Efficacy and safety of tafamidis doses in the Tafamidis in Transthyretin Cardiomyopathy Clinical Trial (ATTR-ACT) and long-term extension study.

Aims Tafamidis is an effective treatment for transthyretin amyloid cardiomyopathy (ATTR-CM) in the Tafamidis in Transthyretin Cardiomyopathy Clinical Trial (ATTR-ACT). While ATTR-ACT was not designed for a dose-specific assessment, further analysis from ATTR-ACT and its long-term extension study (LTE) can guide determination of the optimal dose. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Methods and results In ATTR-ACT, patients were randomized (2:1:2) to tafamidis 80 mg, 20mg, or placebo for 30months. Patients completing ATTR-ACT could enrol in the LTE (with placebo-treated patients randomized to tafamidis 80 or 20 mg; 2:1) and all patients were subsequently switched to high-dose tafamidis. All-cause mortality was assessed in ATTR-ACT combined with the LTE (median follow-up 51 months). In ATTR-ACT, the combination of all-cause mortality and cardiovascular-related hospitalizations over 30 months was significantly reduced with tafamidis 80mg (P = 0.0030) and 20mg (P = 0.0048) vs. placebo. All-cause mortality vs. placebo was reduced with tafamidis 80mg [Cox hazards model (95% confidence interval (CI): 0.690 (0.487–0.979), P = 0.0378] and 20mg [0.715 (0.450–1.137), P = 0.1564]. The mean (standard error) change in N-terminal pro-B-type natriuretic peptide from baseline to Month 30 was −1170.51 (587.31) (P = 0.0468) with tafamidis 80 vs. 20 mg. In ATTR-ACT combined with the LTE there was a significantly greater survival benefit with tafamidis 80 vs. 20 mg [0.700 (0.501–0.979), P = 0.0374]. Incidence of adverse events in both tafamidis doses were comparable to placebo. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Conclusion Tafamidis, both 80 and 20mg, effectively reduced mortality and cardiovascular-related hospitalizations in patients with ATTR-CM. The longer-term survival data and the lack of dose-related safety concerns support tafamidis 80mg as the optimal dose. Clinical Trial Registration: ClinicalTrials.gov NCT01994889; NCT02791230post-print369 K

Application of Quality by Design to the robust preparation of a liposomal GLA formulation by DELOS-susp method

Fabry disease is a lysosomal storage disease arising from a deficiency of the enzyme α-galactosidase A (GLA). The enzyme deficiency results in an accumulation of glycolipids, which over time, leads to cardiovascular, cerebrovascular, and renal disease, ultimately leading to death in the fourth or fifth decade of life. Currently, lysosomal storage disorders are treated by enzyme replacement therapy (ERT) through the direct administration of the missing enzyme to the patients. In view of their advantages as drug delivery systems, liposomes are increasingly being researched and utilized in the pharmaceutical, food and cosmetic industries, but one of the main barriers to market is their scalability. Depressurization of an Expanded Liquid Organic Solution into aqueous solution (DELOS-susp) is a compressed fluid-based method that allows the reproducible and scalable production of nanovesicular systems with remarkable physicochemical characteristics, in terms of homogeneity, morphology, and particle size. The objective of this work was to optimize and reach a suitable formulation for in vivo preclinical studies by implementing a Quality by Design (QbD) approach, a methodology recommended by the FDA and the EMA to develop robust drug manufacturing and control methods, to the preparation of α-galactosidase-loaded nanoliposomes (nanoGLA) for the treatment of Fabry disease. Through a risk analysis and a Design of Experiments (DoE), we obtained the Design Space in which GLA concentration and lipid concentration were found as critical parameters for achieving a stable nanoformulation. This Design Space allowed the optimization of the process to produce a nanoformulation suitable for in vivo preclinical testing

Highly Versatile Polyelectrolyte Complexes for Improving the Enzyme Replacement Therapy of Lysosomal Storage Disorders

Lysosomal storage disorders are currently treated by enzyme replacement therapy (ERT) through the direct administration of the unprotected recombinant protein to the patients. Herein we present an ionically cross-linked polyelectrolyte complex (PEC) composed of trimethyl chitosan (TMC) and α-galactosidase A (GLA), the defective enzyme in Fabry disease, with the capability of directly targeting endothelial cells by incorporating peptide ligands containing the RGD sequence. We assessed the physicochemical properties, cytotoxicity, and hemocompatibility of RGD-targeted and untargeted PECs, the uptake by endothelial cells and the intracellular activity of PECs in cell culture models of Fabry disease. Moreover, we also explored the effect of different freeze-drying procedures in the overall activity of the PECs. Our results indicate that the use of integrin-binding RGD moiety within the PEC increases their uptake and the efficacy of the GLA enzyme, while the freeze-drying allows the activity of the therapeutic protein to remain intact. Overall, these results highlight the potential of TMC-based PECs as a highly versatile and feasible drug delivery system for improving the ERT of lysosomal storage disorders

