Blood-brain barrier transport kinetics of NOTA-modified proteins : the somatropin case

BACKGROUND: Chemical modifications such as PEG, polyamine and radiolabeling on proteins can alter their pharmacokinetic behaviour and their blood-brain barrier (BBB) transport characteristics. NOTA, i.e. 1,4,7-triazacyclononane-1,4,7-triacetic acid, is a bifunctional chelating agent that has attracted the interest of the scientific community for its high complexation constant with metals like gallium. Until now, the comparative BBB transport characteristics of NOTA-modified proteins versus unmodified proteins are not yet described. METHODS: Somatropin (i.e. recombinant human growth hormone), NOTA-conjugated somatropin and gallium-labelled NOTA-conjugated somatropin were investigated for their brain penetration characteristics (multiple time regression and capillary depletion) in an in vivo mice model to determine the blood-brain transfer properties. RESULTS: The three compounds showed comparable initial brain influx, with Kin = 0.38 ± 0.14 µL/(g×min), 0.36 ± 0.16 µL/(g×min) and 0.28 ± 0.18 µL/(g×min), respectively. Capillary depletion indicated that more than 80% of the influxed compounds reached the brain parenchyma. All three compounds were in vivo stable in serum and brain during the time frame of the experiments. CONCLUSIONS: Our results show that modification of NOTA as well as gallium chelation onto proteins, in casu somatropin, does not lead to a significantly changed pharmacokinetic profile at the blood-brain barrier

Quorumpeps database : chemical space, microbial origin and functionality of quorum sensing peptides

Quorum-sensing (QS) peptides are biologically attractive molecules, with a wide diversity of structures and prone to modifications altering or presenting new functionalities. Therefore, the Quorumpeps database (http://quorumpeps.ugent.be) is developed to give a structured overview of the QS oligopeptides, describing their microbial origin (species), functionality (method, result and receptor), peptide links and chemical characteristics (3D-structure-derived physicochemical properties). The chemical diversity observed within this group of QS signalling molecules can be used to develop new synthetic bio-active compounds

Gut microbiome patterns depending on children's psychosocial stress : reports versus biomarkers

Aim: Chronic stress increases disease vulnerability factors including inflammation, a pathological characteristic potentially regulated by the gut microbiota. We checked the association between the gut microbiome and psychosocial stress in children/adolescents and investigated which stress parameter (negative versus positive emotion, self-report versus parental report, events versus emotions, biomarker cortisol versus parasympathetic activity) is the most relevant indicator herein. Methods: Gut microbiome sequencing was completed in fecal samples from 93 Belgian 8-16y olds. Stress measures included negative events, negative emotions, emotional problems reported by parents, happiness, hair cortisol and heart rate variability (pnn50 parameter reflecting parasympathetic activity). Alpha diversity, beta diversity and linear discriminant analysis were the unadjusted analyses. Age, sex, socio-economic status, diet, physical activity, sleep and weight status were adjusted for via a redundancy analysis and differential abundance via zero-inflated negative binomial regression. Results: High stress as reflected by low pnn50 and more negative events were associated with a lower alpha diversity as indicated by the Simpson index. Happiness and pnn50 showed significant differences between high and low stress groups based on weighted UniFrac distance, and this remained significant after confounder adjustment. Adjusted and unadjusted taxonomic differences were also most pronounced for happiness and pnn50 being associated respectively with 24 OTU (=11.8% of bacterial counts) and 31 OTU (=13.0%). As a general pattern, high stress was associated with lower Firmicutes at the phylum level and higher Bacteroides, Parabacteroides, Rhodococcus, Methanobrevibacter and Roseburia but lower Phascolarctobacterium at genus level. Several genera gave conflicting results between different stress measures e.g. Ruminococcaceae UCG014, Tenericutes, Eubacterium coprostanoligenes, Prevotella 9 and Christensenellaceae R7. Differential results in preadolescents versus adolescents were also evident. Conclusion: Even in this young healthy population, stress parameters were cross-sectionally associated with gut microbial composition but this relationship was instrument specific. Positive emotions and parasympathetic activity appeared the strongest parameters and should be integrated in future microbiota projects amongst other stress measures

Experimental models to study intestinal microbes-mucus interactions in health and disease

A close symbiotic relationship exists between the intestinal microbiota and its host. A critical component of gut homeostasis is the presence of a mucus layer covering the gastrointestinal tract. Mucus is a viscoelastic gel at the interface between the luminal content and the host tissue that provides a habitat to the gut microbiota and protects the intestinal epithelium. The review starts by setting up the biological context underpinning the need for experimental models to study gut bacteria-mucus interactions in the digestive environment. We provide an overview of the structure and function of intestinal mucus and mucins, their interactions with intestinal bacteria (including commensal, probiotics and pathogenic microorganisms) and their role in modulating health and disease states. We then describe the characteristics and potentials of experimental models currently available to study the mechanisms underpinning the interaction of mucus with gut microbes, including in vitro, ex vivo and in vivo models. We then discuss the limitations and challenges facing this field of research

Particle size determines the anti-inflammatory effect of wheat bran in a model of fructose over-consumption : implication of the gut microbiota

