Insights on the molecular mechanisms of cytotoxicity induced by AS1411 linked to folate-functionalized DNA nanocages in cancer cells.

Self-assembled multivalent DNA nanocages are an emerging class of molecules useful for biomedicine applications. Here, we investigated the molecular mechanisms of cytotoxicity induced by AS1411 free aptamer, AS1411-linked nanocages (Apt-NCs) and nanocages harboring both folate and AS1411 functionalization (Fol-Apt-NCs) in HeLa and MDA-MB-231 cancer cell lines. The three treatments showed different cytotoxic efficacy and Fol-Apt-NCs resulted the most effective in inhibiting cell proliferation and inducing apoptotic pathways and ROS activation in both HeLa and MDA-MB-231 cells. RNA-seq analysis allowed to identify biological functions and genes altered by the various treatments, depending on the AS1411 route of intracellular entry, highlighting the different behavior of the two cancer cell lines. Notably, Fol-Apt-NCs altered the expression of a subset of genes associated to cancer chemoresistance in MDA-MB-231, but not in HeLa cells, and this may explain the increased chemosensitivity to drugs delivered through DNA nanocages of the triple-negative breast cancer cells.

Gut microbiota dysbiosis in Parkinson disease: A systematic review and pooled analysis.

BACKGROUND AND PURPOSE: The role of the gut microbiome in the pathogenesis of Parkinson disease (PD) is under intense investigation, and the results presented are still very heterogeneous. These discrepancies arise not only from the highly heterogeneous pathology of PD, but also from widely varying methodologies at all stages of the workflow, from sampling to final statistical analysis. The aim of the present work is to harmonize the workflow across studies to reduce the methodological heterogeneity and to perform a pooled analysis to account for other sources of heterogeneity. METHODS: We performed a systematic review to identify studies comparing the gut microbiota of PD patients to healthy controls. A workflow was designed to harmonize processing across all studies from bioinformatics processing to final statistical analysis using a Bayesian random-effects meta-analysis based on individual patient-level data. RESULTS: The results show that harmonizing workflows minimizes differences between statistical methods and reveals only a small set of taxa being associated with the pathogenesis of PD. Increased shares of the genera Akkermansia and Bifidobacterium and decreased shares of the genera Roseburia and Faecalibacterium were most characteristic for PD-associated microbiota. CONCLUSIONS: Our study summarizes evidence that reduced levels of butyrate-producing taxa in combination with possible degradation of the mucus layer by Akkermansia may promote intestinal inflammation and reduced permeability of the gut mucosal layer. This may allow potentially pathogenic metabolites to transit and enter the enteric nervous system.

Folate-Functionalization Enhances Cytotoxicity of Multivalent DNA Nanocages on Triple-Negative Breast Cancer Cells.

DNA is an excellent programmable polymer for the generation of self-assembled multivalent nanostructures useful for biomedical applications. Herein, we developed (i) folate-functionalized nanocages (Fol-NC), very efficiently internalized by tumor cells overexpressing the α isoform of the folate receptor; (ii) AS1411-linked nanocages (Apt-NC), internalized through nucleolin, a protein overexpressed in the cell surface of many types of cancers; and (iii) nanostructures that harbor both folate and AS1411 aptamer functionalization (Fol-Apt-NC). We analyzed the specific miRNA silencing activity of all types of nanostructures harboring miRNA sequestering sequences complementary to miR-21 and the cytotoxic effect when loaded with doxorubicin in a drug-resistant triple-negative breast cancer cell line. We demonstrate that the presence of folate as a targeting ligand increases the efficiency in miR-21 silencing compared to nanocages functionalized with AS1411. Double-functionalized nanocages (Fol-Apt-NC), loaded with doxorubicin, resulted in an increase of over 51% of the cytotoxic effect on MDA-MB-231 cells compared to free doxorubicin, demonstrating, besides selectivity, the ability of nanocages to overcome Dox chemoresistance. The higher efficiency of the folate-functionalized nanocages is due to the way of entrance, which induces more than four times higher intracellular stability and indicates that the folate-mediated route of cell entry is more efficient than the nucleolin-mediated one when both folate and AS1411 modifications are present.

