Stability Comparison Between Microglassification and Lyophilization Using a Monoclonal Antibody.

Producing solid-state formulations of biologics remains a daunting task despite the prevalent use of lyophilization and spray drying technologies in the biopharmaceutical industry. The challenges include protein stability (temperature stresses), high capital costs, particle design/controllability, shortened processing times and manufacturing considerations (scalability, yield improvements, aseptic operation, etc.). Thus, scientists/engineers are constantly working to improve existing methodologies and exploring novel dehydration/powder-forming technologies. Microglassification™ is a dehydration technology that uses solvent extraction to rapidly dehydrate protein formulations at ambient temperatures, eliminating the temperature stress experienced by biologics in traditional lyophilization and spray drying methods. The process results in microparticles that are spherical, dense, and chemically stable. In this study, we compared the molecular stability of a monoclonal antibody formulation processed by lyophilization to the same formulation processed using Microglassification™. Both powders were placed on stability for 3 months at 40 °C and 6 months at 25 °C. Both dehydration methods showed similar chemical stability, including percent monomer, charge variants, and antigen binding. These results show that Microglassification™ is viable for the production of stable solid-state monoclonal antibody formulations.

Preservatives from food-For food: Pea protein hydrolysate as a novel bio-preservative against Escherichia coli O157:H7 on a lettuce leaf.

Fresh-cut fruits and vegetables are becoming particularly popular as healthy fast-food options; however, they present challenges such as accelerated rates of decay and increased risk for contamination when compared to whole produce. Given that food safety must remain paramount for producers and manufacturers, research into novel, natural food preservation solutions which can help to ensure food safety and protect against spoilage is on the rise. In this work, we investigated the potential of using a novel protein hydrolysate, produced by the enzymatic hydrolysis of Pisum sativum (PSH), as a novel bio-preservative and its abilities to reduce populations of Escherichia coli O157:H7 after inoculation on a lettuce leaf. While unhydrolyzed P. sativum proteins show no antimicrobial activity, once digested, and purified, the enzymatically released peptides induced in vitro bactericidal effects on the foodborne pathogen at 8 mg/ml. When applied on an infected lettuce leaf, the PSH significantly reduced the number of bacteria recovered after 2 hr of treatment. PSH may be preferred over other preservation strategies based on its natural, inexpensive, sustainable source, environmentally friendly process, nontoxic nature, good batch to batch consistency, and ability to significantly reduce counts of E. coli both in vitro and in a lettuce leaf.

Measuring particle concentration of multimodal synthetic reference materials and extracellular vesicles with orthogonal techniques: Who is up to the challenge?

The measurement of physicochemical properties of polydisperse complex biological samples, for example, extracellular vesicles, is critical to assess their quality, for example, resulting from their production and isolation methods. The community is gradually becoming aware of the need to combine multiple orthogonal techniques to perform a robust characterization of complex biological samples. Three pillars of critical quality attribute characterization of EVs are sizing, concentration measurement and phenotyping. The repeatable measurement of vesicle concentration is one of the key-challenges that requires further efforts, in order to obtain comparable results by using different techniques and assure reproducibility. In this study, the performance of measuring the concentration of particles in the size range of 50-300 nm with complementary techniques is thoroughly investigated in a step-by step approach of incremental complexity. The six applied techniques include multi-angle dynamic light scattering (MADLS), asymmetric flow field flow fractionation coupled with multi-angle light scattering (AF4-MALS), centrifugal liquid sedimentation (CLS), nanoparticle tracking analysis (NTA), tunable resistive pulse sensing (TRPS), and high-sensitivity nano flow cytometry (nFCM). To achieve comparability, monomodal samples and complex polystyrene mixtures were used as particles of metrological interest, in order to check the suitability of each technique in the size and concentration range of interest, and to develop reliable post-processing data protocols for the analysis. Subsequent complexity was introduced by testing liposomes as validation of the developed approaches with a known sample of physicochemical properties closer to EVs. Finally, the vesicles in EV containing plasma samples were analysed with all the tested techniques. The results presented here aim to shed some light into the requirements for the complex characterization of biological samples, as this is a critical need for quality assurance by the EV and regulatory community. Such efforts go with the view to contribute to both, set-up reproducible and reliable characterization protocols, and comply with the Minimal Information for Studies of Extracellular Vesicles (MISEV) requirements.

