7196 research outputs found
Sort by
Engineering an omega-Transaminase for the Efficient Production of a Chiral Sacubitril Precursor
No full textAn omega-transaminase was engineered for the efficient production of a chiral precursor to sacubitril, (2R,4S)-5-([1,1'-biphenyl]-4-yl)-4-amino-2-methylpentanoic acid, a key component in the blockbuster heart failure drug Entresto®. Starting from an enzyme with trace activity and preference for the undesired diastereoisomer, eleven rounds of enzyme evolution were performed. The resultant variant, CDX-043, showed high productivity giving 90% conversion at 75 g/L substrate concentration with 1% enzyme loading with respect to the substrate in 24 h and without the use of an organic co-solvent. The product diastereomeric purity towards the desired (2R,4S)-stereoisomer was > 99:1 d.r. This variant also exhibited high process robustness and could tolerate reaction temperatures up to 65 °C and isopropylamine concentrations of at least 2 M. A structural analysis of the enzyme variants gave insight into how the mutations affected activity and selectivity. This new enzyme variant allows for the efficient and cost-effective production of sacubitril at large scale
Scientific and Regulatory Policy Committee Best Practices*: Documentation of Sexual Maturity by Microscopic Evaluation in Nonclinical Safety Studies
No full textThe sexual maturity status of animals in nonclinical safety studies can have a significant impact on the microscopic assessment of the reproductive system, the interpretation of potential test article–related findings, and ultimately the assessment of potential risk to humans. However, the assessment and documentation of sexual maturity for animals in nonclinical safety studies is not conducted in a consistent manner across the pharmaceutical and chemical industries. The Scientific and Regulatory Policy Committee of the Society of Toxicologic Pathology convened an international working group of pathologists and nonclinical safety scientists with expertise in the reproductive system, pathology nomenclature, and Standard for Exchange of Nonclinical Data requirements. This article describes the best practices for documentation of the light microscopic assessment of sexual maturity in males and females for both rodent and nonrodent nonclinical safety studies. In addition, a review of the microscopic features of the immature, peripubertal, and mature male and female reproductive system and general considerations for study types and reporting are provided to aid the study pathologist tasked with documentation of sexual maturity
Publication Categories in Toxicologic Pathology
No full textToxicologic Pathology is the official journal of the Society of Toxicologic Pathology (STP), the British Society of Toxicological Pathology, and the European STP (ESTP). Toxicologic Pathology publishes articles related to topics in various aspects of toxicologic pathology such as anatomic pathology, clinical pathology, experimental pathology, and biomarker research. Publications include society-endorsed Best Practice/Position and Points to Consider publications and ESTP Expert Workshop articles that are relevant to toxicologic pathology and scientific regulatory processes, Opinion articles under the banner of the STP Toxicologic Pathology Forum, Original Articles, Review Articles (unsolicited/contributed, mini, and invited), Brief Communications, Letters to the Editor, Meeting Reports, and Book Reviews. This article provides details on the various publication categories in Toxicologic Pathology and will serve as a reference for authors and readers
Clinical investigation of metabolic and renal clearance pathways contributing to the elimination of fevipiprant using probenecid as perpetrator
No full textFevipiprant, an oral, non-steroidal, highly selective, reversible, and competitive antagonist of the prostaglandin D2 receptor 2, is eliminated by glucuronidation, and by direct renal excretion. This study aimed to assess the effect of simultaneous UDP-glucuronosyltransferase (UGT) and organic anion transporter (OAT) 3 inhibition by probenecid on the pharmacokinetics of fevipiprant and its acyl glucuronide (AG) metabolite.. This was a single-center, open-label, single sequence, two-period, crossover study in healthy subjects. Liquid chromatography with tandem mass spectrometry was used to measure concentrations of fevipiprant and its AG metabolite in plasma and urine. In the presence of probenecid, the mean maximum concentration of fevipiprant increased approximately 1.7-fold, and the area under the curve (AUC)last and AUCinf increased approximately 2.5-fold, while the mean apparent volume of distribution as well as the AG metabolite-fevipiprant ratio decreased. The apparent systemic clearance decreased by approximately 60% and the renal clearance decreased by approximately 88% in the presence of probenecid. Using the data from this study and previous studies, the relative contribution of OAT and UGT inhibition to the overall effect of probenecid was determined. This allowed to establish a general disposition scheme for fevipiprant and to estimate quantitative contributions for the involved pathways: OATP1B1-mediated hepatic uptake, OAT3-mediated renal excretion and glucuronidation via UGT1A3, UGT2B7 and UGT2B17
Comprehensive review of genotoxicity data for diclofenac
No full textDiclofenac is a non-steroidal anti-inflammatory drug (NSAID) discovered decades ago, which has since been used by an estimated one billion patients, and demonstrated a well acceptable safety profile. In support of its marketing approval, a comprehensive set of genotoxicity studies had been conducted in vitro and in vivo. Despite the fact that these studies preceded both GLP requirements and ICH guidelines on genotoxicity testing, they were conducted using the best scientific principles and are considered appropriate by contemporary standards. In addition to bacterial mutagenicity and mammalian in vitro assays, repeat-dose, germ cell and dominant lethal assays had been conducted. These data are made available for the first time to offer researchers an opportunity to review the existing data set which unequivocally demonstrates that diclofenac is not genotoxic. The lack of a genotoxic potential is further substantiated by long-term bioassay data demonstrating that diclofenac has no carcinogenic potential in rodents. However, more recently, new studies were published showing a genotoxic potential for diclofenac in novel or modified in vitro test systems. These new data are discussed in the context of the existing comprehensive data package
Flow management strategies for a connected purification process
No full textThis paper describes different flow management strategies for a connected purification process which includes two polishing steps, virus filtration (VF) and Tangential Flow Filtration (TFF). Connecting these unit operations avoids introducing large intermediate product pool vessels in small manufacturing facilities. However, a connected-downstream process requires an elaborate control strategy enabling multiple unit operations to function as a single unit. The key strategy to enable the connected-downstream process is a robust management of flow disparities among unit operations. During a typical Ultrafiltration (UF) step, product concentration increases as mass is added to the retentate tank, leading to a permeate flux decline. In a connected-downstream process, the inlet stream is directly connected to the prior unit operation and any decrease in permeate flow rate could cause a flow disparity. Four different flow management approaches are proposed to manage potential flow disparities and their advantages and challenges are discussed. Bench-scale results of these strategies are presented and evaluated. This article is protected by copyright. All rights reserved
Online information discrepancies regarding safety of medicine use during pregnancy and lactation: a ConcePTION study
No full textAbstract
Introduction
Inconsistencies in information concerning the safety of medicines during pregnancy and lactation might result in non-optimal treatment to pregnant and lactating women, subsequent risks to the fetus and to unnecessary weaning from breastfeeding.
