Engineering and evaluation of FXa bypassing agents that restore hemostasis following Apixaban associated bleeding.

Direct oral anticoagulants (DOACs) targeting activated factor Xa (FXa) are used to prevent or treat thromboembolic disorders. DOACs reversibly bind to FXa and inhibit its enzymatic activity. However, DOAC treatment carries the risk of anticoagulant-associated bleeding.

Septins Enable T Cell Contact Guidance Amoeboid-Mesenchymal Switch.

Lymphocytes exit circulation and enter in-tissue guided migration toward sites of tissue pathologies, damage, infection, or inflammation. By continuously sensing and adapting to the guiding chemo-mechano-structural properties of the tissues, lymphocytes dynamically alternate and combine their amoeboid (non-adhesive) and mesenchymal (adhesive) migration modes. However, which mechanisms guide and balance different migration modes are largely unclear.

Heparan sulfate promotes ACE2 super-cluster assembly to enhance SARS-CoV-2-associated syncytium formation.

The mechanism of syncytium formation, caused by spike-induced cell-cell fusion in severe COVID-19, is largely unclear. Here we combine chemical genetics with 4D confocal imaging to establish the cell surface heparan sulfate (HS) as a critical host factor exploited by SARS-CoV-2 to enhance spike’s fusogenic activity. HS binds spike to facilitate ACE2 clustering, generating synapse-like cell-cell contacts to promote fusion pore formation.

Implementing computational methods in tandem with synonymous gene recoding for therapeutic development.

Synonymous gene recoding, the substitution of synonymous variants into the genetic sequence, has been used to overcome many production limitations in therapeutic development. However, the safety and efficacy of recoded therapeutics can be difficult to evaluate because synonymous codon substitutions can result in subtle, yet impactful changes in protein features and require sensitive methods for detection. Given that computational approaches have made significant leaps in recent years, we propose that machine-learning (ML) tools may be leveraged to assess gene-recoded therapeutics and foresee an opportunity to adapt codon contexts to enhance some powerful existing tools.

Synonymous Variants: Necessary Nuance in Our Understanding of Cancer Drivers and Treatment Outcomes.

Once called "silent mutations" and assumed to have no effect on protein structure and function, synonymous variants are now recognized to be drivers for some cancers. There have been significant advances in our understanding of the numerous mechanisms by which synonymous single nucleotide variants (sSNVs) can affect protein structure and function by affecting pre-mRNA splicing, mRNA expression, stability, folding, micro-RNA binding, translation kinetics, and co-translational folding. This review highlights the need for considering sSNVs in cancer biology to gain a better understanding of the genetic determinants of human cancers and to improve their diagnosis and treatment.

Structural, functional, and immunogenicity implications of F9 gene recoding.

Hemophilia B is a blood clotting disorder caused by deficient activity of coagulation factor IX (FIX). Multiple recombinant FIX proteins are currently approved to treat hemophilia B, and several gene therapy products are currently being developed. Codon optimization is a frequently used technique in the pharmaceutical industry to improve recombinant protein expression by recoding a coding sequence using multiple synonymous codon substitutions.

Analyzing the Role of Gut Microbiota on the Onset of Autoimmune Diseases Using TNF Murine Model.

Very little is known about disease transmission via the gut microbiome. We hypothesized that certain inflammatory features could be transmitted via the gut microbiome and tested this hypothesis using an animal model of inflammatory diseases. Twelve-week-old healthy C57 Bl/6 and Germ-Free (GF) female and male mice were fecal matter transplanted (FMT) under anaerobic conditions with TNF donors exhibiting spontaneous Rheumatoid Arthritis (RA) and Inflammatory Bowel Disease (IBD) or with conventional healthy mice control donors.

Heterogeneity in treatment effects across diverse populations.

Differences in patient characteristics, including age, sex, and race influence the safety and effectiveness of drugs, biologic products, and medical devices. Here we provide a summary of the topics discussed during the opening panel at the 2018 Johns Hopkins Center for Excellence in Regulatory Science and Innovation symposium on Assessing and Communicating Heterogeneity of Treatment Effects for Patient Subpopulations: Challenges and Opportunities. The goal of this session was to provide a brief overview of FDA-regulated therapeutics, including drugs, biologics and medical devices, and some of the major sources of heterogeneity of treatment effects (HTE) related to patient demographics, such as age, sex and race.

Distinct signatures of codon and codon pair usage in 32 primary tumor types in the novel database CancerCoCoPUTs for cancer-specific codon usage.

Background: Gene expression is highly variable across tissues of multi-cellular organisms, influencing the codon usage of the tissue-specific transcriptome. Cancer disrupts the gene expression pattern of healthy tissue resulting in altered codon usage preferences. The topic of codon usage changes as they relate to codon demand, and tRNA supply in cancer is of growing interest.

Characterization of the β-KDO Transferase KpsS, the Initiating Enzyme in the Biosynthesis of the Lipid Acceptor for Polysialic Acid.

Antiphagocytic capsular polysaccharides are key components of effective vaccines against pathogenic bacteria. groups B and C, as well as serogroups K1 and K92, are coated with polysialic acid capsules. Although the chemical structure of these polysaccharides and the organization of the associated gene clusters have been described for many years, only recently have the details of the biosynthetic pathways been discovered.

Statistical methods of screening cut point determination in immunogenicity studies.

Currently, screening cut point (CP) calculated from an assay validation with replicates are applied to an immunogenicity study with nonreplicates, for which the antidrug antibodies rate is determined. IID treats the replicate of a sample as coming from another independent sample. AVE uses average results from each sample across runs but inter-assay variability is reduced.

