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Pulmonary Delivery of Voriconazole Loaded Nanoparticles Providing a Prolonged Drug Level in Lungs: A Promise for Treating Fungal Infection
No full textCurrent therapies are insufficient to prevent recurrent fungal infection especially in the lower part of the
lung. A careful and systematic understanding of the properties of nanoparticles plays a significant role in the design,development, optimization, and in vivo performances of the nanoparticles. In the present study, PLGA nanoparticles containing the antifungal drug voriconazole was prepared and two best formulations were selected for further characterization and in vivo studies. The nanoparticles and the free drug were radiolabeled with technetium-99m with 90% labelling efficiency, and the radiolabeled particles were administered to investigate the effect on their blood clearance, biodistribution, and
in vivo gamma imaging. In vivo deposition of the drug in the lobes of the lung was studied by LC−MS/MS study. The particles were found to be spherical and had an average hydrodynamic diameter of 300 nm with a smooth surface. The radiolabeled particles and the free drug were found to accumulate in various major organs. Drug accumulation was more pronounced in the lung in the case of administration of the nanoparticles than that of the free drug. The free drug was found to be excreted more rapidly than the nanoparticle containing drug following the inhalation route as assessed by gamma scintigraphy study. Thus, the
study reveals that pulmonary administration of nanoparticles containing voriconazole could be a better therapeutic choice even as compared to the iv route of administration of the free drug and/or the drug loaded nanoparticles
Sulfonoquinovosyl diacylglyceride selectively targets acute lymphoblastic leukemia cells and exerts potent anti-leukemic effects in vivo
No full textDNA topoisomerase II inhibitors e.g. doxorubicin and etoposide are currently used in the chemotherapy for acute lymphoblastic leukemia (ALL). These inhibitors have serious side effects during the chemotherapy e.g. cardiotoxicity and secondary malignancies. In this study we show
that sulfonoquinovosyl diacylglyceride (SQDG) isolated from Azadirachta indica exerts potent anti-ALL activity both in vitro and in vivo in nude mice and it synergizes with doxorubicin and etoposide. SQDG selectively targets ALL MOLT-4 cells by inhibiting catalytic activity of topoisomerase I enzyme and inducing p53 dependent apoptotic pathway. SQDG treatment induces recruitment of ATR at chromatin and arrests the cells in S-phase. Down-regulation of topoisomerase I or p53 renders the cells less sensitive for SQDG, while ectopic expression of wild type p53 protein in p53 deficient K562 cells results in chemosensitization of the cells for SQDG. We also show that constant
ratio combinations of SQDG and etoposide or SDQG and doxorubicin exert synergistic effects on MOLT-4 cell killing. This study suggests that doses of etoposide/ doxorubicin can be substantially reduced by combining SQDG with these agents during ALL chemotherapy and side effects caused can be minimized. Thus dual targeting of topoisomerase I and II enzymes is a promising strategy for
improving ALL chemotherap
Synthesis of Unsymmetrical Sulfides and Their Oxidation to Sulfones to Discover Potent Antileishmanial Agents
No full textUnsymmetrical sulfides were first synthesized using
combinations of a 1,3-dicarbonyl, an aromatic aldehyde and a thiol in the presence of 10 mol % ethanolic piperidine. These sulfides derivatives were subsequently converted into corresponding sulfones via oxidation in the presence of m-chloroperoxybenzoic acid (m-CPBA) at ice-bath to room temperature. The former reaction was achieved at
room temperature through one-pot three-component. The later was obtained in good yields using mild reaction conditions with flexibility in choice from a range of substrates. The antimicrobial properties of the newly synthesized sulfone derivatives were investigated against the protozoan parasite, Leishmania donovani, a causative agent of
visceral leishmaniasis (VL). Nine sulfone derivatives were found to be efficacious and exhibited significant antimicrobial activity. Further, these compounds were nontoxic on murine peritoneal macrophages thus eliminating potential cytoxicity in the host cells. These compounds may be indicated as potential leads in the treatment of visceral leishmaniasi
