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Early restoration of parvalbumin interneuron activity prevents memory loss and network hyperexcitability in a mouse model of Alzheimer’s disease
No full textNeuronal network dysfunction is increasingly recognized as an early symptom in Alzheimer’s disease (AD) and may provide new entry points for diagnosis and intervention. Here, we show that amyloid-beta (Aβ)-induced hyperexcitability of hippocampal inhibitory parvalbumin (PV) interneurons contributes importantly to neuronal network dysfunction and memory impairment in APP/PS1 mice, a mouse model of increased amyloidosis. We demonstrate that hippocampal PV interneurons become hyperexcitable at ~16 weeks of age, when no changes are observed yet in the intrinsic properties of pyramidal cells. This hyperexcitable state of PV interneurons coincides with increased inhibitory transmission onto hippocampal pyramidal neurons and deficits in spatial learning and memory. We show that treatment aimed at preventing PV interneurons from becoming hyperexcitable is sufficient to restore PV interneuron properties to wildtype levels, reduce inhibitory input onto pyramidal cells, and rescue memory deficits in APP/PS1 mice. Importantly, we demonstrate that early intervention aimed at restoring PV interneuron activity has long-term beneficial effects on memory and hippocampal network activity, and reduces amyloid plaque deposition, a hallmark of AD pathology. Taken together, these findings suggest that early treatment of PV interneuron hyperactivity might be clinically relevant in preventing memory decline and potentially delaying AD progression
New potent DOT1L inhibitors for in vivo evaluation in mouse
No full textIn MLL-rearranged cancer cells, disruptor of telomeric silencing 1-like protein (DOT1L) is aberrantly recruited to ectop-ic loci leading to local hypermethylation of H3K79 and consequently misexpression of leukemogenic genes. A struc-ture-guided optimization of a HTS hit led to the discovery of DOT1L inhibitors with subnanomolar potency, allowing to test the therapeutic principle of DOT1L inhibition in a preclinical mouse tumor xenograft model. Compounds dis-playing good exposure in mouse and nanomolar inhibition of target gene expression in cell were obtained and tested in viv
Structural basis of indisulam-mediated RBM39 recruitment to DCAF15 E3 ligase complex
No full textThe anticancer agent indisulam inhibits cell proliferation by causing degradation of RBM39, an essential mRNA splicing factor. Indisulam promotes an interaction between RBM39 and the DCAF15 E3 ligase substrate receptor, leading to RBM39 ubiquitination and proteasome-mediated degradation. To delineate the precise mechanism by which indisulam mediates the DCAF15–RBM39 interaction, we solved the DCAF15–DDB1–DDA1–indisulam–RBM39(RRM2) complex structure to a resolution of 2.3 Å. DCAF15 has a distinct topology that embraces the RBM39(RRM2) domain largely via non-polar interactions, and indisulam binds between DCAF15 and RBM39(RRM2), coordinating additional interactions between the two proteins. Studies with RBM39 point mutants and indisulam analogs validated the structural model and defined the RBM39 α-helical degron motif. The degron is found only in RBM23 and RBM39, and only these proteins were detectably downregulated in indisulam-treated HCT116 cells. This work further explains how indisulam induces RBM39 degradation and defines the challenge of harnessing DCAF15 to degrade additional targets
Study of the critical points in combined matrix tablets containing both inert and swelling excipients
No full textThis work estimates for the first time critical points in combined matrices containing varying concentrations of the hydrophilic polymer Hydroxypropyl methylcellulose (HPMC) K100 M CR in presence of a constant percentage of the inert matrix forming polymer Eudragit RS-PO as well as varying concentrations of the inert polymer in presence of a constant percentage of the hydrophilic excipient. Drug release assays, water uptake studies and calculation of the Exicipient Efficiency (EE) have been carried out to study the interaction between the polymers. Surprisingly, an increase in the drug release rate occurs as the percentage of the hydrophobic polymer increases in the formulations. This fact is supported by the EE values which indicate a negative interaction between the two excipients. Moreover the HPMC percolation threshold estimated is higher than the one observed in pure HPMC matrices. It can be concluded that the HPMC creates pores in the inert skeleton, destabilizing the system. Moreover, the inert excipient destabilizes the gel layer formed by HPMC, changing its critical point. This information is essential for a rational estimation of the Design Space of a formulation and provides new knowledge on the behavior of the polymers in combined matrices, which contributes to the science based design
USP 800: Protecting Site Staff from Hazardous Drugs
No full textUSP 800: Protecting site staff from hazardous drugs
- Review the USP 800 regulation and application to clinical research
- Describe proper handling of hazardous drugs
- Develop a plan for handling a hazardous produc
Gut microbiome structure and metabolic activity in inflammatory bowel disease
