Search

Background Retinal microglial cells (RMCs) play crucial roles in maintaining normal visual functions in a healthy eye. However, the underlying mechanisms of RMCs over-activation manifesting the alterations of sensome profile and inflammation state, which contribute to various retinal neurodegenerative diseases, remain elusive. Here, we aimed to identify the core set of sensome and pro-inflammatory genes and their regulators using transcriptome and data mining approaches. Methods We performed paired-end RNA-sequencing in primary microglial cell cultures treated with TNFα/IFNϒ (10 ng/ml for 12 h) and PBS as a control. Gene enrichment analysis and hierarchical clustering for the differentially expressed transcripts highlight functional pathways and network perturbations. We examined overlaps of the mouse microglial gene expression profiles with the data-mined human sensome and pro-inflammatory marker genes. The core sets of sensome and pro-inflammatory genes were selected and predicted for transcription factors (TFs). The identified TFs in RNA-Seq are validated by the quantitative PCR method. Results TNFα/IFNϒ induced 668 differentially expressed transcripts in retinal microglial cells relative to the control. Furthermore, gene enrichment analysis and the gene expression network revealed activated microglial genes, biological, molecular and inflammatory pathways. The overlapping analysis of the TNFα/IFNϒ-activated microglia genes and the data-mined human gene sets revealed 22 sensome and 61 pro-inflammatory genes. Based on network analysis, we determined 10 genes as the core sets of sensome and pro-inflammatory genes and predicted the top ten TFs that regulate them. The SP110, IRF1 , FLI1 , SP140 (sensome) and RELB, BATF2 , NFKB2 , TRAFD1 , SP100, NFKB1 (inflammation) are differentially expressed between the TNFα/IFNϒ activated and the non-activated microglia which were validated by quantitative PCR. The outcomes indicate that these transcriptional regulators are highly expressed and may regulate the sensome and inflammatory genes of RMCs and switch them to over-activation. Conclusion Our results comprise a powerful, cross-species functional genomics resource for sensome and inflammation of RMCs, which may provide novel therapeutic approaches to prevent retinal neurodegenerative diseases.

Homeostasis of the retinal pigment epithelium (RPE) is essential for the health and proper function of the retina. Regulation of RPE homeostasis is, however, largely unexplored, yet dysfunction of this process may lead to retinal degenerative diseases, including age-related macular degeneration (AMD). Here, we report that chemokine receptor CXCR5 regulates RPE homeostasis through PI3K/AKT signaling and by suppression of FOXO1 activation. We used primary RPE cells isolated from CXCR5-deficient mice and wild type controls, as well as ex vivo RPE-choroidal-scleral complexes (RCSC) to investigate the regulation of homeostasis. CXCR5 expression in mouse RPE cells was diminished by treatment with hydrogen peroxide. Lack of CXCR5 expression leads to an abnormal cellular shape, pigmentation, decreased expression of the RPE differentiation marker RPE65, an increase in the undifferentiated progenitor marker MITF, and compromised RPE barrier function, as well as compromised cell-to-cell interaction. An increase in epithelial-mesenchymal transition (EMT) markers (αSMA, N-cadherin, and vimentin) was noted in CXCR5-deficient RPE cells both in vitro and in age-progression specimens of CXCR5-/- mice (6, 12, 24-months old). Deregulated autophagy in CXCR5-deficient RPE cells was observed by decreased LC3B-II, increased p62, abnormal autophagosomes, and impaired lysosome enzymatic activity as shown by GFP-LC3-RFP reporter plasmid. Mechanistically, deficiency in CXCR5 resulted in the downregulation of PI3K and AKT signaling, but upregulation and nuclear localization of FOXO1. Additionally, inhibition of PI3K in RPE cells resulted in an increased expression of FOXO1. Inhibition of FOXO1, however, reverts the degradation of ZO-1 caused by CXCR5 deficiency. Collectively, these findings suggest that CXCR5 maintains PI3K/AKT signaling, which controls FOXO1 activation, thereby regulating the expression of genes involved in RPE EMT and autophagy deregulation.

