Fabrication, characterization and evaluation of a new designed botulinum toxin-cell penetrating peptide nanoparticulate complex.

BACKGROUND: To have a better and longer effect, botulinum neurotoxin (BoNT) is injected several times in a treatment course, which could increase side effects and cost. Some of the most cutting-edge strategies being investigated for proteins to their physiologic targets involve the reformulation of BoNT based on peptide-based delivery systems. For this purpose, cell-penetrating peptides (CPPs) are of particular interest because of their capacity to cross the biological membranes. OBJECTIVES: A short and simple CPP sequence was used as a carrier to create nanocomplex particles from BoNT/A, with the purpose of increasing toxin entrapment by target cells, reducing diffusion, and increasing the duration of the effect. METHOD: CPP-BoNT/A nanocomplexes were formed by polyelectrolyte complex (PEC) method, considering the anionic structure of botulinum toxin and the cationic CPP sequence. The cellular toxicity, and absorption profile of the complex nanoparticles were evaluated, and the digit abduction score (DAS) was used to assess the local muscle weakening efficacy of BoNT/A and CPP-BoNT/A. RESULTS: The provided optimized polyelectrolyte complex nanoparticles had a 244 ± 20 nm particle size and 0.28 ± 0.04 PdI. In cellular toxicity, CPP-BoNT/A nanocomplexes as extended-release formulations of BoNT/A showed that nanocomplexes had a more toxic effect than BoNT/A. Furthermore, the comparison of weakening effectiveness on muscle was done among nanoparticles and free toxin on mice based on the digit abduction score (DAS) method, and nanocomplexes had a slower onset effect and a longer duration of action than toxin. CONCLUSION: Using PEC method allowed us to form nanocomplex from proteins, and peptides without a covalent bond and harsh conditions. The muscle-weakening effect of toxin in CPP-BoNT/A nanocomplexes showed acceptable efficacy and extended-release pattern.

Engineered PLGA microspheres for extended-release of naltrexone: in vitro, in vivo, and IVIVR.

Poly(lactide-co-glycolide) (PLGA)-based formulation is one of the most often used parenteral extended-release forms to deliver various therapeutics. VIVITROL® as a commercialized PLGA microsphere formulation encapsulates naltrexone, a narcotic antagonist for opioid addiction and alcohol dependency. However, no U.S. Food and Drug Administration-approved generic product of naltrexone PLGA microsphere formulation has entered the market. The availability of generic naltrexone PLGA microspheres in low-income countries will broaden patients' accessibility to the safe, effective, and more affordable drug. A major challenge in developing such generic forms is the sensitivity of the drug-loaded microspheres' critical characteristics to the small manufacturing changes, even in formulations with the same compositions as the reference product. In this study, we evaluated the different key manufacturing parameters on the physicochemical, in vitro and in vivo release characteristics of naltrexone microspheres to develop a generic form of naltrexone PLGA microspheres. The selected formulations demonstrated a significant similarity in physicochemical characteristics and release profiles (f2 > 50) to the reference product, VIVITROL®. A strong relationship was observed between in vitro release profile of naltrexone as against its corresponding in vivo profile. It helped to roughly predict the in vivo release behavior of the different manufactured formulations by their corresponding in vitro release profiles.

The effects of a combination oral spray (Mucosamin®) for the prevention of oral mucositis in pediatric patients undergoing hematopoietic stem cell transplantation: a double blind randomized clinical trial.

PURPOSE: Oral mucositis (OM) is a frequent complication of conditioning regimens for hematopoietic stem cell transplantation (HSCT). Damage to the nuclear and non-nuclear materials of the mucosal cells by the production of Reactive Oxygen Species (ROS) and proinflammatory cytokines could result to development and progression of OM. Previous studies have shown the effectiveness of Mucosamin® oral spray in the management of pain and acceleration of OM healing. The aims of the current study were to evaluate prophylactic effects of Mucosamin® oral spray in reducing the incidence and severity of OM in pediatric patients undergoing allogeneic HSCT. METHOD: The current study was designed as a double-blind, placebo-controlled randomized clinical trial. Sixty patients were enrolled in the study and received placebo or Mucosamin® spray. Patients in both groups used sprays 4 times daily. Product application was begun at the time of initiation of conditioning regimen and was continued for 14 days. RESULTS: Mucosamin® significantly reduced incidence and severity of OM compared to the placebo (P values: 0.027 and 0.035, respectively). This product could also decrease OM duration and delay OM onset (P values: 0.007 and 0.006, respectively). CONCLUSION: Mucosamin® could effectively reduce incidence, severity, and duration of OM and delay OM onset in pediatric patients undergoing allogeneic HSCT. TRIAL REGISTRATION: The study protocol was registered in the Iranian Registry of Clinical Trials under the registry number IRCT20190917044805N1.

