MicroRNA-195-5p mediates arsenic-induced cytotoxicity in human lung epithelial cells: Beneficial role of plant-derived tannic acid.

Arsenic (As), a highly toxic metalloid, which causes environmental lung diseases and affects millions of people worldwide. Respiratory epithelial cells are essential for maintaining lung homeostasis, aberrant epithelial damage and death due to exposure to a wide range of environmental pollutants, which are considered to be the initial trigger for many pulmonary diseases. Accumulating evidence has shown that microRNAs (miRNAs) appear to be important players in various normal physiological and pathological processes. Therefore, the present study was carried out to examine the cytotoxic effects of a trivalent form of As (As3+) in normal human bronchial (BEAS-2B) and adenocarcinoma alveolar basal (A549) epithelial cells and the role of miR-195-5p. Further, we also explored the protective effects of a natural dietary polyphenol tannic acid (TA). As3+ (1 µM) treatment in BEAS-2B cells for 24 h induced cytotoxicity by decreasing the cell viability, mitochondrial membrane potential (ΔΨm) and inducing reactive oxygen species (ROS) generation, lipid peroxidation (LPO), cell cycle arrest, and apoptosis, which was associated with a significantly higher level of miR-195-5p expression compared with vehicle control. Forced expression of miR-195-5p alone suppressed cell survival, ΔΨm, regulated cell cycle distribution and induced ROS generation in BEAS-2B cells. As expected, miR-195-5p inhibition effectively rescued BEAS-2B cells from As3+-mediated toxicity, confirming the involvement of miR-195-5p in the cytotoxic effects of As3+. Further, TA pre-treatment expressively alleviated As3+-induced toxicity by suppressing ROS production, miR-195-5p expression, and increasing ΔΨm. These in vitro results indicate that miR-195-5p may be useful as a therapeutic target for treating As3+ toxicity.

DDX3 modulates cisplatin resistance in OSCC through ALKBH5-mediated m6A-demethylation of FOXM1 and NANOG.

Platinum based drugs alone or in combination with 5FU and docetaxel are common regimen chemotherapeutics for the treatment of advanced OSCC. Chemoresistance is one of the major factors of treatment failure in OSCC. Human RNA helicase DDX3 plays an important role in cell proliferation, invasion, and metastasis in several neoplasms. The potential role of DDX3 in chemoresistance is yet to be explored. Enhanced cancer stem cells (CSCs) population significantly contributes to chemoresistance and recurrence. A recent study showed that m6A RNA regulates self-renewal and tumorigenesis property in cancer. In this study we found genetic (shRNA) or pharmacological (ketorolac salt) inhibition of DDX3 reduced CSC population by suppressing the expression of FOXM1 and NANOG. We also found that m6A demethylase ALKBH5 is directly regulated by DDX3 which leads to decreased m6A methylation in FOXM1 and NANOG nascent transcript that contribute to chemoresistance. Here, we found DDX3 expression was upregulated in both cisplatin-resistant OSCC lines and chemoresistant tumors when compared with their respective sensitive counterparts. In a patient-derived cell xenograft model of chemoresistant OSCC, ketorolac salt restores cisplatin-mediated cell death and facilitates a significant reduction of tumor burdens. Our work uncovers a critical function of DDX3 and provides a new role in m6 demethylation of RNA. A combination regimen of ketorolac salt with cisplatin deserves further clinical investigation in advanced OSCC.

Impact of naturopathy, yoga, and dietary interventions as adjuvant chemotherapy in the management of stage II and III adenocarcinoma of the colon.

INTRODUCTION: Naturopathy, Yoga and Dietary interventions are known to improve the quality of life in cancer patients. We aim to evaluate the effect of naturopathy interventions along with adjuvant chemotherapy in patients who underwent surgery for Adenocarcinoma of the Colon. METHODS: A total of 116 adult patients were randomised in to one of the two groups; the experimental group received naturopathy, Yoga and Dietary interventions and the control group received psycho-social counselling in addition to standard chemotherapy. Haematological, biochemical and psychological evaluations were performed at set intervals during a total period of eighteen months starting from the first cycle of adjuvant chemotherapy. RESULTS: Results showed that the overall hemoglobin (p < 0.0001) and carcinoembryonic antigen (CEA) (p = 0.0038) levels were statistically significant in patients on the experimental arm. The rest of the laboratory parameters, viz. total leukocyte count, platelet counts, and serum creatinine levels, for overall data was not statistically significant in both the groups. Psychological attributes such as anxiety, depression, symptom severity, and Functional Living Index: Cancer (FLIC) were found to be statistically significant (p < 0.0001) in the experimental subjects as compared with those in the control. On the whole, men benefited more than women from the study interventions. CONCLUSIONS: We conclude that Yoga and Naturopathy interventions in addition to chemotherapy show improvement in overall functional life index along with improvement in haemoglobin in patients with stages II and III Adenocarcinoma of Colon.

