Showing papers by "Amit Singh published in 2015"

Hyaluronic acid targeting of CD44 for cancer therapy: from receptor biology to nanomedicine

Abstract: Cluster of differentiation-44 (CD44) is a ubiquitously present glycoprotein on the surface of mammalian cells that plays a significant role in a number of biological functions. Since the discovery that the receptor is over-expressed in a variety of solid tumors, such as pancreatic, breast and lung cancer, many studies have focused on methods for targeting CD44 in an attempt to improve drug delivery and discrimination between healthy and malignant tissue, while reducing residual toxicity and off-target accumulation. In this review, we describe CD44 receptor biology and its involvement in the different stages of tumor growth and metastasis, as well as methods currently used for targeting the receptor. Hyaluronic acid, the primary CD44 binding molecule, has proved a significant ally in developing nanocarriers that demonstrate preferential tumor accumulation and increased cell uptake. We outline a number of research approaches from the current literature that take advantage of hyaluronic acid's targeting ability and describe the possible advantages for each approach. The value of CD44 targeting can be easily appreciated from the number of different approaches that have reached clinical trials.

MDR1 siRNA loaded hyaluronic acid-based CD44 targeted nanoparticle systems circumvent paclitaxel resistance in ovarian cancer.

Abstract: Development of multidrug resistance (MDR) is an almost universal phenomenon in patients with ovarian cancer, and this severely limits the ultimate success of chemotherapy in the clinic. Overexpression of the MDR1 gene and corresponding P-glycoprotein (Pgp) is one of the best known MDR mechanisms. MDR1 siRNA based strategies were proposed to circumvent MDR, however, systemic, safe, and effective targeted delivery is still a major challenge. Cluster of differentiation 44 (CD44) targeted hyaluronic acid (HA) based nanoparticle has been shown to successfully deliver chemotherapy agents or siRNAs into tumor cells. The goal of this study is to evaluate the ability of HA-PEI/HA-PEG to deliver MDR1 siRNA and the efficacy of the combination of HA-PEI/HA-PEG/MDR1 siRNA with paclitaxel to suppress growth of ovarian cancer. We observed that HA-PEI/HA-PEG nanoparticles can efficiently deliver MDR1 siRNA into MDR ovarian cancer cells, resulting in down-regulation of MDR1 and Pgp expression. Administration of HA-PEI/HA-PEG/MDR1 siRNA nanoparticles followed by paclitaxel treatment induced a significant inhibitory effect on the tumor growth, decreased Pgp expression and increased apoptosis in MDR ovarian cancer mice model. Our findings suggest that CD44 targeted HA-PEI/HA-PEG/MDR1 siRNA nanoparticles can serve as a therapeutic tool with great potentials to circumvent MDR in ovarian cancer.

Robust and Imperceptible Dual Watermarking for Telemedicine Applications

Abstract: In this paper, the effects of different error correction codes on the robustness and imperceptibility of discrete wavelet transform and singular value decomposition based dual watermarking scheme is investigated. Text and image watermarks are embedded into cover radiological image for their potential application in secure and compact medical data transmission. Four different error correcting codes such as Hamming, the Bose, Ray-Chaudhuri, Hocquenghem (BCH), the Reed---Solomon and hybrid error correcting (BCH and repetition code) codes are considered for encoding of text watermark in order to achieve additional robustness for sensitive text data such as patient identification code. Performance of the proposed algorithm is evaluated against number of signal processing attacks by varying the strength of watermarking and covers image modalities. The experimental results demonstrate that this algorithm provides better robustness without affecting the quality of watermarked image.This algorithm combines the advantages and removes the disadvantages of the two transform techniques. Out of the three error correcting codes tested, it has been found that Reed---Solomon shows the best performance. Further, a hybrid model of two of the error correcting codes (BCH and repetition code) is concatenated and implemented. It is found that the hybrid code achieves better results in terms of robustness. This paper provides a detailed analysis of the obtained experimental results.

