72-h Treatment Research Articles

Single agent efficacy of the HDAC inhibitor DATS in preclinical models of glioblastoma.

Glioblastoma (GB) remains incurable despite aggressive chemotherapy, radiotherapy, and surgical interventions; immunotherapies remain experimental in clinical practice. Relevant preclinical models that can accurately predict tumor response to therapy are equally challenging. This study aimed to validate the effect of the naturally occurring agent diallyl trisulfide (DATS) in human GB in relevant pre-clinical models. Ex vivo slice culture, in vivo cell line derived orthotopic xenograft and patient-derived orthotopic xenograft (PDX) animal models of GB were utilized to assess efficacy of treatment with DATS. Our results showed 72-h treatments of 25µM DATS induced cell death in ex vivo human GB slice culture. We treated U87MG orthotopic xenograft models (U87MGOX) and patient-derived orthotopic xenograft models (PDX) with daily intraperitoneal injections of DATS for 14 days. Magnetic resonance (MR) imaging of mice treated with DATS (10mg/kg) demonstrated reduced tumor size at 5 weeks when compared with saline-treated U87MGOX and PDX controls. Hematoxylin (H&E) staining demonstrated dose-dependent reduction in gross tumor volume with decreased proliferation and decreased angiogenesis. Western blotting showed that DATS was associated with increases in histone acetylation (Ac-Histone H3/H4) and activated caspase-3 in this novel preclinical model. Histological assessment and enzyme assays showed that even the highest dose of DATS did not negatively impact hepatic function. DATS may be an effective and well-tolerated therapeutic agent in preventing tumor progression and inducing apoptosis in human GB.

Selenious-β-lactoglobulin induces the apoptosis of human lung cancer A549 cells via an intrinsic mitochondrial pathway.

In this study, the cytotoxic activity of selenious-β-lactoglobulin (Se-β-Lg) and the anticancer mechanism were investigated in human lung cancer A549 cells in vitro. MTT assay showed that Se-β-Lg at 200μg/mL exhibited a significant suppression effect on A549 cells and the maximum inhibition rate reached 90% after 72h treatment. Flow cytometry analysis revealed that 200μg/mL of Se-β-Lg induced cell cycle arrest at G0/G1 phase. Cell apoptosis was induced via the generation of reactive oxygen species (ROS) and the decrease of mitochondrial membrane potential (ΔΨm) in a time-dependent manner. Furthermore, Se-β-Lg suppressed the expression of Bcl-2 and improved the level of Bax, leading to the release of cytochrome c and a higher expression of caspase-3 in A549 cells. In summary, Se-β-Lg could induce apoptosis in A549 cells via an intrinsic mitochondrial pathway and it might serve as a potential therapeutic agent for human lung cancer.

Prodigiosin produced by Serratia marcescens inhibits expression of MMP-9 and survivin and promotes caspase-3 activation with induction of apoptosis in acute lymphoblastic leukaemia cells.

Matrix metalloproteinase-9 (MMP-9) and survivin are involved in several steps of carcinogenesis in acute lymphoblastic leukaemia (ALL). Yet, no MMP-9 and survivin-modulating drugs with low toxicity on normal cells but high efficacy against high MMP-9- and survivin-expressing leukaemia cells have been approved for clinical application in ALL. Prodigiosin, a secondary metabolite of Serratia marcescens, induces apoptosis in different kinds of cancer cells with low toxicity on normal cells. However, little is known about the effects of this compound on the high MMP-9- and survivin-expressing leukaemia cells. CCRF-CEM cells as a model for high MMP-9- and survivin-expressing ALL cells were treated with 100, 200 and 400nmoll-1 prodigiosin after which cell number, proliferation rate, MMP-9 and survivin expression, caspase-3 activation and apoptosis were evaluated. After 24-, 48-, and 72-h treatments with 100, 200 and 400nmoll-1 prodigiosin, proliferation rates were measured to be 92·3-76·7%, 82-63% and 63·7-46·6% respectively. Treatment with prodigiosin for 48h decreased MMP-9 mRNA levels followed by decreases in secreted (S) and intracellular (I) MMP-9 protein levels by 20-22% and 69-72% for 100-400nmoll-1 prodigiosin respectively. Prodigiosin decreased survivin protein levels from 40 to 26% followed by 3·7-5·6-fold increases in caspase-3 activation for the aforementioned prodigiosin concentration ranges. Treatment with 100-400nmoll-1 prodigiosin increased the caspase-3/survivin, caspase-3/I-MMP-9 and caspase-3/S-MMP-9 ratios by 6-7·3-, 11·5-19·1- and 4·9-6·8-fold increases respectively. A dramatic increase in the number of apoptotic cells was also observed with increasing prodigiosin concentrations. The inhibitory effects of prodigiosin on MMP-9 and survivin expression, as well as its pro-apoptotic capacity, represent a novel therapeutic avenue against ALL cells. These findings provide an important and interesting basis to develop a new therapeutic compound with high potential against ALL cells.