Colorectal cancer (CRC) is the third most common cancer diagnosed worldwide making it a serious global challenge. CRC progression results from dysregulated cytoplasmic transcription factors, including signal transducer and activator of transcription (STAT) proteins that are involved in JAK-STAT pathway. The STAT proteins contain a conserved SH2 domain that facilitates the initiation of STAT activation via binding to tyrosine motifs followed by STAT dimerization. The STAT proteins include STAT1, STAT2 and STAT3 which all facilitate therapeutic targets for many drugs, since they are associated with pathogenesis in various cancers such as CRC. Genistein is an efficient chemopreventive phytochemical drug against CRC. The current investigation presents a computational study performed to investigate the molecular interaction between STAT1, STAT2 and STAT3 proteins with genistein. The molecular dynamic simulation was conducted for STAT2 protein. The studies from molecular docking revealed that the interaction of STAT proteins and genistein is predicted to be effective with better binding energies. Furthermore, targeting STAT3 could be an efficient therapeutic target and understanding the interaction between STAT3 and genistein can help to contribute to a better inhibition process for CRC progression. Treatment with genistein led to significant suppression of cell proliferation and STAT3 protein expression in both CRC (HCT 116 and HT-29) cell lines. This further provides development of efficient STAT inhibitors with better potency and bioavailability.

Placental growth factor (PlGF or PGF) is a member of the VEGF (vascular endothelial growth factor) family. It plays a pathological role in inflammation, vascular permeability, and pathological angiogenesis. The molecular signaling by which PlGF mediates its effects in nonproliferative diabetic retinopathy (DR) remains elusive. This study aims to characterize the transcriptome changes of human retinal endothelial cells (HRECs) with the presence and the absence of PlGF signaling. Primary HRECs were treated with the PlGF antibody (ab) to block its activity. The total RNA was isolated and subjected to deep sequencing to quantify the transcripts and their changes in both groups. We performed transcriptome-wide analysis, gene ontology, pathway enrichment, and gene-gene network analyses. The results showed that a total of 3760 genes were significantly differentially expressed and were categorized into cell adhesion molecules, cell junction proteins, chaperone, calcium-binding proteins, and membrane traffic proteins. Functional pathway analyses revealed that the TGF-β pathway, pentose phosphate pathway, and cell adhesion pathway play pivotal roles in the blood-retina barrier (BRB) and antioxidant defense system. Collectively, the data provide new insights into the molecular mechanisms of PlGF’s biological functions in HRECs relevant to DR and diabetic macular edema (DME). The newly identified genes and pathways may act as disease markers and target molecules for therapeutic interventions for the patients with DR and DME refractory to the current anti-VEGF therapy.

Placental growth factor (PlGF) and Angiopoietin (Ang)-1 are two proteins that are involved in the regulation of endothelial cell (EC) growth and vasculature formation. In the retina and endothelial cells, pericytes are the major source of both molecules. The purpose of this study is to examine the association of PlGF and Ang-1 with human EC/pericyte co-cultures and iPSC-derived vascular organoids. In this study, we used co-cultures of human primary retinal endothelial cells (HREC) and primary human retinal pericytes (HRP), western blotting, immunofluorescent analysis, TUNEL staining, LDH-assays, and RNA seq analysis, as well as human-induced pluripotent stem cells (iPSC), derived organoids (VO) to study the association between PlGF and Ang-1. Inhibition of PlGF by PlGF neutralizing antibody in HREC-HRP co-cultures resulted in the increased expression of Ang-1 and Tie-2 in a dose-dependent manner. This upregulation was not observed in HREC and HRP monocultures but only in co-cultures suggesting the association of pericytes and endothelial cells. Furthermore, Vascular endothelial growth factor receptor 1 (VEGFR1) inhibition abolished the Ang-1 and Tie-2 upregulation by PlGF inhibition. The pericyte viability in high-glucose conditions was also reduced by VEGFR1 neutralization. Immunofluorescent analysis showed that Ang-1 and Ang-2 were expressed mainly by perivascular cells in the VO. RNA seq analysis of the RNA isolated from VO in high glucose conditions indicated increased PlGF and Ang-2 expressions in the VO. PlGF inhibition increased the expression of Ang-1 and Tie-2 in VO, increasing the pericyte coverage of the VO microvascular network. Combined, these results suggest PlGF's role in the regulation of Ang-1 and Tie-2 expression through VEGFR1. These findings provide new insights into the neovascularization process in diabetic retinopathy and new targets for potential therapeutic intervention.

Our previous studies found that the C-X-C motif chemokine receptor 5 (CXCR5) loss leads to retinal pigment epithelium (RPE) dysfunction and AMD pathogenesis. The current study aimed to characterize the G protein-coupled receptor (GPCR) structure of CXCR5 and analyze its interactions with AMD-related risk genes. The sequence alignments, homology model of CXCR5 and structural assessment analysis were performed. Data and text mining were then performed to identify AMD-related risk genes and their interaction with CXCR5 using statistical and mathematical algorithms. Sequence alignment and phylogenetic tree analysis revealed that human CXCR5 was highly similar (85.4839%) to the rabbit. The least similarity (33.871%) was found to be in zebrafish compared to the other species. The CXCR5 model structural assessment and secondary structure analysis exhibited an excellent model. Network analysis revealed that IL10, TNF, ICAM1, CXCL1, CXCL8, APP, TLR4, SELL, C3, IL17A and CCR2 were the most connected genes CXCR5. These findings suggest that CXCR5 signaling may regulate the biological function of RPE and modulate AMD pathophysiology via GPCR signaling and interacting with identified AMD risk genes. In summary, the data presented here provide novel and crucial insights into the molecular mechanisms of CXCR5 involvement in AMD.Communicated by Ramaswamy H. Sarma.