Cybersecurity Convergence: Digital Human and National Security.

Over the past decade, people everywhere have become as dependent on the virtual world for their daily activities as they are dependent on the physical world for human activities. Global fiber optic networks have enabled communication in an unprecedented manner, connecting people in unique ways, propelling global supply chains, and giving consumers access to a variety of data from around the world. The online world is threatened by interstate rivals that engage in influence operations, economic espionage, and intelligence gathering and criminal groups that steal identities, ransom data, and grow their enterprises. Insiders facilitate intrusions wittingly and unwittingly raising the importance of corporations' roles in cybersecurity. This convergence between the virtual and physical worlds with the government and the corporate upends the entire frame of reference for national security, which is tilted toward physical attack and strict jurisdictional lines. As cybersecurity integrates further into U.S. national security, a new approach is needed to incorporate a human security construct at the user level. This article is adapted from their forthcoming book, Security in the Cyber Age from Georgetown University Press. The views expressed are their own.

Primary Breast Carcinoma of the Vulva Metastatic to Lymph Nodes and Bones: A Case Report and Literature Review.

INTRODUCTION: Primary breast carcinoma can occur at ectopic sites. The axilla is the most common site of ectopic primary breast cancer, but presentation in the vulva is rare. We discuss a rare presentation of primary breast carcinoma of the vulva with distant lymph node and bone metastases in a premenopausal woman. CASE PRESENTATION: A vulvar malignancy consistent with adenocarcinoma of the mammary gland type was diagnosed in a 47-year-old premenopausal woman. The patient underwent radical vulvectomy with bilateral superficial and deep inguinal lymphadenectomy. The tumor was positive for estrogen receptor and negative for progesterone receptor and human epidermal growth factor receptor 2/neu on immunohistochemical findings. A positron emission tomography-computed tomography scan demonstrated lymph node and bone metastases. Her disease was treated as stage IV breast cancer with metastases to the bone. Palliative treatment with ovarian suppression, aromatase inhibitor, and cyclin-dependent kinase 4/6 inhibitor was recommended. DISCUSSION: For a diagnosis of primary breast cancer of the vulva, a thorough metastatic workup should be performed, with attention directed toward detecting a breast primary disease by results of the history, physical examination, and radiologic examination of the breasts mainly to help confirm that the vulvar lesion is the primary site as opposed to metastasis from a breast primary cancer and also for staging. Management of this rare entity is challenging because of a lack of specific guidelines, and treatment, therefore, is similar to that of breast cancer.Treatment should consist of an individualized combination of surgery, radiotherapy, chemotherapy, and antiestrogen hormonal therapy.

Persistent Subdiffusive Proton Transport in Perfluorosulfonic Acid Membranes.

Proton transport (PT) in solutions of small amphiphiles in water has previously been shown to be subdiffusive for long times. The present study analyzes simulations of hydrated perfluorosulfonic acid (PFSA) membranes in order to determine whether PT is also subdiffusive in these important amphiphilic systems. We show that PT is indeed subdiffusive for several hundred picoseconds for all hydration levels examined, and the subdiffusive behavior is highly dependent on water concentration. We also investigate the caging of the excess proton using a recently developed technique and show that the excess proton exhibits caging effects up to at least 1 ns in PFSA systems. In order to fully characterize the long-time behavior of PT in PFSAs, these results demonstrate that multiple nanosecond trajectories are needed, well beyond the current capabilities of ab initio molecular dynamics.

Entropy-driven crystal formation on highly strained substrates.