mRNA Targeting to Endoplasmic Reticulum Precedes Ago Protein Interaction and MicroRNA (miRNA)-mediated Translation Repression in Mammalian Cells
No full textmiRNA, Ago2, and target messages are enriched on ER in metazoan cells. Polysome association of newly formed mRNA
precedes Ago2 binding on ER membrane and its translation
repression. miRNA repression process is compartmentalized
in mammalian cells, and polysome association of
mRNA happens before the repression. This work provides a detailed understanding of the miRNA-mediated gene repression and mRNA compartmentalization in mammalian cells
Studies on deamidation mediated inactivation of cellular proteins and the role of protein isoaspartyl methyltransferase towards their repair
No full textThe spontaneous deamidation of asparaginyl residues at biological pH of 7.4
causes experimentally and biologically important changes resulting in the
formation of inactive peptides and proteins [1]. In asparaginyl deamidation, the
primary reaction products are aspartyl and isoaspartyl residues [1]. Deamidation
at neutral pH introduces a negative charge at the deamidation site and sometimes
also leads to β isomerization [2]. These alterations in structure affect the
properties of peptides and proteins in chemically and biologically important
pathways [1]. It has been suggested that isoaspartates in proteins serves as a
molecular timer of biological events and as a mechanism for postsynthetic
production of unique proteins of biological significance [3-5]. In the case of in
vivo protein turnover, the use of deamidation as a molecular timer has been
experimentally demonstrated [6-8].
Deamidation has been primarily observed to progressively increase in aging cells
and tissues[9]. Cellular aging is a fundamental phenomenon that affects all
somatic cells at definite points of time in their longevity cycle and whose
manifestation varies within cell types. Aging in its essence encompasses all those
processes that lead to a decline in the ability of the organism to cope with
environmental stresses and is defined as the accumulation of diverse deleterious
changes occurring in cells and tissues with time that are responsible for the
increased risk of disease and death. The major theories of aging eg, the free
radical theory [10], the immunologic theory[11], the inflammation theory[12],
and mitochondrial theory [13] are all specific of a particular cause of aging,
providing useful and important insights for the understanding of physiological
changes occurring with aging. With progressive cellular aging, the vital
biochemical process slows down and biologically relevant macromolecules
accumulate undesirable modifications. Such modifications further impair cellular
functions thus hastening the progression of aging
PI3K-Mediated Proliferation of Fibroblasts by Calendula officinalis Tincture: Implication in Wound Healing
No full textCalendula officinalis, a member of theAsteraceae family, is a flowering plant and has been used for its antibacterial,
antifungal, antiviral, antiinflammatory, anticancer and wound healing activity. The mode of action of C. officinalis
tincture on wound healing is poorly understood.Here, we investigated the role of C. officinalis tincture (CDOT) on
cell viability and wound closure. C. officinalis tincture stimulated both proliferation and migration of fibroblasts in a statistically significant manner in a PI3K-dependent pathway. The increase in phosphorylation of FAK(Tyr 397) and
Akt (Ser 473) was detected after treatment of CDOT. Inhibition of the PI3K pathway by wortmannin and LY294002
decreased both cell proliferation and cell migration. HPLC-ESIMS revealed the presence of flavonol glycosides as
themajor compounds of CDOT.Altogether, our results showed that CDOT potentiated wound healing by stimulating
proliferation and migration of fibroblast in a PI3K-dependent pathway, and the identified compounds are likely to be responsible for wound healing activit
Synthetic Studies towards Medium ring Heterocycles using Chiral Precursors
No full textAll reactions were performed in flame- or oven-dried glassware under a positive pressure of nitrogen or argon. Pd(OAc)2, Pd2(dba)3, BINAP, DPPF, CAN, and imiazole-2-carbaldehyde were purchased from Aldrich. Solvents were distilled and dried immediately prior to use.
