Biodegradable nanocarriers coated with polymyxin B: Evaluation of leishmanicidal and antibacterial potential.

Most treatments of leishmaniasis require hospitalization and present side effects or parasite resistance; innovations in drug formulation/reposition can overcome these barriers and must be pursued to increase therapeutic alternatives. Therefore, we tested polymyxin B (polB) potential to kill Leishmania amazonensis, adsorbed or not in PBCA nanoparticles (PBCAnp), which could augment polB internalization in infected macrophages. PBCAnps were fabricated by anionic polymerization and analyzed by Dynamic Light Scattering (size, ζ potential), Nanoparticle Tracking Analysis (size/concentration), vertical diffusion cell (release rate), drug incorporation (indirect method, protein determination) and in vitro cell viability. Nanoparticles coated with polB (PBCAnp-polB) presented an adequate size of 261.5 ± 25.9 nm, low PDI and ζ of 1.79 ± 0.17 mV (stable for 45 days, at least). The 50% drug release from PBCAnp-polB was 6-7 times slower than the free polB, which favors a prolonged and desired release profile. Concerning in vitro evaluations, polB alone reduced in vitro amastigote infection of macrophages (10 µg/mL) without complete parasite elimination, even at higher concentrations. This behavior limits its future application to adjuvant leishmanicidal therapy or antimicrobial coating of carriers. The nanocarrier PBCAnp also presented leishmanicidal effect and surpassed polB activity; however, no antimicrobial activity was detected. PolB maintained its activity against E. coli, Pseudomonas and Klebsiella, adding antimicrobial properties to the nanoparticles. Thus, this coated drug delivery system, described for the first time, demonstrated antileishmanial and antimicrobial properties. The bactericidal feature helps with concomitant prevention/treatment of secondary infections that worst ulcers induced by cutaneous L. amazonensis, ultimately ending in disfiguring or disabling lesions.

Polymyxin Binding to the Bacterial Outer Membrane Reveals Cation Displacement and Increasing Membrane Curvature in Susceptible but Not in Resistant Lipopolysaccharide Chemotypes.

Lipid-A is the causative agent of Gram-negative sepsis and is responsible for an increasingly high mortality rate among hospitalized patients. Compounds that bind Lipid-A can limit this inflammatory process. The cationic antimicrobial peptide polymyxin B (Pmx-B) is one of the simplest molecules capable of selectively binding to Lipid-A and may serve as a model for further development of Lipid-A binding agents. Gram-negative bacteria resistance to Pmx-B relies on the upregulation of a number of regulatory systems, which promote chemical modifications of the lipopolysaccharide (LPS) structure and leads to major changes in the physical-chemical properties of the outer membrane. A detailed understanding of how the chemical structure of the LPS modulates macroscopic properties of the outer membrane is paramount for the design and optimization of novel drugs targeting clinically relevant strains. We have performed a systematic investigation of Pmx-B binding to outer membrane models composed of distinct LPS chemotypes experimentally shown to be either resistant or susceptible to the peptide. Molecular dynamics simulations were carried out for Pmx-B bound to the penta- and hexa-acylated forms of Lipid-A (more susceptible) and Lipid-A modified with 4-amino-4-deoxy-l-arabinose (resistant) as well as the penta-acylated form of LPS Re (less susceptible). The present simulations show that upon binding to the bacterial outer membrane surface, Pmx-B promotes cation displacement and structural changes in membrane curvature and integrity as a function of the LPS chemotype susceptibility or resistance to the antimicrobial peptide.

Phosphoethanolamine addition to the Heptose I of the Lipopolysaccharide modifies the inner core structure and has an impact on the binding of Polymyxin B to the Escherichia coli outer membrane.