Extracellular histones activate autophagy and apoptosis via mTOR signaling in human endothelial cells

Circulating histones have been proposed as targets for therapy in sepsis and hyperinflammatory symptoms. However, the proposed strategies have failed in clinical trials. Although different mechanisms for histone-related cytotoxicity are being explored, those mediated by circulating histones are not fully understood. Extracellular histones induce endothelial cell death, thereby contributing to the pathogenesis of complex diseases such as sepsis and septic shock. Therefore, the comprehension of cellular responses triggered by histones is capital to design effective therapeutic strategies. Here we report how extracellular histones induce autophagy and apoptosis in a dose-dependent manner in cultured human endothelial cells. In addition, we describe how histones regulate these pathways via Sestrin2/AMPK/ULK1-mTOR and AKT/mTOR. Furthermore, we evaluate the effect of Toll-like receptors in mediating autophagy and apoptosis demonstrating how TLR inhibitors do not prevent apoptosis and/or autophagy induced by histones. Our results confirm that histones and autophagic pathways can be considered as novel targets to design therapeutic strategies in endothelial damage

Highly versatile polyelectrolyte complexes for improving the enzyme replacement therapy of lysosomal storage disorders

Lysosomal storage disorders are currently treated by enzyme replacement therapy (ERT) through the direct administration of the unprotected recombinant protein to the patients. Herein we present an ionically crosslinked polyelectrolyte complex (PEC) composed of trimethyl chitosan (TMC) and -galactosidase A (GLA), the defective enzyme in Fabry disease, with the capability of directly targeting endothelial cells by incorporating peptide ligands containing the RGD sequence. We assessed the physicochemical properties, cytotoxicity and hemocompatibility of RGD-targeted and un-targeted PECs, the uptake by endothelial cells and the intracellular activity of PECs in cell culture models of Fabry disease. Moreover, we also explored the effect of different freezedrying procedures in the overall activity of the PECs. Our results indicate that the use of integrin-binding RGD moiety within the PEC increases their uptake and the efficacy of the GLA enzyme, while the freeze-drying allows keeping intact the activity of the therapeutic protein. Overall, these results highlight the potential of TMC-based PECs as a highly versatile and feasible drug delivery system for improving the ERT of lysosomal storage disorders

Levodopa-refractory hyperprolactinemia and pituitary findings in inherited disorders of biogenic amine metabolism

Elevated serum prolactin concentrations occur in inherited disorders of biogenic amine metabolism because dopamine deficiency leads to insufficient inhibition of prolactin secretion. This work from the International Working Group on Neurotransmitter Related Disorders (iNTD) presents the results of the first standardized study on levodopa-refractory hyperprolactinemia (LRHP; >1000 mU/L) and pituitary magnetic resonance imaging (MRI) abnormalities in patients with inherited disorders of biogenic amine metabolism. Twenty-six individuals had LRHP or abnormal pituitary findings on MRI. Tetrahydrobiopterin deficiencies were the most common diagnoses (n = 22). The median age at diagnosis of LRHP was 16 years (range: 2.5-30, 1st-3rd quartiles: 12.25-17 years). Twelve individuals (nine females) had symptoms attributed to hyperprolactinemia: menstruation-related abnormalities (n = 7), pubertal delay or arrest (n = 5), galactorrhea (n = 3), and decreased sexual functions (n = 2). MRI of the pituitary gland was obtained in 21 individuals; six had heterogeneity/hyperplasia of the gland, five had adenoma, and 10 had normal findings. Eleven individuals were treated with the dopamine agonist cabergoline, ameliorating the hyperprolactinemia-related symptoms in all those assessed. Routine monitoring of these symptoms together with prolactin concentrations, especially after the first decade of life, should be taken into consideration during follow-up evaluations. The potential of slow-release levodopa formulations and low-dose dopamine agonists as part of first-line therapy in the prevention and treatment of hyperprolactinemia should be investigated further in animal studies and human trials. This work adds hyperprolactinemia-related findings to the current knowledge of the phenotypic spectrum of inherited disorders of biogenic amine metabolism