Tetrahydrofuran (THF) and ether were distilled from sodium and benzophenone. Methylene chloride (CH2Cl2) was distilled from phosphorous pentoxide. Ethyl acetate, ethanol,
methanol, dimethylformamide (DMF), and toluene were distilled from potassium carbonate, Mg(OEt)2, Mg(OMe)2, CaH2, and sodium respectively. Petroleum ether used generally refers to the fraction having boiling point range 60-80 °C unless otherwise stated. Reactions were
monitored by thin layer chromatography (TLC) using Merck 60 F254 precoated silica gel plates (0.2 mm thickness). Components were visualized by illumination with ultraviolet light (254 nm) or staining with iodine. For routine column chromatography, silica gel of mesh sizes 60-120 or 100-200 (SRL, India) was used and for flash chromatography, product of mash size 230-400 (SRL, India) was employed. Reactions at room temperature generally imply a temperature of 25 °C. Cooling was performed using ice-water (0 °C) bath (unless
otherwise mentioned). Organic extracts were dried with anhydrous sodium sulphate. Solvents were concentrated in a rotary evaporator using water aspirator. All melting points (m.p.) were checked on a Thomas-Hoover capillary melting point apparatus and are uncorrected. NMR spectra of CDCl3 solutions of compounds were recorded with Bruker DPX-300 or Bruker AVANCE-600 spectrometer. NMR data are reported as chemical shifts in parts per million(ppm) using Tetramethyl silane (δ 0.0) as internal standard on the δ scale, J values are given in Hz, and multiplicity is quoted as follows: s, singlet; d, doublet; t, triplet; dd, doublet of
doublets; dt, doublet of triplets; q, quartet; br, broad; m, multiplet; etc. 13C NMR spectra were recorded on a Bruker DPX model (75 MHz) or Bruker AVANCE (150 MHz) spectrometer and chemical shift values are given in ppm using the mid-point of CDCl3 peaks as internal
standard (δ 77.0). DI-EIMS were recorded on a Shimadzu GCMS (model no QP5050A) and ESIMS were done on a Waters Micromass Q-TOF microTM Mass Spectrometer. IR spectra
were obtained using JASCO FT/IR model 410 instruments. Elemental analysis was done in a C, H, N analyzer. A SHIMADZU (model UV-1700) UV-vis Spectrophotometer was used for recording UV-vis Spectra. The corrected fluorescence spectra were recorded with SPEX Fluorolog II spectrofluorometer at right angle configuration. Fluorescence lifetimes were determined from the time resolved fluorescence decays by the method of time-correlated single photon counting (TCSPC) using Horiba Jobin Yvon Fluorocube spectrofluorometer with excitation source NanoLED-03 at 370 nm and TBX-04 detector. The instrument response time was ~ 1 ns
Development of New Classes of Peptide Therapeutics
No full textSignaling cascades that propagate biological information in the cell consist of complex networks of protein-protein interactions which play a central role in a large number of cellular processes like cellular homeostasis, proliferation, migration, angiogenesis, immune responses, apoptosis and cell death. De-regulation of these interactions lead to different diseases, most prominent of which is cancer. Protein–protein interactions, being part of a large number of signaling networks, are potential targets for drug development and peptides are better inhibitors of protein-protein interactions than small molecules. The major aim of this work has been to develop peptide inhibitors of a particular protein-protein interaction and test their therapeutic potential, especially in malignant melanoma cells. S100B, a calcium-regulated protein, plays a crucial role in the proliferation of malignant melanoma cells through protein–protein interactions. The particular protein-protein interaction that has been chosen in this study is the S100B-p53 interaction. S100B is a member of the S100 protein family of Ca2+ binding proteins of the EF-hand type, comprising more than 20 members and is expressed in many cancers like malignant melanoma, glioblastoma etc. Recently S100B has come into focus as it binds directly to the tumor suppressor p53, reduces p53 protein levels and inhibits wild-type p53 functions in malignant melanoma. TRTK-12, a peptide derived from the actin-binding protein Cap Z and the C-terminal region of p53 (residues 367-388) have overlapping binding sites on S100B. Also each monomeric subunit of S100B binds to one peptide. Both the peptides are unstructured in solution but become helical upon binding to S100B. Hence, dimeric helically constrained peptides derived from TRTK-12 and the p53 C-terminal end have been designed, synthesized and purified, which induce cell death in malignant melanoma cells where S100B is overexpressed and downregulates wild type p53 by binding to the latter.The peptide derived from TRTK-12, Arg-Br-aib-TRTK, induces rapid apoptosis