Dimethylmyricacene: An In Vitro and In Silico Study of a Semisynthetic Non-Camptothecin Derivative Compound, Targeting Human DNA Topoisomerase 1B.

Human topoisomerase 1B regulates the topological state of supercoiled DNA enabling all fundamental cell processes. This enzyme, which is the unique molecular target of the natural anticancer compound camptothecin, acts by nicking one DNA strand and forming a transient protein-DNA covalent complex. The interaction of human topoisomerase 1B and dimethylmyricacene, a compound prepared semisynthetically from myricanol extracted from Myrica cerifera root bark, was investigated using enzymatic activity assays and molecular docking procedures. Dimethylmyricacene was shown to inhibit both the cleavage and the religation steps of the enzymatic reaction, and cell viability of A-253, FaDu, MCF-7, HeLa and HCT-116 tumor cell lines.

Machine Learning Data Analysis Highlights the Role of Parasutterella and Alloprevotella in Autism Spectrum Disorders.

In recent years, the involvement of the gut microbiota in disease and health has been investigated by sequencing the 16S gene from fecal samples. Dysbiotic gut microbiota was also observed in Autism Spectrum Disorder (ASD), a neurodevelopmental disorder characterized by gastrointestinal symptoms. However, despite the relevant number of studies, it is still difficult to identify a typical dysbiotic profile in ASD patients. The discrepancies among these studies are due to technical factors (i.e., experimental procedures) and external parameters (i.e., dietary habits). In this paper, we collected 959 samples from eight available projects (540 ASD and 419 Healthy Controls, HC) and reduced the observed bias among studies. Then, we applied a Machine Learning (ML) approach to create a predictor able to discriminate between ASD and HC. We tested and optimized three algorithms: Random Forest, Support Vector Machine and Gradient Boosting Machine. All three algorithms confirmed the importance of five different genera, including Parasutterella and Alloprevotella. Furthermore, our results show that ML algorithms could identify common taxonomic features by comparing datasets obtained from countries characterized by latent confounding variables.

From Antarctica to cancer research: a novel human DNA topoisomerase 1B inhibitor from Antarctic sponge Dendrilla antarctica.

Nature has been always a great source of possible lead compounds to develop new drugs against several diseases. Here we report the identification of a natural compound, membranoid G, derived from the Antarctic sponge Dendrilla antarctica displaying an in vitro inhibitory activity against human DNA topoisomerase 1B. The experiments indicate that membranoid G, when pre-incubated with the enzyme, strongly and irreversibly inhibits the relaxation of supercoiled DNA. This compound completely inhibits the cleavage step of the enzyme catalytic mechanism by preventing protein binding to the DNA. Membranoid G displays also a cytotoxic effect on tumour cell lines, suggesting its use as a possible lead compound to develop new anticancer drugs.

Intestinal Taxa Abundance and Diversity in Inflammatory Bowel Disease Patients: An Analysis including Covariates and Confounders.

Intestinal dysbiosis has been widely documented in inflammatory bowel diseases (IBDs) and is thought to influence the onset and perpetuation of gut inflammation. However, it remains unclear whether such bacterial changes rely in part on the modification of an IBD-associated lifestyle (e.g., smoking and physical activity) and diet (e.g., rich in dairy products, cereals, meat and vegetables). In this study, we investigated the impact of these habits, which we defined as confounders and covariates, on the modulation of intestinal taxa abundance and diversity in IBD patients. 16S rRNA gene sequence analysis was performed using genomic DNA extracted from the faecal samples of 52 patients with Crohn's disease (CD) and 58 with ulcerative colitis (UC), which are the two main types of IBD, as well as 42 healthy controls (HC). A reduced microbial diversity was documented in the IBD patients compared with the HC. Moreover, we identified specific confounders and covariates that influenced the association between some bacterial taxa and disease extent (in UC patients) or behaviour (in CD patients) compared with the HC. In particular, a PERMANOVA stepwise regression identified the variables "age", "eat yogurt at least four days per week" and "eat dairy products at least 4 days per week" as covariates when comparing the HC and patients affected by ulcerative proctitis (E1), left-sided UC (distal UC) (E2) and extensive UC (pancolitis) (E3). Instead, the variables "age", "gender", "eat meat at least four days per week" and "eat bread at least 4 days per week" were considered as covariates when comparing the HC with the CD patients affected by non-stricturing, non-penetrating (B1), stricturing (B2) and penetrating (B3) diseases. Considering such variables, our analysis indicated that the UC extent differentially modulated the abundance of the Bifidobacteriaceae, Rikenellaceae, Christensenellaceae, Marinifilaceae, Desulfovibrionaceae, Lactobacillaceae, Streptococcaceae and Peptostreptococcaceae families, while the CD behaviour influenced the abundance of Christensenellaceae, Marinifilaceae, Rikenellaceae, Ruminococcaceae, Barnesiellaceae and Coriobacteriaceae families. In conclusion, our study indicated that some covariates and confounders related to an IBD-associated lifestyle and dietary habits influenced the intestinal taxa diversity and relative abundance in the CD and UC patients compared with the HC. Indeed, such variables should be identified and excluded from the analysis to characterize the bacterial families whose abundance is directly modulated by IBD status, as well as disease extent or behaviour.

