Precision analysis of mutant U2AF1 activity reveals deployment of stress granules in myeloid malignancies.

Splicing factor mutations are common among cancers, recently emerging as drivers of myeloid malignancies. U2AF1 carries hotspot mutations in its RNA-binding motifs; however, how they affect splicing and promote cancer remain unclear. The U2AF1/U2AF2 heterodimer is critical for 3' splice site (3'SS) definition. To specifically unmask changes in U2AF1 function in vivo, we developed a crosslinking and immunoprecipitation procedure that detects contacts between U2AF1 and the 3'SS AG at single-nucleotide resolution. Our data reveal that the U2AF1 S34F and Q157R mutants establish new 3'SS contacts at -3 and +1 nucleotides, respectively. These effects compromise U2AF2-RNA interactions, resulting predominantly in intron retention and exon exclusion. Integrating RNA binding, splicing, and turnover data, we predicted that U2AF1 mutations directly affect stress granule components, which was corroborated by single-cell RNA-seq. Remarkably, U2AF1-mutant cell lines and patient-derived MDS/AML blasts displayed a heightened stress granule response, pointing to a novel role for biomolecular condensates in adaptive oncogenic strategies.

Disease modification in chronic spontaneous urticaria.

Chronic spontaneous urticaria (CSU) is a debilitating, inflammatory skin condition characterized by infiltrating immune cells. Available treatments are limited to improving the signs and symptoms. There is an unmet need to develop therapies that target disease-driving pathways upstream of mast cell activation to inhibit or delay the progression of CSU and associated comorbidities. Here, we aim to define disease modification due to a treatment intervention and criteria that disease-modifying treatments (DMTs) must meet in CSU. We have defined disease modification in CSU as a favorable treatment-induced change in the underlying pathophysiology and, therefore, the disease course, which is clinically beneficial and enduring. A DMT must fulfil the following criteria: (1) prevents or delays the progression of CSU, (2) induces long-term, therapy-free clinical remission, which is the sustained absence of CSU signs and symptoms without the need for treatment, and (3) affects the underlying mechanism of CSU, as demonstrated by an effect on disease-driving signals and/or a biomarker. DMTs in CSU should slow disease progression, achieve long-lasting disease remission, target disease-driving mechanisms, reduce mast cell-activating IgE autoantibodies, target cytokine profile polarization, and normalize the gut microbiome and barrier. Treating CSU at the immune system level could provide valuable alternatives to pharmacotherapy in CSU management. Specific DMTs in CSU are yet to be developed, but some show potential benefits, such as inhibitors of Bruton's Tyrosine Kinase, IL-4 and IL-13. Future therapies could prevent CSU signs and symptoms, achieve long-term clinical benefits after discontinuing treatment, and prevent associated concomitant disorders.

New insights into chronic inducible urticaria.

PURPOSE OF REVIEW: Chronic inducible urticaria (CIndU) is a group of long-persisting and challenging to manage diseases, characterized by recurrent wheals and angioedema induced by definite triggers. In this review, we address recent findings on CIndU pathogenesis, diagnosis as well as its treatment, and we discuss novel potential targets that may lead to the development of more effective therapies for CIndU patients. RECENT ADVANCES: Meaningful advances in the understanding of its pathogenesis have been reported in the last decades. Novel CIndU-specific patient-reported outcome measures enable a closer and better evaluation of patients. CIndU is a hard-to-treat disease that highly impairs quality of life (QoL) of affected patients. Provocation tests allow to diagnose CIndU subtypes. The only licensed and recommended treatment for CIndU are second generation non-sedating H1-antihistamines, which lack efficacy in many cases. Omalizumab off-label use has been assessed in all types of CIndU with overall good outcomes. Promising emerging therapies currently assessed in chronic spontaneous urticaria are paving the path for novel treatments for CIndU.

Internally controlled RNA sequencing comparisons using nucleoside recoding chemistry.

