Combating planktonic and biofilm growth of Serratia marcescens by repurposing ebselen.

AIM OF THE STUDY: The rising instances of multidrug-resistant pathogens are rapidly evolving into a global healthcare crisis. Identifying new ways of synthesis of antibiotics is both time-consuming and expensive. Repurposing existing drugs for the treatment of such antimicrobial-resistant pathogens has also been explored. METHODS AND RESULTS: In the current study, ebselen was screened for antibacterial and antibiofilm activity against Serratia marcescens. Various antibacterial studies such as minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), time-kill curves, intracellular reactive oxygen species (ROS) quantification, and colony-forming unit assays were performed. The antibiofilm potential was assayed by biofilm inhibition, cell surface hydrophobicity assay, eradication, quantification of extracellular DNA (eDNA), and extracellular polymeric substance (EPS) layer and scanning electron microscopy (SEM) analysis were performed. Anti-quorum sensing assay was validated by quantifying the virulence factors production. Further molecular docking of ebselen with two quorum sensing (QS) specific proteins was also carried out. Antibacterial susceptibility tests showed potent antimicrobial activity of ebselen against S. marcescens with MIC50 of 14 µg/mL. Ebselen's ability to disturb the redox environment by inducing significant ROS generation led to bacterial death. It also showed concentration-dependent bactericidal activity as indicated by reduced bacterial growth and colony-forming unit propagation. Ebselen was also found to prevent biofilm attachment by altering the cell surface hydrophobicity while also being effective against preformed biofilms as validated by scanning electron microscopy (SEM) analysis. Additionally, ebselen showed reduced virulence factors like urease enzyme activity and prodigiosin pigment production indicating its promising anti-quorum sensing potential. Molecular docking analysis validated the strong binding of ebselen with QS-specific proteins (1Joe and PigG) with binding energies of - 6.6 and - 8.1kj/mol through hydrogen bonds and aromatic interactions. These results show that ebselen has potent antibiofilm potential that can be explored to identify treatment against bacterial infections.

Elucidating the anti-biofilm and anti-quorum sensing potential of selenocystine against respiratory tract infections causing bacteria: in vitro and in silico studies.

Bacteria are increasingly relying on biofilms to develop resistance to antibiotics thereby resulting in their failure in treating many infections. In spite of continuous research on many synthetic and natural compounds, ideal anti-biofilm molecule is still not found thereby warranting search for new class of molecules. The current study focuses on exploring anti-biofilm potential of selenocystine against respiratory tract infection (RTI)-causing bacteria. Anti-bacterial and anti-biofilm assays demonstrated that selenocystine inhibits the growth of bacteria in their planktonic state, and formation of biofilms while eradicating preformed-biofilm effectively. Selenocystine at a MIC50 as low as 42 and 28 µg/mL effectively inhibited the growth of Klebsiella pneumonia and Pseudomonas aeruginosa. The antibacterial effect is further reconfirmed by agar cup diffusion assay and growth-kill assay. Selenocystine showed 30-60% inhibition of biofilm formation in K. pneumonia, and 44-70% in P. aeruginosa respectively. It also distorted the preformed-biofilms by degrading the eDNA component of the Extracellular Polymeric Substance matrix. Molecular docking studies of selenocystine with quorum sensing specific proteins clearly showed that through the carboxylic acid moiety it interacts and inhibits the protein function, thereby confirming its anti-biofilm potential. With further validation selenocystine can be explored as a potential candidate for the treatment of RTIs.

Electrostatically bound lanreotide peptide - gold nanoparticle conjugates for enhanced uptake in SSTR2-positive cancer cells.

Lanreotide peptide (LP) has high affinity to somatostatin receptors like SSTR2 and is commonly used in the treatment of neuro-endocrine tumors. The main objective of this study is to target gold nanoparticles (AuNPs) towards SSTR2-positive cancer cells using lanreotide peptide (LP) as the targeting agent for enhanced tumor uptake and antitumor activity. pH mediated changes in the surface potential of LP and AuNP is used to prepare electrostatically bound AuNP-LP complexes. AuNP-LP complex formation was demonstrated by UV-Visible spectroscopy, surface potential, dynamic light scattering (DLS), small angle X-ray scattering and HR-TEM. Confocal microscopy and flow cytometric studies show that AuNP-LP complex has higher cellular uptake in SSTR2 expressed cancer cells (MCF-7 and AR42J) than in CHO cells. The enhanced cellular uptake of LP coated AuNPs lead to ~1.5 to 2-fold GSH depletion and enhanced ROS generation in MCF-7 cells. The preferential cytotoxicity of the AuNP-LP complex towards MCF-7 and AR42J cells, as revealed by MTT assay, is consistent with the increased cellular uptake. Our studies demonstrate that LP coated AuNP can be used as an effective platform to selectively target SSTR2 positive cancer cells for combination therapy approaches involving gold nanoparticles.