Glutamate-urea-based PSMA-targeted PLGA nanoparticles for prostate cancer delivery of docetaxel.

Targeted drug delivery is a tool to make treatment more specific, selective, and effective and to prevent unwanted complications. Prostate specific membrane antigen (PSMA) is a useful biomarker in order to monitor and control prostate cancer. Glutamate-Urea-R (Glu-Urea-R) is a PSMA enzyme inhibitor capable of binding to this surface marker of prostate cancer cell in an efficient and special manner. The aim of this project was to develop a docetaxel-loaded nanoparticle of poly (lactic-co-glycolic acid) polyethylene glycol which is cojugated to a urea-based anti-PSMA ligand named glutamate-urea-lysine (glu-urea-lys) for targeted delivery of docetaxel in prostate cancer. The obtained nanoparticles, prepared by nanoprecipitation method, were spheres with a particle size of around 150 nm and zeta potential of -7.08 mV. Uptake studies on the PC3 (as PSMA negative) and LNCaP (as PSMA positive) cells demonstrated that drug uptake was efficient by the PSMA positive cells. IC50 of targeted NPs on LNCaP cell line compared to non-targeted ones was reduced by more than 70% in three different incubation times of 24, 48, and 72 h. In conclusion, the nanoparticles are expected to specifically transport docetaxel to PSMA-positive prostate cancer cells and consequently, enhance the antitumor efficacy of docetaxel on these cells.

Does Pharmacodynamics of Drugs Change After Presenting them as Nanoparticles Like their Pharmacokinetics?

Nowadays, the breakthrough in different medical branches makes it feasible to designate new methods of drug delivery to achieve the most cost-effective and the least unpleasant consequenceimposing solutions to overcome a wide range of diseases. Nanoparticle (NP) drugs entered the therapeutic system, especially in cancer chemotherapy. These drugs are quite well-known for two traits of being long-acting and less toxic. For a long time, it has been investigated how NPs will change the kinetics of drugs. However, there are a few studies that inclined their attention to how NPs affect the dynamics of drugs. In this review, the latter point will mainly be discussed in an example-based manner. Besides, other particular features of NPs will be briefly noted. NPs are capable of affecting the biologic system as much as a drug. Moreover, NPs could arise a wide variety of effects by triggering their own receptors. NPs are able to change a receptor function and manipulate its downstream signaling cascade.

Preparation and Characterization of Albumin Nanoparticles of Paclitaxel-Triphenylphosphonium Conjugates: New Approach to Subcellular Targeting.

Mitochondria have been recognized as important targets in cancer therapy due to their role in the respiratory process of cells. One approach employed for mitochondrion targeting is conjugation of a delocalized cation such as triphenylphosphonium (TPP), with antineoplastic agents, for instance paclitaxel (PTX). In cell cytoplasm, TPP-PTX can come close to mitochondria due to its high positive charge, which has a strong tendency toward the enhanced negative charge of mitochondria. The esteric bond of TPP-PTX can break down in the acidic environment of tumor cells and release the PTX, which can act directly on mitochondria to kill tumor cells. TPP-PTX was synthesized in three steps: Succinic anhydride (SUC) reacted with PTX to achieve succinyl paclitaxel (SUC-PTX), which has an acid-labile esteric bond. Then 2-triphenylphosphonium ethylammonium (ATPP) was prepared by attaching 2-bromoethylammunium bromide to TPP. Finally, a TPP-PTX prodrug was synthesized by attaching these materials. The products of all steps were characterized by thin-layer chromatography (TLC), infrared spectroscopy (IR), and nuclear magnetic resonance (1H NMR, 13C NMR). The purity of the products was determined by HPLC methods. TPP-PTX, as a prodrug, was loaded in to human serum albumin (HSA) nanoparticles by a method inspired by nab-technology with 130-160 nm particle size distribution, PdI=0.166 and Zeta potential -12.6 mV.

