Understanding the Multi-Functional Role of TCTP in the Regeneration Process of Earthworm, Perionyx excavatus.

BACKGROUND: Regeneration is a highly complex process that requires the coordination of numerous molecular events, and identifying the key ruler that governs is important to investigate. While it has been shown that TCTP is a multi-functional protein that regulates cell proliferation, differentiation, apoptosis, anti-apoptosis, stem cell maintenance, and immune responses, but only a few studies associated to regeneration have been reported. To investigate the multi-functional role of TCTP in regeneration, the earthworm Perionyx excavatus was chosen. METHODS: Through pharmacological suppression of TCTP, amputation, histology, molecular docking, and western blotting, the multi-function role of TCTP involved in regeneration is revealed. RESULTS: Amputational studies show that P. excavatus is a clitellum-independent regenerating earthworm resulting in two functional worms upon amputation. Arresting cell cycle at the G1/S boundary using 2 mM Thymidine confirms that P. excavatus execute both epimorphosis and morphallaxis regeneration mode. The pharmacological suppression of TCTP using buclizine results in regeneration suppression. Following the combinatorial injection of 2 mM Thymidine and buclizine, the earthworm regeneration is completely blocked, which suggests a critical functional role of TCTP in morphallaxis. The pharmacological inhibition of TCTP also suppresses the key proteins involved in regeneration: Wnt3a (stem cell marker), PCNA (cell proliferation) and YAP1 (Hippo signalling) but augments the expression of cellular stress protein p53. CONCLUSION: The collective results indicate that TCTP synchronously is involved in the process of stem cell activation, cell proliferation, morphallaxis, and organ development in the regeneration event.

Buclizine crystal forms: First Structural Determinations, counter-ion stoichiometry, hydration, and physicochemical properties of pharmaceutical relevance.

Buclizine (BCZ) is a chiral synthetic piperazine derivative which has antihistaminic, anti-muscarinic and antiemetic properties, and has been reintroduced as an appetite stimulant, especially for pediatric patients. Structural information about this drug, as well as other buclizine crystalline forms (solvates, salts and co-crystals) including the BCZ free-base (BCZ-FB), is non-existent. Here, we present for the first time the crystal structure of the monohydrochloride monohydrate salt of BCZ (BCZHCl·H2O), and of its anhydrous form, BCZHCl. Interestingly, BCZHCl·H2O was obtained by recrystallization from the raw material (BCZH2Cl2) in ethanol:water solution showing that BCZ anhydrous dihydrochloride salt changes easily to a monohydrochloride monohydrate salt modification, which raise concerns about formulation quality control. BCZHCl·H2O and BCZHCl crystallize in the orthorhombic space groups (Pna21 and Pca21) belonging to the mm2 point group and are thus classified as non-centrosymmetric achiral structures (NA). Intuitively, we expect these salts to crystallize in a space group with a center of symmetry, since less than 5% of the known racemic compounds crystallize in the NA type. The crystal structures of BCZH2Cl2 and BCZ-FB were not determined, but their existence was verified by other techniques (chloride ion analysis, PXRD, HPLC, FT-IR, DSC, TGA) and by comparison of the obtained results with those found for BCZHCl. Additionally, we have also performed an evaluation of the equilibrium solubility (at six different aqueous media) and the dissolution profile of the BCZHCl salt compared to the raw material and BCZ-FB. Different equilibrium solubility values were found comparing the three forms in acidic and neutral pH ranges and all of them were insoluble at pH > 7.0. Moreover, tablets prepared with BCZH2Cl2, BCZHCl or BCZ-FB show significant differences in terms of dissolution profile.

Drug Repurposing Approach for Developing Novel Therapy Against Mupirocin-Resistant Staphylococcus aureus.

Multidrug resistance (MDR) is a major health issue for the treatment of infectious diseases throughout the world. Staphylococcus aureus (S. aureus) is a Gram-positive bacteria, responsible for various local and systemic infections in humans. The continuous and abrupt use of antibiotics against bacteria such as S. aureus results in the development of resistant strains. Presently, mupirocin (MUP) is the drug of choice against S. aureus and MDR (methicillin-resistant). However, S. aureus has acquired resistance against MUP as well due to isoleucyl-tRNA synthetase (IleS) mutation at sites 588 and 631. Thus, the aim of the present study was to discover novel bioactives against MUP-resistant S. aureus using in silico drug repurposing approaches. In silico drug repurposing techniques were used to obtain suitable bioactive lead molecules such as buclizine, tasosartan, emetine, medrysone, and so on. These lead molecules might be able to resolve this issue. These leads were obtained through molecular docking simulation based virtual screening, which could be promising for the treatment of MUP-resistant S. aureus. The findings of the present work need to be validated further through in vitro and in vivo studies for their clinical application.

Interaction of antihistaminic drugs with human translationally controlled tumor protein (TCTP) as novel approach for differentiation therapy.

Translationally controlled tumor protein (TCTP) represents an exquisite target for cancer differentiation therapy, because it was most strikingly down-regulated in tumor reversion experiments. Since TCTP is identical with the histamine releasing factor, antihistamic drugs may inhibit TCTP. Indeed, antihistaminics, such as promethazine, thioridazine, perphemazine and chlorpromazine reveal antiproliferative effects. The aim of this investigation was to study antihistaminic drugs as new TCTP inhibitors to inhibit tumor growth. Levomepromazine and buclizine showed higher in silico binding affinities to TCTP among 12 different antihistaminic compounds including the control drugs, promethazine and hydroxyzine by using Autodock4 and AutodockTools-1.5.7.rc1. Recombinant human TCTP was codon-optimized, expressed in E. coli and purified by chitin affinity chromatography. For experimental validation of in silico data, we applied microscale thermophoresis. Levomepromazine bound with a Kd of 57.2 µM (p < 0.01) and buclizine with a Kd of 433µM (p < 0.01) to recombinant TCTP. Both drugs inhibited MCF-7 breast cancer cell growth in resazurin assays. TCTP expression was down-regulated after treatment with the two drugs. Cell cycle was arrested in the G1 phase without apoptosis as confirmed by the expression of cell cycle and apoptosis-regulating proteins. Annexin V-PI staining and Trypan blue exclusion assay supported that the two drugs are cytostatic rather than cytotoxic. Induction of differentiation with two drugs was detected by the increased appearance of lipid droplets. In conclusion, levomepromazine and buclizine inhibited cancer cell growth by binding to TCTP and induction of cell differentiation. These compounds may serve as lead compounds for cancer differentiation therapy.

Caracteristica da buclizina e determinação por titulação espectrofotométrica em produtos farmacêuticos / Characteristics of buclizine and determination by spectrophotometric titration in pharmaceutical products

Descrevem-se neste artigo algumas características do dicloridrato de buclizina, um tranquilizante e anti-histamínico usado em, composição farmacêuticas. Curvas de titulação e espectros de absorção da substância pura são apresentados. Descreva-se, tambem, uma técnica para determinação da substância, quando livre, por simples titulação em meio não aquoso, com ácido perclórico e cristal violeta como indicador, e técnica para determinação por titulação espectrofotométrica, com ou sem purificação prévia por lavagem com éter dietílico, e por cromtografia em camada delgada e com leitura espectrifotométrica a 239nm, em solução clorofórmica, ou 274 nm, em bezeno (AU).