Combined toxicity of chlorantraniliprole, lambda-cyhalothrin, and imidacloprid to the silkworm Bombyx mori (Lepidoptera: Bombycidae).

Insecticides with different modes of action may act in combination, in ways such as drifting, spray equipment residual, or utilizing concurrently in mulberry orchards or nearby agricultural fields. Silkworms may suffer from a diverse impact on the survival. In this study, the toxicity of chlorantraniliprole, lambda-cyhalothrin, and imidacloprid and their combinations to the second instar of silkworms (Bombyx mori (L.)(Lepidoptera: Bombycidae)) were evaluated after 48 and 72h treatment by the leaf-dipping method and the combination index (CI)-isobologram equation. After 48h treatment, results indicated that (1) the increasing order of toxicity was imidacloprid < chlorantraniliprole < lambda-cyhalothrin, and that (2) synergism was predominated in most combinations excepted for the lambda-cyhalothrin + imidacloprid combination which displayed an additive effect at fa value 0.5. Then, after 72h treatment, results exhibited that (1) the increasing order of toxicity was imidacloprid < lambda-cyhalothrin < chlorantraniliprole, and that (2) only the chlorantraniliprole + imidacloprid mixture yielded antagonism at fa value 0.5; the other combinations performed an additive effect at least. Consequently, combined toxicity of mixtures may pose a worse effect on silkworm than single toxicity of insecticides. Therefore, we suggest that insecticide mixtures should be added into ecotoxicological risk assessment.

Omega-3 fatty acid DHA modulates p53, survivin, and microRNA-16-1 expression in KRAS-mutant colorectal cancer stem-like cells

BackgroundThe presence of chemotherapy-resistant colorectal cancer stem cells (CCSCs) with KRAS mutation is thought to be one of the primary causes for treatment failure in colorectal cancer (CRC). P53, survivin, and microRNA-16-1 are challenging targets for anticancer drugs which are associated with chemoresistance in CRC. Yet, no p53-, survivin-, and microRNA-16-1-modulating drug with low toxicity but high efficacy against KRAS-mutant CCSCs have been approved for clinical application in CRC. Here, we investigated whether in vitro concentrations of DHA equal to human plasma levels, are able to modulate, Wt-p53, survivin, and microRNA-16-1 in CRC cells with stem cell-like properties.MethodsWt-p53/KRAS-mutant CRC cells (HCT-116) with stem cell-like properties were treated with 100-, 150- and 200-μM/L DHA, after which cell number, viability, growth inhibition, Wt-p53, survivin and microRNA-16-1 expression, caspase-3 activation and apoptotic-rate were evaluated by different cellular and molecular techniques.ResultsAfter 24-, 48-, and 72-h treatments with 100- to 200-μM/L DHA, growth inhibition- rates were measured to be 54.7% to 59.7%, 73.% to 75.8%, and 63.3% to 97.7%, respectively. Treatment for 48 h with indicated DHA concentrations decreased cell number and viability. In addition, we observed a decrease in both the transcript and protein levels of survivin followed by 1.3- to 1.7- and 1.1- to 4.7-fold increases in the Wt-p53 accumulation and caspase-3 activation levels respectively. Treatment with 100 and 150 μM/L DHA increased microRNA-16-1 expression levels by 1.3- to 1.7-fold and enhanced the microRNA-16-1/survivin mRNA, p53/survivin, and caspase-3/survivin protein ratios by 1.7- to 1.8-, 1.3- to 2.6-, and 1.3- to 2-fold increases respectively. A decrease in the number of live cells and an increase in the number of apoptotic cells were also observed with increasing DHA concentrations.ConclusionWt-p53, survivin, and microRNA-16-1 appear to be promising molecular targets of DHA. Thus, DHA might represent an attractive anti-tumor agent directed against KRAS-mutant CCSCs.