GATA6 Deficiency Leads to Epithelial Barrier Dysfunction and Enhances Susceptibility to Gut Inflammation.

BACKGROUND AND AIMS: Intestinal barrier dysfunction is a hallmark of inflammatory bowel diseases [IBD], but the mechanisms that lead to such a defect are not fully understood. This study was aimed at characterising the factors involved in the defective barrier function in IBD. METHODS: Transcriptome analysis was performed on colon samples taken from healthy controls [CTR] and IBD patients. Expression of GATA-binding factor 6 [GATA6], a transcription factor involved in intestinal epithelial cell differentiation, was evaluated in colon samples taken from CTR and IBD patients by real-time polymerase chain reaction [PCR] and immunohistochemistry. Intestinal sections of wild-type and Gata6del mice, which exhibit a conditional Gata6 deletion in intestinal epithelial cells and which are either left untreated or receive subcutaneous indomethacin or rectal trinitrobenzene sulphonic acid, were stained with haematoxylin and eosin. In parallel, some Gata6del mice received antibiotics to deplete intestinal flora. Mucosal inflammatory cell infiltration and cytokine production were evaluated by flow cytometry and real-time PCR, respectively, and tight junction proteins were examined by immunofluorescence. Intestinal barrier integrity was assessed by fluorescein isothiocyanate [FITC]-dextran assay. RESULTS: Multiple genes involved in cell commitment/proliferation and wound healing were differentially expressed in IBD compared with CTR. Among these, GATA6 was significantly decreased in the IBD epithelium compared with CTR. In mice, conditional deletion of GATA6 in the intestinal epithelium induced primarily epithelial damage, diminished zonula occludens-1 expression, and enhanced intestinal permeability, ultimately resulting in bacteria-driven local immune response and enhanced susceptibility to gut inflammation. CONCLUSIONS: Reduced expression of GATA6 promotes intestinal barrier dysfunction, thus amplifying intestinal inflammatory pathology.

Validity of Anti-PSMA ScFvD2B as a Theranostic Tool: A Narrative-Focused Review.