Quantitative comparisons of RNA levels from different samples can lead to new biological understanding if they are able to distinguish biological variation from variable sample preparation. These challenges are pronounced in comparisons that require complex biochemical manipulations (e.g. isolating polysomes to study translation). Here, we present Transcript Regulation Identified by Labeling with Nucleoside Analogues in Cell Culture (TILAC), an internally controlled approach for quantitative comparisons of RNA content. TILAC uses two metabolic labels, 4-thiouridine (s4U) and 6-thioguanosine (s6G), to differentially label RNAs in cells, allowing experimental and control samples to be pooled prior to downstream biochemical manipulations. TILAC leverages nucleoside recoding chemistry to generate characteristic sequencing signatures for each label and uses statistical modeling to compare the abundance of RNA transcripts between samples. We verified the performance of TILAC in transcriptome-scale experiments involving RNA polymerase II inhibition and heat shock. We then applied TILAC to quantify changes in mRNA association with actively translating ribosomes during sodium arsenite stress and discovered a set of transcripts that are translationally upregulated, including MCM2 and DDX5. TILAC is broadly applicable to uncover differences between samples leading to improved biological insights.

Implementation of patient priorities-aligned care in a home-based primary care program.

BACKGROUND: Older adults may be limited in their ability to access care that meets their health goals owing to disease burden, financial instability, and psychosocial barriers. A home-based primary care (HBPC) program established in 2020 within a large family medicine practice uses the Patient Priorities Care (PPC) approach to identify and address patients' health priorities. When incorporated as part of the HBPC model of care, the PPC approach has the potential to enhance person-centered care for older adults in a way that best supports their health goals. OBJECTIVE: The objective of this study is to summarize common recommendations for alignment of care with patients' health outcome goals after implementation of the PPC approach in an HBPC population. METHODS: This retrospective study was exempt from review by an institutional review board. After enrollment in the HBPC program, patients participated in a PPC priorities identification conversation to identify their health outcome goals and care preferences. Through chart review, 2 researchers independently categorized these goals based on the set of values they most reflect: connecting, managing health, enjoying life, and functioning. Aspects of care in place before enrollment in HBPC were considered to determine any adjustments that needed to be made to align care with patients' identified priorities. RESULTS: The most common value associated with patients' most desired health outcome goal was functioning (n = 33, 66%). For secondary and tertiary health outcome goals, the most common value identified was managing health (secondary, n = 28, 56%; tertiary, n = 22, 44%). Common recommendations made to align care with patients' identified priorities included stopping potentially harmful medications, starting medications for untreated conditions, starting physical or occupational therapy, and adjusting medications. CONCLUSION: Through the PPC approach, patients' values were identified and care was assessed to aid in attainment of individualized health outcome goals and tailor care to What Matters most.

Anti-KIT antibody, barzolvolimab, reduces skin mast cells and disease activity in chronic inducible urticaria.

BACKGROUND: Chronic inducible urticaria (CIndU) is characterized by mast cell (MC)-mediated wheals in response to triggers: cold in cold urticaria (ColdU) and friction in symptomatic dermographism (SD). KIT receptor activation by stem cell factor (SCF) is essential for MC function. Barzolvolimab (CDX-0159) is a humanized antibody that inhibits KIT activation by SCF and was well tolerated in healthy volunteers with dose-dependent plasma tryptase suppression indicative of systemic mast cell ablation. METHODS: This is an open-label, trial in patients with antihistamine refractory ColdU or SD, receiving one IV dose of barzolvolimab (3 mg/kg), with a 12-week follow-up. Primary endpoint was safety/tolerability; pharmacodynamic (PD)/clinical endpoints included serum tryptase, plasma SCF, skin MC histology, provocation tests, urticaria control test (UCT), and dermatology life quality index (DLQI). RESULTS: Analysis populations were safety (n = 21) and pharmacodynamics/clinical activity (n = 20). Barzolvolimab was well tolerated; most adverse events were mild and resolved. Treatment resulted in significant depletion of skin MCs, decreased tryptase (

Using Wearable Sensors to Measure Goal Achievement in Older Veterans with Dementia.