No full textThe inflammatory bowel diseases (IBDs), which include Crohn’s disease (CD) and ulcerative colitis (UC), are multifactorial chronic conditions of the gastrointestinal tract. While IBD has been associated with dramatic changes in the gut microbiota, changes in the gut metabolome—the molecular interface between host and microbiota—are less well understood. To address this gap, we performed untargeted metabolomic and shotgun metagenomic profiling of cross-sectional stool samples from discovery (n = 155) and validation (n = 65) cohorts of CD, UC and non-IBD control patients. Metabolomic and metagenomic profiles were broadly correlated with faecal calprotectin levels (a measure of gut inflammation). Across >8,000 measured metabolite features, we identified chemicals and chemical classes that were differentially abundant in IBD, including enrichments for sphingolipids and bile acids, and depletions for triacylglycerols and tetrapyrroles. While > 50% of differentially abundant metabolite features were uncharacterized, many could be assigned putative roles through metabolomic ‘guilt by association’ (covariation with known metabolites). Differentially abundant species and functions from the metagenomic profiles reflected adaptation to oxidative stress in the IBD gut, and were individually consistent with previous findings. Integrating these data, however, we identified 122 robust associations between differentially abundant species and well-characterized differentially abundant metabolites, indicating possible mechanistic relationships that are perturbed in IBD. Finally, we found that metabolome- and metagenome-based classifiers of IBD status were highly accurate and, like the vast majority of individual trends, generalized well to the independent validation cohort. Our findings thus provide an improved understanding of perturbations of the microbiome–metabolome interface in IBD, including identification of many potential diagnostic and therapeutic targets
Real-time monitoring of particle size distribution in a continuous granulation and drying process by near infrared spectroscopy
No full textIn continuous granulation, it can be important to control granules particle size distribution (PSD), as it may affect final product quality. Near infrared spectroscopy (NIRS) is already a routine analytical procedure within pharmaceutical continuous manufacturing for the in-line analysis of chemical material-characteristics. Consequently, the extraction of additional information related to granules’ physical properties like particle size distribution is tempting, as it would enhance process knowledge without the need for new capital investments. Three in-line NIRS methods were developed via partial least squares regression, to predict dried granules PSD-fractions X10, X50, and X90 within a GMP-qualified continuous twin-screw wet granulation and fluid-bed drying process. Methods were developed for the size range of 20–234 µm (X10), 98–1017 µm (X50), and 748–2297 µm (X90) and assessed with one internal and three external validation datasets in agreement with current guidelines on NIRS. Internal validation indicated root mean square error of predictions (RMSEPs) of 17 µm, 97 µm, and 174 µm, for PSD X10, X50, and X90 respectively, with acceptable linearity, slope, and bias. Furthermore, the ratio of prediction to deviation (RPD), the ratio of prediction error to laboratory error (PRL), and the range error ratio (RER) were evaluated, with all values within the acceptance range for adequate to good NIR methods (1.75 > RPD < 3, PRL ≤ 2, RER ≥ 10). Methods applicability to in-line processes and their robustness towards water content and active pharmaceutical ingredient content was further demonstrated with three independent in-line datasets in real-time, showing good agreement between predicted and reference values. In summary, methods demonstrated to be sufficient for their intended purpose to monitor trends and sudden changes in dried granules PSD during continuous granulation and drying. Because of their fast response time, they are unique tools to characterize the dynamic behavior and navigate the agglomeration state of the material in static and transient process conditions during continuous granulation and drying
Pharmacokinetics and pharmacodynamics of the novel monobactam LYS228 in a neutropenic murine thigh model of infection
No full textObjectives: The neutropenic murine thigh infection model and a dose-fractionation approach were used to determine the pharmacokinetic/pharmacodynamic (PK/PD) relationship of LYS228, a novel monobactamantibiotic with activity against Enterobacteriaceae including carbapenem-resistant strains. Methods: Mice (n"4 per group) were inoculated with Enterobacteriaceae strains via intramuscular injection. Two hours post-bacterial inoculation, treatment with LYS228 was initiated. Animals were euthanized with CO2 24 h after the start of therapy and bacterial counts (log10 cfu) per thigh were determined. PK parameters were calculated using free (f) plasma drug levels. Results: Following a dose-fractionation study, non-linear regression analysis determined that the predominant PK/PD parameter associated with antibacterial efficacy of LYS228 was the percentage of the dosing interval that free drug concentrations remained above the MIC (%fT.MIC). In a dose-dependent manner, LYS228 reduced the thigh bacterial burden in models established with Enterobacteriaceae producing b-lactamase enzymes of all classes (e.g. ESBLs, NDM-1, KPC, CMY-2 and OXA-48). The range of the calculated static dose was 86-649 mg/kg/ day for the isolates tested, and the magnitude of the driver of efficacy was 37-83%fT.MIC. %fT.MIC was confirmed as the parameter predominantly driving efficacy as evidenced by a strong coefficient of determination (r2"0.68). Neutrophils had minimal impact on the effect of LYS228 in the murine thigh infection model. Conclusions: LYS228 is efficacious in murine thigh infection models using b-lactamase-producing strains of Enterobacteriaceae, including those expressing metallo-b-lactamases, ESBLs and serine carbapenemases, with the PK/PD driver of efficacy identified as%T.MIC