Prostate cancer (PCa) is the second leading cause of cancer among men, and its diagnosis and adequate staging are fundamental. Among the biomarkers identified in recent years for PCa management, prostate-specific-membrane-antigen (PSMA), physiologically expressed at a low level on healthy prostate and in other normal tissues and highly overexpressed in PCa, represents a reliable marker ideal for imaging and therapy. The development of anti-PSMA antibodies, such as D2B, demonstrated slow clearance of intact antibodies compared with fragments resulting in low tumor-to-blood ratios; however, the modular structural and functional nature of antibodies allowed the generation of smaller fragments, such as scFvs. In this review of the anti-PSMA antibody fragment scFvD2B, we combined further characterization of its biomolecular and tissue cross-reactivity characteristics with a comprehensive summary of what has already been performed in preclinical models to evaluate imaging and therapeutic activities. A molecular dynamics study was performed, and ScFvD2B occupied a limited conformational space, characterized by low-energy conformational basins, confirming the high stability of the protein structure. In the cross-reactivity study, the weak/absent immunoreactivity in non-tumor tissues was comparable to the PSMA expression reported in the literature. Biodistribution studies and therapeutic treatments were conducted in different animal models obtained by subcutaneous or locoregional injection of PSMA-positive-versus-negative xenografts. The maximum tumor uptake was observed for 123I(SPECT), 124I(PET), and optical imaging, which avoids kidney accumulation (compared with radiometals) and leads to an optimal tumor-to-kidney and tumor-to-background ratios. Regarding its possible use in therapy, experimental data suggested a strong and specific antitumor activity, in vitro and in vivo, obtained using CAR-T or NK-92/CAR cells expressing scFvD2B. Based on presented/reviewed data, we consider that scFvD2B, due to its versatility and robustness, seems to: (i) overcome some problems observed in other studied scFvs, very often relatively unstable and prone to form aggregates; (ii) have sufficient tumor-to-background ratios for targeting and imaging PSMA-expressing cancer; (iii) significantly redirect immune killing cells to PSMA-positive tumors when inserted in second-generation CAR-T or NK-92/CAR cells. These data suggest that our product can be considered the right reagent to fill the gap that still exists in PCa diagnosis and treatment.

AS1411 Aptamer Linked to DNA Nanostructures Diverts Its Traffic Inside Cancer Cells and Improves Its Therapeutic Efficacy.

The nucleolin-binding G-quadruplex AS1411 aptamer has been widely used for cancer therapy and diagnosis and linked to nanoparticles for its selective targeting activity. We applied a computational and experimental integrated approach to study the effect of engineering AS1411 aptamer on an octahedral truncated DNA nanocage to obtain a nanostructure able to combine selective cancer-targeting and anti-tumor activity. The nanocages functionalized with one aptamer molecule (Apt-NC) displayed high stability in serum, were rapidly and selectively internalized in cancer cells through an AS1411-dependent mechanism, and showed over 200-fold increase in anti-cancer activity when compared with the free aptamer. Comparison of Apt-NCs and free AS1411 intracellular distribution showed that they traffic differently inside cells: Apt-NCs distributed through the endo-lysosomal pathway and were never found in the nuclei, while the free AS1411 was mostly found in the perinuclear region and in nucleoli. Molecular dynamics simulations indicated that the aptamer, when linked to the nanocage, sampled a limited conformational space, more confined than in the free state, which is characterized by a large number of metastable conformations. A different intracellular trafficking of Apt-NCs compared with free aptamer and the confined aptamer conformations induced by the nanocage were likely correlated with the high cytotoxic enhancement, suggesting a structure-function relationship for the AS1411 aptamer activity.

Ruthenium(II) Diphosphine Complexes with Mercapto Ligands That Inhibit Topoisomerase IB and Suppress Tumor Growth In Vivo.

Ruthenium(II) complexes (Ru1-Ru5), with the general formula [Ru(N-S)(dppe)2]PF6, bearing two 1,2-bis(diphenylphosphino)ethane (dppe) ligands and a series of mercapto ligands (N-S), have been developed. The combination of these ligands in the complexes endowed hydrophobic species with high cytotoxic activity against five cancer cell lines. For the A549 (lung) and MDA-MB-231 (breast) cancer cell lines, the IC50 values of the complexes were 288- to 14-fold lower when compared to cisplatin. Furthermore, the complexes were selective for the A549 and MDA-MB-231 cancer cell lines compared to the MRC-5 nontumor cell line. The multitarget character of the complexes was investigated by using calf thymus DNA (CT DNA), human serum albumin, and human topoisomerase IB (hTopIB). The complexes potently inhibited hTopIB. In particular, complex [Ru(dmp)(dppe)2]PF6 (Ru3), bearing the 4,6-diamino-2-mercaptopyrimidine (dmp) ligand, effectively inhibited hTopIB by acting on both the cleavage and religation steps of the catalytic cycle of this enzyme. Molecular docking showed that the Ru1-Ru5 complexes have binding affinity by active sites on the hTopI and hTopI-DNA, mainly via π-alkyl and alkyl hydrophobic interactions, as well as through hydrogen bonds. Complex Ru3 displayed significant antitumor activity against murine melanoma in mouse xenograph models, but this complex did not damage DNA, as revealed by Ames and micronucleus tests.