Fibroblast growth factor 23 does not directly influence skeletal muscle cell proliferation and differentiation or ex vivo muscle contractility.

Skeletal muscle dysfunction accompanies the clinical disorders of chronic kidney disease (CKD) and hereditary hypophosphatemic rickets. In both disorders, fibroblast growth factor 23 (FGF23), a bone-derived hormone regulating phosphate and vitamin D metabolism, becomes chronically elevated. FGF23 has been shown to play a direct role in cardiac muscle dysfunction; however, it is unknown whether FGF23 signaling can also directly induce skeletal muscle dysfunction. We found expression of potential FGF23 receptors ( Fgfr1-4) and α-Klotho in muscles of two animal models (CD-1 and Cy/+ rat, a naturally occurring rat model of chronic kidney disease-mineral bone disorder) as well as C2C12 myoblasts and myotubes. C2C12 proliferation, myogenic gene expression, oxidative stress marker 8-OHdG, intracellular Ca2+ ([Ca2+]i), and ex vivo contractility of extensor digitorum longus (EDL) or soleus muscles were assessed after treatment with various amounts of FGF23. FGF23 (2-100 ng/ml) did not alter C2C12 proliferation, expression of myogenic genes, or oxidative stress after 24- to 72-h treatment. Acute or prolonged FGF23 treatment up to 6 days did not alter C2C12 [Ca2+]i handling, nor did acute treatment with FGF23 (9-100 ng/ml) affect EDL and soleus muscle contractility. In conclusion, although skeletal muscles express the receptors involved in FGF23-mediated signaling, in vitro FGF23 treatments failed to directly alter skeletal muscle development or function under the conditions tested. We hypothesize that other endogenous substances may be required to act in concert with FGF23 or apart from FGF23 to promote muscle dysfunction in hereditary hypophosphatemic rickets and CKD.

Low-Temperature Fumigation of Harvested Lettuce Using a Phosphine Generator.

A research-scale phosphine generator, QuickPHlo-R, from United Phosphorus Ltd. (Mumbai, India) was tested to determine whether it was suitable for low-temperature fumigation and oxygenated phosphine fumigation of harvested lettuce. Vacuum cooled Iceberg and Romaine lettuce (Lactuca sativa) were fumigated in 442-liter chambers at 2°C for 24 and 72 h for control of western flower thrips [Frankliniella occidentalis (Pergande) (Thysanoptera: Thripidae)] and lettuce aphid [Nasonovia ribisnigri (Mosely) (Homoptera: Aphididae)]. Oxygenated phosphine fumigation for 48 h under 60% O2 was also conducted at 2°C with Iceberg and Romaine lettuce for control of lettuce aphid. The generator completed phosphine generation in 60-90 min. Complete control of western flower thrips was achieved in 24-h treatment, and the 48-h oxygenated fumigation, and 72-h regular fumigation treatments completely controlled lettuce aphid. Lettuce quality was evaluated 14 d after fumigation. There was increased incidence of brown stains on fumigated Iceberg lettuce, and the increases were more obvious in longer (≥48 h) treatments. Both Iceberg and Romaine lettuce from all treatments and controls had good visual quality even though there was significantly higher brown stain incidence on fumigated Iceberg lettuce in ≥48-h treatment and significant differences in quality score for both Iceberg and Romaine lettuce in the 72-h treatment. The brown stains were likely due to the high sensitivity of lettuce to carbon dioxide. The study indicated that QuiPHlo-R phosphine generator has potential in low-temperature phosphine fumigation due to the quick establishment of desired phosphine levels, efficacy in pest control, and reasonable safety to product quality.

New HSP90 selective inhibitors as therapeutic agents for prostate and bladder cancer.