Neuronal pentraxin 1 expression is regulated by hypoxia inducible factor-1α.

Neuronal pentraxins (NPs) are belong to sub family of long pentraxin proteins consist of neuronal pentraxin 1 (NP1), neuronal pentraxin 2 (NP2), and neuronal pentraxin receptor (NPR). Enhanced expression of NP1 in hypoxic conditions has shown to induce cell death in neuronal cells, however, the underlying mechanism of NP1 regulation by hypoxia remains elusive. To demonstrate that, we have cloned human NP1 gene promoter upstream of the luciferase gene and the activity of NP1 promoter was studied using HEK cell lines. Within the promoter region of the human NP1 gene, we identified six putative hypoxia inducible factor (HIF) responsive elements. By luciferase reporter assays we determined that the hypoxia inducible factor responsive element is located between -332 to -215 positions relative to the translation start site are essential for transcriptional activation of NP1 under hypoxic conditions. To further confirm the activity is solely due to hypoxia, we transiently transfected green fluorescent protein (EGFP) under transcriptional control of five copies of a hypoxia response element (HRE). The intensity of GFP was recorded at normal and hypoxic conditions. Taken together, our results demonstrate that NP1 gene is a target of as a hypoxia-inducible factor and it regulate NP1 expression by binding to hypoxia responsive elements (HREs) in its promoter region.

Characterization of vitamin-cisplatin-loaded chitosan nano-particles for chemoprevention and cancer fatigue.

CONTEXT: Vitamins have been shown to reduce chemotherapy-related fatigue (CRF) by conserving energy loss both during and after cancer treatment. However, it remains unknown whether this reduction of fatigue interferes with the cancer drugs or alters the effectiveness of these agents. OBJECTIVES: The objective was to synthesize vitamin-cisplatin-loaded chitosan nano-particles for chemoprevention and cancer fatigue. MATERIALS AND METHODS: Multi-vitamin (C, D3, and B12)-cisplatin composite nano-formulation called NanoCisVital (NCV) to overcome CRF. The interactions between vitamins and NCV were characterized using scanning electron microscopy (SEM), Fourier transform infrared (FT-IR) spectroscopy, and a particle size analyser. The chemo-preventive activity was performed by in vitro bio assays. RESULTS: SEM analysis showed spherical shape and the size is < 225 nm. NCV inhibited the proliferating yeast cells as well as denaturation of bovine serum albumin, and it also reduced the sprouting of new blood vessels in dose-dependent manner. CONCLUSION: Collectively, these results demonstrate that the NCV particles can be used to reduce CRF without much affecting the anti-cancer properties of cisplatin.

Classification and diagnostic prediction of breast cancer metastasis on clinical data using machine learning algorithms.

Metastatic Breast Cancer (MBC) is one of the primary causes of cancer-related deaths in women. Despite several limitations, histopathological information about the malignancy is used for the classification of cancer. The objective of our study is to develop a non-invasive breast cancer classification system for the diagnosis of cancer metastases. The anaconda-Jupyter notebook is used to develop various python programming modules for text mining, data processing, and Machine Learning (ML) methods. Utilizing classification model cross-validation criteria, including accuracy, AUC, and ROC, the prediction performance of the ML models is assessed. Welch Unpaired t-test was used to ascertain the statistical significance of the datasets. Text mining framework from the Electronic Medical Records (EMR) made it easier to separate the blood profile data and identify MBC patients. Monocytes revealed a noticeable mean difference between MBC patients as compared to healthy individuals. The accuracy of ML models was dramatically improved by removing outliers from the blood profile data. A Decision Tree (DT) classifier displayed an accuracy of 83% with an AUC of 0.87. Next, we deployed DT classifiers using Flask to create a web application for robust diagnosis of MBC patients. Taken together, we conclude that ML models based on blood profile data may assist physicians in selecting intensive-care MBC patients to enhance the overall survival outcome.