In Vitro and In Silico Characterization of an Antimalarial Compound with Antitumor Activity Targeting Human DNA Topoisomerase IB.

Human DNA topoisomerase IB controls the topological state of supercoiled DNA through a complex catalytic cycle that consists of cleavage and religation reactions, allowing the progression of fundamental DNA metabolism. The catalytic steps of human DNA topoisomerase IB were analyzed in the presence of a drug, obtained by the open-access drug bank Medicines for Malaria Venture. The experiments indicate that the compound strongly and irreversibly inhibits the cleavage step of the enzyme reaction and reduces the cell viability of three different cancer cell lines. Molecular docking and molecular dynamics simulations suggest that the drug binds to the human DNA topoisomerase IB-DNA complex sitting inside the catalytic site of the enzyme, providing a molecular explanation for the cleavage-inhibition effect. For all these reasons, the aforementioned drug could be a possible lead compound for the development of an efficient anti-tumor molecule targeting human DNA topoisomerase IB.

Natural Compounds as Therapeutic Agents: The Case of Human Topoisomerase IB.

Natural products are widely used as source for drugs development. An interesting example is represented by natural drugs developed against human topoisomerase IB, a ubiquitous enzyme involved in many cellular processes where several topological problems occur due the formation of supercoiled DNA. Human topoisomerase IB, involved in the solution of such problems relaxing the DNA cleaving and religating a single DNA strand, represents an important target in anticancer therapy. Several natural compounds inhibiting or poisoning this enzyme are under investigation as possible new drugs. This review summarizes the natural products that target human topoisomerase IB that may be used as the lead compounds to develop new anticancer drugs. Moreover, the natural compounds and their derivatives that are in clinical trial are also commented on.

Combined and selective miR-21 silencing and doxorubicin delivery in cancer cells using tailored DNA nanostructures.

MicroRNAs play an important role in tumorigenesis and, among them, miR-21 is found to be aberrantly up-regulated in various tumors. The tumor-associated antigen, folate receptor alpha is a GPI-membrane protein overexpressed in many malignant tumors of epithelial origin, including ovarian and cervical cancers. Covalently bound octahedral DNA nanocages were functionalized with folate molecules and utilized as scaffolds to engineer four sequestering units with a miR-21 complementary sequence for obtaining biocompatible Fol-miR21-NC non-toxic nanostructures, to be able to selectively recognize folate receptor alpha-overexpressing cancer cells and sequester the oncogenic miR-21. qPCR assays showed that Fol-miR21-NCs reduce the miR-21 expression up to 80% in cancer cells in the first 2 days of treatment. Functional assays demonstrated that miR-21 sequestering leads to up-regulation of miR-21 tumor suppressor targets (i.e., PTEN and Pdcd4), reduction in cancer cell migration, reduction in proliferation, and increase in cell death. Fol-miR21-NCs can be efficiently loaded with the chemotherapeutic agent doxorubicin. Co-delivery of anti-miR-21 and doxorubicin showed additive cytotoxic effects on tumor cells, paving the way for their use as selective nucleic acid drugs.

Can Gut Microbiota Be a Good Predictor for Parkinson's Disease? A Machine Learning Approach.

The involvement of the gut microbiota in Parkinson's disease (PD), investigated in several studies, identified some common alterations of the microbial community, such as a decrease in Lachnospiraceae and an increase in Verrucomicrobiaceae families in PD patients. However, the results of other bacterial families are often contradictory. Machine learning is a promising tool for building predictive models for the classification of biological data, such as those produced in metagenomic studies. We tested three different machine learning algorithms (random forest, neural networks and support vector machines), analyzing 846 metagenomic samples (472 from PD patients and 374 from healthy controls), including our published data and those downloaded from public databases. Prediction performance was evaluated by the area under curve, accuracy, precision, recall and F-score metrics. The random forest algorithm provided the best results. Bacterial families were sorted according to their importance in the classification, and a subset of 22 families has been identified for the prediction of patient status. Although the results are promising, it is necessary to train the algorithm with a larger number of samples in order to increase the accuracy of the procedure.