285 Background: Hsp90 is a molecular chaperone responsible for folding many of the proteins directly associated with cancer progression and consequently, inhibition of the Hsp90 protein folding machinery results in a combinatorial attack on numerous oncogenic pathways. Hsp90 family consist of four isoforms; Hsp90α, Hsp90β, Grp94 and Trap-1. The development of Hsp90 isoform-selective inhibitors represent an alternative approach towards the treatment of cancer that may limit some of the detriments. We demonstrate novel Hsp90 inhibitors, on prostate and bladder cancer cells, which shows both potent antiproliferative effects and specific selectivity for Hsp90β. Methods: PC3MM2, LNCap-LN3, C4-2b, LAPC4 (prostate cancer) and T24, UC3 (bladder cancer) cancer cells were utilized. Cell Titer-Glo luminescent anti-proliferative assay was used to determine the IC50 numbers after 72h treatment. Trypan Blue Cytotoxicity assay was performed for 24h treatment with increasing concentrations of KUNB inhibitors. Effects of KUNB inhibitors on Hsp90’s client protein degradation were investigated by Western Blot. Results: KUNB31 manifested an IC50 of 3.00 µM against UC3 bladder cancer cells, UC3 cells were then evaluated via western blot analyses of known Hsp90α- and Hsp90β-dependent client proteins following treatment with KUNB31 for 24 hours. The data showed that, KUNB31 would not induce the heat shock response like 17AAG, and did cause Hsp90β related protein degradation (CXCR4). Moreover, Hsp27, PKM2, Her2, Hsf-1and Akt all showed degradation to different extent. KUNB105 exhibited potent anti-proliferative in both prostate and bladder cancer cells. IC50 number was determined as 1.24 µM for PC3MM2, 1.18 µM for LNCap-LN3, 1.03 µM for C4-2b, 2.56 µM for LAPC4, 0.20 µM for T24, and 0.30 µM for UC3 cancer cells. Conclusions: KUNB novel Hsp90β selective inhibitors, exhibit potent anti-proliferative and cytotoxic activity along with client protein degradation, without induction of HSR in prostate and bladder cancer cell lines. KUNB compound’s selective inhibition on Hsp90β isomers supports the development of Hsp90-selective inhibitors as a method to overcome the detriments associated with pan-inhibition in cancer treatment.

Effects of α-zearalenol on the metabolome of two breast cancer cell lines by 1H-NMR approach.

Zearalenone (ZEN) is one of the most widely distributed toxins that contaminates many crops and foods. Its major metabolites are α-Zearalenol (α-zol) and β-Zearalenol. Previous studies showed that ZEN and α-zol have estrogenic properties and are able to induce growth promoting effect in breast tissues. Considering that tumorigenesis is dependent on the reprogramming of cellular metabolism and that the evaluation of the cellular metabolome is useful to understand the metabolic changes that can occur during the cancer development and progression or after treatments, aim of our work is to study, for the first time, the effects of α-zol on the metabolomic profile of an estrogen positive breast cancer cell line, MCF-7, and of an estrogen negative breast cancer cell lines MDA-MB231. Firstly, we tested the effects of α-zol on the cell viability after 24, 48 and 72h of treatments with 10-10, 10-8 and 10-6M concentrations on breast cancer MCF-7 and MDA-MB231 cell lines in comparison to human non-cancerous breast MCF10A cell line. Then, we evaluated cell cycle progression, levels of reactive oxygen species (ROS) and the metabolomic profiling by 1H-NMR approach on MCF-7 and MDA-MB231 before and after 72h treatments. Principal component analysis was used to compare the obtained spectra. α-zol is resulted able to induce: (i) an increase of the cell viability on MCF-7 cells mainly after 72h treatment, (ii) a slight decrease of the cell viability on MDA-MB231 cells, and (iii) an increase of cells in S phase of the cell cycle and of ROS only in MCF-7 cells. Moreover, the evaluation of metabolomics profile evidenced that after treatment with α-zol the levels of some metabolites increased in MCF-7 cells whereas decreased slightly in MDA-MB231 cells. Our results showed that α-zol was able to increase the protein biosynthesis as well as the lipid metabolism in MCF-7 cells, and, hence, to induce an estrogen positive breast cancer progression.