Identification of prognostic hub genes and therapeutic targets for selenium deficiency in chicks model through transcriptome profiling.

Selenium deficiency is a prevalent micronutrient deficiency that poses a major health concern worldwide. This study aimed to shed light on the molecular mechanisms underlying selenium deficiency using a chick model. Chickens were divided into control and selenium deficient groups. Plasma samples were collected to measure selenium concentration and transcriptome analyse were performed on oviduct samples. The results showed that selenium deficiency led to a significant reduction in plasma selenium levels and altered the expression of 10,266 differentially expressed genes (DEGs). These DEGs primarily regulated signal transduction and cell motility. The molecular function includes GTPase regulatory activity, and KEGG pathway analysis showed that they were mainly involved in the signal transduction. By using Cytoscape and CancerGeneNet tool, we identified 8 modules and 10 hub genes (FRK, JUN, PTPRC, ACTA2, MST1R, SDC4, SDC1, CXCL12, MX1 and EZR) associated with receptor tyrosine kinase pathway, Wnt and mTOR signaling pathways that may be closely related to cancer. These hub genes could be served as precise diagnostic and prognostic candidate biomarkers of selenium deficiency and potential targets for treatment strategies in both animals and humans. This study sheds light on the molecular basis of selenium deficiency and its potential impact on public health.

Molecular docking, synthesis, and biological evaluation of 7-azaindole-derivative (7AID) as novel anti-cancer agent and potent DDX3 inhibitor:-an in silico and in vitro approach.

The DEAD-box helicase family member DDX3 is involved in many diseases, such as viral infection, inflammation, and cancer. Many studies in the last decade have revealed the role of DDX3 in tumorigenesis and metastasis. DDX3 has both tumour suppressor and oncogenic effect, in the present study we have evaluated the expression levels of DDX3 in cervical squamous cell carcinoma at mRNA level via real-time PCR and protein level via Immunohistochemistry. DDX3 has become a molecule of interest in cancer biology that promotes drug resistance by adaptive response inevitably leading to treatment failure. One approach to avoid the development of resistant to disease is to create novel drugs that target the overexpressed proteins, we designed and synthesized a novel 7-azaindole derivative (7-AID) compound, {5-[1H-pyrrolo (2, 3-b) pyridin-5-yl] pyridin-2-ol]} that could lodge within the adenosine-binding pocket of the DDX3 (PDB ID: 2I4I). The binding efficacy of 7-AID compound with DDX3 was analysed by molecular docking studies. 7-AID was found to interact with the key residues Tyr200 and Arg202 from the Q-motif rendered by π-interactions and hydrogen bonds within the binding pocket with good docking score - 7.99 kcal/mol. The cytotoxicity effect of 7-AID compound was evaluated using MTT assay on human cervical carcinoma cells (HeLa) and breast cancer cells (MCF-7 and MDA MB-231) and the compound shown effective inhibitory concentration (IC50) on Hela cells 16.96 µM/ml and 14.12 and 12.69 µM/ml on MCF-7 and MDA MB-231, respectively. Further, the in-vitro, in-vivo anti-cancer and anti-angiogenic assessment of 7-AID compound was evaluated on Hela cells using scratch wound-healing assay, DAPI staining, cell cycle analysis, immunoblotting, and chorioallontoic membrane assay. Furthermore, the inhibitory effect of derivative compound on DDX3 was investigated in HeLa, MCF-7, and MDA MB-231 cells at the mRNA and protein levels. The results showed that the 7-AID compound effectively inhibited DDX3 in a dose-dependent manner, and the findings suggest that the compound could be used as a potential DDX3 inhibitor.

Identification of Differentially Expressed Genes in Cervical Cancer Patients by Comparative Transcriptome Analysis.

