Current Drug Discovery Technologies - Volume 7, Issue 2, 2010

The nightmare of multi-drug resistant bacteria will still haunt if no panacea is ever found. Efforts on seeking desirable natural products with bactericidal property and screening chemically modified derivatives of traditional antibiotics have lagged behind the emergence of new multi-drug resistant bacteria. The concept of using antisense antibiotics, now as revolutionary as is on threshold has experienced ups and downs in the past decade. In the past five years, however, significant technology advances in the fields of microbial genomics, structural modification of oligonucleotides and efficient delivery system have led to fundamental progress in the research and in vivo application of this paradigm. The wealthy information provided in the microbial genomics era has allowed the identification and/or validation of a number of essential genes that may serve as possible targets for antisense inhibition; antisense oligodeoxynucleotides (ODNs) based on the 3rd generation of modified structures, e.g., peptide nucleic acids (PNAs) and phosphorodiamidate morpholino oligomers (PMOs) have shown great potency in gene expression inhibition in a sequence-specific and dosedependent manner at low micromolar concentrations; and cell penetrating peptide mediated delivery system has enabled the effective display of intracellular antisense inhibition of targeted genes both in vitro and in vivo. The new methods show promise in the discovery of novel gene-specific antisense antibiotics that will be useful in the future battle against drug-resistant bacterial infections. This review describes this promising paradigm, the targets that have been identified and the recent technologies on which it is delivered.

The rampant use of antibiotics in the last half-century has imposed an unforeseen biological cost, the unprecedented acceleration of bacterial evolution to produce drug-resistant strains to practically every approved antibiotic. This rise in antimicrobial drug resistance, alongside the failure of conventional research efforts to discover new antibiotics, may eventually lead to a public health crisis that can drastically curtail our ability to combat infectious disease. To address this public health need for novel countermeasure strategies, research efforts have recently focused on identification of genes in the host, rather than the pathogen, that are essential for successful pathogen infection, as potential targets for drug discovery. In the past decade, RNA interference (RNAi) has emerged as a powerful tool for analyzing gene function by silencing target genes through the specific destruction of their mRNAs. Based on RNAi methodology, high-throughput genome- wide assay platforms have been developed to identify candidate host genes that are manipulated by pathogens during infection. In this review, we will discuss recent strategies for RNAi-based genomic screens to investigate hostpathogen mechanisms in human cell models using both bacterial pathogens, including Salmonella typhimurium, Mycobacterium tuberculosis, and Listeria monocytogenes, and viruses, such as Human Immunodeficiency Virus (HIV), Hepatitis C Virus (HCV) and influenza. These functional genomics studies have begun to elucidate novel pathogen virulence mechanisms and thus, may serve as the basis for the design of novel host-based inhibitor therapeutics that can block or alleviate the downstream effects of pathogen infection.

Over the past several years, microRNAs (miRNAs) have been identified as a fine-tuner in a wide array of biological processes, including development, cell growth and metabolism. Recent studies have shown that many kinds of miRNAs act as oncomirs or tumor suppressors in tumors where the miRNA genes are up- or down- regulated, respectively. These dysregulations occur through a variety of mechanisms, such as genetic alterations, epigenetic repression or altered expression of transcription factors which target miRNAs. The aberrant expressions of miRNAs are observed not only in tumor lesions but also in plasma and serum of cancer patients. These characteristics of miRNAs have created extensive interest in tapping into them for diagnosis and prognosis as well as drug discovery in cancer therapy. In this literature, the significance of miRNAs in tumor initiation and development is first reviewed. Second topic is extracellular miRNAs as biomarkers for cancer classification and prediction. Further, we focus on secretory machinery of miRNAs and share new evidence suggesting that extracellular miRNAs can play biological roles beyond mere biomarkers. Extending this concept, our hypothetical model that extracellular miRNAs may function as a signaling molecule in a crosstalk between cancer cells and their surrounding cells is presented. Finally, we discuss the potential of miRNAs for therapeutic applications in clinical oncology.

