Enzyme stability in nanoparticle preparations part 1: Bovine serum albumin improves enzyme function

Enzymes have gained attention for their role in numerous disease states, calling for research for their efficient delivery. Loading enzymes into polymeric nanoparticles to improve biodistribution, stability, and targeting in vivo has led the field with promising results, but these enzymes still suffer from a degradation effect during the formulation process that leads to lower kinetics and specific activity leading to a loss of therapeutic potential. Stabilizers, such as bovine serum albumin (BSA), can be beneficial, but the knowledge and understanding of their interaction with enzymes are not fully elucidated. To this end, the interaction of BSA with a model enzyme B-Glu, part of the hydrolase class and linked to Gaucher disease, was analyzed. To quantify the natural interaction of beta-glucosidase (B-Glu,) and BSA in solution, isothermal titration calorimetry (ITC) analysis was performed. Afterwards, polymeric nanoparticles encapsulating these complexes were fully characterized, and the encapsulation efficiency, activity of the encapsulated enzyme, and release kinetics of the enzyme were compared. ITC results showed that a natural binding of 1:1 was seen between B-Glu and BSA. Complex concentrations did not affect nanoparticle characteristics which maintained a size between 250 and 350 nm, but increased loading capacity (from 6% to 30%), enzyme activity, and extended-release kinetics (from less than one day to six days) were observed for particles containing higher B-Glu:BSA ratios. These results highlight the importance of understanding enzyme:stabilizer interactions in various nanoparticle systems to improve not only enzyme activity but also biodistribution and release kinetics for improved therapeutic effects. These results will be critical to fully characterize and compare the effect of stabilizers, such as BSA with other, more relevant therapeutic enzymes for central nervous system (CNS) disease treatments

Pyroprocess Experiments at ENEA Laboratories

A new facility, known as Pyrel III, has been installed at ENEA laboratories for pyrochemical process studies under inactive conditions. It is a pilot plant which allows electrorening and electroreduction experiments to be conducted on simulated fuel. The main component of the plant is a zirconia crucible. The crucible is heated by a furnace which is supported in an externally water-cooled well under the oor of a steel glove-box, where an argon atmosphere is maintained by a continual purge of about 10 L·min-1. The vessel is loaded with LiCl-KCl eutectic salt (59-41 mol%) and is currently operated at 460 °C. Several improvements on Pyrel II (the previous operating plant) have been introduced into Pyrel III. They are described in detail, together with the results from the rst experimental campaign which used lanthanum metal.Moreover, studies about the treatment of chloride salt wastes from pyroprocesses have been conducted in parallel. They follow two main routes: on one hand, a matrix termed sodalite, a naturally occurring mineral containing chlorine, has been synthesized from a mix of nepheline, simulated exhausted salts and glass frit; on the other hand, a novel method proposed by Korea Atomic Energy Research Institute (KAERI) is under assessment. The nal waste forms have been fully characterized with the support of the Politechnique of Milan, by means of density measurements, thermal analysis, and stereomicroscopy observations, FTIR, XRD, and RAMAN spectra, as well as leach tests under static condition

4-PAM Dispersion-Uncompensated Transmission with Micro-Ring Resonator Enhanced 1.55-µm DML

International audienceReal-time transmission of 14-GBd 4-PAM signal is demonstrated by combining a commercial 1.55-µm DML with a silicon MRR. BER below the HD-FEC threshold is measured after 26-km SSMF transmission without offline digital signal processing

Collimated dual species oven source and its characterisation via spatially resolved fluorescence spectroscopy

We describe the design, construction and characterisation of a collimated, dual-species oven source for generating intense beams of lithium and caesium in UHV environments. Our design produces full beam overlap for the two species. Using an aligned microtube array the FWHM of the output beam is restricted to ~ 75 milliradians, with an estimated axial brightness of 3.6x10[superscript]14 atoms s[superscript]-1 sr[superscript]-1 for Li and 7.4x10[superscript]15 atoms s[superscript]-1 sr[superscript]-1 for Cs. We measure the properties of the output beam using a spatially-resolved fluorescence technique, which allows for the extraction of additional information not accessible without spatial resolution

Plasma-Based Longitudinal Evaluation of ESR1 Epigenetic Status in Hormone Receptor-Positive HER2-Negative Metastatic Breast Cancer