Cervical cancer is one of the most malignant reproductive diseases seen in women worldwide. The identification of dysregulated genes in clinical samples of cervical cancer may pave the way for development of better prognostic markers and therapeutic targets. To identify the dysregulated genes (DEGs), we have retrospectively collected 10 biopsies, seven from cervical cancer patients and three from normal subjects who underwent a hysterectomy. Total RNA isolated from biopsies was subjected to microarray analysis using the human Clariom D Affymetrix platform. Based on the results of principal component analysis (PCA), only eight samples are qualified for further studies; GO and KEGG were used to identify the key genes and were compared with TCGA and GEO datasets. Identified genes were further validated by quantitative real-time PCR and receiver operating characteristic (ROC) curves, and the highest Youden index was calculated in order to evaluate cutoff points (COPs) that allowed distinguishing of tissue samples of cervical squamous carcinoma patients from those of healthy individuals. By comparative microarray analysis, a total of 108 genes common across the six patients' samples were chosen; among these, 78 genes were upregulated and 26 genes were downregulated. The key genes identified were SPP1, LYN, ARRB2, COL6A3, FOXM1, CCL21, TTK, and MELK. Based on their relative expression, the genes were ordered as follows: TTK > ARRB2 > SPP1 > FOXM1 > LYN > MELK > CCL21 > COL6A3; this generated data is in sync with the TCGA datasets, except for ARRB2. Protein-protein interaction network analysis revealed that TTK and MELK are closely associated with SMC4, AURKA, PLK4, and KIF18A. The candidate genes SPP1, FOXM1, LYN, COL6A3, CCL21, TTK and MELK at mRNA level, emerge as promising candidate markers for cervical cancer prognosis and also emerge as potential therapeutic drug targets.

Neoplastic transformation of breast epithelial cells by genotoxic stress.

BACKGROUND: Exposure to genotoxic stresses such as radiation and tobacco smoke can cause increased cancer incidence rate as reflected in an in depth meta-analysis of data for women and breast cancer incidence. Published reports have indicated that exposures to low dose radiation and tobacco smoke are factors that contribute to the development of breast cancer. However, there is a scarcity of information on the combinatorial effects of low dose radiation and tobacco smoke on formation and progression of breast cancer. The combination of these two genotoxic insults can induce significant damage to the genetic material of the cells resulting in neoplastic transformation. METHODS: To study the effects of low dose ionizing radiation and tobacco smoke on breast cells, MCF 10A cells were treated either with radiation (Rad - 0.1 Gray) or cigarette smoke condensate (Csc - 10 microgram/ml of medium) or a combination of Rad + Csc. Following treatments, cells were analyzed for cell cycle distribution patterns and the ability to extrude the Hoechst 33342 dye. In addition, in vitro invasion and migration as well as mammosphere formation assays were performed. Finally, differential gene expression profiles were generated from the individual and combination treatment. RESULTS: Exposure of MCF 10A cells to the combination of radiation plus cigarette smoke condensate generated a neoplastic phenotype. The transformed phenotype promoted increased mammosphere numbers, altered cell cycle phases with a doubling of the population in S phase, and increased invasion and motility. Also, exclusion of Hoechst 33342 dye, a surrogate marker for increased ABC transporters, was observed, which indicates a possible increase in drug resistance. In addition, changes in gene expression include the up regulation of genes encoding proteins involved in metabolic pathways and inflammation. CONCLUSIONS: The results indicate that when normal breast cells are exposed to low dose radiation in combination with cigarette smoke condensate a phenotype is generated that exhibits traits indicative of neoplastic transformation. More importantly, this is the first study to provide a new insight into a possible etiology for breast cancer formation in individuals exposed to low dose radiation and tobacco smoke.

In vitro and in silico characterization of angiogenic inhibitors from Sophora interrupta.

Sophora interrupta Bedd, (Fabaceae) is used in Indian folk medicine to treat cancer. Angiogenesis is one of the crucial characteristics of cancer metastasis and is regulated by vascular endothelial growth factor (VEGF). In this study, we examined the antiangiogenic properties of the root ethyl acetate extract of Sophora interrupta by various methods. In vitro antioxidant activity (100-600 µg/ml) of S. interrupta ethyl acetate (SEA) extract was evaluated by DPPH and ABTS, anti-inflammatory activity (50, 100 and 150 µg/ml) by estimating nitric oxide (NO) levels, anti-angiogenic activity (200 and 500 µg/ml) was validated by chorio allantoic membrane (CAM) assay and in silico molecular dynamic (MD) simulations analyses (25 ns) were performed to identify the anti-angiogenic compounds extracted from root extract. The antioxidative activity of SEA extract at IC50 (200 ± 0.6 µg/mL) is equal to that of ascorbic acid at IC50 (50 ± 0.6 µg/mL), and the anti-inflammatory activity of SEA extract at IC50 (150 ± 0.2 µg/mL) was inhibited significantly by nitric oxide (NO) production. The SEA extract significantly reduced the sprouting of new blood vessels at ID50 500 ± 0.13 µg/mL in the CAM assay. Gas chromatography-mass spectrometry analysis of the SEA extract detected 34 secondary metabolites, of which 6a,12a-dihydro-6H-(1,3)dioxolo(5,6)benzofuro(3,2-c)chromen-3-ol (maackiain) and funiculosin formed strong hydrogen bond interactions with Lys 920, Thr 916 and Cys 919 (2H), as well as Glu 917 of VEGFR2, and these interactions were similar to those of the anti-angiogenic compound axitinib. Significant findings in all the assays performed indicate that SEA extract has potential anti-angiogenic compounds that may interfere with VEGF-induced cancer malignancy.