Mycobacterium tuberculosis Inhibits RAB7 Recruitment to Selectively Modulate Autophagy Flux in Macrophages

Abstract: Here we report a novel regulatory mechanism for autophagy-mediated degradation of Mycobacterium tuberculosis (Mtb) and specific strategy exploited by the virulent Mtb to evade it. We show while both avirulent (H37Ra) and virulent (H37Rv) mycobacteria could readily localize to autophagosomes, their maturation into autolysosomes (flux) was significantly inhibited by the latter strain. The inhibition of autophagy flux by the virulent strain was highly selective, as it did not perturb the basal autophagy flux in the macrophages. Selective inhibition of flux of Mtb-containing autophagosomes required virulence regulators PhoP and ESAT-6. We show that the maturation of Mtb-containing autophagosomes into autolysosomes required recruitment of the late endosome marker RAB7, forming the intermediate compartment amphisomes. Virulent Mtb selectively evaded their targeting to the amphisomes. Thus we report a crosstalk between autophagy and phagosome maturation pathway and highlight the adaptability of Mtb, manifested by selective regulation of autophagy flux.

Multiple Watermarking on Medical Images Using Selective Discrete Wavelet Transform Coefficients

Abstract: This paper presents a new spread-spectrum based secure multiple watermarking scheme on medical images in wavelet transform domain by using selective discrete wavelet transform (DWT) coefficients for embedding. The proposed algorithm is applied for embedding text watermarks like patient identification/source identification represented in binary arrays using ASCII code and doctor’s signature or telemedicine centre name represented in binary image format into host digital radiological image for potential telemedicine applications. The algorithm is based on secure spread-spectrum technique where pseudo-noise (PN) sequences are generated corresponding to each watermarking bit and embedding of these sequences is done column wise into the selected DWT coefficients in the subband. Selection of DWT coefficient for embedding is done by thresholding the coefficient values present in that column. In the embedding process, the cover image is decomposed at second level DWT. The image and text watermark is embedded into the selective coefficients of the first level and second level DWT respectively. In order to enhance the robustness of text watermarks like patient identity code, error correcting code (ECC) is applied to the ASCII representation of the text watermark before embedding. Results are obtained by varying the gain factor, subband decomposition levels, size of watermark, and medical image modalities. Performace of the proposed watermarking algorithm is analyzed against numerous known attacks like compression, filtering, noise, sharpening, scaling and histogram equalization and desired outcome is obtained without much degradation in extracted watermarks and watermarked image quality. The method is compared with other reported techniques and has been found to be giving superior performance for robustness and imperceptibility suggested by other authors.

Identification and characterization of a small heat shock protein 17.9-CII gene from faba bean (Vicia faba L.)

Abstract: We cloned and characterized the full-length coding sequence of a small heat shock protein 17.9 gene from faba bean encoding 160 amino acids and containing the conserved α-crystallin domain at the C-terminus. Homology and phylogenetic analysis suggested its proximity with the class II sHsp members of fabaceae family. Therefore, we name this gene as VfHsp17.9-CII. The VfHsp17.9-CII transcript showed a clear heat stress induction pattern in leaves of young seedlings and flowering plants. Transient expression of VfHsp17.9-CII fused with green fluorescent protein reporter indicated its nuclear localization. Overexpression of recombinant VfHsp17.9-CII protein in Escherichia coli cells increased tolerance of the bacterial cells to heat and arsenic stresses. The reduction of faba bean pollen viability in response to heat stress correlated with the accumulation pattern of VfHsp17.9-CII transcript in heat stressed pollen. It is suggested that VfHsp17.9-CII protein plays a key role in heat and heavy metal stress tolerance.

Cluster of Differentiation 44 Targeted Hyaluronic Acid Based Nanoparticles for MDR1 siRNA Delivery to Overcome Drug Resistance in Ovarian Cancer

Abstract: Purpose Approaches for the synthesis of biomaterials to facilitate the delivery of “biologics” is a major area of research in cancer therapy. Here we designed and characterized a hyaluronic acid (HA) based self-assembling nanoparticles that can target CD44 receptors overexpressed on multidrug resistance (MDR) ovarian cancer. The nanoparticle system is composed of HA-poly(ethyleneimine)/HA-poly(ethylene glycol) (HA-PEI/HA-PEG) designed to deliver MDR1 siRNA for the treatment of MDR in an ovarian cancer model.