Background: Endocrine therapy (ET) is the mainstay of treatment for hormone receptor-positive human epidermal growth factor receptor 2 (HER2)-negative metastatic breast cancer; however, adaptive mechanisms emerge in about 25\u201330% of cases through alterations in the estrogen receptor ligand-binding domain, with a consequent ligand-independent estrogen receptor activity. Epigenetic-mediated events are less known and potentially involved in alternative mechanisms of resistance. The aim of this study was to test the feasibility of estrogen receptor 1 (ESR1) epigenetic characterization through liquid biopsy and to show its potential longitudinal application for an early ET sensitivity assessment. Methods: A cohort of 49 women with hormone receptor-positive HER2-negative MBC was prospectively enrolled and characterized through circulating tumor DNA using methylation-specific droplet digital PCR (MS-ddPCR) before treatment start (BL) and after 3 months concomitantly with computed tomography (CT) scan restaging (EV1). ESR1 epigenetic status was defined by assessing the methylation of its main promoters (promA and promB). The most established cell-free tumor DNA (ctDNA) factors associated with ET resistance [ESR1 and phosphatidylinositol-4,5-bisphosphate 3-kinase, catalytic subunit alpha (PIK3CA) mutations] were assessed through next-generation sequencing. Associations were tested through Mann\u2013Whitney U test, matched pairs variations through Wilcoxon signed rank test, and survival was analyzed by log-rank test. Results: The ET backbone was mainly based on aromatase inhibitors (AIs) (70.83%) in association with CDK4/6 inhibitors (93.75%). Significantly lower promA levels at baseline were observed in patients with liver metastases (P = 0.0212) and in patients with ESR1 mutations (P = 0.0091). No significant impact on PFS was observed for promA (P = 0.3777) and promB (P = 0.7455) dichotomized at the median while a 652-fold increase in promB or in either promA or promB at EV1 resulted in a significantly worse prognosis (respectively P = 0.0189, P = 0.0294). A significant increase at EV1 was observed for promB among patients with PIK3CA mutation (P = 0.0173). A trend was observed for promB in ESR1 wild-type patients and for promA in the ESR1 mutant subgroup. Conclusion: The study proofed the concept of an epigenetic characterization strategy based on ctDNA and is capable of being integrated in the current clinical workflow to give useful insights on treatment sensitivity

An atlas of chromatoid body components

The genome of male germ cells is actively transcribed during spermatogenesis to produce phase-specific protein-coding mRNAs and a considerable amount of different noncoding RNAs. Ribonucleoprotein (RNP) granule-mediated RNA regulation provides a powerful means to secure the quality and correct expression of the requisite transcripts. Haploid spermatids are characterized by a unique, unusually large cytoplasmic granule, the chromatoid body (CB), which emerges during the switch between the meiotic and post-meiotic phases of spermatogenesis. To better understand the role of the CB in male germ cell differentiation, we isolated CBs from mouse testes and revealed its full RNA and protein composition. We showed that the CB is mainly composed of RNA-binding proteins and other proteins involved RNA regulation. The CB was loaded with RNA, including pachytene piRNAs, a diverse set of mRNAs, and a number of uncharacterized long noncoding transcripts. The CB was demonstrated to accumulate nascent RNA during all the steps of round spermatid differentiation. Our results revealed the CB as a large germ cell-specific RNP platform that is involved in the control of the highly complex transcriptome of haploid male germ cells

First- and second-line treatment strategies for hormone-receptor (HR)-positive HER2-negative metastatic breast cancer: A real-world study

Background: Endocrine therapy (ET) plus cyclin-dependent-kinases 4/6 inhibitors (CDK4/6i) represents the standard treatment for luminal-metastatic breast cancer (MBC). However, prospective head-to-head comparisons are still lacking for 1st line (L) options, and it is still crucial to define the best strategy between 1st and 2nd L. Materials and methods: 717 consecutive luminal-MBC pts treated between 2008 and 2020 were analyzed at the Oncology Department of Aviano and Udine, Italy. Differences about survival outcomes (OS, PFS and PPS) were tested by log-rank test. The attrition rate (AR) between 1st and 2ndL was calculated. Results: At 1stL, pts were treated with ET (49%), chemotherapy (CT) (31%) and ET-CDKi (20%) while, at 2ndL, 33% received ET, 33% CT and 8% ET-CDKi. Overall AR was 10%, 7% for CT, 8% for ET and 17% for ET-CDKi. By multivariate analysis, 1stL ET-CDK4/6i showed a better mPFS1 and OS. Moreover, 2ndL ET-CDK4/6i demonstrated better mPFS2 compared to ET and CT. Notably, 1stL ET-CDKi resulted in higher mPFS than 2ndL ET-CDKi. Intriguingly, 1stL ET-CDK4/6i was associated with worse mPPS compared to CT and ET. Secondarily, 1stL ET-CDK4/6i followed by CT had worse OS compared to 1stL ET-CDK4/6i followed by ET. Notably, none of baseline characteristics at 2ndL influenced 2ndL treatment choice (ET vs. CT) after ET-CDKi. Conclusion: Our real-world data demonstrated that ET-CDKi represents the best option for 1stL luminal-MBC compared to ET and CT. Also, the present study pointed out that 2ndL ET, potentially combined with other molecules, could be a feasible option after CDK4/6i failure, postponing CT on later lines