Biochemistry And Microbial Sciences - Research Publications

Permanent URI for this collectionhttps://kr.cup.edu.in/handle/32116/27

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Author: search.filters.author.Prajapati, Kumari Sunita

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    Piper chaba, an Indian spice plant extract, inhibits cell cycle G1/S phase transition and induces intrinsic apoptotic pathway in luminal breast cancer cells
    (John Wiley and Sons Ltd, 2023-09-15T00:00:00) Prajapati, Kumari Sunita; Kumar, Shashank
    Piper chaba (Piperaceae) is a medicinal spice plant that possesses several pharmacological activities. In the present study, we for the first time studied the effect of P. chaba extract on breast cancer cells. P. chaba stem methanolic (PCSM) extract produced time and dose dependent cytotoxicity in luminal breast cancer cells (MCF-7 and T47D) with a minimal toxicity in breast normal cells (MCF-10A) at 10�100 �g/mL concentration. PCSM extract exerts 16.79 and 31.21 �g/mL IC50 for T47D and MCF-7 cells, respectively, in 48 h treatment. PCSM significantly arrests the T47D cells at the G0/G1 phase by reducing the CCND1 and CDK4 expression at mRNA and protein levels. PCSM extract treatment significantly altered nuclear morphology, mitochondria membrane potential, and production of reactive oxygen species in T47D cells at IC50 concentration. Extract treatment significantly altered the Bax/Bcl-2 ratio and altered caspase 8 and 3 mRNA/protein levels in T47D cells. Confocal microscopy showed an increase in late apoptosis in PCSM extract-treated breast cancer cells at IC50. Further, an increased caspase 9 and caspase 3/7 enzymatic activity was observed in test cells compared with nontreated cells. In conclusion, P. chaba phytocompound possesses the potential to induce cell cycle arrest and induce apoptosis in luminal breast cancer cells. � 2023 John Wiley & Sons Ltd.
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    Loss of miR-6844 alters stemness/self-renewal and cancer hallmark(s) markers through CD44-JAK2-STAT3 signaling axis in breast cancer stem-like cells
    (John Wiley and Sons Inc, 2023-07-12T00:00:00) Prajapati, Kumari Sunita; Kumar, Shashank
    MicroRNAs regulate breast stemness and self-renewal properties in breast cancer cells at the molecular level. Recently we reported the clinical relevance and in vitro expression profile of novel miR-6844 in breast cancer and -derived stem-like cells (mammosphere). In the present study, we first time explore the functional role of loss of miR-6844 in breast cancer cells derived mammosphere. Down expression of miR-6844 significantly decreased cell proliferation in MCF-7 and T47D cells derived mammosphere in a time-dependent manner. MiR-6844 down expression reduced the sphere formation in terms of size and number in test cells. Loss of miR-6844 significantly altered stemness and self-renewal markers (Bmi-1, Nanog, c-Myc, Sox2, and CD44) in mammosphere compared to negative control spheres. Moreover, loss of miR-6844 inhibits the JAK2-STAT3 signaling pathway by decreasing p-JAK2 and p-STAT3 levels in breast cancer cells derived mammosphere. Loss of miR-6844 expression significantly decreased CCND1 and CDK4 mRNA/protein levels and arrested breast cancer stem-like cells in G2/M phase. Reduced expression of miR-6844 increased Bax/Bcl-2 ratio, late apoptotic cell population, and Caspase 9 and 3/7 activity in the mammosphere. Low expression of miR-6844 decreased migratory and invasive cells by altering the expression of Snail, E-cad, and Vimentin at mRNA/protein levels. In conclusion, loss of miR-6844 decreases stemness/self-renewal and other cancer hallmark in breast cancer stem-like cells through CD44-JAK2-STAT3 axis. Thus, downregulation of miR-6844 by therapeutic agents might be a novel strategy to target breast cancer stemness and self-renewal. � 2023 Wiley Periodicals LLC.