Synergistic effect of microRNA and albumin-bound nanoparticles for inhibition of glioblastoma cancer cell proliferation

The functional significance of upregulation miR-34a in combination with albumin-bound paclitaxel nanoparticles in U251 glioblastoma cell line has been evaluated. The MTT assay determined that miR-34a and albumin-bound paclitaxel nanoparticles can reduce cell viability, but the combination of both factors has a stronger effect on cell viability. The application of qRT-PCR has demonstrated that the transduction of miR-34a could lead to exogenous upregulation of miR-34a level and downregulation of SURVIVIN. Moreover, treatment of U251 cells with miR-34a and nanoparticles together considerably inhibit SURVIVIN expression compared to miR-34a and nanoparticles alone. Flow cytometry showed that upon miR-34a overexpression cell cycle arrested in G1 phase, while treatment with nanoparticles increased the cell population in G2 phase. Upregulation of miR-34a along with treatment with nanoparticles elevated the number of cells arrested in G1/ G2 phases of the cell cycle. Expression of miR-34a with albumin-bound paclitaxel nanoparticles reduced cell viability, downregulated SURVIVIN and enhanced cell cycle arrest in G1/G2 phases. Thus, the upregulation of miR-34a with these nanoparticles are potential candidates therapeutic for glioblastoma cancer.

Preparation and characterization of self nano-emulsifying drug delivery system (SNEDDS) for oral delivery of heparin using hydrophobic complexation by cationic polymer of ß-cyclodextrin.

OBJECTIVE: The aim of this study was the preparation of a self nano-emulsifying drug delivery system (SNEDDS) for oral delivery of heparin. SIGNIFICANCE: Preparation of hydrophobic complexes between heparin as the hydrophilic macromolecule and cationic polymer of ß-cyclodextrin (CPßCD) was considered for preparation of orally administered SNEDDS in which the drug incorporated in internal oil phase of O/W nano-droplets. METHODS: Hydrophobic complexes of heparin-CPßCD were prepared by electrostatic interaction. The lipophilic feature of complexes was characterized by determining their partition co-efficients. SNEDDS prototypes were prepared by mixing liquid paraffin, Tween 80, propylene glycol and ethanol, diluted 1:100 in an aqueous medium. Central composite response surface methodology was applied for statistical optimization. Independent variables were the amount of liquid paraffin and the amount of Tween 80, while responses were size and poly dispersity index (PdI). Optimized SNEDDS were studied morphologically using transmission electron microscopy (TEM). In vitro release of heparin was studied in the simulated gastric and simulated intestinal media. RESULTS: The data revealed that in molar ratio 1:3 (heparin:CPßCD), the n-octanol recovery was maximized and reached 67.6 ± 11.86%. Size, PdI, zeta potential, EE% in gastric medium and EE% in intestinal medium for optimized nano-droplets were reported as 307 ± 30.51 nm, 0.236 ± 0.02, +2.1 ± 0.66 mV, 90.2 ± 0.04 and 96.1 ± 0.73%, respectively. Microscopic images revealed spherical nano-droplets. The obtained data revealed no burst release of heparin from nano-droplets. CONCLUSIONS: The obtained results indicate that SNEDDS could be regarded as a good candidate for oral delivery of heparin as the hydrophilic macromolecule.

Fabrication and biological evaluation of chitosan coated hyaluronic acid-docetaxel conjugate nanoparticles in CD44(+) cancer cells.

BACKGROUND: Hyaluronic acid (HA) has been used for target-specific drug delivery because of strong affinity to CD44, a marker in which overexpressed in cancer cells and cancer stem cells. Conjugation of HA to the cytotoxic agents via active targeting can improve efficacy, biodistribution, and water solubility. To be able to benefit from passive targeting as well, a nanoparticulate system by counter ion using a polycation like chitosan may lead to a perfect delivery system. METHODS: Water soluble Hyaluronic acid-Docetaxel (HA-DTX) conjugate was prepared and used to formulate chitosan-coated HA-DTX nanoparticles by polyelectrolyte complex (PEC) method and optimized using Box-Behnken design. Biological evaluation of nanoparticles was done in CD44+ cancer cells. RESULTS AND DISCUSSION: Biological evaluation of optimized formula showed IC50 of nanoparticles for 4 T1 and MCF-7 cell lines were 45.34 µM and 354.25 µM against 233.8 µM and 625.9 µM for DTX, respectively with increased cellular uptake showed by inverted confocal microscope. CONCLUSION: Chitosan-coated HA-DTX nanoparticles were more effective against CD44+ cells than free DTX. Chitosan coated hyaluronic acid-docetaxel conjugate nanoparticles fabricated and evaluated in CD44+ cancer cells.

