting carrier was developed using the T7 peptide and exosomes. The transferrin receptor is overexpressed on the surface of glioblastoma cells, and T7 is a transferrin receptor-binding peptide. A T7 peptide-decorated exosome (T7-exo) was produced by incorporation of T7 into the exosome membrane as a fusion protein of T7 and Lamp2b. As a control, rabies virus glycoprotein (RVG) peptide targeting brain neuron cells was incorporated into the exosome membrane. AMO-21 was loaded into the exosomes by electroporation. In vitro studies of AMO-21 delivery showed that T7-exo had a higher delivery efficiency to C6 glioblastoma cells than unmodified exosome (Unmod-exo) and RVG-decorated exosome (RVG-exo). For in vivo delivery studies, T7-exo with AMO-21 was delivered into intracranial glioblastoma rat models by intravenous injection through the tail vein. The results showed that T7-exo delivered AMO-21 into the brain more efficiently than Unmod-exo and RVG-exo. In addition, delivery of AMO-21 using T7-exo reduced the miR-21 level in the glioblastoma efficiently. Reduction of miR-21 by AMO-21 induced the expression of PDCD4 and PTEN in tumors, resulting in reduction of tumor sizes. Taken together, these findings indicate that T7-exo is an efficient carrier of AMO-21 into the glioblastoma and may be useful in development of glioblastoma therapy.

Synergistic Enhancement of Paclitaxel-induced Inhibition of Cell Growth by Metformin in Melanoma Cells.

Melanoma is one of the most aggressive and treatment-resistant malignancies. Antidiabetic drug metformin has been reported to inhibit cell proliferation and metastasis in many cancers, including melanoma. Metformin suppresses the mammalian target of rapamycin (mTOR) and our previous study showed that it also inhibits the activity of extracellular signal-regulated kinase (ERK). Paclitaxel is currently prescribed for treatment of melanoma. However, paclitaxel induced the activation of ERK/mitogen-activated protein kinase (MAPK) pathway, a cell signaling pathway implicated in cell survival and proliferation. Therefore, we reasoned that combined treatment of paclitaxel with metformin could be more effective in the suppression of cell proliferation than treatment of paclitaxel alone. Here, we investigated the combinatory effect of paclitaxel and metformin on the cell survival in SK-MEL-28 melanoma cell line. Our study shows that the combination of paclitaxel and metformin has synergistic effect on cell survival and suppresses the expression of proteins involved in cancer metastasis. These findings suggest that the combination of paclitaxel and metformin can be a possible therapeutic option for treatment of melanoma.

Effects of aspirin resistance and mean platelet volume on vascular access failure in hemodialysis patients.

BACKGROUND/AIMS: Maintaining the patency of vascular access (VA) in hemodialysis (HD) patients is important and can be life-saving. We investigated the effects of aspirin resistance and mean platelet volume (MPV) on VA failure in HD patients. METHODS: We enrolled 163 patients on maintenance HD. VA failure was defined as thrombosis or a decrease of > 50% of the normal vessel diameter, as revealed by angiography. RESULTS: Aspirin resistance was observed in 17 of 109 patients in whom this parameter was measured, and was not significantly associated with VA failure (p = 0.051). The mean MPV was 9.15 ± 0.05 fL. The 163 patients were grouped by the median MPV value (9.08 fL) at baseline; patients with higher MPVs (n = 82) had lower platelet counts (p = 0.002) and albumin levels (p = 0.009). During 34 months of follow-up, 65 VA failures (39.9%) occurred. The Kaplan-Meier curve revealed significant differences between the two groups in terms of cumulative VA failure (54.1% vs. 35.3%, p = 0.018). On multivariate analysis, the MPV (hazard ratio [HR], 1.794; 95% confidence interval [CI], 1.066 to 3.020; p = 0.028), platelet count (HR, 1.003; 95% CI, 1.001 to 1.006; p = 0.01), and smoking status (HR, 1.894; 95% CI, 1.019 to 3.519; p = 0.043) independently predicted VA failure. CONCLUSION: A high MPV was associated with an increased risk of VA failure, whereas aspirin resistance showed only a weak association. The MPV may predict VA survival in HD patients.

Combination of BEZ235 and Metformin Has Synergistic Effect on Cell Viability in Colorectal Cancer Cells.