The main objective of this study was to compare the behaviour of drug release among the famotidine polymorphs prepared by using various additives and solvents, by solvent evaporation method. The famotidine polyvinyl pyrrolidone polymorphs with different concentrations (0.5, 1 and 1.5%) were prepared by using solvent evaporation method. In these polymorphs of different concentrations 1% w/v polymorphs showed better release. Similarly, famotidine polymorphs of Tween 80 with different concentrations, polyethylene glycol 1% w/v and methanol was prepared. Famotidine polymorphs prepared the PVP (1% w/v) showed better drug release and solubility. DSC, FTIR, SEM and XRD studies were carried out. DSC studies revealed that PVP polymorphs were found to stable compared to other polymorphs. FTIR studies of the polymorphs prepared indicated that there was an interaction found in all polymorphs except PVP polymorphs indicating the absence of drug-additive interaction. SEM studies of PVP and methanol polymorphs revealed that they are tabular and prismatic and columnar respectively. These changes in morphology were due to variations in face dimensions and also properties of additives and solvent used in the preparation. XRD studies revealed that there is an increase in crystallinity in methanol polymorphs when compared to PVP polymorphs and pure drug. The mechanism of drug release was determined using zero order, first order and Hixon-Crowel equations. From the drug release kinetics these polymorphs followed first order and Hixon-Crowel release kinetics, exhibited fair linearity in their dissolution data. Further, in vivo studies were carried out for the evaluation of antiulcer activity. Based upon the drug release pattern and its kinetics only two of the prepared polymorphs of famotidine i.e. famotidine PVP polymorphs and famotidine methanol polymorphs were selected for animal studies. Antiulcer studies were carried out using pylorus ligation model and estimation of antioxidant parameters was also done. In these studies also polymorphs of PVP showed better antiulcer activity and also significant antioxidant activity when compared to famotidine (pure) and famotidine methanol polymorphs. Hence in the present investigation, amongst the various polymorphs of famotidine prepared, PVP polymorphs were found to possess good dissolution behaviour, stability and absence of drug additive interactions. Further, in vivo studies confirmed the better therapeutic action of these PVP polymorphs over the pure drug and famotidine methanol polymorphs.

Groundnut bud necrosis virus (GBNV) is recognized as one of the most economically important viruses and is known to affect several crops including peanut, potato, tomato and soybean. For managing plant virus diseases, determination of their causal agents’ identity at an early stage of crop is a pre-requisite. In the present study, NSm protein of GBNV has been used to predict out MHC binding peptides and epitopes that are highly suitable for antigenicity. Eighteen peptide regions were found to have high affinity. Few of these NSm protein TAP transporters are 126- RRYMHISRL with score 11.638, 125- NRRYMHISR with score 10.280, 46- AIMNKAKTL with score 7.762, 120- PTWNSNRRY with score 7.632 and 171- ASLKDPMCF with score 7.277. The support vector machine (SVM) based approach predicted MHCII-IAb peptide regions, 45- SAIMNKAKT, 151- ASLIDPNKM, 23- PAVKKENNR, 229- PIAAENNTC, (optimal score 0.938); MHCII-IAd peptide regions, 208- YAKGVGFAS, 101- NDSLVGNGN, 55- NGKQYVSSG, 63-GDSSVLGTY, (optimal score 0.852); MHCII-IAg7 peptide regions, 277- LQKAAERLA, 145- SKNNVKASL, 228-TPIAAENNT, 276- SLQKAAERL, (optimal score 1.640); and MHCII- RT1.B peptide regions, 193- TPKQCMQLN, 195- KQCMQLNLT, 246- KVIQSAALI, 166- IISRQASLK, (optimal score 0.800) as binders from NSm protein. Themost suitable predicted segments in NSm protein of GBNV virus found in the study are 164 KIIISRQASLKDPMCFIFHLNWS-186 and 237-CDVVPINRAKVIQSAALIEACKLMIP-262. These two fragments, obtained from non-structural movement protein with average propensity 1.016, are high-efficiency binders and may, thereforebe used in cross protection to provide resistance against GBNV and develop GBNV specific antibodies that can be exploited in sero-diagnostics.

The complementary mixing and de-mixing of phases is seen as a methodology for use in ‘smart’ drug delivery systems. Surface tensions were routinely lowered to 50-55 mN/m in drug mixtures and this is compared to low molecular weight emulsifiers such as Tween 80. In the research reported here, mixing of a heterocyclic drug and an amphiphilic polymer is responsible for structural inconsistency at the air-water (A/W) interface and presents itself as a means of release of drug from a series of complex drug formulations. The work was extended to look at the behavior of both “soft” and “hard” particle hydrophobic and hydrophilic particle stabilization of emulsions of 20 and 0.7 μm. The work is based on spread interfacial monolayers, thin liquid films emulsion droplets and model latex nanoparticle beads (60 and 500 nm) in the presence and absence of poloxamer (Pluronic) and methotrexate. The interfacial rheological characteristics and AFM-based nano-rheology were used to aid the elucidation of surface structure and nano-architectures within the plane of the adsorbed interfacial layer and for use in the fabrication of solid nano-particle stabilized micron and nano-sized oil-inwater (O/W) emulsions.

The protease, human kallikrein-related peptidase 6 (hK6) is derived from activated macrophages in the central nervous system (CNS) and may contribute to pathology observed in multiple sclerosis (MS). In the present study, we compared serum and cerebrospinal fluid (CSF) protein concentrations of human kallikrein-related peptidase 6 derived from neurological controls and patients diagnosed with advanced multiple sclerotic disease. Mean serum levels of human kallikrein-related peptidase 6 were similar in neurological controls and patients diagnosed with relapsing-remitting (RR), secondary progressive (SP) and primary progressive (PP) multiple sclerosis with mean levels ranging from 3.5 to 3.75 ng/ml. Patients diagnosed with advanced multiple sclerosis showed mean CSF levels (29 ng/ml) that were significantly higher than neurological controls (25.5 ng/ml). Determining CSF concentrations of human kallikrein-related peptidase 6 may therefore have diagnostic value in MS.