Evaluation of Glyphosate Ecotoxicity and Biodegradability in the Municipal Wastewaters

Glyphosate (G) is a broad-spectrum systemic organophosphate herbicide being widely used to control weeds in agricultural fields and urban areas. Its safety for both human health and aquatic biomes is a subject of wide debate. This study was aimed at evaluating the removal efficiency and ecotoxicity of G based herbicide (GBH) Klinik® (Nufarm, Austria) added to the raw municipal wastewater (WW) in a lab-scale model column system. The effect of oxide ceramics as a filtering medium (treatment „B”), as well as activated sludge and nutrients (treatment „C”) was compared with the control columns, which contained only WW (treatment „A”). After 72h treatment of WW spiked with 100 mg/L G, the lowest G concentration was detected in the treatment „B”, i.e., 79.4±0.6 mg/L. Treatments „A” and „C” resulted in the remaining G concentrations of 83.3±3.8 and 89.6±3.7 mg/l, respectively. The second addition of 100 mg/L G to the columns followed by 72h incubation also showed the advantage of oxide ceramics. Most probably, this effect could be explained by coupled sorption and biodegradation processes. Experiments were accompanied by microbiological (colony forming units; biological oxygen demand) and ecotoxicological (Daphtoxkit F magna, MicroBioTests) testing. The results indicated that WW-derived microorganisms resist the presence of GBH in the tested concentration range of G, i.e., up to 300 mg/L. As for Daphnia magna, the 24-h EC50 for the GBH Klinik® under standard conditions and in raw WW was found to be 22 mg/L and 6 mg/L G, respectively.

Quercetin exerts an inhibitory effect on cellular bioenergetics of the B164A5 murine melanoma cell line

Modulation of mitochondrial bioenergetics and glycolysis in malignancies has recently emerged a potential chemotherapeutic strategy since numerous malignant cells have overcome inhibition of the glycolytic pathway by increasing mitochondrial ATP production. Quercetin is a flavonoid with antioxidant, antiangiogenic, and chemoprotective properties but the mitochondrial effects are less characterized. The present study was purported to assess the effects of quercetin on the bioenergetic profile of B164A5 murine melanoma cell line. The oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) were measured at 24, 48, and 72h of treatment of B164A5 murine melanoma cells with increasing concentrations (25, 50, 100, and 150µM) of quercetin using the extracellular flux analyzer Seahorse XF24e (Seahorse Agilent). Analysis of mitochondrial function was performed in the presence of the classic modulators of the electron transport chain: oligomycin, FCCP, and rotenone. 72-h treatment with quercetin induced a dose-dependent decrease of all OCR parameters (basal respiration, proton leak, ATP turnover, maximal respiration, reserve capacity) as well as of ECAR. At variance, 48-h treatment induced a decrease of OCR and ECAR when quercetin was applied at 50, 100, and 150µM, while the 24-h treatment induced a decrease of bioenergetic parameters only for the highest concentrations (100 and 150µM) of the compound. Our data clearly demonstrated that quercetin elicited dose-dependent inhibitory effect on examined parameters of cellular bioenergetics that was most potent at 72h of treatment. Thereby quercetin, modulating both glycolytic and mitochondrial pathways for ATP production, might be an efficient approach in killing cancer cells.

Abstract B037: The establishment and evaluation of anti-ASC amino acid transporter 2 (ASCT2) antibody as a novel therapeutic antibody for gastric cancer