Placental growth factor (PlGF) and Angiopoietin (Ang)-1 are two angiogenic factors that play vital roles in vascular cell growth and stabilization. The present study's objective is to examine PlGF's regulation of Ang-1 and its Tie-2 receptor expression in human vascular cells and vasculature. We exploited the cocultures of human primary retinal endothelial cells (HREC) and pericytes (HRP) and the blood vessel or vascular organoids derived from human-induced pluripotent stem cells (iPSC) as experimental models. In the HREC-HRP cocultures, PlGF blockage upregulated the expressions of Ang-1 and Tie-2 in an antibody dose-dependent manner. Upregulation of Ang-1 and Tie-2 by PlGF blockade did not occur in HREC and HRP monocultures but only in HREC-HRP cocultures, indicating the interactions of the two cell types. VEGFR1 inhibition diminished Ang-1 and Tie-2 upregulation caused by PlGF blockade and reduced pericyte variability in high glucose conditions. In the iPSC-derived vascular organoids (VO), PlGF, Ang-1, and Ang-2 were expressed mainly by perivascular cells. Bioinformatics analysis of RNA sequencing data revealed that diabetes-mimicking conditions upregulated PlGF and Ang-2 expressions in the VO cultures. PlGF blockade upregulated Ang-1 and Tie-2 expression and promoted pericyte coverage and association with ECs in the VO. Together, the data suggest that PlGF regulates Ang-1 and Tie-2 expression in part through VEGFR1, which is involved in vascular cell function and stabilization. The findings may help design new therapeutic interventions for diabetic vasculopathy, such as diabetic macular edema and proliferative diabetic retinopathy, by targeting PlGF and Ang signaling pathways.

We report that placental growth factor (PlGF) negatively affects the endothelial cell (EC) barrier function through a novel regulatory mechanism. The PlGF mAb promotes (but recombinant protein disrupts) EC barrier function, thus affecting the barrier-forming protein levels, membrane distribution, and EC monolayer impedance by the electrical cell-impedance sensing system, Western blot, and immunofluorescence staining. RNA sequencing–based transcriptome analysis identified the up-regulation of the pentose phosphate pathway (PPP) and the antioxidant defense protein by PlGF blockade. The PlGF and PlGF/VEGF dimers (but not VEGF-A) down-regulated the protein expression of glucose-6-phosphate dehydrogenase (G6PD) and peroxiredoxin (PRDX). G6PD inhibition and gene silencing (small interfering RNA) abolished the beneficial effects of PlGF inhibition on EC barrier function and PRDX3/6 protein expression. VEGF receptor (VEGFR)1 or VEGFR2 blockade prevented the inhibitory effect of PlGF on G6PD protein expression and EC barrier function. The PRDX6 played dual roles in EC barrier function through glutathione peroxidase and phospholipase A(2) activity. In sum, PlGF negatively regulates EC barrier function through the activation of VEGFR1 and VEGFR2 and the suppression of the G6PD/PPP and the antioxidant pathways.—Huang, H., Lennikov, A., Saddala, M. S., Gozal, D., Grab, D. J., Khalyfa, A., Fan, L. Placental growth factor negatively regulates endothelial cell barrier function through suppression of glucose-6-phosphate dehydrogenase and antioxidant defense systems.

Therapeutic inhibition of hypoxia inducible factor-1α (HIF-1α) action has emerged as a potential approach for managing several diseases including breast cancer (BC). Genistein has been found to exert anti-malignant activity. However, its mechanisms of action remain unknown. Studies indicate that it could act by downregulating HIF-1α. Based on these findings, we investigated whether genistein could reduce HIF-1α in BC cell lines. Furthermore, we performed molecular docking studies to characterize the sites of interaction between genistein and HIF-1α. In the present investigation, we prove, for the first time, that genistein downregulates HIF-1α in BC cells. Molecular docking analysis also revealed that genistein binds to the FIH-1 binding site of HIF-1α protein. These findings thus indicate that genistein and/or HIF-1α antagonists could be a potential treatment for BC.