Objectives
To analyze information discrepancies concerning medicines during pregnancy and lactation between different on-line sources for patients and health care professionals (HCPs) in four European languages.
Methods
The medicines analyzed were ibuprofen, ondansetron, olanzapine, fingolimod, methylphenidate and adalimumab. A standardized google search was performed in Swedish, Dutch, French and English. The identified recommendations were classified into data source categories, e.g regulatory sources, scientific sources and blogs/forums/social media for patients and for HCPs. The recommendations were compared between the data source categories for each medicine and language: 24 comparisons for pregnancy and 24 for lactation.
Results
For patients, 46% (11/24 comparisons) of the pregnancy recommendations were consistent between all information sources, while for lactation, 17% (4/24) were consistent. The corresponding figures for HCP data sources were 54% (13/24) and 21% (5/24). The regulatory sources were generally more conservative than other sources. Recommendations from five TIS centers (Teratology Information Services) were consistent in 93% (25/27) of the comparisons for pregnancy and 68% (15/22) for lactation.
Conclusion
Discrepancies in online information sources regarding medicines during pregnancy and lactation are common. These differences are more pronounced for lactation than for pregnancy. Recommendations from TIS centers showed better consistency, indicating more consensus on a scientific level. Additional work is needed to harmonize information within and between countries, to avoid conflicting messages
Systematic dissection of transcriptional regulatory networks by genome-scale and singlecell CRISPR screens
No full textNo Abstrac
In vitro transformation assays for non-clinical safety assessment of CRISPR/Cas9 genome-edited cells
No full textOff-target editing is one of the main safety concerns for the use of CRISPR/Cas9 genome editing in gene therapy. Although theoretically rare, these unwanted modifications could lead to malignant transformation, which renders tumorigenicity assessment of the cell therapy product indispensable. Here, we establish two in vitro assays, the soft agar colony forming assay (SACF) and growth in low attachment plates (GILA), as valid, quick and cost-efficient methods for tumorigenicity assessment of genome-edited cells. Using a CRISPR/Cas9 based approach to transform immortalized MCF10A cells, we identified PTPN12, a known tumor suppressor, as first true positive control in GILA and SACF. Next, we assessed the limit of detection for both assays and found that SACF is more sensitive than GILA (0.8% vs. 3.2% transformed cells). We further validated SACF and GILA by identifying a set of positive and negative controls. In contrast to SACF and GILA, an in vivo tumorigenicity study failed to detect the known tumorigenic potential of PTPN12-/- demonstrating the importance of including GILA and SACF in tumorigenicity testing. In conclusion, SACF and GILA are both attractive and valuable additions to non-clinical safety assessment of genome-edited cells
Effect of longevity genetic variants on the molecular aging rate
No full textWe conducted a genome-wide association study of 1320 centenarians from the New England Centenarian Study (median age = 104 years) and 2899 unrelated controls using >9 M genetic variants imputed to the HRC panel of ~65,000 haplotypes. The genetic variants with the most significant associations were correlated to 4131 proteins that were profiled in the serum of a subset of 224 study participants using a SOMAscan array. The genetic associations were replicated in a genome-wide association study of 480 centenarians and ~800 controls of Ashkenazi Jewish descent. The proteomic associations were replicated in a proteomic scan of approximately 1000 Ashkenazi Jewish participants from a third cohort. The analysis replicated a protein signature associated with APOE genotypes and confirmed strong overexpression of BIRC2 (p < 5E−16) and under-expression of APOB in carriers of the APOE2 allele (p < 0.05). The analysis also discovered and replicated associations between longevity variants and slower changes of protein biomarkers of aging, including a novel protein signature of rs2184061 (CDKN2A/CDKN2B in chromosome 9) that suggests a genetic regulation of GDF15. The analyses showed that longevity variants correlate with proteome signatures that could be manipulated to discover healthy-aging targets