Coagulopathy and Thrombosis as a Result of Severe COVID-19 Infection: A Microvascular Focus.

Coronavirus disease of 2019 (COVID-19) is the clinical manifestation of the respiratory infection caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). While primarily recognized as a respiratory disease, it is clear that COVID-19 is systemic illness impacting multiple organ systems. One defining clinical feature of COVID-19 has been the high incidence of thrombotic events.

In silico features of ADAMTS13 contributing to plasmatic ADAMTS13 levels in neonates with congenital heart disease.

Introduction: Risk factors contributing to heightened thrombosis in pediatric congenital heart disease (CHD) patients are not fully understood. Among the neonatal CHD population, those presenting with single ventricular physiology are at the highest risk for perioperative thrombosis. The von Willebrand factor and ADAMTS13 interactions have emerged as causative risk factors for pediatric stroke and could contribute to heightened thrombosis in CHD neonates.

Amyloid precursor-like protein 2 interacts with claudin-7 and affects ovarian cancer cell survival.

Aim: In our previous report, we identified roles of CLDN7 in regulation of cell signaling. The goal of this study was to identify proteins interacting with CLDN7 in ovarian cancer.

Methods: The yeast two-hybrid system was used to identify proteins directly interacting with CLDN7 and cell survival was tested using colony formation assay.

A Single Synonymous Variant (c.354G>A [p.P118P]) in Confers Enhanced Specific Activity.

Synonymous variants within coding regions may influence protein expression and function. We have previously reported increased protein expression levels ex vivo (~120% in comparison to wild-type) from a synonymous polymorphism variant, c.354G>A [p.

Development of a high-throughput assay to measure measles neutralizing antibodies.

Measles virus is highly infectious and remains a leading cause of vaccine preventable deaths in children. Neutralizing antibody responses elicited by measles virus infection or immunization are a serological correlate of protection. We describe a high-throughput neutralization assay to improve surveillance for measles immunity.

Splicing dysregulation contributes to the pathogenicity of several F9 exonic point variants.

Background: Pre-mRNA splicing is a complex process requiring the identification of donor site, acceptor site, and branch point site with an adjacent polypyrimidine tract sequence. Splicing is regulated by splicing regulatory elements (SREs) with both enhancer and suppressor functions. Variants located in exonic regions can impact splicing through dysregulation of native splice sites, SREs, and cryptic splice site activation.

Subcellular compartmentalization of PKM2 identifies anti-PKM2 therapy response in vitro and in vivo mouse model of human non-small-cell lung cancer.

Pyruvate kinase M2 (PKM2) is an alternatively spliced variant, which mediates the conversion of glucose to lactate in cancer cells under normoxic conditions, known as the Warburg effect. Previously, we demonstrated that PKM2 is one of 97 genes that are overexpressed in non-small-cell lung cancer (NSCLC) cell lines. Herein, we demonstrate a novel role of subcellular PKM2 expression as a biomarker of therapeutic response after targeting this gene by shRNA or small molecule inhibitor (SMI) of PKM2 enzyme activity in vitro and in vivo.

Contradictory to its effects on thrombin, C1-inhibitor reduces plasmin generation in the presence of thrombomodulin.

C1-inhibitor (C1INH) was shown to enhance thrombin generation (TG) in the presence of thrombomodulin (TM) by reducing production of activated protein C. Because C1INH is known to inhibit fibrinolytic system proteases, the objective of this study was to evaluate the effect of moderate (3 IU/ml) and high (16 IU/ml) C1INH concentrations on TG and plasmin generation (PG) in the presence of TM. These concentrations were evaluated based on expected maximum plasma levels following C1INH replacement therapy and recently suggested supraphysiologic dosing.

Combination immunotherapy with IL-4 Pseudomonas exotoxin and IFN-α and IFN-γ mediate antitumor effects in vitro and in a mouse model of human ovarian cancer.

Aim: We have shown that IL-4 fused to Pseudomonas exotoxin (IL-4-PE) is cytotoxic to ovarian cancer cell lines. The antineoplastic properties of IFN-α, IFN-γ and IL-4-PE have been studied and showed some promise in the clinical trials. Here, we investigated whether the combination of IL-4-PE, IFN-α and IFN-γ will result in increased ovarian cancer cell death in vitro and in vivo.

Rapid selection of single-stranded DNA aptamers binding Staphylococcus epidermidis in platelet concentrates.

Staphylococcus epidermidis is the most common transfusion-associated pathogen contaminating platelet concentrates. Methods to reduce or eliminate contaminating bacteria from platelet units are critical for improving the safety of blood transfusions. We used rapid isolation of DNA aptamers (RIDA) to identify single-stranded (ss)DNA aptamers as ligands that specifically bind to S.

Livestock abundance predicts vampire bat demography, immune profiles and bacterial infection risk.

Human activities create novel food resources that can alter wildlife-pathogen interactions. If resources amplify or dampen, pathogen transmission probably depends on both host ecology and pathogen biology, but studies that measure responses to provisioning across both scales are rare. We tested these relationships with a 4-year study of 369 common vampire bats across 10 sites in Peru and Belize that differ in the abundance of livestock, an important anthropogenic food source.

Novel hemotropic mycoplasmas are widespread and genetically diverse in vampire bats.

Bats (Order: Chiroptera) have been widely studied as reservoir hosts for viruses of concern for human and animal health. However, whether bats are equally competent hosts of non-viral pathogens such as bacteria remains an important open question. Here, we surveyed blood and saliva samples of vampire bats from Peru and Belize for hemotropic Mycoplasma spp.