Patients with type II diabetes mellitus are more susceptible to colorectal cancer (CRC) incidence than non-diabetics. The anti-diabetic drug metformin is most commonly prescribed for the treatment of this disease and has recently shown antitumor effect in preclinical studies. The aberrant mutational activation in the components of RAS/RAF/MEK/ERK and PI3K/AKT/mTOR signaling pathway is very frequently observed in CRC. We previously reported that metformin inhibits the phosphorylation of ERK and BEZ235, a dual inhibitor of PI3K and mTOR, has anti-tumor activity against HCT15 CRC cells harboring mutations of KRAS and PIK3CA. Therefore, we hypothesized that simultaneous inhibition of two pathways by combining metformin with BEZ235 could be more effective in the suppression of proliferation than single agent treatment in HCT15 CRC cells. Here, we investigated the combinatory effect of metformin and BEZ235 on the cell survival in HCT15 CRC cells. Our study shows that both of the two signaling pathways can be blocked by this combinational strategy: metformin suppressed both pathways by inhibiting the phosphorylation of ERK, 4E-BP1 and S6, and BEZ235 suppressed PI3K/AKT/ mTOR pathway by reducing the phosphorylation of 4E-BP1 and S6. This combination treatment synergistically reduced cell viability. The combination index (CI) values ranged from 0.44 to 0.88, indicating synergism for the combination. These results offer a preclinical rationale for the potential therapeutic option for the treatment of CRC.

Metformin Synergistically Potentiates the Antitumor Effects of Imatinib in Colorectal Cancer Cells.

Metformin is the most commonly prescribed anti-diabetic drug with relatively minor side effect. Substantial evidence has suggested that metformin is associated with decreased cancer risk and anticancer activity against diverse cancer cells. The tyrosine kinase inhibitor imatinib has shown powerful activity for treatment of chronic myeloid leukemia and also induces growth arrest and apoptosis in colorectal cancer cells. In this study, we tested the combination of imatinib and metformin against HCT15 colorectal cancer cells for effects on cell viability, cell cycle and autophagy. Our data show that metformin synergistically enhances the imatinib cytotoxicity in HCT15 cells as indicated by combination and drug reduction indices. We also demonstrate that the combination causes synergistic down-regulation of pERK, cell cycle arrest in S and G2/M phases via reduction of cyclin B1 level. Moreover, the combination resulted in autophagy induction as revealed by increased acidic vesicular organelles and cleaved form of LC3-II. Inhibition of autophagic process by chloroquine led to decreased cell viability, suggesting that induction of autophagy seems to play a cell protective role that may act against anticancer effects. In conclusion, our present data suggest that metformin in combination with imatinib might be a promising therapeutic option in colorectal cancer.

Blockage of Autophagy Rescues the Dual PI3K/mTOR Inhibitor BEZ235-induced Growth Inhibition of Colorectal Cancer Cells.

Molecular targeting for the altered signaling pathways has been proven to be effective for the treatment ofmany types of human cancer, including colorectal cancer (CRC). The dual phosphatidylinositol-3-kinase (PI3K) and mammalian target of rapamycin (mTOR) inhibitor BEZ235 has shown to exhibit potent antitumor activity against solid tumors. Autophagy is a cellular lysosomal catabolic process to maintain metabolic homeostasis, which has been known to be induced in response to many therapeutic agents in cancer cells. This process is negatively regulated by mTOR and often acts as prosurvival or prodeath mechanism following cancer therapeutics. The current study was designed to investigate the antiproliferation activity of BEZ235 and to evaluate the role of autophagy induced by BEZ235 using HCT15 CRC cells bearing ras oncogene mutation. We found that BEZ235 decreases cell viability, which was mostly dependent on G1 arrest of cell cycle via suppression of cyclin A expression. BEZ235 affects PI3K/Akt/mTOR signaling pathway by increasing the phosphorylation of AKT at Ser(473) and RAS/RAF/MEK/ERK pathway by decreasing the phosphorylation of ERK at Tyr(204). BEZ235 also stimulated autophagy induction as evidenced by the increased expression of LC3-II and abundant acidic vesicular organelles (AVOs) in the cytoplasm. In addition, the combination of BEZ235 with autophagy inhibitor chloroquine, a known antagonist of autophagy, counteracted the antiproliferation effect of BEZ235. Thus, our study indicates that autophagy induced in response to BEZ235 treatment appears to act as cell death mechanism in HCT15 CRC cells.

Determination of matrix effects occurred during the analysis of organochlorine pesticides in agricultural products using GC-ECD.

Matrix effects observed during the multiresidue analysis of seven organochlorine pesticides in six different agricultural products with GC-ECD were assessed. The presence of matrix coextractives, a major cause of observed matrix effects, directly and/or indirectly influenced the chromatographic responses of some pesticides. Two types of external calibrations, solvent calibration (SC) and matrixmatched calibration (MC), were used to assess matrix effects. Greater matrix effects were observed at the lower concentrations of each pesticide. The extent of matrix effects varied unpredictably with matrix type. Among the analyzed pesticides, iprodione, cyhalothrin, and cypermethrin exhibited greater matrix effects (>150%) for almost all matrices. The pesticide recovery rates obtained with MC were not statistically different from a 100% recovery rate in most samples, which indicates that MC may diminish the overestimates occurred due to matrix effects in GC analysis.