Glutamic acid-coated Fe3O4 nanoparticles for tumor-targeted imaging and therapeutics.

We have developed surface functionalised Fe3O4 magnetic nanoparticles (MNPs) based system that can be used for tumor-targeted multimodal therapies and MR imaging. Biocompatible, non-essential amino acid (glutamic acid) was introduced onto the surface of Fe3O4 MNPs to provide functional sites for binding of chemotherapeutic drugs. These glutamic acid-coated Fe3O4 MNPs (GAMNPs) exhibit good water-dispersibility, magnetic responsivity and pH dependent charge conversal feature. The magnetic core as well as organic shell of GAMNPs was characterized by XRD, TEM, DLS, FTIR, PPMS and UV-visible spectroscopy and zeta-potential analyzer etc. The broad spectrum anticancer drugs, doxorubicin hydrochloride (DOX) and methotrexate (MTX) were electrostatically and covalently conjugated to the surface of GAMNPs, respectively for combination chemotherapy. These dual drugs loaded system (DOX-MTX-GAMNPs) shows pH dependent release behaviour of both the drugs and enhanced toxicity towards breast cancer cell line (MCF-7) as compared to their individual treatment. Fluorescence microscopy and flow cytometric analyses confirmed the successful uptake of drug loaded system into MCF-7 cell lines. Further MTX being analogue of folic acid, its co-delivery with DOX would help in internalization of both the drugs into MCF-7 cells. These GAMNPs also show good heating efficiency under AC magnetic field (Intrinsic loss power, ILP = 0.95 and 0.73 and 0.48 nHm2/Kg at Fe concentration of 0.5, 1 and 2 mg/ml, respectively) and transverse relaxivity (r2 = 152 mM-1 s-1) indicating their potential capability for hyperthermia therapy and MRI tracking. Furthermore, it has been observed that the combination of chemotherapeutic drugs and hyperthermia leads to an enhancement of cytotoxicity in MCF-7 cells.

Multitargeting by curcumin as revealed by molecular interaction studies.

Curcumin (diferuloylmethane), the active ingredient in turmeric (Curcuma longa), is a highly pleiotropic molecule with anti-inflammatory, anti-oxidant, chemopreventive, chemosensitization, and radiosensitization activities. The pleiotropic activities attributed to curcumin come from its complex molecular structure and chemistry, as well as its ability to influence multiple signaling molecules. Curcumin has been shown to bind by multiple forces directly to numerous signaling molecules, such as inflammatory molecules, cell survival proteins, protein kinases, protein reductases, histone acetyltransferase, histone deacetylase, glyoxalase I, xanthine oxidase, proteasome, HIV1 integrase, HIV1 protease, sarco (endo) plasmic reticulum Ca(2+) ATPase, DNA methyltransferases 1, FtsZ protofilaments, carrier proteins, and metal ions. Curcumin can also bind directly to DNA and RNA. Owing to its ß-diketone moiety, curcumin undergoes keto-enol tautomerism that has been reported as a favorable state for direct binding. The functional groups on curcumin found suitable for interaction with other macromolecules include the α, ß-unsaturated ß-diketone moiety, carbonyl and enolic groups of the ß-diketone moiety, methoxy and phenolic hydroxyl groups, and the phenyl rings. Various biophysical tools have been used to monitor direct interaction of curcumin with other proteins, including absorption, fluorescence, Fourier transform infrared (FTIR) and circular dichroism (CD) spectroscopy, surface plasmon resonance, competitive ligand binding, Forster type fluorescence resonance energy transfer (FRET), radiolabeling, site-directed mutagenesis, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), immunoprecipitation, phage display biopanning, electron microscopy, 1-anilino-8-naphthalene-sulfonate (ANS) displacement, and co-localization. Molecular docking, the most commonly employed computational tool for calculating binding affinities and predicting binding sites, has also been used to further characterize curcumin's binding sites. Furthermore, the ability of curcumin to bind directly to carrier proteins improves its solubility and bioavailability. In this review, we focus on how curcumin directly targets signaling molecules, as well as the different forces that bind the curcumin-protein complex and how this interaction affects the biological properties of proteins. We will also discuss various analogues of curcumin designed to bind selective targets with increased affinity.