In heteroepitaxy, lattice mismatch between the deposited material and the underlying surface strongly affects nucleation and growth processes. The effect of mismatch is well studied in atoms with growth kinetics typically dominated by bond formation with interaction lengths on the order of one lattice spacing. In contrast, less is understood about how mismatch affects crystallization of larger particles, such as globular proteins and nanoparticles, where interparticle interaction energies are often comparable to thermal fluctuations and are short ranged, extending only a fraction of the particle size. Here, using colloidal experiments and simulations, we find particles with short-range attractive interactions form crystals on isotropically strained lattices with spacings significantly larger than the interaction length scale. By measuring the free-energy cost of dimer formation on monolayers of increasing uniaxial strain, we show the underlying mismatched substrate mediates an entropy-driven attractive interaction extending well beyond the interaction length scale. Remarkably, because this interaction arises from thermal fluctuations, lowering temperature causes such substrate-mediated attractive crystals to dissolve. Such counterintuitive results underscore the crucial role of entropy in heteroepitaxy in this technologically important regime. Ultimately, this entropic component of lattice mismatched crystal growth could be used to develop unique methods for heterogeneous nucleation and growth of single crystals for applications ranging from protein crystallization to controlling the assembly of nanoparticles into ordered, functional superstructures. In particular, the construction of substrates with spatially modulated strain profiles would exploit this effect to direct self-assembly, whereby nucleation sites and resulting crystal morphology can be controlled directly through modifications of the substrate.

Temperature dependence of droplet breakup in 8CB and 5CB liquid crystals.

Droplet breakup of many Newtonian fluids is well described by current experiments, theory, and simulations. Breakup in complex fluids where interactions between mesoscopic structural features can affect the flows remains poorly understood and a burgeoning area of research. Here, we report on our investigations of droplet breakup in thermotropic liquid crystals. We investigate breakup in the smectic, nematic, and isotropic phases of 4-cyano 4-octylbiphenyl (8CB) and the nematic and isotropic phases of 4-cyano 4-pentylbiphenyl (5CB). The experiment consists of varying the ambient temperature to control liquid crystalline phase and imaging breakup using a fast video camera at up to 110000 frames/s. We expand on previous work [John R. Savage et al., Soft Matter 6, 892 (2010)] that shows breakup in the smectic phase is symmetric, producing no satellite droplets, and is well described by a similarity solution for a shear-thinning power-law fluid. We show that in the nematic phase the breakup occurs in two stages. In the first stage, the breakup is symmetric and the power-law exponent for the minimum radius dependence on the time left to breakup is 1.2

Proton conduction in exchange membranes across multiple length scales.

Concerns over global climate change associated with fossil-fuel consumption continue to drive the development of electrochemical alternatives for energy technology. Proton exchange fuel cells are a particularly promising technology for stationary power generation, mobile electronics, and hybrid engines in automobiles. For these devices to work efficiently, direct electrical contacts between the anode and cathode must be avoided; hence, the separator material must be electronically insulating but highly proton conductive. As a result, researchers have examined a variety of polymer electrolyte materials for use as membranes in these systems. In the optimization of the membrane, researchers are seeking high proton conductivity, low electronic conduction, and mechanical stability with the inclusion of water in the polymer matrix. A considerable number of potential polymer backbone and side chain combinations have been synthesized to meet these requirements, and computational studies can assist in the challenge of designing the next generation of technologically relevant membranes. Such studies can also be integrated in a feedback loop with experiment to improve fuel cell performance. However, to accurately simulate the currently favored class of membranes, perfluorosulfonic acid containing moieties, several difficulties must be addressed including a proper treatment of the proton-hopping mechanism through the membrane and the formation of nanophase-separated water networks. We discuss our recent efforts to address these difficulties using methods that push the limits of computer simulation and expand on previous theoretical developments. We describe recent advances in the multistate empirical valence bond (MS-EVB) method that can probe proton diffusion at the nanometer-length scale and accurately model the so-called Grotthuss shuttling mechanism for proton diffusion in water. Using both classical molecular dynamics and coarse-grained descriptions that replace atomistic representations with collective coordinates, we investigated the proton conductivity of polymer membrane structure as a function of hydration level. Nanometer-sized water channels form torturous pathways that are traversed by the charges during fuel cell operation. Using a combination of coarse-grained membrane structure and novel multiscale methods, we demonstrate emerging approaches to treat proton motion at the mesoscale in these complex materials.