Aligning treatment with patients' self-determined goals and health priorities is challenging in dementia care. Wearable-based remote health monitoring may facilitate determining the active participation of individuals with dementia towards achieving the determined goals. The present study aimed to demonstrate the feasibility of using wearables to assess healthcare goals set by older adults with cognitive impairment. We present four specific cases that assess (1) the feasibility of using wearables to monitor healthcare goals, (2) differences in function after goal-setting visits, and (3) goal achievement. Older veterans (n = 17) with cognitive impairment completed self-report assessments of mobility, then had an audio-recorded encounter with a geriatrician and wore a pendant sensor for 48 h. Follow-up was conducted at 4-6 months. Data obtained by wearables augments self-reported data and assessed function over time. Four patient cases illustrate the utility of combining sensors, self-report, notes from electronic health records, and visit transcripts at baseline and follow-up to assess goal achievement. Using data from multiple sources, we showed that the use of wearable devices could support clinical communication, mainly when patients, clinicians, and caregivers work to align care with the patient's priorities.

TimeLapse-seq: adding a temporal dimension to RNA sequencing through nucleoside recoding.

RNA sequencing (RNA-seq) offers a snapshot of cellular RNA populations, but not temporal information about the sequenced RNA. Here we report TimeLapse-seq, which uses oxidative-nucleophilic-aromatic substitution to convert 4-thiouridine into cytidine analogs, yielding apparent U-to-C mutations that mark new transcripts upon sequencing. TimeLapse-seq is a single-molecule approach that is adaptable to many applications and reveals RNA dynamics and induced differential expression concealed in traditional RNA-seq.

Expanding the Nucleoside Recoding Toolkit: Revealing RNA Population Dynamics with 6-Thioguanosine.

RNA-sequencing (RNA-seq) measures RNA abundance in a biological sample but does not provide temporal information about the sequenced RNAs. Metabolic labeling can be used to distinguish newly made RNAs from pre-existing RNAs. Mutations induced from chemical recoding of the hydrogen bonding pattern of the metabolic label can reveal which RNAs are new in the context of a sequencing experiment. These nucleotide recoding strategies have been developed for a single uridine analogue, 4-thiouridine (s4U), limiting the scope of these experiments. Here we report the first use of nucleoside recoding with a guanosine analogue, 6-thioguanosine (s6G). Using TimeLapse sequencing (TimeLapse-seq), s6G can be recoded under RNA-friendly oxidative nucleophilic-aromatic substitution conditions to produce adenine analogues (substituted 2-aminoadenosines). We demonstrate the first use of s6G recoding experiments to reveal transcriptome-wide RNA population dynamics.

Impact of Patient-Centered Medical Home Implementation on Diabetes Control in the Veterans Health Administration.

BACKGROUND: Given its widespread dissemination across primary care, the Veterans Health Administration (VA) is an ideal setting to examine the impact of the patient-centered medical home (PCMH) on diabetes outcomes. OBJECTIVE: To assess the impact of PCMH implementation on diabetes outcomes among patients receiving care in the Veterans Health Administration. DESIGN: Retrospective cohort analysis and multilevel logistic regression. PATIENTS: Twenty thousand eight hundred fifty-eight patients in one Midwest VA network who had a diabetes diagnosis in both 2009 and 2012 and who received primary care between October 1, 2008 and September 30, 2009. MAIN MEASURES: Glycemic and lipid control using VA quality indicators [hemoglobin (Hb) A1c < 9%, low-density lipoprotein cholesterol (LDL-C) < 100 mg/dL]. KEY RESULTS: Odds of glycemic control were lower in 2012 than 2009 (OR = 0.72, 95% CI = 0.67-0.77, p < 0.001), and this change in control over time varied by race (OR of the interaction between time and race = 1.18, 95% CI = 1.02-1.36, p = 0.028). While the disparity in glycemic control between white and black patients persisted post-PCMH, the magnitude of the disparity was smaller in 2012 compared to 2009 (2012: OR = 1.32, 95% CI = 1.18-1.47, p < 0.0001 and 2009: OR = 1.59, 95% CI = 1.39-1.82, p < 0.0001). Odds of lipid control did not significantly change between 2009 and 2012 and change did not vary by race and/or gender. CONCLUSIONS: Although there were no significant improvements in odds of lipid control, and odds of glycemic control decreased following PCMH implementation, there was evidence of reduced racial disparities in glycemic control post-PCMH implementation.