Biowaiver Monographs for Immediate Release Solid Oral Dosage Forms: Ribavirin.

Literature data relevant to the decision to allow a waiver of in vivo bioequivalence (BE) testing for the approval of immediate release solid oral dosage forms containing ribavirin are reviewed. Ribavirin is highly soluble, but its permeability characteristics are not well defined. Therefore according to the Biopharmaceutical Classification System, and taking a "worst case" approach, ribavirin should be assigned to class III. As ribavirin is transported across the brush border membrane of the human jejunum by hCNT2, it shows saturable uptake in the intestine. However, no common excipients have been shown to compete for ribavirin absorption, nor have problems with BE of immediate release ribavirin formulations containing different excipients and produced by different manufacturing methods been reported in the open literature. So the risk of bioinequivalence caused by these factors appears to be low. Ribavirin is considered a narrow therapeutic index drug, as judged by comparing the minimum effective concentration and minimum toxic concentrations in blood. Although ribavirin would not be eligible for approval via a Biopharmaceutical Classification System-based biowaiver procedure according to today's guidances due to its narrow therapeutic index, the risks of biowaiving should be weighed against the considerable risks associated with studying BE of ribavirin products in healthy subjects.

Biotin/Folate-decorated Human Serum Albumin Nanoparticles of Docetaxel: Comparison of Chemically Conjugated Nanostructures and Physically Loaded Nanoparticles for Targeting of Breast Cancer.

Docetaxel (DTX) is a widely used chemotherapeutic agent with very low water solubility. Conjugation of DTX to human serum albumin (HSA) is an effective way to increase its water solubility. Attachment of folic acid (FA) or biotin as targeting moieties to DTX-HSA conjugates may lead to active targeting and specific uptake by cancer cells with overexpressed FA or biotin receptors. In this study, FA or biotin molecules were attached to DTX-HSA conjugates by two different methods. In one method, FA or biotin molecules were attached to remaining NH2 residues of HSA in DTX-HSA conjugate by covalent bonds. In the second method, HSA-FA or HSA-biotin conjugates were synthesized separately and then combined by DTX-HSA conjugate in proper ratio to prepare nanoparticles containing DTX-HSA plus HSA-FA or HSA-biotin. Cell viability of different nanoparticle was evaluated on MDA-MB-231 (folate receptor positive), A549 (folate receptor negative), and 4T1 (biotin receptor positive) and showed superior cytotoxicity compared with free docetaxel (Taxotere). In vivo studies of DTX-HSA-FA and DTX-HSA-biotin conjugates in BULB/c mice, tumorized by 4T1 cell line, showed the conjugates prepared in this study were more powerful in the reduction in tumor size and increasing the survival rate when compared to free docetaxel.

cis-Dichlorodiamminoplatinum (II) glyconanoparticles by drug-induced ionic gelation technique targeted to prostate cancer: preparation, optimization and in vitro characterization.