We investigated the impact of the particle size of wheat bran on gut dysbiosis and inflammation induced by a fructose overload. Mice received drinking water with or without fructose (30%) and a standard diet supplemented with or without 5% of wheat bran fractions characterized by different average particle sizes (1690 pm versus 150 um) for 8 weeks. Fructose increased Enterobacteriaceae associated with higher expression of key inflammatory genes in the liver. The two wheat bran fractions differently affected specific gut bacteria known to be involved in the regulation of the gut barrier function and/or inflammatory processes. Moreover, wheat bran with small particle size was the sole fibre that reduced hepatic and systemic inflammatory markers upon high fructose intake. The anti-inflammatory effects of wheat bran may be dependent on their particle size and could be related to the changes in caecal Enterobacteriaceae

Probes for Non-invasive Matrix Metalloproteinase-targeted Imaging with PET and SPECT

Dysregulation of matrix metalloproteinase (MMP) activity can lead to a wide range of disease states such as atherosclerosis, inflammation or cancer. The ability to image MMP activity non-invasively in vivo, by radiolabelled synthetic inhibitors, would allow the characterization of atherosclerotic plaques, inflammatory lesions or tumors. Here we present an overview of radiolabelled MMP inhibitors (MMPIs) and MMP peptides for positron emission tomography (PET) and single photon emission computed tomography (SPECT) for the detection of proteolytic activity of MMPs. So far, most studies are at a preliminary stage; however, some hydroxamate-based tracers such as the peptidomimetics [In-111]-DTPA-RP782, [Tc-99m]-(HYNIC-RP805)(tricine)(TPPTS), or Marimastat-ArB[F-18]F-3 and the picolyl-benzenesulfonamide [I-123]I-HO-CGS 27023A identified specifically the enzymatic action of MMPs in animal models of various pathologies. The development of new compounds that may lead to novel tracers (e.g. modification of zinc-binding group, variation of substituents attached to the S1', S2' and S3' pockets of the MMP inhibitors) and the use of antibodies and cell penetrating peptides are also discussed. In general, preclinical studies with atherosclerosis models proved to be more successful than those with oncological models

Grid of analysis supporting the participative design methodology

PALETTE deliverable - report number D.PAR.01This deliverable presents the participatory design methodology of the PALETTE project and some first results of the implementation of this process. Some principles of the Actor Network Theory and of the Agile Methodology are embedded in the different stages of this methodology whose twelve stages (described in details in the last section) will be the basis of the participative development of services and scenarios of use

In vitro functional quality characterization of NOTA-modified somatropins

Chemical modifications on protein biopharmaceuticals introduce extra variability in addition to their inherent complexity, hence require more comprehensive analytical and functional characterization during their discovery, development, and manufacturing. Somatropin (i.e., recombinant human growth hormone, rhGH) modified with the chelating agent S-2-(4-isothiocyanatobenzyl)-1,4,7-triazacyclononane-1,4,7-triacetic acid (p-SCN-Bn-NOTA) allows the incorporation of radiometals for research and possible theranostic purposes. We previously demonstrated that this conjugation leads to multiple substitution degrees and positional isomers within the product. In vitro techniques at the molecular and cellular levels were now applied to assess their functional quality: (i) size exclusion chromatography (SEC) demonstrated functional complexation with human growth hormone binding protein (hGHBp) to the different NOTA-modified somatropins as well as to gallium chelated NOTA-functionalities (Ga-10:1 NOTA-somatropin); (ii) native mass spectrometry (MS) offered in-depth information, a substitution degree up to four NOTAs was still functional; (iii) circular dichroism (CD) analysis confirmed the complexation of unmodified and NOTA-modified somatropin to hGHBp; and (iv) a hGHR bioassay demonstrated initiation of the signal transduction cascade, after binding of all investigated products to the receptor presented on cells with a similar potency (pEC50 values between 9.53 and 9.78) and efficacy (Emax values between 130 and 160%). We conclude that the NOTA-modified somatropins do not possess a significantly different in vitro functionality profile compared to unmodified somatropin. Techniques such as SEC, MS, and CD, traditionally used in the physicochemical characterization of proteins have a demonstrated potential use in the functionality evaluation not only in drug discovery and development but also in quality control settings.Grants from the Institute for the Promotion of Innovation through Science and Technology in Flanders (IWT-Vlaanderen) to B.G. (Grant 121512) and F.V. (Grant 131356), and the PhD research grants from China Scholarship Council (CSC) to H.Y. and X.X.http://pubs.acs.org/loi/ancham2018-03-07hj2017Nuclear Medicin

MycoBank gearing up for new horizons.

MycoBank, a registration system for fungi established in 2004 to capture all taxonomic novelties, acts as a coordination hub between repositories such as Index Fungorum and Fungal Names. Since January 2013, registration of fungal names is a mandatory requirement for valid publication under the International Code of Nomenclature for algae, fungi and plants (ICN). This review explains the database innovations that have been implemented over the past few years, and discusses new features such as advanced queries, registration of typification events (MBT numbers for lecto, epi- and neotypes), the multi-lingual database interface, the nomenclature discussion forum, annotation system, and web services with links to third parties. MycoBank has also introduced novel identification services, linking DNA sequence data to numerous related databases to enable intelligent search queries. Although MycoBank fills an important void for taxon registration, challenges for the future remain to improve links between taxonomic names and DNA data, and to also introduce a formal system for naming fungi known from DNA sequence data only. To further improve the quality of MycoBank data, remote access will now allow registered mycologists to act as MycoBank curators, using Citrix software