Background Age-related macular degeneration (AMD) is the most common, progressive, and polygenic cause of irreversible visual impairment in the world. The molecular pathogenesis of the primary events of AMD is poorly understood. We have investigated a transcriptome-wide analysis of differential gene expression, single-nucleotide polymorphisms (SNPs), indels, and simple sequence repeats (SSRs) in datasets of the human peripheral retina and RPE-choroid-sclera control and AMD. Methods and results Adaptors and unbiased components were removed and checked to ensure the quality of the data sets. Molecular function, biological process, cellular component, and pathway analyses were performed on differentially expressed genes. Analysis of the gene expression datasets identified 5011 upregulated genes, 11,800 downregulated genes, 42,016 SNPs, 1141 indels, and 6668 SRRs between healthy controls and AMD donor material. Enrichment categories for gene ontology included chemokine activity, cytokine activity, cytokine receptor binding, immune system process, and signal transduction respectively. A functional pathways analysis identified that chemokine receptors bind chemokines, complement cascade genes, and create cytokine signaling in immune system pathway genes (p value

The molecular mechanisms whereby placental growth factor (PlGF) mediates its effects in nonproliferative diabetic retinopathy (DR) are unknown. To better understand the role of PlGF in DR, we used tandem mass tags (TMT)-labeled quantitative proteomics to human retinal endothelial cells (HRECs), treated anti-PlGF antibody, and PBS as a control. Functional annotation and pathway enrichments were performed, which suggested that the differentially expressed proteins (DEPs) were involved in key metabolic processes, protein binding, and membrane, pentose phosphate pathway (PPP) and adherens junction. We conducted integrated gene profiles of our previously published transcriptomic data to the TMT-labeled proteomics data. The results showed the sixty genes were found to be changed at the proteome level. The functional annotation conducted for the sixty proteins suggested that 58.3% of proteins were involved in PPP, 25% of proteins were in interleukin-12 singling and 16.7% of proteins were involved in glycolysis and gluconeogenesis pathway. Mass spectrometry results were validated by transendothelial electrical resistance measurement by an electrical cell-impedance sensing (ECIS) and western blot analysis of VE-cadherin, G6PD. These findings suggest that the PPP proteins and antioxidants may act as a downstream target of PlGF and may play a decisive role in HREC biological functions in DR. SIGNIFICANCE: PlGF (Placental growth factor) is known to play a pivotal role in pathological angiogenesis and inflammation by stimulating endothelial cell migration and by recruiting pericytes and inflammatory cells such as microglia and macrophages. Despite the well-defined pathophysiological roles of PlGF, the underlying molecular and cellular mechanisms are not completely understood, especially the exact relationships between biochemical events and molecular pathways regulated by PlGF, whose inhibition exhibits a protective role in DR. This study provides new insights into protein expression patterns and enables the identification of many attractive candidates for investigation of PPP pathway role in the activation of the antioxidant defense system in DR. Our findings suggest that the PPP proteins and antioxidants (PRDX6, HMOX1, NQO1 and YES1) may act as downstream targets of PlGF and may play a decisive role in HREC biological functions in DR.

Glucose-6-Phosphate Dehydrogenase (G6PD) is a ubiquitous cytoplasmic enzyme converting glucose-6-phosphate into 6-phosphogluconate in the pentose phosphate pathway (PPP). The G6PD deficiency renders the inability to regenerate glutathione due to lack of Nicotine Adenosine Dinucleotide Phosphate (NADPH) and produces stress conditions that can cause oxidative injury to photoreceptors, retinal cells, and blood barrier function. In this study, we constructed pharmacophore-based models based on the complex of G6PD with compound AG1 (G6PD activator) followed by virtual screening. Fifty-three hit molecules were mapped with core pharmacophore features. We performed molecular descriptor calculation, clustering, and principal component analysis (PCA) to pharmacophore hit molecules and further applied statistical machine learning methods. Optimal performance of pharmacophore modeling and machine learning approaches classified the 53 hits as drug-like (18) and nondrug-like (35) compounds. The drug-like compounds further evaluated our established cheminformatics pipeline (molecular docking and in silico ADMET (absorption, distribution, metabolism, excretion and toxicity) analysis). Finally, five lead molecules with different scaffolds were selected by binding energies and in silico ADMET properties. This study proposes that the combination of machine learning methods with traditional structure-based virtual screening can effectively strengthen the ability to find potential G6PD activators used for G6PD deficiency diseases. Moreover, these compounds can be considered as safe agents for further validation studies at the cell level, animal model, and even clinic setting.