Multilevel Encrypted Text Watermarking on Medical Images Using Spread-Spectrum in DWT Domain

Abstract: This paper presents a secure multilevel watermarking scheme in which the encrypted text acts as a watermark. The algorithm is based on secure spread-spectrum technique for digital images in discrete wavelet transform (DWT) domain. Potential application of the proposed watermarking scheme is successfully demonstrated for embedding various medical watermarks in text format at different subband decomposition levels depending upon their performance requirements. In the embedding process, the cover CT Scan image is decomposed up to third level of DWT coefficients. Different text watermarks such as personal and medical record of the patient, diagnostic/image codes and doctor code/signature are embedded into the selective coefficients of the second and third level DWT for potential telemedicine applications. Selection of DWT coefficients for embedding is done by column wise thresholding of coefficients values. Also, encryption is applied to the ASCII representation of the text and the encoded text watermark is embedded. The algorithm correctly extracts the embedded watermarks without error and is robust against numerous known attacks without much degradation of the medical image quality of the watermarked image.

Genetic approaches for breeding heat stress tolerance in faba bean ( Vicia faba L.)

Abstract: Vicia faba L. (faba bean) is an important legume and is cultivated essentially as a cool-season crop. Changes in sowing dates and lack of precipitation expose faba bean crop to drought and heat stresses. The gradual rise in global temperatures owing to climate change is likely to exacerbate the detrimental effects of hot and dry climatic conditions on faba bean cultivation. High temperature stress is particularly damaging to faba bean during the flowering period, when the viability of pollen is critical for successful reproduction. Recent studies have shown that maintenance of protein homeostasis through synthesis of heat shock proteins plays a key role in the heat response of plants. To date, there has been no significant work linking the heat response of faba bean to the repertoire of its heat shock proteins. While quantitative trait loci have been identified for resistance against biotic stresses in faba bean, there is no parallel success with abiotic stresses in this species. Programs aiming at genetic improvement of the heat/drought resistance of this crop by both conventional breeding and molecular breeding methods are hampered because of the large and majorly ill-analyzed genome of faba bean plants. Likewise, molecular and biotechnology-related tools are poorly developed for faba bean; as a result, the fruits of transgenic research developed with model plant species are not reaching this crop. While specifically discussing the prospects for the genetic improvement of faba bean against heat and drought stresses, we highlight the areas of research which need to be strengthened on faba bean.

EGFR Targeted Theranostic Nanoemulsion for Image-Guided Ovarian Cancer Therapy

Abstract: Purpose Platinum-based therapies are the first line treatments for most types of cancer including ovarian cancer. However, their use is associated with dose-limiting toxicities and resistance. We report initial translational studies of a theranostic nanoemulsion loaded with a cisplatin derivative, myrisplatin and pro-apoptotic agent, C6-ceramide.

Inhibition of hexokinase-2 with targeted liposomal 3-bromopyruvate in an ovarian tumor spheroid model of aerobic glycolysis.

Abstract: Background: The objective of this study was to evaluate the expression levels of glycolytic markers, especially hexokinase-2 (HK2), using a three-dimensional multicellular spheroid model of human ovarian adenocarcinoma (SKOV-3) cells and to develop an epidermal growth factor receptor-targeted liposomal formulation for improving inhibition of HK2 and the cytotoxicity of 3-bromopyruvate (3-BPA). Methods: Multicellular SKOV-3 tumor spheroids were developed using the hanging drop method and expression levels of glycolytic markers were examined. Non-targeted and epidermal growth factor receptor-targeted liposomal formulations of 3-BPA were formulated and characterized. Permeability and cellular uptake of the liposomal formulations in three-dimensional SKOV-3 spheroids was evaluated using confocal microscopy. The cytotoxicity and HK2 inhibition potential of solution form of 3-BPA was compared to the corresponding liposomal formulation by using cell proliferation and HK2 enzymatic assays. Results: SKOV-3 spheroids were reproducibly developed using the 96-well hanging drop method, with an average size of 900 μm by day 5. HK2 enzyme activity levels under hypoxic conditions were found to be higher than under normoxic conditions (P<0.0001, Student’s t-test, unpaired and two-tailed). Liposomal formulations (both non-targeted and targeted) of 3-BPA showed a more potent inhibitory effect (P<0.001, Student’s t-test, unpaired and two-tailed) at a dose of 50 μM than the aqueous solution form at 3, 6, and 24 hours post administration. Similarly, the cytotoxic activity 3-BPA at various concentrations (10 μM–100 μM) showed that the liposomal formulations had an enhanced cytotoxic effect of 2–5-fold (P<0.0001, Student’s t-test, unpaired and two-tailed) when compared to the aqueous solution form for both 10 μM and 25 μM concentrations. Conclusion: SKOV-3 spheroids developed by the hanging drop method can be used as a tumor aerobic glycolysis model for evaluation of therapies targeting the glycolytic pathway in cancer cells. Encapsulation of 3-BPA in a liposomal formulation improved permeability, HK2 inhibition, and cytotoxicity in the multicellular spheroid model.