Free radical scavenging, α-glucosidase inhibitory and anti-inflammatory constituents from Indian sedges, Cyperus scariosus R.Br and Cyperus rotundus L.

BACKGROUND: Cyperus scariosus R. Br and Cyperus rotundus L are widely used in ayurvedic preparation for the treatment of diabetes and other diseases. The early literature, so far, does not indicate the presence of any bioactive principle isolated from these plants. OBJECTIVE: To identify free radical scavenging, anti-diabetic and anti- inflammatory principles from these two species. MATERIALS AND METHODS: The bioassay guided fractionation and isolation of active constituents was done by chromatographic techniques. They also evaluated their anti-oxidant activity by DPPH and ABTS. The anti-diabetic activity was screened by α- glucosidase and α- amylase assays. Also, the further evaluation of in vitro anti-inflammatory activity using THP-1 monocytic cells and in vivo anti- inflammatory activity, was confirmed by carrageenan induced rat paw edema as model. RESULTS: The activity guided isolation led to isolation of twelve compounds Which are: Stigmasterol[1], ß- sitosterol[2], Lupeol[3], Gallic acid[4], Quercetin[5], ß- amyrin[6], Oleanolic acid[7], ß- amyrin acetate[8], 4- hydroxyl butyl cinnamate[9], 4- hydroxyl cinnamic acid[10], Caffeic acid,[11] and Kaempferol[12] respectively. Among the isolates, the compounds 4 and 5 displayed potent radical scavenging activity with an IC50 values of 0.43 and 0.067 ΅g/ml. The compounds 4, 5 and 10 showed significant anti-diabetic activities. while lupeol[3] showed potent IL-1 ß activity inhibition in THP-1 monocytic cells and also displayed significant (p<0.0025) in vivo anti-inflammatory activity. CONCLUSION: Inbrief, we isolated twelve compounds from both the species. Collectively, our results suggested that aromatic compounds showed good anti-oxidant and anti-diabetic activities. SUMMARY: The study investigates the free radical scavenging, α-glucosidase inhibitory and anti-inflammatory effects of constituents isolated from Indian sedges viz. C. scariosus and C. rotundus. The results indicated that phenolic compounds displayed potent fee radical scavenging activty and alpha-glucosidase inhibition activity. While terpene constituent, Lupeol[3] showed good IL-1ß activity inhibition in THP-1 monocytic cells and also displayed significant (p<0.0025) in vivo anti inflammatory activity in carrageenan induced rat paw edema. However, further studies are required to know the exact molecular mechanism. Abbreviations used: DPPH: 2,2- Diphenyl-1-1-picryl hydrazyl, ABTS: 2,2-Azinobis-3-ethylbenzo thiazoline-6-sulfonic acid, THP-1: Human leukaemia monocytic cell line, IL-1ß: Interleukin-1ß, IC50-Inhibitory concentration 50%.

In vitro anti-cancer activity of doxorubicin against human RNA helicase, DDX3.

RNA helicase, DDX3 is a multifunctional enzyme and is known to be associated with several diseases like HIV progression, brain and breast cancer. Some of the ring expanded nucleoside compounds such as REN: NZ51, fused di imidazodiazepine ring (RK33), (Z)-3-(5- (3-bromo benzylidene)-4-oxo-2-thioxothiazolidin-3-yl)-N-(2- hydroxy phenyl) propanamide compound (FE15) have been documented to inhibit DDX3 helicase activity. However, synthesis of these drugs is limited to few research groups. Prevalence of literature study, we found that doxorubicin form strong hydrogen bond interactions with crystallized form of DDX3 using in-silico molecular docking approach. To evaluate the biological inhibitory action of doxorubicin, we performed the ATPase activity assay and anti-cancer activity using H357 cancer cell lines. Results showed that doxorubicin continually declined the inorganic phosphate (Pi) release and inhibited the ATP hydrolysis by directly interacting with DDX3. Anticancer activity was detected by MTT assay. The half maximal inhibitory concentrations of doxorubicin (IC50) for H357 cancer cell line is 50 µM and also doxorubicin significantly down regulated the expression of DDX3. Taken together, our results demonstrate, that inhibition of DDX3 expression by using doxorubicin can be used as an ideal drug candidate to treat DDX3 associated cancer disorder by interacting with unique amino acid residues (Thr 198) and common amino acid residues (Tyr 200 and Thr 201).