Age-related macular degeneration (AMD) is the most common cause of irreversible blindness in the elderly population. In our previous studies, we found that deficiency of CXCR5 causes AMD-like pathological phenotypes in mice, characterized by abnormalities and dysfunction of the retinal pigment epithelium (RPE) cells. The abnormalities included abnormal cellular shape and impaired barrier function. In the present study, primary RPE cells were derived separately from CXCR5 knockout (KO) mice and from C57BL6 wild type (WT). The isolated primary cells were cultured for several days, and then total RNA was isolated and used for library preparation, sequencing, and the resultant raw data analyzed. Relative to the WT, a total of 1392 differentially expressed genes (DEG) were identified. Gene ontology analysis showed various biological processes, cellular components, and molecular functions were enriched. Pathway enrichment analysis revealed several pathways, including the PI3K-Akt signaling, mTOR signaling, FoxO, focal adhesion, endocytosis, ubiquitin-mediated proteolysis, TNF&alpha, NF-kB Signaling, adipogenesis genes, p53 signaling, Ras, autophagy, epithelial&ndash, mesenchymal transition (EMT), and mitochondrial pathway. This study explores molecular signatures associated with deficiency of CXCR5 in RPE cells. Many of these signatures are important for homeostasis of this tissue. The identified pathways and genes require further evaluation to better understand the pathophysiology of AMD.

Background Retinal degenerative diseases affect millions of people and represent the leading cause of vision loss around the world. Retinal degeneration has been attributed to a wide variety of causes, such as disruption of genes involved in phototransduction, biosynthesis, folding of the rhodopsin molecule, and the structural support of the retina. The molecular pathogenesis of the biological events in retinal degeneration is unclear; however, the molecular basis of the retinal pathological defect can be potentially determined by gene-expression profiling of the whole retina. In the present study, we analyzed the differential gene expression profile of the retina from a wild-type zebrafish and phosphodiesterase 6c (pde6c) mutant. Results The datasets were downloaded from the Sequence Read Archive (SRA), and adaptors and unbiased bases were removed, and sequences were checked to ensure the quality. The reads were further aligned to the reference genome of zebrafish, and the gene expression was calculated. The differentially expressed genes (DEGs) were filtered based on the log fold change (logFC) (±4) and p-values (p Conclusions Our data strongly indicate that, among these genes, the above-mentioned pathways’ genes as well as calcium-binding, neural damage, peptidase, immunological, and apoptosis proteins are mostly involved in the retinal and neural degeneration that cause abnormalities in photoreceptors or retinal pigment epithelium (RPE) cells.

Synthesis of a series of new ethyl phosphoramidates 9a–j of acyclovir through phosphorylation of hydroxy group followed by substitution of numerous amines/amino acid esters 7a–j was accomplished. The structures of newly synthesized compounds were elucidated by spectroscopic data such as IR, NMR (1H, 13C, and 31P) and mass spectrophotometery, and elemental analyses. The synthesized products were screened for their antiviral activity against Newcastle disease virus, antioxidant potency by α, α-diphenyl-β-picrylhydrazyl free radical scavenging and nitric oxide radical scavenging methods, and antioxidant capacity by Ferric reducing antioxidant power method. The compound bearing 1-hydroxy 2-butylamine, 9c and amino acid ester derivatives, bonded with leucine methyl ester 9h and tyrosine methyl ester 9j were found to be potent inhibiters of Newcastle disease virus than that of acyclovir drug. The compounds, 9c (IC50 value, 22.5 ± 1.00 μg/mL), 9i (IC50 value, 21.5 ± 1.00 μg/mL) and 9j (IC50 value, 19.0 ± 1.50 μg/mL) were exhibited promising antioxidant activity while other compounds displayed moderate to good antioxidant activity. Docking study using molecular docking environment into haemaglutinin–neuraminidase protein has been carried out to find the potential selectivity and binding orientation of these new compounds into protein binding site. Interestingly, all the title compounds except 9a and 9g docked well into HN protein with considerable binding affinities better than that of acyclovir drug.

Placental growth factor (PlGF or PGF) is a member of the VEGF family, which is known to play a critical role in pathological angiogenesis, inflammation, and endothelial cell barrier function. However, the molecular mechanisms by which PlGF mediates its effects in non-proliferative diabetic retinopathy (DR) remain elusive. In this study, we performed transcriptome-wide profiling of differential gene expression for human retinal endothelial cells (HRECs) treated with PlGF antibody. The effect of antibody treatment on the samples was validated using trans-endothelial electric resistance (TEER), and western blot. A total of 3760 genes (1750 upregulated and 2010 downregulated) were found to be differentially expressed between the control and PlGF antibody treatment group. These differentially expressed genes (DEGs) were used for gene ontology and enrichment analysis to identify gene function, signal pathway, and interaction networks. The gene ontology results revealed that catalytic activity (GO:0003824) of molecular function, cell (GO:0005623) of the cellular component, and cellular process (GO:0009987) were among the most enriched biological processes. Pathways such as TGF-β, VEGF-VEGFR2, p53, apoptosis, pentose phosphate pathway, and ubiquitin-proteasome pathway, were among the most enriched, and TGF-β1 was identified as a primary upstream regulator. These data provide new insights into the underlying molecular mechanisms of PlGF in mediating biological functions, in relation to DR.

