Nitric Oxide Production and Effects in Group B Streptococcus Chorioamnionitis.

Intrauterine infection, or chorioamnionitis, due to group B Streptococcus (GBS) is a common cause of miscarriage and preterm birth. To cause chorioamnionitis, GBS must bypass maternal-fetal innate immune defenses including nitric oxide (NO), a microbicidal gas produced by nitric oxide synthases (NOS). This study examined placental NO production and its role in host-pathogen interactions in GBS chorioamnionitis. In a murine model of ascending GBS chorioamnionitis, placental NOS isoform expression quantified by RT-qPCR revealed a four-fold expression increase in inducible NOS, no significant change in expression of endothelial NOS, and decreased expression of neuronal NOS. These NOS expression results were recapitulated ex vivo in freshly collected human placental samples that were co-incubated with GBS. Immunohistochemistry of wild type C57BL/6 murine placentas with GBS chorioamnionitis demonstrated diffuse inducible NOS expression with high-expression foci in the junctional zone and areas of abscess. Pregnancy outcomes between wild type and inducible NOS-deficient mice did not differ significantly although wild type dams had a trend toward more frequent preterm delivery. We also identified possible molecular mechanisms that GBS uses to survive in a NO-rich environment. In vitro exposure of GBS to NO resulted in dose-dependent growth inhibition that varied by serovar. RNA-seq on two GBS strains with distinct NO resistance phenotypes revealed that both GBS strains shared several detoxification pathways that were differentially expressed during NO exposure. These results demonstrate that the placental immune response to GBS chorioamnionitis includes induced NO production and indicate that GBS activates conserved stress pathways in response to NO exposure.

Intranasal delivery of a synthetic Entamoeba histolytica vaccine containing adjuvant (LecA + GLA-3 M-052 liposomes): In vitro characterization.

Amebiasis, a disease caused by the parasite Entamoeba histolytica, is estimated to cause millions of infections and at least 55,000 deaths globally each year. With no vaccine currently available, there is an urgent need for an accessible means of stimulating protective mucosal immunity. The objective of this study was to characterize the nasal spray of a novel amebiasis vaccine candidate from a syringe-based liquid atomization device, the Teleflex MAD Nasal™, in both adult and infant nasal airways. Human ergonomic testing was completed to determine realistic actuation parameters. Spray pattern, plume geometry, and droplet size distribution were measured to evaluate reproducibility of free plume characteristics. The Alberta Idealized Nasal Inlet (AINI) and three realistic infant nasal airways were used to determine the in vitro deposition profile in adult and infant airways, respectively. Collectively, in vitro results demonstrated the feasibility of delivering the vaccine candidate to target sites within the nasal airways. Penetration through the nasal airways that could lead to deposition in the lungs was below the limit of quantification for both adult and infant geometries, indicating a low likelihood of adverse events due to lung exposure. These results support continued investigation of intranasal delivery of the synthetic Entamoeba histolytica vaccine.

First-in-Human Studies of MW01-6-189WH, a Brain-Penetrant, Antineuroinflammatory Small-Molecule Drug Candidate: Phase 1 Safety, Tolerability, Pharmacokinetic, and Pharmacodynamic Studies in Healthy Adult Volunteers.

MW01-6-189WH (MW189) is a novel central nervous system-penetrant small-molecule drug candidate that selectively attenuates stressor-induced proinflammatory cytokine overproduction and is efficacious in intracerebral hemorrhage and traumatic brain injury animal models. We report first-in-human, randomized, double-blind, placebo-controlled phase 1 studies to evaluate the safety, tolerability, and pharmacokinetics (PK) of single and multiple ascending intravenous doses of MW189 in healthy adult volunteers. MW189 was safe and well tolerated in single and multiple doses up to 0.25 mg/kg, with no clinically significant concerns. The most common drug-related treatment-emergent adverse event was infusion-site reactions, likely related to drug solution acidity. No clinically concerning changes were seen in vital signs, electrocardiograms, physical or neurological examinations, or safety laboratory results. PK analysis showed dose-proportional increases in plasma concentrations of MW189 after single or multiple doses, with approximately linear kinetics and no significant drug accumulation. Steady state was achieved by dose 3 for all dosing cohorts. A pilot pharmacodynamic study administering low-dose endotoxin to induce a systemic inflammatory response was done to evaluate the effects of a single intravenous dose of MW189 on plasma cytokine levels. MW189 treatment resulted in lower levels of the proinflammatory cytokine TNF-α and higher levels of the anti-inflammatory cytokine IL-10 compared with placebo treatment. The outcomes are consistent with the pharmacological mechanism of MW189. Overall, the safety profile, PK properties, and pharmacodynamic effect support further development of MW189 for patients with acute brain injury.

Automated small-scale protein purification and analysis for accelerated development of protein therapeutics.

Small-scale protein purification presents opportunities for accelerated process development of biotherapeutic molecules. Miniaturization of purification conditions reduces time and allows for parallel processing of samples, thus offering increased statistical significance and greater breadth of variables. The ability of the miniaturized platform to be predictive of larger scale purification schemes is of critical importance. The PerkinElmer JANUS BioTx Pro and Pro-Plus workstations were developed as intuitive, flexible, and automated devices capable of performing parallel small-scale analytical protein purification. Preprogrammed methods automate a variety of commercially available ion exchange and affinity chromatography solutions, including miniaturized chromatography columns, resin-packed pipette tips, and resin-filled microtiter vacuum filtration plates. Here, we present a comparison of microscale chromatography versus standard fast protein LC (FPLC) methods for process optimization. In this study, we evaluated the capabilities of the JANUS BioTx Pro-Plus robotic platform for miniaturized chromatographic purification of proteins with the GE ӒKTA Express system. We were able to demonstrate predictive analysis similar to that of larger scale purification platforms, while offering advantages in speed and number of samples processed. This approach is predictive of scale-up conditions, resulting in shorter biotherapeutic development cycles and less consumed material than traditional FPLC methods, thus reducing time-to-market from discovery to manufacturing.