Abstract Introduction: Glutamine is the most abundant amino acid in blood, serving as a major biosynthetic substrate in nucleic acids and proteins. Recent studies have shown that glutamine contributes to cancer cell proliferation, invasion, and metastasis, so targeting the glutamine pathway is expected to be a feasible approach to cancer treatment. In cancer cells, the Na+-dependent neutral amino acid exchanger alanine-serine-cysteine (ASC) transporter 2 (ASCT2) plays a key role in the glutamine uptake. ASCT2 has been shown to be overexpressed in a variety of cancerous tissues, including gastric cancer. In this study, we established a defucosylated humanized anti-human ASCT2 monoclonal antibody, KM8094, that specifically binds to the natural configuration of an extracellular domain of ASCT2. We further evaluated the efficacy of KM8094 as a therapeutic agent for gastric cancer treatment using gastric cancer cell lines. Materials and Methods: Immunohistochemistry was conducted on tumor tissue specimens. The ASCT2 expression on tumor cell-surface was analyzed by flow cytometry using KM8094 and PE-conjugated anti-human IgG monoclonal antibody. The glutamine uptake in human gastric cancer cells (SNU-16) was examined using 14C-labeled glutamine. Cell growth was measured by a CellTiter-Glo Luminescent Cell Viability Assay after 72-h treatment with KM8094 or control IgG. The in vivo anti-tumor activity of KM8094 combined with docetaxel was examined using gastric cancer xenograft models. Results: In the immunohistochemical analysis, we observed the high expression of ASCT2 in human tumor tissues, including gastric cancer. To evaluate the inhibitory activity on the ASCT2 transporter function and tumor cell growth of KM8094, we performed a glutamine uptake assay using 14C-glutamine. As a result, KM8094 inhibited the 14C-glutamine uptake in SNU-16 gastric cancer cells expressing ASCT2. KM8094 also suppressed cell growth of multiple gastric cancer cell lines both in vitro and in vivo. These results suggested that KM8094 has therapeutic potential for gastric cancer by inhibiting ASCT2 transporter activity. Furthermore, in an SNU-16 gastric cancer xenograft model, combination therapy of KM8094 and docetaxel, a therapeutic drug for gastric cancer, showed a more potent antitumor effect than either agent alone. Conclusions: KM8094 may be a potential new therapeutic agent for gastric cancer expressing ASCT2, as it exhibited antitumor growth activities both in vitro and in vivo. Citation Format: Aya Sasakawa, Kenta Hosomi, Toshihiko Ishii, Hiroshi Ando, Masayo Suzuki, Minami Imaizumi, Jian Zou, Noriyuki Kasai, Kazuya Yamano, Kazuyasu Nakamura, Ryuichiro Nakai. The establishment and evaluation of anti-ASC amino acid transporter 2 (ASCT2) antibody as a novel therapeutic antibody for gastric cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr B037.

The Cytotoxic Effects of Benzoic Acid and Coumarin on Allium cepa var. cepa

Benzoic acid and coumarin are found naturally in many plants and can be used as food additives in various food products. They are also synthetic allelopathic chemicals that inhibit plant growth and development. The cytotoxic effects of different concentrations of benzoic acid (30 ppm and 60 ppm) and coumarin (0.2 ppm and 0.4 ppm) were investigated on Allium cepa. EC50 (effective concentration) values were calculated according to relative reduction in root length (T/C %) after 72-h treatment. Germination percentages of primary A. cepa roots decreased with increasing concentrations of benzoic acid but not for coumarin. This is maybe a cause of hormesis effect. Cytological observations showed that the mitotic frequency in root meristematic cells decreased with increasing concentration of benzoic acid. Mitotic frequency increased with increasing concentration with coumarin treatment. Abnormality frequency increased in all concentrations of benzoic acid and coumarin when compared with the control group.

Cytotoxic, proapoptotic and antioxidative potential of flavonoids isolated from propolis against colon (HCT-116) and breast (MDA-MB-231) cancer cell lines

Isolated and structurally confirmed, eleven flavonoids from propolis were examined for their cytotoxicity toward human colon cancer and human breast cancer cells. Their effect on induction of apoptosis and their antioxidative activities were also evaluated. Six flavonoids induced cytotoxic effects in both cell lines. Luteolin had a marked effect on both cell lines, especially on HCT-116 cells (IC50 72h, 66.86μM). Also, luteolin was observed to have the highest apoptotic potential after 72h treatment of examined cell lines (27.13% and 37.09%, respectively). Myricetin exhibited selective inhibition of cell growth (IC50 114.75μM) and induced apoptosis in MDA-MB-231 cells only. Luteolin and galangin exhibited prooxidative properties 24h after the treatment in HCT-116 cells, while myricetin induced prooxidative effects in MDA-MB-231 cells. On the other hand, selected flavonoids exhibited antioxidative properties 72h after the treatment, decreasing superoxide anion radical and nitrite levels in both cell lines. Cytotoxic and proapoptotic effects on colon and breast cancer cell lines and the influence on their redox status make tested flavonoids good candidates for developing new anticancer drugs.

Increased Loading, Efficacy and Sustained Release of Silibinin, a Poorly Soluble Drug Using Hydrophobically-Modified Chitosan Nanoparticles for Enhanced Delivery of Anticancer Drug Delivery Systems.