BACKGROUND: Cancer stem cells (CSC) have been proposed as the reason of cancer relapse which are characterized mainly based on CD44+ phenotype with other supplementary markers. The aim of the present study is to fabricate cis-dichlorodiamminoplatinum (II) (CDDP) loaded glyconanoparticles using hyaluronic acid (HA) which is also known as the endogenous substrate for CD44 in vivo. METHODS: For this purpose, a drug-induced ionic gelation technique has been used to prepare CDDP-incorporated nanoparticles. To optimize the fabrication technique, stirring rate, stirring time, and HA/CDDP ratio have been selected as the main factors from other factors and subjected to face-centered central composite design for optimization purposes. The optimized nanoparticles were further characterized using different complementary methods including FTIR, SEM, AFM and DSC. To evaluate the biological effectiveness of CDDP nanoparticles release study, MTS assay, tumor cell clonogenicity and sphere formation assay have been performed as well. RESULTS: Spherical CDDP nanoparticles with Z-average approx. 150nm with low PdI were prepared by adjusting the selected variables. FTIR results indicated the presence of inclusion complexes between CDDP and HA which lead to preparing nanoparticles with high entrapment efficiency and drug content of 87.4 and 43.74 percentage respectively. In vitro release study showed a sustained release of CDDP up to 4 days, and cellular studies confirmed that nanoparticles formation keeps the anticancer activity of formulated CDDP while moderate increase in cancer stem cell suppression. CONCLUSION: It seems hyaluronic acid could be successfully exploited as carrier in cancer-targeted drug delivery with a look at targeting the CSCs.

Compounding practices in Iran.

This article has been presented by pharmacists of the Razi Hospital Pharmacy, which is located within the Dermatology Center of Tehran University of Medical Sciences in Tehran, Iran, and by members of the staff at the Tehran University of Medical Sciences. Discussed within this article are the legalization and regulation of pharmaceutical compounding in Iran, the restrictions on pharmaceutical compounding, the general equipment used in Iran pharmacies, beyond-use dating/expiration dating, the required pharmacy education, as well as information related specifically to Razi Hospital Pharmacy.

CD44-targeted docetaxel conjugate for cancer cells and cancer stem-like cells: a novel hyaluronic acid-based drug delivery system.

A CD44-targeted macromolecular conjugate of docetaxel was prepared via a pH-sensitive linkage to hyaluronic acid and was characterized using NMR, gel permeation chromatography, and differential scanning calorimetry. The conjugated species were further evaluated in terms of drug release, cytotoxicity, cellular uptake, cell cycle inhibition, and subacute toxicity in mice. Cellular microscopic studies revealed that CD44-expressing cells including MCF-7 cancer stem cells and MDA-MB-231 metastatic breast cancer cells had internalized the conjugates via a selective receptor-mediated mechanism, leading to cell cycle arrest in the G2/M phase. Hyaluronic acid-docetaxel conjugates showed specific toxicity only in CD44-expressing cells in vitro, along with a decreased risk of neutropenia and dose-dependent mortality in vivo. Hyaluronic acid-drug conjugates represent a promising and efficient platform for solubilization of sparingly soluble molecules as well as active and selective targeted delivery to cancer cells and cancer stem cells.

Coumarin compounds of Biebersteinia multifida roots show potential anxiolytic effects in mice.

BACKGROUND: Traditional preparations of the root of Biebersteinia multifida DC (Geraniaceae), a native medicinal plant of Irano-Turanian floristic region, have been used for the treatment of phobias as anxiolytic herbal preparation. METHODS: We utilized the phobic behavior of mice in an elevated plus-maze as a model to evaluate the anxiolytic effect of the plant extract and bio-guided fractionation was applied to isolate the active compounds. Total root extract, alkaline and ether fraction were administered to mice at different doses 30 and 90 min prior to the maze test. Saline and diazepam were administered as negative and positive controls, respectively. The time spent in open and closed arms, an index of anxiety behavior and entry time, was measured as an index of animal activity. RESULTS: The total root extract exhibited anxiolytic effect which was comparable to diazepam but with longer duration. This sustained effect of the crude extract was sustained for 90 min and was even more after injection of 45 mg/kg while the effect of diazepam had been reduced by 90 min. The anxiolytic effect factor was only present in the alkaline fraction and displayed its effect at lower doses than diazepam while pure vasicinone as the previously known alkaloid did not shown anxiolytic effect. The effect of the alkaline fraction was in a dose dependent manner starting at 0.2 mg/kg with a maximum at 1.0 mg/kg. Bio-guided fractionation using a variety of chromatographic methods led to isolation and purification of three coumarin derivatives from the bioactive fraction, including umbelliferone, scopoletin, and ferulic acid. CONCLUSION: For the first time, bio-guided fractionation of the root extract of B. multifida indicates significant sustained anxiolytic effects which led to isolation of three coumarin derivatives with well-known potent MAO inhibitory and anti-anxiety effects. These data contribute to evidence-based traditional use of B. multifida root for anxiety disorders.