Modified Gravity and Cosmology

In this review we present a thoroughly comprehensive survey of recent work on modified theories of gravity and their cosmological consequences. Amongst other things, we cover General Relativity, Scalar-Tensor, Einstein-Aether, and Bimetric theories, as well as TeVeS, f(R), general higher-order theories, Horava-Lifschitz gravity, Galileons, Ghost Condensates, and models of extra dimensions including Kaluza-Klein, Randall-Sundrum, DGP, and higher co-dimension braneworlds. We also review attempts to construct a Parameterised Post-Friedmannian formalism, that can be used to constrain deviations from General Relativity in cosmology, and that is suitable for comparison with data on the largest scales. These subjects have been intensively studied over the past decade, largely motivated by rapid progress in the field of observational cosmology that now allows, for the first time, precision tests of fundamental physics on the scale of the observable Universe. The purpose of this review is to provide a reference tool for researchers and students in cosmology and gravitational physics, as well as a self-contained, comprehensive and up-to-date introduction to the subject as a whole.Comment: 312 pages, 15 figure

Multifunctional nanovesicle-bioactive conjugates prepared by a one-step scalable method using CO2-expanded solvents

The integration of therapeutic biomolecules, such as proteins and peptides, in nanovesicles is a widely used strategy to improve their stability and efficacy. However, the translation of these promising nanotherapeutics to clinical tests is still challenged by the complexity involved in the preparation of functional nanovesicles and their reproducibility, scalability, and cost production. Here we introduce a simple one-step methodology based on the use of CO2-expanded solvents to prepare multifunctional nanovesicle-bioactive conjugates. We demonstrate high vesicle-to-vesicle homogeneity in terms of size and lamellarity, batch-to-batch consistency, and reproducibility upon scaling-up. Importantly, the procedure is readily amenable to the integration/encapsulation of multiple components into the nanovesicles in a single step and yields sufficient quantities for clinical research. The simplicity, reproducibility, and scalability render this one-step fabrication process ideal for the rapid and low-cost translation of nanomedicine candidates from the bench to the clinic.We acknowledge financial support from Instituto de Salud Carlos III, through “Acciones CIBER”. The Networking Research Center on Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN) is an initiative funded by the VI National R&D&I Plan 2008-2011, Iniciativa Ingenio 2010, Consolider Program, CIBER Actions and financed by the Instituto de Salud Carlos III with assistance from the European Regional Development Fund. The authors appreciate the financial support through the project “Development of nanomedicines for enzymatic replacement therapy in Fabry disease”, granted by the Fundació Marató TV3 and projects POMAS (CTQ2010-019501), granted by DGI (Spain), 2009SGR0516 and 2009SGR0108, financed by DGR (Catalunya). The authors wish also to thank the Microscopy Service of UAB, especially Pablo Castro and Ana Tarruella, for the technical support in taking the Cryo-TEM images. A.V. is a recipient of an ICREA Academia (Generalitat de Catalunya) award. We acknowledge BERENICE Project (305937-2) supported by the European Comission under the Health Innovation Work Programme of the 7th Framework Programme.Peer reviewe

The Level of DING Proteins Is Increased in HIV-Infected Patients: In Vitro and In Vivo Studies

DING proteins constitute an interesting family, owing to their intriguing and important activities. However, after a decade of research, little is known about these proteins. In humans, at least five different DING proteins have been identified, which were implicated in important biological processes and diseases, including HIV. Indeed, recent data from different research groups have highlighted the anti-HIV activity of some DING representatives. These proteins share the ability to inhibit the transcriptional step of HIV-1, a key step of the viral cycle that is not yet targeted by the current therapies. Since such proteins have been isolated from humans, we undertook a comprehensive study that focuses on the relationship between these proteins and HIV-infection in an infectious context. Hence, we developed a home-made ELISA for the quantification of the concentration of DING proteins in human serum. Using this method, we were able to determine the concentration of DING proteins in healthy and HIV-infected patients. Interestingly, we observed a significant increase of the concentration of DING proteins in non treated and treated HIV-infected patients compared to controls. In addition, cell cultures infected with HIV also show an increased expression of DING proteins, ruling out the possible role of antiretroviral treatment in the increase of the expression of DING proteins. In conclusion, results from this study show that the organism reacts to HIV-infection by an overexpression of DING proteins