Conventional delivery of anticancer drugs is less effective due to pharmacological drawbacks such as lack of aqueous solubility and poor cellular accumulation. This study reports the increased drug loading, therapeutic delivery, and cellular accumulation of silibinin (SLB), a poorly water-soluble phenolic compound using a hydrophobically-modified chitosan nanoparticle (pCNP) system. In this study, chitosan nanoparticles were hydrophobically-modified to confer a palmitoyl group as confirmed by 2,4,6-Trinitrobenzenesulfonic acid (TNBS) assay. Physicochemical features of the nanoparticles were studied using the TNBS assay, and Attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) analyses. The FTIR profile and electron microscopy correlated the successful formation of pCNP and pCNP-SLB as nano-sized particles, while Dynamic Light Scattering (DLS) and Field Emission-Scanning Electron Microscopy (FESEM) results exhibited an expansion in size between pCNP and pCNP-SLB to accommodate the drug within its particle core. To evaluate the cytotoxicity of the nanoparticles, a Methylthiazolyldiphenyl-tetrazolium bromide (MTT) cytotoxicity assay was subsequently performed using the A549 lung cancer cell line. Cytotoxicity assays exhibited an enhanced efficacy of SLB when delivered by CNP and pCNP. Interestingly, controlled release delivery of SLB was achieved using the pCNP-SLB system, conferring higher cytotoxic effects and lower IC50 values in 72-h treatments compared to CNP-SLB, which was attributed to the hydrophobic modification of the CNP system.

Rationale and Design of Low-dose Administration of Carperitide for Acute Heart Failure (LASCAR-AHF).

Despite current therapies, acute heart failure (AHF) remains a major public health burden with high rates of in-hospital and post-discharge morbidity and mortality. Carperitide is a recombinantly produced intravenous formulation of human atrial natriuretic peptide that promotes vasodilation with increased salt and water excretion, which leads to reduction of cardiac filling pressures. A previous open-label randomized controlled study showed that carperitide improved long-term cardiovascular mortality and heart failure (HF) hospitalization for patients with AHF, when adding to standard therapy. However, the study was underpowered to detect a difference in mortality because of the small sample size. Low-dose Administration of Carperitide for Acute Heart Failure (LASCAR-AHF) is a multicenter, randomized, open-label, controlled study designed to evaluate the efficacy of intravenous carperitide in hospitalized patients with AHF. Patients hospitalized for AHF will be randomly assigned to receive either intravenous carperitide (0.02μg/kg/min) in addition to standard treatment or matching standard treatment for 72h. The primary end point is death or rehospitalization for HF within 2years. A total of 260 patients will be enrolled between 2013 and 2018. The design of LASCAR-AHF will provide data of whether carperitide reduces the risk of mortality and rehospitalization for HF in selected patients with AHF.

Preparation, characterization and cytotoxic evaluation of bovine serum albumin nanoparticles encapsulating 5-methylmellein: A secondary metabolite isolated from Xylaria psidii

In this study, 5-methylmellein (5-MM) loaded bovine serum albumin nanoparticles (BSA NPs) were developed using desolvation technique. The developed nanoparticles were characterized for their mean particle size, polydispersity, zeta potential, loading efficiency, X-ray diffractometry (XRD), differential scanning calorimetry (DSC) and release profile. The developed nanoparticles were spherical in shape under transmission electron microscopy (TEM) and atomic force microscopy (AFM). The developed 5-MM loaded BSA NPs demonstrated a mean particle size with a diameter of 154.95 ± 4.44 nm. The results from XRD and DSC studies demonstrated that the crystal state of the 5-MM was converted to an amorphous state in polymeric matrix. The encapsulation and loading efficiency was found to be 73.26 ± 4.48% and 7.09 ± 0.43%. The in vitro cytotoxicity in human prostate cancer cell line (PC-3), human colon cancer cells (HCT-116) and human breast adenocarcinoma cell line (MCF-7) cells demonstrated enhanced cytotoxicity of 5-MM BSA NPs as compared to native 5-MM after 72-h treatment. The enhancement in cytotoxicity of 5-MM BSA NPs was also supported by increase in cellular apoptosis, mitochondrial membrane potential loss and generation of high reactive oxygen species (ROS). In conclusion, these findings collectively indicated that BSA nanoparticles may serve as promising drug delivery system for improving the efficacy of 5-methylmellein.