Robust and Imperceptible Spread-Spectrum Watermarking for Telemedicine Applications

Abstract: This paper presents a new robust and secure digital watermarking scheme for its potential application in Telemedicine. The algorithm embeds medical text watermarks such as patient’s identity, image identity code and doctor’s identity into selected sub-band discrete wavelet transform (DWT) coefficients of the cover medical image using spread-spectrum technique. In the embedding process, the cover image is decomposed up to third level DWT coefficients. Three different text watermarks are embedded into the selected horizontal and vertical sub band DWT coefficients of the first, second and third level respectively. Selection of these coefficients for embedding purpose is based on threshold criteria defined in this paper. Robustness of the proposed watermarking scheme is further enhanced by applying error correcting code to the ASCII representation of the text watermark and the encoded text watermark is finally embedded into the cover medical image. It is found that the proposed scheme correctly extracts the embedded watermarks without error and provides high degree robustness against numerous known attacks while maintaining the imperceptibility of watermarked image.

Complex disruption effect of natural polyphenols on Bcl-2-Bax: molecular dynamics simulation and essential dynamics study

Abstract: Apoptosis (programmed cell death) is a process by which cells died after completing physiological function or after a severe genetic damage. Apoptosis is mainly regulated by the Bcl-2 family of proteins. Anti apoptotic protein Bcl-2 prevents the Bax activation/oligomerization to form heterodimer which is responsible for release of the cytochrome c from mitochondria to the cytosol in response to death signal. Quercetin and taxifolin (natural polyphenols) efficiently bound to hydrophobic groove of Bcl-2 and altered the structure by inducing conformational changes. Taxifolin was found more efficient when compared to quercetin in terms of interaction energy and collapse of hydrophobic groove. Taxifolin and quercetin were found to dissociate the Bcl-2-Bax complex during 12 ns MD simulation. The effect of taxifolin and quercetin was, further validated by the MD simulation of ligand-unbound Bcl-2-Bax which showed stability during the simulation. Obatoclax (an inhibitor of Bcl-2) had no significant dissociation e...

Image data-deduplication using the block truncation coding technique

Abstract: In this paper, we have CBIR, which is known as Content based Image Retrieval is the application of computer vision technique to the image retrieval problem from the large databases. In this paper de-duplication of electricity bills is done through CBIR. The technique for comparing the images is Block truncation coding; it is a lossy compression technique for grayscale image. So Photograph of electricity was divided into different block size using a compression technique known as Block Truncation coding. On the basis of block size, we will put images of the same block size in the same cluster. As there will be large block, so they will be divided into chunks of data. Through this experiment we will try to remove duplicates in the same cluster. In this paper the block of data is also divided into different chunks, so that image of single instance can be stored in these chunks. So duplication will be improved with the help of compression techniques discussed in detail.

Combinatorial-Designed Epidermal Growth Factor Receptor-Targeted Chitosan Nanoparticles for Encapsulation and Delivery of Lipid-Modified Platinum Derivatives in Wild-Type and Resistant Non-Small-Cell Lung Cancer Cells.