Retinal degenerative diseases affect millions of people and are the leading cause of vision loss. Retinal degeneration has been attributed to a wide variety of causes, such as disruption of genes that are involved in phototransduction, biosynthesis, folding of the rhodopsin molecule and the structural support of the retina. The molecular pathogenesis of the biological events in retinal degeneration is unclear. The molecular basis of the retinal pathologies defect can be potentially determined by gene-expression profiling of the whole retina. In the present study, we have analyzed the differential gene expression profile of retina from a wild-type zebrafish and pde6c mutant. The datasets were downloaded from the Sequence Read Archive (SRA), removed adaptors and unbiased bases and checked to ensure the quality. The reads were further aligned to the reference genome of zebrafish and calculated gene expression. The differentially expressed genes (DEGs) were filtered based on the FDR (±4) and p-values (p

A new series of α-thiazolyl aminomethylene bisphosphonates were synthesized by a three component reaction of 4-aryl substituted thiazol-2-amine with different dialkyl/aryl phosphites and triethyl orthoformate in the presence of Ag NPs (nano particles) as a catalyst under solvent free conditions. All the synthesized target compounds were characterized by 1H, 13C, 31P, mass and elemental analysis. The target compounds were screened for their in vitro antioxidant, antibacterial and antifungal activity. Molecular docking studies were also performed. The results revealed that among the synthesized compounds tetramethyl(((4-(4-methoxyphenyl)thiazol-2-yl)amino) methylene)bis(phosphonate) (5d), tetramethyl(((4-(4-fluorophenyl)thiazol-2-yl)amino) methylene) bis(phosphonate) (5h), and tetramethyl(((4-(4-bromophenyl)thiazol-2-yl)amino)methylene) bis (phosphonate) (5j) showed remarkably higher antioxidant activity by DPPH and H2O2 than the standard ascorbic acid. Compounds tetramethyl(((4-phenyl thiazol-2-yl)amino) methylene) bis(phosphonate) (5a), 5d, 5h and tetraethyl(((4-(4-bromophenyl)thiazol-2-yl) amino)methylene)bis (phosphonate) (5k) showed good antibacterial activity. 5a, 5d, and 5h also showed rather higher antifungal activity than the standard flucanozole. Computational docking methods have been used to predict how several aminomethylene bisphosphonate derivatives compete against the inhibitor BPH-1330 at the crystal enzyme structure of the 4H3A protein active site and how R and R1 influence their binding ability.

A series of new class of P-heterocycle encompassing urea and thiourea derivatives, N-(substitutedphenyl)-N′-(2-oxo-2,3-dihydro-1H-2λ5-[1,3,2]diazaphospholo[4,5-b]pyridin-2-yl)ureas 11a–e/thioureas 11f–k, was accomplished from the precursor intermediate, 2-amino-2,3-dihydro-1H-2λ5-[1,3,2]diazaphospholo[4,5-b]pyridin-2-one, 9. The compound 9 was obtained by cyclization of pyridine-2,3-diamine, 6 with POCl3 followed by amidation with NaNH2. The products were tested for their in vitro and in vivo anti-inflammatory activity, and in vitro antimicrobial activity including minimum inhibitory concentration. Compounds 11a, 11d and 11j exhibited comparable anti-inflammatory activity to the standard drug, diclofenac, both in in vitro and in vivo assays, which might be due to the presence of lipophilic functional groups, F, NO2 and CF3. The compounds 11c and 11j exhibited potential growth of inhibition against selected bacterial and fungal strains at lower minimum inhibitory concentrations, while most of the thiourea-linked analogues exhibited good antimicrobial activity. A molecular modelling study was performed on cyclooxygenase isoenzyme (COX-2) to investigate the hypothetical binding mode of the most active anti-inflammatory agents, and binding conformers were proposed.