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    Discovery of differentially expressed novel miRNAs in breast normal cells and their putative targets
    (Springer, 2023-01-21T00:00:00) Shuaib, Mohd; Prajapati, Kumari Sunita; Singh, Atul Kumar; Kumar, Shashank
    MicroRNAs (miRNAs) play critical role in normal breast development and their altered expression may lead to breast cancer. Identification of new miRNAs allows us to understand the normal physiological process and associated disease pathophysiology. In the present study we identify the novel miRNAs in withaferin A treated breast normal cells (MCF-10A) using small RNA sequencing. The pathophysiological potential of the identified miRNAs was checked by studying their expression pattern in MDA-MB-231 and MCF-7 breast cancer cells using qRT-PCR technique. The secondary/tertiary structure of the identified miRNAs, target gene enrichment in Gene Ontology terms and KEGG pathway, miRNA-mRNA interaction of the sorted target genes, miRNA-mRNA/miRNA-argonaute protein/miRNA-mRNA-argonaute protein interaction and stability, were studied using bioinformatics tools/software, and molecular dynamics simulations. Hsa-miR-N88585 and hsa-miR-N461089 were identified and validated as novel miRNAs in normal breast cells. Up-expression of identified miRNAs in MDA-MB-231 and MCF-7 cells indicates their oncogenic nature. Identified target genes were enriched in classical signaling pathways (AMPK and Ras) and important GO terms. PLXDC2, BHLHE40, ARMC8, and PECAM1, CDC27, KCNK3 genes were sorted as putative targets for hsa-miR-N88585 and hsa-miR-N461089, respectively. MD simulation revealed stable hsa-miR-N88585/hsa-miR-N461089-AGO protein complex formation which indicates their further processing. In conclusion, the study identifies hsa-miR-N88585 and hsa-miR-N461089 as novel miRNAs in breast normal cells which are significantly inversely expressed in breast cancer cells. Further experiments are required to study the role of identified novel miRNAs in normal breast development and pathophysiology of breast cancer. Graphical abstract: [Figure not available: see fulltext.]. � 2023, The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.
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    Natural Steroidal Lactone Induces G1/S Phase Cell Cycle Arrest and Intrinsic Apoptotic Pathway by Up-Regulating Tumor Suppressive miRNA in Triple-Negative Breast Cancer Cells
    (MDPI, 2022-12-27T00:00:00) Shuaib, Mohd; Prajapati, Kumari Sunita; Gupta, Sanjay; Kumar, Shashank
    Triple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer with minimal treatment options. In the present work, Withaferin A (WA), a natural steroidal lactone found in Withania somnifera (Solanaceae), was studied to deduce the miRNA expression modulation mediated anticancer mode of action in TNBC cells. Small RNA next generation sequencing (NGS) of WA (2 �M) and vehicle (0.1% DMSO)-treated MDA-MB-231 cells revealed a total of 413 differentially expressed miRNAs (DEMs) and demonstrated that WA potentially up-regulates the miR-181c-5p, miR-15a-5p, miR-500b-5p, miR-191-3p, and miR-34a-5p and down-regulates miR-1275, miR-326, miR-1908-5p, and miR-3940-3p among total DEMs. The NGS and qRT-PCR expression analysis revealed a significantly higher expression of miR-181c-5p among the top 10 DEMs. Predicted target genes of the DEMs showed enrichment in cancer-associated gene ontology terms and KEGG signaling pathways. Transient up-expression of mir-181c-5p showed a time-dependent decrease in MDA-MB-231 and MDA-MB-453 cell viability. Co-treatment of miR-181c-5p mimic and WA (at varying concentration) down-regulated cell cycle progression markers (CDK4 and Cyclin D1) at mRNA and protein levels. The treatment induced apoptosis in MDA-MB-231 cells by modulating the expression/activity of Bax, Bcl2, Caspase 3, Caspase 8, Caspase 3/7, and PARP at mRNA and protein levels. Confocal microscopy and Annexin PI assays revealed apoptotic induction in miRNA- and steroidal-lactone-treated MDA-MB-231 cells. Results indicate that the Withaferin A and miRNA mimic co-treatment strategy may be utilized as a newer therapeutic strategy to treat triple-negative breast cancer. � 2022 by the authors.
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    Hesperidin potentially interacts with the catalytic site of gamma-secretase and modifies notch sensitive genes and cancer stemness marker expression in colon cancer cells and colonosphere
    (Taylor and Francis Ltd., 2022-10-14T00:00:00) Singh, Atul Kumar; Prajapati, Kumari Sunita; Kumar, Shashank
    Gamma secretase (GS) produces Notch Intracellular Domain (NICD) by trans-membrane cleavage of notch receptor. The NICD enters the nucleus and activates the notch signaling pathway (NSP) by activating notch-responsive gene transcription. Hyperactivation of NSP is related to cancer aggressiveness, therapy resistance, and poor therapy outcome, and decreased overall disease-free survival in patients. Till date, none of the GS inhibitors (GSI) has been clinically approved due to their toxicity in patients. Thus in the present study, we explored the GS catalytic site binding potential of hesperidin (natural flavone glycoside) and its effect on notch responsive gene expression in HCT-116 cells. Molecular docking, MM-GBSA binding energy calculations, and molecular dynamics (MD) simulation experiments were performed to study the GS catalytic site binding potential of hesperidin. The compound showed better GS catalytic site binding potential at the active site compared to experimentally validated GSI, N-N-(3, 5-Difluorophenacetyl)-L-alanyl-S-phenylglycine t-butyl ester (DAPT) in molecular docking and MM-GBSA experiments. MD simulation results showed that hesperidin forms stable and energetically favorable complex with gamma secretase in comparison to standard inhibitor (DAPT)-GS complex. Further, in�vitro experiments showed that hesperidin inhibited cell growth and sphere formation potential in HCT-116 cells. Further, hesperidin treatment altered notch responsive genes (Hes1, Hey1, and E-cad) and cancer stemness/self-renewal markers expression at transcription levels. In conclusion, hesperidin produces toxicity in HCT-116 cells and decreases colonosphere formation by inhibiting transcription of notch signaling pathway target genes and stemness markers. Communicated by Ramaswamy H. Sarma. � 2022 Informa UK Limited, trading as Taylor & Francis Group.
