Rapid LC-MS/MS Evaluation of Collagen and Elastin Crosslinks in Human and Mouse Lung Tissue with a Novel Bioanalytical Surrogate Matrix Approach.

Alterations to post-translational crosslinking modifications in the extracellular matrix (ECM) are known to drive the pathogenesis of fibrotic diseases, including idiopathic pulmonary fibrosis (IPF). Thus, the methodology for measuring crosslinking dynamics is valuable for understanding disease progression. The existing crosslinking analysis sample preparation and liquid chromatography tandem mass spectrometry (LC-MS/MS) methods are typically labor-intensive and time-consuming which limits throughput.

Trends and Challenges of the Modern Pathology Laboratory for Biopharmaceutical Research Excellence.

Pathology, a fundamental discipline that bridges basic scientific discovery to the clinic, is integral to successful drug development. Intrinsically multimodal and multidimensional, anatomic pathology continues to be empowered by advancements in molecular and digital technologies enabling the spatial tissue detection of biomolecules such as genes, transcripts, and proteins. Over the past two decades, breakthroughs in spatial molecular biology technologies and advancements in automation and digitization of laboratory processes have enabled the implementation of higher throughput assays and the generation of extensive molecular data sets from tissue sections in biopharmaceutical research and development research units.

Enhancing aggregate reduction using anion exchange hybrid filter in an immunocytokine diabody fusion protein purification process.

Significant aggregation challenges were encountered during the manufacturing processing of an immunocytokine diabody fusion protein. To mitigate this issue and reduce aggregates in the purification process, a variety of depth filters were evaluated for harvest clarification and post-Protein A intermediate filtration. Emphaze™ AEX Hybrid Purifier, a fully synthetic quaternary amine functionalized anion exchange (AEX) nonwoven filter, was found to be particularly effective in removing aggregates during both harvest clarification and post-Protein A intermediate filtration steps.

Limitation of anion exchange chromatography and potential application of hydrophobic interaction chromatography for monitoring AAV9 capsid degradation upon thermal stress.

Adeno-Associated Virus (AAV) is often selected as the vector of choice for gene therapy due to its superior clinical performance compared to other gene delivery systems. Currently the characterization of AAV degradation, especially the chemical degradation of capsid, has been limited due to lack of suitable methods. Our study using AAV9 as a model molecule shows that anion exchange chromatography (AEX) as a charge-based separation method has limitations in monitoring the chemical degradation of AAV9 capsid due to a confounding effect from DNA cargo ejection.

Use of MM-SES-CD Endoscopic Improvement Thresholds Enhances Effect Size Differentiation between Adalimumab vs. Placebo: A Post-hoc Analysis of the EXTEND Trial.

Introduction: The Modified Multiplier of the SES-CD (MM-SES-CD) refines the assessment of endoscopic Crohn's Disease (CD) severity by differentially weighting parameters in the original SES-CD. A threshold of <22.5 for MM-SES-CD suggests endoscopic remission and correlates with a low risk of long-term disease progression.

Development and validation of an ultra-performance liquid chromatography with tandem mass spectrometry method for determination of soluble repulsive guidance molecule A in human serum and cerebrospinal fluid.

Repulsive guidance molecule A (RGMa) is upregulated in neurodegenerative diseases. To assess RGMa levels in human serum and cerebrospinal fluid (CSF), a quantification method was developed and validated according to ICH M10 guideline. Sample preparation consisted of immunoprecipitation (IP, only for serum), digestion and purification followed by MS.

A Protocol for the Inclusion of Minoritized Persons in Alzheimer Disease Research From the ADNI3 Diversity Taskforce.

Importance: Black or African American (hereinafter, Black) and Hispanic or Latino/a/x (hereinafter, Latinx) adults are disproportionally affected by Alzheimer disease, but most research studies do not enroll adequate numbers of both of these populations. The Alzheimer's Disease Neuroimaging Initiative-3 (ADNI3) launched a diversity taskforce to pilot a multipronged effort to increase the study inclusion of Black and Latinx older adults.