Placental growth factor (PlGF or PGF), a member of the vascular endothelial growth factor (VEGF) sub-family, plays a crucial role in pathological angiogenesis and inflammation. However, the underlying molecular mechanisms that PlGF mediates regarding the complications of non-proliferative diabetic retinopathy (DR) remain elusive. Using an LC-MS/MS-based label-free quantification proteomic approach we characterized the alterations in protein expression caused by PlGF ablation in the retinas obtained from C57BL6, Akita, PlGF−/− and Akita.PlGF−/− mice. After extraction and enzymatic digestion with Trypsin/LysC, the retinal proteins were analyzed by Q-Exactive hybrid Quadrupole-Orbitrap mass spectrometry. Differentially expressed proteins (DEPs) were identified in four comparisons based on Z-score normalization and reproducibility by Pearson’s correlation coefficient. The gene ontology (GO), functional pathways, and protein-protein network interaction analysis suggested that several proteins involved in insulin resistance pathways (Gnb1, Gnb2, Gnb4, Gnai2, Gnao1, Snap2, and Gngt1) were significantly down-regulated in PlGF ablated Akita diabetic mice (Akita.PlGF−/− vs. Akita) but up-regulated in Akita vs. C57 and PlGF−/− vs. C57 conditions. Two proteins involved in the antioxidant activity and neural protection pathways, Prdx6 and Map2 respectively, were up-regulated in the Akita.PlGF−/− vs. Akita condition. Overall, we predict that down-regulation of proteins essential for insulin resistance, together with the up-regulation of antioxidant and neuroprotection proteins highlight and epitomize the potential mechanisms important for future anti-PlGF therapies in the treatment of DR.

Pancreatic cancer (PC) is an aggressive carcinoma and the fourth cause of cancer deaths in Western countries. Although surgery is the most effective therapeutic option for PC, the management of unresectable, locally advanced disease is highly challenging. Our improved understanding of pancreatic tumor biology and associated pathways has led to the development of various treatment modalities that can control the metastatic spread of PC. This review intends to present trials of small molecule tyrosine kinase inhibitors (TKIs) in PC management and the troubles encountered due to inevitable acquired resistance to TKIs.

Vivapain-3(VP-3) protein is a family of cysteine rich proteases of malaria parasite is extensively reported to participate in a range of wide cellular processes including survival. VP-3 of plasmodium recognized as an attractive drug target in vector-borne diseases like malaria. In the present study we robust a homology model of VP-3 protein and generated the pharmacophore based models adapted to screen the best drug like compounds from PubChem database. Our results finds the fourteen best lead molecules were mapped with core pharmacophore features of VP-3 and top hits were further evaluated by molecular dynamics simulation and docking studies. Based on the molecular dynamics simulation and docking results and binding vicinity of ligand molecules, top five i.e., CID 74427945, CID 74427946, CID 360883, CID193721 and CID 51416859 showed the best docking scores with good molecular interactions against VP-3. Furthermore in silico ADMET and in vitro assays clearly exhibited that out of five three CID74427946, CID74427945 and CID360883 ligand molecules showed the best promising inhibition against VP-3. The present study believed to provide significant information of potential ligand inhibitors against VP-3 to design and develop the next generation malaria therapeutics through computational approach.

Hypoxia inducible factor-1(HIF-1) is a bHLH-family transcription factor that control genes involved in glucolysis, angiogenesis, migration, as well as invasion factors that are important for tumor progression and metastasis. HIF-1, a hetero dimer of HIF-1α and HIF-1β, binds to the hypoxia responsive genes, such as vascular endothelial growth factor (VEGF). It is one the molecular target for angiogenesis. A series of Chalcone - like compounds described that preferentially inhibit HIF-1 dimer, which can interact with amino acids within the active site of the protein. It is of interest model the HIF-1 dimer protein and protein was subjected to molecular dynamics simulations using NAMD 2.9 software with CHARMM27 force field in water and the protein structure was minimized with 25000 steps for 500 ps and simulation with 1000000 steps for 2ns. 2500 compounds were screened from Zinc database through structure based virtual screening with references to Chalcone natural drug compound. The screened compounds were docked into the active site of the protein using AutoDock Vina in PyRx Virtual screening tool. The docking result showed the compounds Zinc04280532, Zinc04280533, Zinc04280469, Zinc04280534, Zinc16405915, Zinc04277060, Zinc04280538, Zinc04582923, Zinc05280554 and Zinc05943723 have high binding affinities then query compound. The lead hit compounds were also testing for toxicity and bioavailability using Osiris and Molinspiration online server. The active site amino acids such as TYR-21, ASN-34, VAL-35, MET-18, LYS-17, SER-36, ARG- 46 and ARG-14 are key role in the inhibitors activity. This is useful in the design of small molecule therapeutics or the treatment of different abnormalities associated with impaired HIF-1α.