Interaction of a curcumin analogue dimethoxycurcumin with DNA.

Dimethoxycurcumin (Dimc), a synthetic analogue of curcumin, that has been reported to exhibit better in vivo stability and anti-tumour activity, was investigated for its interaction with DNA, employing spectroscopic methods based on absorption, fluorescence, circular dichroism (CD), ethidium bromide (EtBr) competitive binding assay, 4'-6-diamidino-2-phenylindole (DAPI) displacement assay and fluorescence resonance energy transfer (FRET) assay. The mean binding constant for its interaction with calf thymus DNA (ct-DNA) was estimated to be 4.4±0.8 × 10(4) m(-1) . The studies using CD revealed that Dimc did not cause unwinding of the ct-DNA helix or induce major conformational changes. The EtBr and DAPI assays indicated that Dimc is not an intercalator but a minor groove binder. FRET assay also confirmed that Dimc interacts with DNA strands. Furthermore, viscosity measurements of ct-DNA solutions in the presence of Dimc supported these spectroscopic observations. Addition of Dimc to MCF-7 cells showed nuclear localization as visualized by confocal microscopy. In conclusion, the present studies addressed the mode of interaction of Dimc with biomolecules, which may have implications in developing Dimc as a DNA-targeted drug.

Biological activities of curcumin and its analogues (Congeners) made by man and Mother Nature.

Curcumin, a yellow pigment present in the Indian spice turmeric (associated with curry powder), has been linked with suppression of inflammation; angiogenesis; tumorigenesis; diabetes; diseases of the cardiovascular, pulmonary, and neurological systems, of skin, and of liver; loss of bone and muscle; depression; chronic fatigue; and neuropathic pain. The utility of curcumin is limited by its color, lack of water solubility, and relatively low in vivo bioavailability. Because of the multiple therapeutic activities attributed to curcumin, however, there is an intense search for a "super curcumin" without these problems. Multiple approaches are being sought to overcome these limitations. These include discovery of natural curcumin analogues from turmeric; discovery of natural curcumin analogues made by Mother Nature; synthesis of "man-made" curcumin analogues; reformulation of curcumin with various oils and with inhibitors of metabolism (e.g., piperine); development of liposomal and nanoparticle formulations of curcumin; conjugation of curcumin prodrugs; and linking curcumin with polyethylene glycol. Curcumin is a homodimer of feruloylmethane containing a methoxy group and a hydroxyl group, a heptadiene with two Michael acceptors, and an alpha,beta-diketone. Structural homologues involving modification of all these groups are being considered. This review focuses on the status of all these approaches in generating a "super curcumin.".

Effect of curcumin and curcumin copper complex (1:1) on radiation-induced changes of anti-oxidant enzymes levels in the livers of Swiss albino mice.

The effect of mononuclear copper (II) complex of curcumin in 1:1 stoichiometry (hereafter referred to as complex) administered 30 min before gamma-irradiation (4.5 Gy) on alterations in antioxidant and Thiobarbituric acid reactive substances (TBARS) levels in livers was studied in comparison to curcumin at a dose of 50 mg/kg. The different antioxidants like GSH, GST, catalase, SOD, TBARS and total thiols were estimated in the liver homogenates excised at different time intervals (1, 2 and 4 h) post irradiation using colorimetric methods. There was a radiation-induced decrease in the levels of all the studied enzymes at 1 h post irradiation, while an increase was observed at later time points. Both curcumin and complex treatment in sham-irradiated mice decreased the levels of GSH and total thiols, whereas there was an increase in the levels of catalase, GST and SOD compared to normal control. Under the influence of irradiation, both curcumin and complex treatment protected the decline in the levels of GSH, GST, SOD, catalase and total thiols, and inhibited radiation-induced lipid peroxidation. Further, the complex was found to be more effective in protecting the enzymes at 1 h post irradiation compared to curcumin treated group. This may be due to the higher rate constants of the complex compared to curcumin for their reactions with various free radicals.