Abstract: Development of efficient and versatile drug delivery platforms to overcome the physical and biological challenges in cancer therapeutics is an area of great interest, and novel materials are actively sought for such applications. Recent strides in polymer science have led to a combinatorial approach for generating a library of materials with different functional identities that can be "mixed and matched" to attain desired characteristics of a delivery vector. We have applied the combinatorial design to chitosan (CS), where the polymer backbone has been modified with polyethylene glycol, epidermal growth factor receptor-binding peptide, and lipid derivatives of varying chain length to encapsulate hydrophobic drugs. Cisplatin, cis-([PtCl2(NH3)2]), is one of the most potent chemotherapy drugs broadly administered for cancer treatment. Cisplatin is a hydrophilic drug, and in order for it to be encapsulated in the developed nanosystems, it was modified with lipids of varying chain length. The library of four CS derivatives and six platinum derivatives was self-assembled in aqueous medium and evaluated for physicochemical characteristics and cytotoxic effects in platinum-sensitive and -resistant lung cancer cells. The results show that the lipid-modified platinate encapsulation into CS nanoparticles significantly improved cellular cytotoxicity of the drug. In this work, we have also reinforced the idea that CS is a multifaceted system that can be as successful in delivering small molecules as it has been as a nucleic acids carrier.

Analysis of the hierarchical structure of the B. subtilis transcriptional regulatory network

Abstract: The transcriptional regulation of gene expression is orchestrated by complex networks of interacting genes. Increasing evidence indicates that these ‘transcriptional regulatory networks’ (TRNs) in bacteria have an inherently hierarchical architecture, although the design principles and the specific advantages offered by this type of organization have not yet been fully elucidated. In this study, we focussed on the hierarchical structure of the TRN of the gram-positive bacterium Bacillus subtilis and performed a comparative analysis with the TRN of the gram-negative bacterium Escherichia coli. Using a graph–theoretic approach, we organized the transcription factors (TFs) and σ-factors in the TRNs of B. subtilis and E. coli into three hierarchical levels (Top, Middle and Bottom) and studied several structural and functional properties across them. In addition to many similarities, we found also specific differences, explaining the majority of them with variations in the distribution of σ-factors across the hierarchical levels in the two organisms. We then investigated the control of target metabolic genes by transcriptional regulators to characterize the differential regulation of three distinct metabolic subsystems (catabolism, anabolism and central energy metabolism). These results suggest that the hierarchical architecture that we observed in B. subtilis represents an effective organization of its TRN to achieve flexibility in response to a wide range of diverse stimuli.

Antigiardial activity of novel triazolyl-quinolone-based chalcone derivatives: when oxygen makes the difference.

Abstract: Giardiasis is a common diarrheal disease worldwide caused by the protozoan parasite Giardia (G.) intestinalis. It is urgent to develop novel drugs to treat giardiasis, due to increasing clinical resistance to the gold standard drug metronidazole (MTZ). New potential antiparasitic compounds are usually tested for their killing efficacy against G. intestinalis under anaerobic conditions, in which MTZ is maximally effective. On the other hand, though commonly regarded as an ‘anaerobic pathogen’, G. intestinalis is exposed to relatively high O2 levels in vivo, living attached to the mucosa of the proximal small intestine. It is thus important to test the effect of O2 when searching for novel potential antigiardial agents, as outlined in a previous study (Bahadur, Mastronicola et al. (2014) Antimicrob. Agents Chemother. 58, 543). Here, forty-five novel chalcone derivatives with triazolyl-quinolone scaffold were synthesized, purified and characterized by high resolution mass spectrometry, 1H and 13C nuclear magnetic resonance and infrared spectroscopy. Efficacy of the compounds against G. intestinalis trophozoites was tested under both anaerobic and microaerobic conditions, and selectivity was assessed in a counter-screen on human epithelial colorectal adenocarcinoma cells. MTZ was used as a positive control in the assays. All the tested compounds proved to be more effective against the parasite in the presence of O2, with the exception of MTZ that was less effective. Under anaerobiosis eighteen compounds were found to be as effective as MTZ or more (up to 3-4 fold); the same compounds proved to be up to > 100 fold more effective than MTZ under microaerobic conditions. Four of them represent potential candidates for the design of novel antigiardial drugs, being highly selective against Giardia trophozoites. This study further underlines the importance of taking O2 into account when testing novel potential antigiardial compounds.