in SK-Mel- 5 melanoma cells. This peptide binds to its target S100B in vivo. Apoptosis induced by Arg-Br-aib-TRTK peptide occurs due to the mitochondrial translocation of cytosolic p53. This apoptotic cell death is independent of p53-dependent gene expression as well as caspase activation. No ROS generation is involved in the cell death induced by Arg-Braib- TRTK peptide. This peptide is a better therapeutic molecule than either pentamidine or Nutlin-3 in melanoma cells. Thus, this peptide has the potential to become a candidate drug for malignant melanoma. Monomeric and dimeric helically constrained peptides have been derived from the Cterminal negative regulatory domain of p53 and their efficacy has been studied in malignant melanoma cells. The branched dimeric peptide p53CT-B-Br-6DR binds tightly to S100B in vitro and induces cell death in melanoma cells. It is more potent than the corresponding helically constrained monomeric peptide p53CT-B-6DR. However, the mutant branched peptide p53CT-B-Br-6DR-AAA also induces cell death to some extent. This may be attributed to p53 C-terminus binding to other protein targets apart from S100B. The other part of the work deals with peptides which specifically target to cancerousb tumors and/or blood vessels associated with the tumors. These peptides are called tumor homing peptides and their major application is targeted delivery of a drug conjugated to such peptides. They have also been used for delivery of oligonucleotides, imaging agents, inorganic nanoparticles, liposomes and viruses apart from drugs. A tumor homing peptide HPM and its designed mutant HPV, have been studied for their tumor targeting efficacy in a syngenic mouse melanoma model in vivo by animal biodistribution studies. Both the HPM and HPV peptides are deemed unsuitable for targeting Arg-Br-aib-TRTK peptide to this particular syngenic mouse melanoma model
Divergences in gene repertoire among the reference Prevotella genomes derived from distinct body sites of human
No full textThe community composition of the human microbiome is known to vary at distinct anatomical niches. But little is known about the nature of variations, if any, at the genome/sub-genome levels of a specific microbial community across different niches. The present report aims to explore, as a case study, the variations in gene repertoire of 28 Prevotella reference genomes derived from different body-sites of human, as reported earlier by the Human Microbiome Consortium.
Results: The pan-genome for Prevotella remains “open”. On an average, 17% of predicted protein-coding genes of
any particular Prevotella genome represent the conserved core genes, while the remaining 83% contribute to the
flexible and singletons. The study reveals exclusive presence of 11798, 3673, 3348 and 934 gene families and exclusive absence of 17, 221, 115 and 645 gene families in Prevotella genomes derived from human oral cavity, gastro-intestinal tracts (GIT), urogenital tract (UGT) and skin, respectively. Distribution of various functional COG categories differs significantly among the habitat-specific genes. No niche-specific variations could be observed in distribution of KEGG pathways.
Conclusions: Prevotella genomes derived from different body sites differ appreciably in gene repertoire, suggesting
that these microbiome components might have developed distinct genetic strategies for niche adaptation within the
host. Each individual microbe might also have a component of its own genetic machinery for host adaptation,
as appeared from the huge number of singleton
Tannery Effluent Treatment by Microfiltration through Ceramic Membrane for Water Reuse: Assessment of Environmental Impacts
No full textThe performance evaluation of an indigenously developed ceramic membrane from a clay–alumina mixture was evaluated toward microfiltration treatment of tannery effluent from a secondary clarifier. The study was aimed at observing the reuse efficiency of the membrane treated effluent using Pistia sp. as plant model and Poecilia sp. as fish
model. About 70–86% chemical oxygen demand removal and 85% total organic carbon removal was achieved in the ceramic membrane based microfiltration process. The comet assay of Pistia sp. leaves showed formation of DNA tail in the untreated effluent suggesting DNA damage whereas no such observations were noted in treated water. Oxidative stress biomarkers like guaiacol peroxidase, superoxide dismutase, etc., in both Pistia sp. and Poecilia sp. increased considerably in untreated effluent, whereas the
treated water values were close to that of control. The overall process demonstrated that microfiltration by ceramic membranes might prove as effective means of wastewater reuse for aquaculture, agriculture, as well as in industrial sectors