Gold nanoparticles induce cell death and suppress migration of melanoma cells

Diverse nanoparticles have been widely applied in different fields, including industries and biomedical sciences. Recent developments in nanotechnology have broadened the potential applications of nanotechnology in clinical diagnosis and medical therapy of malignant and non-malignant diseases. Gold nanoparticles (AuNPs) have been reported to possess unique physico-chemical properties; they have been investigated in studies on tumor growth and metastasis. Here, we explored the possible pharmacological functions of AuNPs on the mouse melanoma cell line, B16F10. We demonstrated that AuNPs of sizes 1–3, 3–5, and 10–15 nm exerted dose-dependent cytotoxic effects after 72-h treatments. AuNPs (4 and 10 ppm) of 1–3 and 3–5 nm were more cytotoxic than those of 10–15 nm. However, AuNPs of 1–3 nm were more cytotoxic against benign HaCaT keratinocytes than those of 3–5 nm. Thus, 4 ppm of AuNPs of 3–5 nm was considered relatively safe and an ideal candidate for this study. Further investigations revealed that AuNPs could induce sub-G1 phase accumulation, cleavage of caspase-3 and poly-(ADP-ribose) polymerase, and activation of JNK/p38. The expression of pro-apoptotic protein, Bax, was also induced. Furthermore, platelet-derived growth factor BB-induced migration and motility of melanoma cells were reduced after pretreatment with AuNPs of 3–5 nm. Adhesion assay showed that AuNPs of 3–5 nm could significantly suppress the adhesion of melanoma cells to collagen. In summary, AuNPs of 3–5 nm might be potential anti-tumor agents, as they were selectively cytotoxic against and inhibited motility of melanoma cells.

Fish-oil-derived eicosapentaenoic acid decreases survivin expression and induces wt-p53 accumulation with caspase-3 activation in acute lymphoblastic leukemia cells.

Defects in modulating wild-type (wt) p53 and survivin are associated with a resistant disease in acute lymphoblastic leukemia (ALL). Yet, no wt-p53 and survivin modulating drugs have been approved for clinical application in ALL. Here, we investigated if in vitro eicosapentaenoic acid (EPA) concentrations equal to human plasma levels are able to target wt-p53 and survivin. Wt-p53 Molt-4 cells (ALL cell line) were treated with 50, 100, 150, and 200 µM of EPA after which cell number, viability, proliferation rate, survivin expression, wt-p53 accumulation, caspase-3 activation, and apoptosis were evaluated. After 48- and 72-h treatments with EPA at concentrations ranging from 50 to 200 µM, cell proliferation rates were measured to be 71.5-32.6% and 68.2-13.7% and metabolic activities were measured to be 77-44% and 71-26%, respectively. Treatment with 50-200 µM of EPA for 48 h resulted in 14.1-74.6% and 69.5-45.5% decreases in survivin mRNA and protein levels, respectively. EPA induced 1.3-6 and 1.9-20-fold increases in caspase-3 activation and wt-p53 accumulation, respectively. Increase in wt-p53/survivin and caspase-3/survivin ratios from 1 in untreated cells to 20.3 and 5.8 was measured for 150 µM of EPA. Low necrotic rates ranging from 0.3% to 2.8% and an increase in the number of total apoptotic cells (early + late) ranging from 9.8% to 81% were also observed with increasing EPA concentrations. EPA induces strongly wt-p53 with a remarkable decrease in survivin expression, representing an attractive compound to modulate wt-p53 and survivin in ALL cells.

Elimination Rates of Electrolytes, Vitamins, and Trace Elements during Continuous Renal Replacement Therapy with Citrate Continuous Veno-Venous Hemodialysis: Influence of Filter Lifetime

Background/Aims: During continuous renal replacement therapy, relevant losses of nutritional substrates, vitamins, and trace elements via the filter may occur. We investigated filter lifetime efficiency during a 72-h treatment period. Methods: This prospective study included 40 patients undergoing citrate continuous veno-venous hemodialysis (CVVHD). The elimination rates were measured at 24, 48, and 72 h. To assess the influence of filter lifetime, we determined substrate loss every 24 h over a 72-h interval. Results: Filter lifetime did not affect the loss of ionized calcium, inorganic phosphate, magnesium, zinc, folic acid, and vitamin B12. Nevertheless, we did observe clinically significant loss of ionized calcium and inorganic phosphate during CVVHD that required supplementation. Conclusions: CVVHD leads to significant loss of ionized calcium and inorganic phosphate that is independent of the filter lifetime.