Morphological and Chemoprofile (Liquid Chromatography-mass Spectroscopy and Gas Chromatography-mass Spectroscopy) Comparisons of Cyperus scariosus R. Br and Cyperus rotundus L.

BACKGROUND: Cyperus scariosus (CS) R.Br and Cyperus rotundus (CR) L. belongs to Cyperaceae family which is well-reputed in the traditional systems of medicine. Although they grow in different agro-climatic conditions, they are often considered to be synonymous with each other. OBJECTIVE: The present study was aimed to systematically classify both the species CS and CR through their morphological features and chemical profiling using liquid chromatography-mass spectroscopy (LC-MS), gas chromatography-mass spectroscopy (GC-MS) and thin layer chromatography patterns of the rhizome extracts. MATERIALS AND METHODS: A method (LC-MS analysis) has been developed on Agilent LC-MSD Trap SL mass spectrometer equipped with Waters HR C18 column (3.9 mm × 300 mm, 6 µm) using isocratic elution with acetonitrile and water (70:30% v/v ratio). GC-MS analysis was performed on a Shimadzu GC-MS-QP 2010 equipped with DB-5 capillary column (30 m × 0.25 mm × 0.25 µm). RESULTS: Chemoprofiling of CS and CR using LC-MS and GC-MS suggested that these two are different based on their deferential spectral pattern, however, some of the common peaks were found in both the species. In addition, we also performed the preliminary phytochemical investigation of hexane and chloroform extracts of these species, which led to the isolation of stigmasterol, ß-sitosterol and lupeol as major constituents in CS. CONCLUSION: In summary, we have developed optimal chromatographic conditions (LC-MS and GC-MS) and morphological profiles to classify both the species, that is, CS and CR. Collectively, our analytical results coupled with the morphological data clearly suggested that CS and CR are morphologically different. SUMMARY: The huge demand for herbal medicine has put pressure on the supply of natural resources which ultimately results in use of substandard materials or substitution and adulteration. The medicinal plants, Cyperus rotundus L and Cyperus scariosus R.Br which belongs to cyperaceae family and extensively used in the traditional systems of medicine. Although these two species are grown in different soil conditions, Cyperus scariosus R.Br often treated as synonymous of Cyperus rotundus. Thus, the present study was undertaken to classify these two species systematically using the modern analytical techniques as a powerful tools. Further, we also carried out the preliminary phytochemical investigation of hexane and chloroform extracts of cyperus scariosus rhizomes, which resulted in the isolation of three compounds namely Sitosterol, Stigmasterol and Lupeol.

Ketorolac salt is a newly discovered DDX3 inhibitor to treat oral cancer.

DDX3 belongs to DEAD box RNA helicase family and is involved in the progression of several types of cancer. In this work, we employed a High Throughput Virtual screening approach to identify bioactive compounds against DDX3 from ZINC natural database. Ketorolac salt was selected based on its binding free energy less than or equals to -5 Kcal/mol with reference to existing synthetic DDX3 inhibitors and strong hydrogen bond interactions as similar to crystallized DDX3 protein (2I4I). The anti-cancer activity of Ketorolac salt against DDX3 was tested using oral squamous cell carcinoma (OSCC) cell lines. This compound significantly down regulated the expression of DDX3 in human OSCC line (H357) and the half maximal growth inhibitory concentration (IC50) of Ketorolac salt in H357 cell line is 2.6 µM. Ketorolac salt also inhibited the ATP hydrolysis by directly interacting with DDX3. More importantly, we observed decreased number of neoplastic tongue lesions and reduced lesion severity in Ketorolac salt treated groups in a carcinogen induced tongue tumor mouse model. Taken together, our result demonstrates that Ketorolac salt is a newly discovered bioactive compound against DDX3 and this compound can be used as an ideal drug candidate to treat DDX3 associated oral cancer.