Phosphoethanolamine (pEtN) decoration of E. coli Lipopolysaccharide (LPS) provides resistance to the antimicrobial Polymyxin B (PolB). While EptA and EptB enzymes catalyze the addition of pEtN to the Lipid A and Kdo (pEtN-Kdo-Lipid A), EptC catalyzes the pEtN addition to the Heptose I (pEtN-HeptI). In this study, we investigated the contribution of pEtN-HeptI to PolB resistance using eptA/eptB and eptC deficient E. coli K12 and its wild-type parent strains. These mutations were shown to decrease the antimicrobial activity of PolB on cells grown under pEtN-addition inducing conditions. Furthermore, the 1-N-phenylnapthylamine uptake assay revealed that in vivo PolB has a reduced OM-permeabilizing activity on the ΔeptA/eptB strain compared with the ΔeptC strain. In vitro, the changes in size and zeta potential of LPS-vesicles indicate that pEtN-HeptI reduce the PolB binding, but in a minor extent than pEtN-Kdo-Lipid A. Molecular dynamics analysis revealed the structural basis of the PolB resistance promoted by pEtN-HeptI, which generate a new hydrogen-bonding networks and a denser inner core region. Altogether, the experimental and theoretical assays shown herein indicate that pEtN-HeptI addition promote an LPS conformational rearrangement, that could act as a shield by hindering the accession of PolB to inner LPS-targets moieties.

Polymyxin B for the treatment of multidrug-resistant pathogens: a critical review.

Polymyxins have re-emerged in clinical practice owing to the dry antibiotic development pipeline and worldwide increasing prevalence of nosocomial infections caused by multidrug-resistant (MDR) Gram-negative bacteria. Polymyxin B and colistin (polymyxin E) have been ultimately considered as the last-resort treatment of such infections. Microbiological, pharmacokinetic, pharmacodynamic and clinical data available for polymyxin B are reviewed in this paper. Polymyxin B has rapid in vitro bactericidal activity against major MDR Gram-negative bacteria, such as Pseudomonas aeruginosa, Acinetobacter baumannii and Klebsiella pneumoniae. Acquired resistance to this agent is still rare among these pathogens. However, optimized dosage regimens are not known yet. Good clinical outcomes have been observed in the majority of the patients treated with intravenous polymyxin B in recent studies. However, these studies failed to provide definitive conclusions due to limitations of study design and additional clinical trials are required. Although combination therapy may be an attractive option based on some currently available in vitro data, clinical data supporting such recommendations are lacking. Since polymyxins will be increasingly used for the treatment of infections caused by MDR bacteria, clinical pharmacokinetic, pharmacodynamic and toxicodynamic studies underpinning the optimal use of these drugs are urgently required.

Evaluation of preservative systems in opthalmic suspension of polymyxin B and dexamethasone by linear regression.

An ophthalmic suspension of polymyxin B and dexametasone with 13 different preservative systems was evaluated by a linear regression method for rapid screening, using D-values. The products were challenged against Pseudomonas aeruginosa, Burkholderia cepacia, Staphylococcus aureus, Candida albicans and Aspergillus niger. Since linear regression showed correlation coefficients higher than 0.84 the decimal reduction time (D-value) was then calculated. The comparison of D-values obtained for each microorganism and each product led to the selection of the formula 9 = 10, which was selected to be submitted to the efficacy of antimicrobial preservation according to the European Pharmacopoeia.

Análise comparativa do potencial antimicrobiano de três medicações utilizadas no canal radicular / Comparative analysis of antimicrobial potential of three root canal dressing

O objetivo do presente estudo foi analisar o potencial antimicrobiano das medicações NDP, Pred-Fort associado a rifamicina e Otosporin quando em contato com bactéria comumente encontrada no interior do canal radicular,Staphylococcus aureus (ATCC 25923), em meio de cultura sólido Mueller Huinton. A infusão bacteriana foi preparada utilizando meio TSB, correspondente à escala 0,5 de McFarland. Foram utilizadas Placas de Petri, já esterilizadas, para acomodação de duas camadas de meio de cultura. Na base foram colocados 10 mL de Mueller Huinton e esperou-se solidificar. A segunda camada, também de 10 mL, foi preparada utilizando infusão bacteriana, diluída em 1 x 10 a 6ª em meio de cultura TSB e 5 mL de meio sólido, resultando na concentração de 15 X 10 a 5ª UFC/mL, que foi colocada sobre a base. Feito isto, foram acomodados discos de papel previamente embebidos nas modificações testadas nesta pesquisa sobre o meio. As placas foram acondicionadas em estufa durante 24h, a uma temperatura de 36°C, para análise dos resultados. Essa análise foi definida a partir da mensuração do halo de inibição, em milímitros, causados pela medicações, cujos valores foram submetidos à análise estatística. Segundo os resultados, houve formação de maiores halos de inibição quando da utilização, em ordem decrescente, do Pred-Fort + Rifamicina, do Otosporim e , finalmente, do NDP.