Objective: To describe and evaluate the culturally informed and community-engaged inclusion efforts to increase the screening and enrollment of Black and Latinx older adults in ADNI3.

Assessing the Accuracy and Efficiency of Free Energy Differences Obtained from Flow-Based Probabilistic Generative Models.

Computing free energy differences between metastable states characterized by nonoverlapping Boltzmann distributions is often a computationally intensive endeavor, usually requiring chains of intermediate states to connect them. Targeted free energy perturbation (TFEP) can significantly lower the computational cost of FEP calculations by choosing a set of invertible maps used to directly connect the distributions of interest, achieving the necessary statistically significant overlaps without sampling any intermediate states. Probabilistic generative models (PGMs) based on normalizing flow architectures can make it much easier via machine learning to train invertible maps needed for TFEP.

Modulating lung fibroblast activation via senolysis of senescent human alveolar epithelial cells.

Idiopathic pulmonary fibrosis (IPF) is an age-related disease with poor prognosis and limited therapeutic options. Activation of lung fibroblasts and differentiation to myofibroblasts are the principal effectors of disease pathology, but damage and senescence of alveolar epithelial cells, specifically type II (ATII) cells, has recently been identified as a potential trigger event for the progressive disease cycle. Targeting ATII senescence and the senescence-associated secretory phenotype (SASP) is an attractive therapeutic strategy; however, translatable primary human cell models that enable mechanistic studies and drug development are lacking.

Understanding mechanisms of JAK1 inhibition on synovial fibroblasts using combinatorial approaches of bulk and single cell RNAseq analyses.

Objectives: The aim of these studies was to characterise the molecular effects of a tool JAK1 inhibitor on cultured primary fibroblast-like synoviocytes (FLS) from patients with rheumatoid arthritis (RA) through both total and individual cell analysis.

Methods: RA-FLS cultures from 6 (Bulk RNA-seq) or 4 (ScRNA-seq) donors were pre-treated with various concentrations (100 nM and 1μM) of ABT-317 with/without exposure to 25% SEB-conditioned PBMC medium to mimic the RA inflammatory milieu. Cells were subjected to both bulk RNA-seq (36 libraries) and single cell RNA-seq (scRNA-seq; 24 libraries) to identify biological processes impacted by CM and ABT-317 treatments.

Insight Into the Degradation Pathways of an AAV9.

The use of recombinant adeno-associated virus (AAV) vectors is a popular choice for in vivo gene therapy, with hundreds of ongoing clinical trials targeting various genetic diseases. However, due to limited material availability and the complexity of AAV structure, there is a critical lack of comprehensive studies on AAV degradation pathways. In this study, we intended to elucidate the degradation pathways for a model AAV9 with GFP as the transgene under relevant stressed conditions.

Agonistic anti-DCIR antibody inhibits ITAM-mediated inflammatory signaling and promotes immune resolution.

DC inhibitory receptor (DCIR) is a C-type lectin receptor selectively expressed on myeloid cells, including monocytes, macrophages, DCs, and neutrophils. Its role in immune regulation has been implicated in murine models and human genome-wide association studies, suggesting defective DCIR function associates with increased susceptibility to autoimmune diseases such as rheumatoid arthritis, lupus, and Sjögren's syndrome. However, little is known about the mechanisms underlying DCIR activation to dampen inflammation.

Broad proteomics analysis of seeding-induced aggregation of α-synuclein in M83 neurons reveals remodeling of proteostasis mechanisms that might contribute to Parkinson's disease pathogenesis.

Aggregation of misfolded α-synuclein (α-syn) is a key characteristic feature of Parkinson's disease (PD) and related synucleinopathies. The nature of these aggregates and their contribution to cellular dysfunction is still not clearly elucidated. We employed mass spectrometry-based total and phospho-proteomics to characterize the underlying molecular and biological changes due to α-syn aggregation using the M83 mouse primary neuronal model of PD.