Phase II study of sequential doublets: topotecan and carboplatin, followed by paclitaxel and carboplatin, in patients with newly diagnosed advanced ovarian cancer.

OBJECTIVE: Incorporating topotecan into standard platinum/taxane chemotherapy for advanced ovarian cancer has been complicated by myelosuppression. This study evaluated sequential doublets of topotecan and carboplatin, followed by paclitaxel and carboplatin, in newly diagnosed advanced ovarian cancer patients. METHODS: Forty-five patients (median age, 56 years; range, 38-77 years) with stage III/IV disease and GOG performance status <2 were enrolled and received four cycles of topotecan (1.0 mg/m(2)/day on days 1 to 3) and carboplatin (AUC 4 on day 1), followed by four cycles of paclitaxel (175 mg/m(2) via 3-h IV infusion on day 1) and carboplatin (AUC 5 on day 1). All cycles were 21 days. Antitumor response was assessed after four and eight cycles; patients with clinical complete response (CR) underwent second-look laparotomy for determination of pathologic CR (PCR). Dose reductions were instituted for grade 4 neutropenia and thrombocytopenia, and for grade 3/4 nonhematologic toxicity. RESULTS: Among 41 CA-125 evaluable patients, complete and partial responses were observed in 29 (70.7%) and 11 (26.8%) patients, respectively. Of the 12 clinical CRs (43%) in 28 evaluable patients, 10 patients underwent second-look laparotomy, with 3 PCRs (30%). Median time to progression was 14 months and actuarial survival was 23 months. Neutropenia was the primary toxicity and cause of dose adjustments and delays, including two deaths. CONCLUSION: The antitumor activity observed is comparable with other series, although neutropenic complications were increased. Progression-free and actuarial survivals were slightly inferior. A Phase III trial (GOG 182) of sequential doublets in the reverse sequence is ongoing.

Reflections and meditations upon complex chromosomal exchanges.

The application of FISH chromosome painting techniques, especially the recent mFISH (and its equivalents) where all 23 human chromosome pairs can be distinguished, has demonstrated that many chromosome-type structural exchanges are much more complicated (involving more "break-rejoins" and arms) than has hitherto been assumed. It is clear that we have been greatly under-estimating the damage produced in chromatin by such agents as ionising radiation. This article gives a brief historical summary of observations leading up to this conclusion, and after outlining some of the problems surrounding the formation of complex chromosomes exchanges, speculates about possible solutions currently being proposed.

Comparison of the experience with acute and chronic electrically stimulated detrusor myoplasty.

AIMS: To evaluate the acute and chronic urodynamic effects of electrically stimulated detrusor myoplasty in dogs. METHODS: Eight female mongrel dogs were studied acutely and six dogs chronically (0 to 12 weeks postoperatively). Bladders were wrapped with the rectus abdominis muscle, keeping an intact blood supply and at least two intercostal nerves of the flap preserved. Bladders were electrically stimulated with bipolar electrodes inserted into the muscle. Urodynamics and post void residual were measured post operatively in the acute studies and every 2 weeks for 3 months in chronic studies. RESULTS: Acutely, the increase in intravesical pressure was 45+/-7 cm H(2)O, which resulted in a postvoid residual of 26+/-3%. In the chronic study, increases of intravesical pressure sufficient to empty the bladder during myoplasty electrical stimulation were not sustained, although detrusor compliance and flap viability were preserved. CONCLUSIONS: The electrically stimulated detrusor myoplasty worked well acutely to increase vesical pressure sufficient to empty the bladder, but the chronically stimulated myoplasty did not maintain efficient bladder emptying primarily due to electrode problems. Further studies with improved electrode material and placement are required before clinical application of the electrically stimulated detrusor myoplasty can be assessed.

Improvement in neurogenic bladder after the antegrade continence enema procedure.

A child with neurogenic bladder and bowel underwent an antegrade continence enema procedure for fecal incontinence and severe constipation. She subsequently demonstrated an improvement in her neurogenic bladder and urinary incontinence.