Development and validation of different ultraviolet-spectrophotometric methods for the estimation of besifloxacin in different simulated body fluids

Abstract: In the present study a simple, accurate, precise, economical and specific UV-spectrophotometric method for estimation of besifloxacin in bulk and in different pharmaceutical formulation has been developed. The drug shows maximum λmax289 nm in distilled water, simulated tears and phosphate buffer saline. The linearity range of developed methods were in the range of 3-30 μg/ml of drug with a correlation coefficient (r(2)) 0.9992, 0.9989 and 0.9984 with respect to distilled water, simulated tears and phosphate buffer saline, respectively. Reproducibility by repeating methods as %RSD were found to be less than 2%. The limit of detection in different media was found to be 0.62, 0.72 and 0.88 μg/ml, respectively. The limit of quantification was found to be 1.88, 2.10, 2.60 μg/ml, respectively. The proposed method was validated statically according to International Conference on Harmonization guidelines with respect to specificity, linearity, range, accuracy, precision and robustness. The proposed methods of validation were found to be accurate and highly specific for the estimation of besifloxacin in different pharmaceutical formulations.

Solid forms of sofosbuvir

Abstract: The present disclosure provides crystalline sofosbuvir form-M1 and amorphous solid dispersion of sofosbuvir. The present disclosure also provides a process for the preparation of the amorphous form of sofosbuvir, the crystalline form of sofosbuvir, and amorphous solid dispersion of sofosbuvir and one or more pharmaceutically acceptable carriers.

Stercoral perforation of the sigmoid colon in a schizophrenic patient.

Abstract: Stercoral perforation of the colon is a life threatening condition, due to pressure necrosis by hard fecaloma which, are commonly found in chronic constipated patients. We report the case of a 45-year-old schizophrenic woman, under psychiatric treatment for two years, presenting with a 10-day history of absolute obstipation and distension. On physical examination the patient had signs of generalized peritonitis. Preoperative diagnostic workup gave no clear-cut clues about diagnosis. Emergency laparotomy revealed a single perforation over the antimesentric border of the sigmoid colon with hard fecal matter protruding through perforation margin. A segmental resection of the sigmoid colon with colonic lavage and end to end anastomosis was performed. The patient made an uneventful recovery. We have discussed the diagnostic work-up, and the management of this rare entity. Patient was on antipsychotic, anticholenergic and NSAID’S drugs for long time with longstanding immobilisation and poor oral intake. Association of these etiological factors with chronic constipation which ultimately leads to stercoral perforation have been corroborated.

Reconstructing height of an unknown point release using least-squares data assimilation

Abstract: Reckoning the height of a release in the source term estimation is important since a ground-level approximation of the release leads to errors in capturing the actual extent of a plume. A least-squares inversion technique, free from initial guess, is adapted here for the reconstruction of an elevated point release in a discretized space. Primarily, this involves estimation of the effective height of the release above the ground along with its location and strength from a limited set of noisy concentration measurements. The methodology is evaluated here with the nine runs from the Idaho diffusion experiment (1974) corresponding to low wind, stable conditions. Both real and model-generated synthetic data are used to test the method. With synthetic data, the methodology can exactly reproduce the input source terms. With real data, the average release height is estimated as 3.1 m, which is very close to the effective release height (3 m) reported in the Idaho data. The release location is retrieved with an average error of 30 m, whereas the minimum distance between source and detectors is 100 m; strength is retrieved within a factor of two in all the runs. The deviations in the source parameters from their prescribed values are explained in the context of model representativeness, wind variability and available monitoring network. The sensitivity of the source term estimation is evaluated against several parameters: (i) receptors' height either neglected or duly taken as 0.76 m, (ii) measurement noises and (iii) number of receptors utilized in the inversion. In addition, the limitations and meteorological issues related to the inversion of an elevated release are highlighted.

Processes for the preparation of amorphous tenofovir alafenamide hemifumarate and a premix thereof

Abstract: An amorphous form of tenofovir alafenamide hemifumarate and process for the 5 preparation of the same. A premix of amorphous tenofovir alafenamide hemifumarate with pharmaceutically acceptable excipients and process for the preparation of the same are also disclosed.

Complex of amorphous empagliflozin and a cyclodextrin

Abstract: The present disclosure relates to solid dispersion of a complex of amorphous empagliflozin and a cyclodextrin.