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    Thiolated ?-cyclodextrin modified iron oxide nanoparticles for effective targeted cancer therapy
    (Elsevier Ltd, 2022-10-13T00:00:00) Nayak, Jyotsnamayee; Prajapati, Kumari Sunita; Kumar, Shashank; Vashistha, Vinod Kumar; Sahoo, Suban K.; Kumar, Rajender
    Inspired by the mitochondria iron-sulfur protein, direct functionalization of thiolated ?-cyclodextrin (?-CD-SH) on iron oxide nanoparticles (IONPs) through Fe-S bond was done. The resulting system had an average size of 14 nm with an ellipsoidal shape. The X-Ray photoelectron spectroscopy (XPS) confirmed the formation of the Fe-S bond. Doxorubicin (DOX) was chosen as a model drug, about 12.45 �M/mg entrapped in ?-CD-SH coated IONPs (T?CD-IONPs). The hybrid nanocarrier possessed high stability and drug loading efficiency. The invitro release data revealed an overall sustainable release profile without initial bust. The Higuchi kinetic model best fits the release mechanism, based on diffusion action in dosages proportional to the square root of time. The surface coating and particle size have a crucial role in the cellular responses and effective toxic mechanisms. The cellular internalization of drug-loaded nanoparticles (NPs) into the breast cancer cell line MCF-07 was done using MTT assay and confocal imaging. The prepared system shows high performance with an IC50 value at 67 nM of nanoparticle concentration. The prepared nanoparticles are promising candidates for the effective targeted delivery of hydrophobic drugs with enhanced theragnostic activity. � 2022 Elsevier Ltd
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    Acarbose Potentially Binds to the Type I Peptide Deformylase Catalytic Site and Inhibits Bacterial Growth: An In Silico and In Vitro Study
    (Bentham Science Publishers, 2022-09-23T00:00:00) Singh, Atul Kumar; Prajapati, Kumari Sunita; Kumar, Shashank
    Background: In bacteria, peptide deformylase (PDF), a metalloenzyme, removes N-formyl methio-nine from a nascent protein, which is a critical step in the protein maturation process. The enzyme is ubiqui-tously present in bacteria and possesses therapeutic target potential. Acarbose, an FDA-approved antidiabetic drug, is an alpha-glucosidase inhibitor of microbial origin. Clinical studies indicate that acarbose administration in humans can alter gut microbiota. As per the best of our knowledge, the antibacterial potential of acarbose has not been reported. Objective: The present study aimed to check the binding ability of acarbose to the catalytic site of E. coli PDF and assess its in vitro antibacterial activity. Methods: Molecular docking, molecular dynamic (MD) simulation, and MM-PBSA experiments were per-formed to study the binding potential of the catalytic site, and a disc diffusion assay was also employed to assess the antibacterial potential of acarbose. Results: Acarbose was found to form a hydrogen bond and interact with the metal ion present at the catalytic site. The test compound showed a better docking score in comparison to the standard inhibitor of PDF. MD simulation results showed energetically stable acarbose-PDF complex formation in terms of RMSD, RMSF, Rg, SASA, and hydrogen bond formation throughout the simulation period compared to the actinonin-PDF complex. Furthermore, MM-PBSA calculations showed better binding free energy (?G) of acarbose PDF than the actinonin-PDF complex. Moreover, acarbose showed in vitro antibacterial activity. Conclusion: Acarbose forms conformational and thermodynamically stable interaction with the E. coli peptide deformylase catalytic site. Results of the present work necessitate in-depth antimicrobial potential studies on the effect of acarbose on drug resistance and nonresistant bacteria. � 2022 Bentham Science Publishers.