Isolation and Characterization of the Anticancer Compound Piceatannol from Sophora Interrupta Bedd.

BACKGROUND: Sophora belongs to the family of Fabaceae and the species in this genus are currently used as a folklore medicine for preventing a variety of ailments including cancer. Our aim was to identify and validate an anticancer compound from Sophora interrupta using multi-spectroscopic, anticancer screening, and molecular docking approach. METHODS: The cytotoxicity of the various solvent extracts, petroleum ether, n-butanol, and ethyl acetate (EtOAc) of the S. interrupta root powder was evaluated in a breast cancer cell lines (MCF-7). The extract that had anticancer activity was subjected to column chromatography based on the polarity of the solvents. The anticancer activity of the elution fractions was validated using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The isolated metabolite fraction with anticancer activity was run through a C18 column isocratic and gradient high-performance liquid chromatography (HPLC). The structure of the isolated compound was characterized using (1)H nuclear magnetic resonance (NMR), (13)C-NMR, Fourier transform infrared spectroscopy, and liquid chromatography-mass spectrometer methods. RESULTS: The crude EtAOc extract effectively inhibited the proliferation of MCF-7 cells. The column eluted chloroform and EtOAc (4:6) fraction of the EtOAc extract showed significant anticancer activity in the MCF-7 cells compared with normal mesenchymal stem cells. This fraction showed three major peaks in the HPLC chromatogram and the first major peak with a retention time (RT) of 7.153 was purified using preparative-HPLC. The structure of the compound is a piceatannol, which is a metabolic product of resveratrol. Piceatannol formed direct two hydrogen bond interactions between Cys912 (2H), and Glu878 of vascular endothelial growth factor receptor 1 (VEGFR1) with a glide-score (G-score) of -10.193, and two hydrogen bond interactions between Cys919, and Asp1046 of VEGFR2, with a G-score of -8.359. The structure is similar to that of the crystallized protein for VEGFR1 and R2. CONCLUSIONS: Piceatannol is a secondary metabolite of S. interrupta that has anticancer activity. Moreover, piceatannol has been isolated for the first time from S. interrupta.

NZ51, a ring-expanded nucleoside analog, inhibits motility and viability of breast cancer cells by targeting the RNA helicase DDX3.

DDX3X (DDX3), a human RNA helicase, is over expressed in multiple breast cancer cell lines and its expression levels are directly correlated to cellular aggressiveness. NZ51, a ring-expanded nucleoside analogue (REN) has been reported to inhibit the ATP dependent helicase activity of DDX3. Molecular modeling of NZ51 binding to DDX3 indicated that the 5:7-fused imidazodiazepine ring of NZ51 was incorporated into the ATP binding pocket of DDX3. In this study, we investigated the anticancer properties of NZ51 in MCF-7 and MDA-MB-231 breast cancer cell lines. NZ51 treatment decreased cellular motility and cell viability of MCF-7 and MDA-MB-231 cells with IC50 values in the low micromolar range. Biological knockdown of DDX3 in MCF-7 and MDA-MB-231 cells resulted in decreased proliferation rates and reduced clonogenicity. In addition, NZ51 was effective in killing breast cancer cells under hypoxic conditions with the same potency as observed during normoxia. Mechanistic studies indicated that NZ51 did not cause DDX3 degradation, but greatly diminished its functionality. Moreover, in vivo experiments demonstrated that DDX3 knockdown by shRNA resulted in reduced tumor volume and metastasis without altering tumor vascular volume or permeability-surface area. In initial in vivo experiments, NZ51 treatment did not significantly reduce tumor volume. Further studies are needed to optimize drug formulation, dose and delivery. Continuing work will determine the in vitro-in vivo correlation of NZ51 activity and its utility in a clinical setting.

Identification of human cyclooxegenase-2 inhibitors from Cyperus scariosus (R.Br) rhizomes.