Quercetin pentaacetate inhibits in vitro human respiratory syncytial virus adhesion.

Human respiratory syncytial virus (hRSV) is one of the main etiological agents of diseases of the lower respiratory tract and is often responsible for the hospitalization of children and the elderly. To date, treatments are only palliative and there is no vaccine available. Natural products show exceptional structural diversity and they have played a vital role in drug research. Several investigations focused on applied structural modification of natural products to improved metabolic stability, solubility and biological actions them. Quercetin is a flavonoid that presents several biological activities, including anti-hRSV role. Some works criticize the pharmacological use of Quercetin because it has low solubility and low specificity. In this sense, we acetylated Quercetin structure and we used in vitro and in silico assays to compare anti-hRSV function between Quercetin (Q0) and its derivative molecule (Q1). Q1 shows lower cytotoxic effect than Q0 on HEp-2 cells. In addition, Q1 was more efficient than Q0 to protect HEp-2 cells infected with different multiplicity of infection (0.1-1 MOI). The virucidal effects of Q0 and Q1 suggest interaction between these molecules and viral particle. Dynamic molecular results suggest that Q0 and Q1 may interact with F-protein on hRSV surface in an important region to adhesion and viral infection. Q1 interaction with F-protein showed ΔG= -14.22 kcal/mol and it was more stable than Q0. Additional, MTT and plate assays confirmed that virucidal Q1 effects occurs during adhesion step of cycle hRSV replication. In conclusion, acetylation improves anti-hRSV Quercetin effects because Quercetin pentaacetate could interact with F-protein with lower binding energy and better stability to block viral adhesion. These results show alternative anti-hRSV strategy and contribute to drug discovery and development.

Plant microRNAs from Moringa oleifera Regulate Immune Response and HIV Infection.

Traditional medicine is often chosen due to its affordability, its familiarity with patient's cultural practices, and its wider access to the local community. Plants play an important role in providing indispensable nutrients, while specific small RNAs can regulate human gene expression in a cross-kingdom manner. The aim of the study was to evaluate the effects of plant-enriched purified extract microRNAs from Moringa oleifera seeds (MO) on the immune response and on HIV infection. Bioinformatic analysis shows that plant microRNAs (p-miRs) from MO belonging to 18 conserved families, including p-miR160h, p-miR166, p-miR482b, p-miR159c, p-miR395d, p-miR2118a, p-miR393a, p-miR167f-3p, and p-miR858b are predicted to target with high affinity BCL2, IL2RA, TNF, and VAV1, all these being involved in the cell cycle, apoptosis, immune response and also in the regulation of HIV pathogenesis. The effects of MO p-miRs transfected into HIV+ PBMCs were analyzed and revealed a decrease in viability associated with an increase of apoptosis; an increase of T helper cells expressing Fas and a decrease of intracellular Bcl2 protein expression. Meanwhile no effects were detected in PBMCs from healthy donors. In CD4+ T cells, transfection significantly reduced cell activation and modified the T cell differentiation, thereby decreasing both central and effector memory cells while increasing terminal effector memory cells. Interestingly, the p-miRs transfection induces a reduction of intracellular HIV p24 protein and a reduction of viral DNA integration. Finally, we evaluated the effect of synthetic (mimic) p-miR858b whose sequence is present in the MO p-miR pool and predicted to target VAV1, a protein involved in HIV-Nef binding. This protein plays a pivotal role in T cell antigen receptor (TCR) signaling, so triggering the activation of various pathways. The transfection of HIV+ PBMCs with the synthetic p-miR858b showed a reduced expression of VAV1 and HIV p24 proteins. Overall, our evidence defines putative mechanisms underlying a supplementary benefit of traditional medicine, alongside current antiretroviral therapy, in managing HIV infection in resource-limited settings where MO remains widely available.

Effect of Low-Protein Diet and Inulin on Microbiota and Clinical Parameters in Patients with Chronic Kidney Disease.