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    Rutin Potentially Binds the Gamma Secretase Catalytic Site, Down Regulates the Notch Signaling Pathway and Reduces Sphere Formation in Colonospheres
    (MDPI, 2022-09-30T00:00:00) Singh, Atul Kumar; Shuaib, Mohd; Prajapati, Kumari Sunita; Kumar, Shashank
    Rutin, a natural flavonol, can modulate molecular signaling pathways and has considerable potential in cancer treatment. However, little is known about the effect of rutin on the notch signaling pathway (NSP) in cancer and cancer stem-like cells. In this study, we explored the effect of rutin on gamma secretase (GS, a putative notch signaling target) inhibition mediated NICD (Notch Intracellular Domain) production in colon cancer cells. Molecular docking, MM-GBSA, and Molecular dynamics (MD) simulation experiments were performed to check rutin�s GS catalytic site binding potential. The HCT-116 colon cancer and cancer stem-like cells (colonospheres) were utilized to validate the in silico findings. The NICD production, notch promoter assay, expression of notch target genes, and cancer stemness/self-renewal markers were studied at molecular levels. The results were compared with the Notch-1 siRNA transfected test cells. The in silico study revealed GS catalytic site binding potential in rutin. The in vitro results showed a decreased NICD formation, an altered notch target gene (E-cad, Hes-1, and Hey-1) expression, and a reduction in stemness/self-renewal markers (CD44, c-Myc, Nanog, and Sox2) in test cells in a time and dose-dependent manner. In conclusion, rutin inhibits the notch signaling pathway and reduces the stemness/self-renewal property in colon cancer cells and the colonospheres by targeting gamma secretase. The clinical efficacy of rutin in combination therapy in colon cancer may be studied in the future. � 2022 by the authors.
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    The Multifaceted Role of Signal Peptide-CUB-EGF Domain-Containing Protein (SCUBE) in Cancer
    (MDPI, 2022-09-13T00:00:00) Kumar, Shashank; Prajapati, Kumari Sunita; Gupta, Sanjay
    Signal peptide, CUB, and EGF-like domain-containing proteins (SCUBE) are secretory cell surface glycoproteins that play key roles in the developmental process. SCUBE proteins participate in the progression of several diseases, including cancer, and are recognized for their oncogenic and tumor suppressor functions depending on the cellular context. SCUBE proteins promote cancer cell proliferation, angiogenesis, invasion, or metastasis, stemness or self-renewal, and drug resistance. The association of SCUBE with other proteins alters the expression of signaling pathways, including Hedgehog, Notch, TGF-?/Smad2/3, and ?-catenin. Further, SCUBE proteins function as potential prognostic and diagnostic biomarkers for breast cancer, renal cell carcinoma, endometrial carcinoma, and nasopharyngeal carcinoma. This review presents key features of SCUBE family members, and their structure and functions, and highlights their contribution in the development and progression of cancer. A comprehensive understanding of the role of SCUBE family members offers novel strategies for cancer therapy. � 2022 by the authors.
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    Synthesis of thiolated chlorogenic acid-capped silver nanoparticles for the effective dual action towards antimicrobial and anticancer therapy
    (Springer Science and Business Media Deutschland GmbH, 2022-07-22T00:00:00) Nayak, Jyotsnamayee; Prajapati, Kumari Sunita; Kumar, Shashank; Sahoo, Suban K.; Kumar, Rajender
    Chlorogenic acid (CGA) was thiolated using Traut�s reagent. Then, thiolated chlorogenic acid (CAT) functionalized on the silver nanoparticles (AgNPs). The resulting CAT-functionalized silver nanoparticles (CAT-AgNPs) have higher stability with a uniform spherical shape compared to naked AgNPs. The average particle size is 10�nm, as revealed by transmission electron microscopy (TEM). The antimicrobial activity was higher against both gram-positive and gram-negative bacteria of spherically stable nanoparticles evaluated by the disk diffusion method. Minimal inhibitory concentration (MIC) values for CAT-AgNPs were 130��g/ml for Staphylococcus aureus and 150��g/ml for Escherichia coli. Cytotoxicity studies showed that the CAT-AgNPs show maximum activity against MCF-07 cell lines with an IC50 value of 20�ng/ml. Compared to traditional noncovalent interactions, the Ag�S covalent linkage of CAT with AgNPs helped in highly stable capping. The prepared CAT-AgNPs show anticancer and antibacterial activities, thus providing an efficient antimicrobial nano-drug carrier. Nanoparticles with a dual-mode of action help in preventing fouling of nanocarriers while delivering drugs to targeted cells, opening up many applications in nanobiotechnology and nanomedicine. � 2022, The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.