Cyperus scariosus (R.Br) belongs to the family Cyperaceae and it has a diverse medicinal importance. To identify human cyclooxegenase-2 (COX-2) inhibitors from C. scariosus, the rhizome powder was exhaustively extracted with various solvents based on the increasing polarity. Based on the presence and absence of secondary metabolites, we have selected the methanolic extract to evaluate the anti-oxidant and anti-inflammatory activity. The same extract was further subjected to gas chromatography-mass spectroscopy (GC-MS) analysis to identify the active compounds. Binding affinities of these compounds towards anti-inflammatory protein COX-2 were analyzed using molecular docking interaction studies. Phytochemical analysis showed that methanol extract is positive for all secondary metabolites. The antioxidant activity of the C. scariosus rhizomes methanolic extract (CSRME) is half to that of ascorbic acid at 50 µg/ml. The anti-inflammatory activity of CSRME is higher than that of diclofenac sodium salt at high concentration, which is evident from the dose dependent inhibition of bovine serum albumin denaturation at 40 µg/ml-5 mg/ml. GC-MS analysis showed the presence of nine compounds, among all N-methyl-1-adamantaneacetamide and 1,5,diphenyl-2H-1,2,4- triazine form a hydrogen bond interactions with Ser-530 and Tyr-385 respectively and found similar interactions with crystal structure of diclofenac bound COX-2 protein. Benzene-1, 2-diol, 4-(4-bromo-3 chlorophenyl iminomethyl forms hydrogen bond interactions with Thr-199 and Thr-200 as similar to crystallized COX-2 protein with valdecoxib. Collectively our results suggest that CSRME contains medicinally important anti-inflammatory compounds and this justifies the use of this plant as a folklore medicine for preventing inflammation associated disorders.

In-Vitro and in-Silico characterization of Sophora interrupta plant extract as an anticancer activity.

Sophora interrupta belongs to the family of Fabaceae and the species in this genus have a diverse medicinal importance as a folk medicine for preventing many ailments including cancer. In order to evaluate the anticancer activity of S.interrupta, we have performed in vitro anti-oxidant, anti-inflammatory, anti-proliferative, and cell based anticancer activity in MCF-7 and PC-3 cell lines. Secondary metabolites of S.interrupta were used to identify anticancer compounds using Open Eye software. The antioxidant activity of the S.interrupta root ethylacetate (SEA) extract at 100 µg/ml is equal to that of ascorbic acid at 50 µg/ml. The antiinflammatory activity of SEA is half of that of diclofenac at 50 µg/ml. Anticancer activity was detected by measuring the mitochondrial dehydrogenase activity (MTT assay). The half maximal inhibitory concentrations (IC50) for MCF-7 and PC-3 cell lines are 250 and 700 µg/ml respectively. This was supported by the morphological changes such as membrane blebbing, cell detachment and rounded cell morphology when compared to the parental cells. In addition, we observed few green cells (live) over red cells (dead) based on the uptake of acridine orange and ethidium bromide dyes. Kaempferol-3-O-b-D-glucopyranoside, a Secondary metabolite of S.interrupta form 6 hydrogen bond interactions with Arg 202, Gln 207, Gly 227, Gly 229, Thr 231 and Ala 232 human DEAD box RNA helicase, DDX3 protein and is equivalent to crystal structure of adenosine mono phosphate to DDX3. Overall, it suggests that the SEA extract has anticancer compounds, and it can be used to enhance death receptor mediated cancer cell death.

Invited review nonmulberry silk biopolymers.

The silk produced by silkworms are biopolymers and can be classified into two types--mulberry and nonmulberry. Mulberry silk of silkworm Bombyx mori has been extensively explored and used for century old textiles and sutures. But for the last few decades it is being extensively exploited for biomedical applications. However, the transformation of nonmulberry silk from being a textile commodity to biomaterials is relatively new. Within a very short period of time, the combination of load bearing capability and tensile strength of nonmulberry silk has been equally envisioned for bone, cartilage, adipose, and other tissue regeneration. Adding to its advantage is its diverse morphology, including macro to nano architectures with controllable degradation and biocompatibility yields novel natural material systems in vitro. Its follow on applications involve sustained release of model compounds and anticancer drugs. Its 3D cancer models provide compatible microenvironment systems for better understanding of the cancer progression mechanism and screening of anticancer compounds. Diversely designed nonmulberry matrices thus provide an array of new cutting age technologies, which is unattainable with the current synthetic materials that lack biodegradability and biocompatibility. Scientific exploration of nonmulberry silk in tissue engineering, regenerative medicine, and biotechnological applications promises advancement of sericulture industries in India and China, largest nonmulberry silk producers of the world. This review discusses the prospective biomedical applications of nonmulberry silk proteins as natural biomaterials.