The present study, we design and synthesize the novel dihydropyridine derivatives, i.e., 3 (a-e) and 5 (a-e) and evaluated, anticonvulsant activity. Initially due to the lacuna of LCC, we modeled the protein through modeller 9.15v and evaluated through servers. Docking studies were performed with the synthesized compounds and resulted two best compounds, i.e., 5a, 5e showed the best binding energies. The activity of intracellular Ca 2+ measurements was performed on two cell lines: A7r5 (rat aortic smooth muscle cells) and SH-SY5Y (human neuroblastoma cells). The 5a and 5e compounds was showing the more specific activity on L-type calcium channels, i.e. A7r5 (IC 50 = 0.18 ± 0.02 and 0.25 ± 0.63 μg/ml, respectively) (containing only L-type channels) than SH-SY5Y (i.e. both L-type and T-type channels) (IC 50 = 8 ± 0.23 and 10 ± 0.18 μg/ml, respectively) with intracellular calcium mobility similar to amlodipine. Finally, both in silico and in vitro results exploring two derivatives 5a and 5e succeeded to treat cadmium toxicity. © The Author(s) 2017. Author, Madhu Sudhana Saddala is especially grateful to University Grants Commission, New Delhi for their financial assistance with the award of BSR-Meritorious fellowship, One of the authors SBS is highly grateful to FCT, Portugal (SFRH / BPD / 81501/2011).

Cadmium (Cd) is a heavy metal present in air, water, soils and sediments. It is well known that long-term exposure to Cd causes various toxic effects in various organ systems such as cardiovascular, kidneys, liver, brain, lung, bones, immune/haemopoietic, endocrine and reproductive systems. Cd influx mediates voltage-gated L-type calcium channels (LCC) in excitable cells including mammalian neurons and also Cd uptake in non-excitable tissues. Therefore, LCC has been recognized as an attractive metal toxicity target. We construct a homology model of LCC in addition to the generated pharmacophore models then used to retrieve 50,500 molecules from Zinc database. There are 18 best reliable molecules mapped with core pharmacophore model of LCC. These hits were retrieved and further evaluated by molecular dynamics (MD) simulation, molecular docking and protein–ligand interactions, and binding affinity predictions as well as in silico ADMET properties were tested. Our work results focus on homology modelling, structure-based pharmacophore mapping, molecular docking, MD simulation, protein–ligand interactions and binding affinity predictions which were used in virtual screening strategy to spot new hits for blockade of LCC. Finally, the outcome results, priming the five best lead compounds, were expected to be the potential lead scaffolds for developing novel and potent blockers of LCC against metal toxicity.

Background: Signal transducer and activator of transcription (STAT) proteins are a seven-member family of cytoplasmic proteins. A conserved SH2 domain facilitates STAT initiation by allowing binding to phosphotyrosine motifs for STAT-receptor interactions and STAT dimerization. STAT proteins (STAT1, STAT2 and STAT3) are promising targets for anti-cancer drugs, because irregular facilitation of STAT initiation may result in cancer, inflammation and auto-immunity. Methods: The crystal structure of the phosphotyrosine STAT1 and STAT3 dimers bound to DNA was obtained from a protein data bank (PDB), while STAT2 was modelled through Swiss-Model workspace by using 1YVL as a template. The stereochemical quality and integrity of STAT2 was tested by RAMPAGE server, ERRAT, ProSA, and Verify3D programs. The active sites were predicted using CASTp 3.0 with a new algorithm, which identifies and measures surface accessible pockets and interior inaccessible cavities on STAT1, STAT2, and STAT3. MD (molecular dynamics) simulation was conducted with STAT2 protein. Molecular Docking was used to predict the binding modes and approximate binding free energies of genistein to STAT1, STAT2 and STAT3 SH2 dimerization sites. Genistein was docked into the active site with AutoDock Vina in PyRx Virtual Screening tool. Results: Genistein showed best binding energies. The most successful binding energies were -5.6 (STAT1), -6.7 (STAT2), -6.3 (STAT3) kcal/mol respectively. The binding interactions of this compound with the active site of STAT proteins suggested that amino acid residues (Tyr631, Leu639, Tyr640, Glu643, His647, Met648, Gln649, Ile659, Met660 and LYS16) play a crucial role in anti-cancer activity. Conclusion: These observations increase the understanding of the functional role of STAT proteins in cancer and allow better development for additional druggable STAT inhibitors with high potency, specificity and outstanding bioavailability. Citation Format: Sneha Govardhanagiri, Shipra Reddy Bethi, Madhu Sudhana Saddala, Bassel F. El-Rayes, Ganji Purnachandra Nagaraju. Interaction of STAT proteins with genistein: A computational analysis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3398.

A series of new phosphoramidate derivatives 7a-l were synthesized by reacting 5-nitroindazole with 4-chlorophenyl dichlorophosphate (4), and then further with various bio-potent aromatic/hetero/alk...