INTRODUCTION: The gut microbiota has coevolved with humans for a mutually beneficial coexistence and plays an important role in health and disease. A dysbiotic gut microbiome may contribute to progression to chronic kidney disease (CKD) and CKD-related complications such as cardiovascular disease. Microbiota modulation through the administration of prebiotics may represent an important therapeutic target. AIM: We sought to evaluate the effects of a low-protein diet (LPD) (0.6 g/kg/day) with or without the intake of the prebiotic inulin (19 g/day) on microbiota and clinical parameters in CKD patients. MATERIALS AND METHODS: We performed a longitudinal, prospective, controlled, and interventional study on 16 patients: 9 patients treated with LPD (0.6 g/kg/day) and inulin (19 g/day) and 7 patients (control group) treated only with LPD (0.6 g/kg/day). Clinical evaluations were performed and fecal samples were collected for a subsequent evaluation of the intestinal microbiota in all patients. These tests were carried out before the initiation of LPD, with or without inulin, at baseline (T0) and at 6 months (T2). The microbiota of 16 healthy control (HC) subjects was also analyzed in order to identify potential dysbiosis between patients and healthy subjects. RESULTS: Gut microbiota of CKD patients was different from that of healthy controls. The LPD was able to significantly increase the frequencies of Akkermansiaceae and Bacteroidaceae and decrease the frequencies of Christensenellaceae, Clostridiaceae, Lactobacillaceae, and Pasteurellaceae. Only Bifidobacteriaceae were increased when the LPD was accompanied by oral inulin intake. We showed a significant reduction of serum uric acid (SUA) and C-reactive protein (CRP) in patients treated with LPD and inulin (p = 0.018 and p = 0.003, respectively), an improvement in SF-36 (physical role functioning and general health perceptions; p = 0.03 and p = 0.01, respectively), and a significant increase of serum bicarbonate both in patients treated with LPD (p = 0.026) or with LPD and inulin (p = 0.01). Moreover, in patients treated with LPD and inulin, we observed a significant reduction in circulating tumor necrosis factor alpha (TNF-α) (p = 0.041) and plasma nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX2) (p = 0.027) levels. We did not find a significant difference in the circulating levels of Interleukin (IL)-1ß (p = 0.529) and IL-6 (p = 0.828) in the two groups. CONCLUSIONS: LPD, associated or not with inulin, modified gut microbiota and modulated inflammatory and metabolic parameters in patients with CKD. Our results suggest that interventions attempting to modulate the gut microbiome may represent novel strategies to improve clinical outcomes in CKD patients and may provide useful therapeutic effects.

In Silico and In Cell Analysis of Openable DNA Nanocages for miRNA Silencing.

A computational and experimental integrated approach was applied in order to study the effect of engineering four DNA hairpins into an octahedral truncated DNA nanocage, to obtain a nanostructure able to recognize and bind specific oligonucleotide sequences. Modeling and classical molecular dynamics simulations show that the new H4-DNA nanocage maintains a stable conformation with the closed hairpins and, when bound to complementary oligonucleotides produces an opened conformation that is even more stable due to the larger hydrogen bond number between the hairpins and the oligonucleotides. The internal volume of the open conformation is much larger than the closed one, switching from 370 to 650 nm3, and the predicted larger conformational change is experimentally detectable by gel electrophoresis. H4-DNA nanocages display high stability in serum, can efficiently enter the cells where they are stable and maintain the ability to bind, and sequester an intracellular-specific oligonucleotide. Moreover, H4-DNA nanocages, modified in order to recognize the oncogenic miR21, are able to seize miRNA molecules inside cells in a selective manner.

Palladium(ii) complexes with thiosemicarbazones derived from pyrene as topoisomerase IB inhibitors.

New palladium complexes with thiosemicarbazonate ligands derived from pyrene exhibit potent antiproliferative activity against A2780 and cisplatin-resistant A2780Cis human ovarian cancer cells, which is dependent on substituent groups of the thiosemicarbazone ligands. Cellular accumulation and distribution studies confirmed that palladium enters the cell nucleus. DNA and topoisomerase IB studies show that one complex is a potent TopIB inhibitor, with selectivity for cancer versus normal cells.