Building a primary care/research partnership: lessons learned from a telehealth intervention for diabetes and depression.

INTRODUCTION: Evidence-based interventions are often poorly translated into primary care settings due to inadequate integration into organizational cultures and clinical workflows. Study designs that blend evaluation of effectiveness and implementation may enhance uptake of interventions into primary care settings. Community-Based Participatory Research (CBPR) models are useful for developing partnerships between research teams and primary care clinical partners to test blended study designs. METHODS: We conducted a formative evaluation of partnership building between a health services research team and a primary care community in US Veterans Affairs Health System to conduct a randomized effectiveness trial of an intervention embedded in routine primary care. The formative evaluation used qualitative data drawn from research/clinical partnership meetings. Data were coded and analysed using qualitative framework analysis. RESULTS: The CBPR model guided development of a research/clinical partnership based on a facilitation team consisting of 'external facilitators' (research team), 'internal facilitators' (primary care leadership) and a 'clinical advisory committee' drawn from the primary care community. Qualitative themes focused on: how the intervention components ('evidence') aligned with local clinical cultures, barriers and facilitators to acceptance and adoption of the intervention processes within the context of clinical workflows and identified 'facilitators' of intervention uptake and sustainability. CONCLUSION: A CBPR model can guide the development of research/clinical partnerships. Partnerships can identify barriers and craft modifications to intervention procedures that promote integration and into primary care workflows. Formative research/clinical partnerships are critical for designing and testing interventions focused on implementation and sustainability of new evidence within routine primary care.

Tuning cohesin trajectories enables differential readout of the Pcdhα cluster across neurons.

Expression of Protocadherin (Pcdh) genes is critical to the generation of neuron identity and wiring of the nervous system. Pcdhα genes are arranged in clusters and exhibit a range of expression profiles, from stochastic to deterministic. Because Pcdhα promoters have high sequence identity and share distal enhancers, how distinct neurons choose which gene to express remains unclear. We show that the interplay between multiple enhancers, epigenetics, and genome folding orchestrates differential readouts of the locus across neurons. The probability of Pcdhα promoter choice depends on enhancer/promoter encounters catalyzed by cohesin, whose extrusion trajectories determine the likelihood that an individual promoter can "escape" heterochromatin-mediated silencing. We propose that tunable locus-specific regulatory elements and cell type-specific cohesin activity underlie the generation of cellular diversity by Pcdh genes.

An innovative approach to aligning healthcare with what matters most to patients: A hybrid type 1 trial protocol of patient priorities care for older adults with multiple chronic conditions.

BACKGROUND: Providing healthcare for older adults with multiple chronic conditions (MCC) is challenging. Polypharmacy and complex treatment plans can lead to high treatment burden and risk for adverse events. For clinicians, managing the complexities of patients with MCC leaves little room to identify what matters and align care options with patients' health priorities. New care approaches are needed to navigate these challenges. In this clinical trial, we evaluate implementation and effectiveness outcomes of an innovative, structured, patient-centered care approach (Patient Priorities Care; PPC) for reducing treatment burden and aligning health care decisions with the health priorities of older adults with MCC. METHODS: This is a multisite, assessor-blind, two-arm, parallel hybrid type 1 randomized controlled trial. We are enrolling 396 older (65+) Veterans with MCC who receive primary care at the Veterans Affairs Medical Center. Veterans are randomly assigned to either PPC or usual care. In the PPC arm, Veterans have a brief telephone call with a study facilitator to identify their personal health priorities. Then, primary care providers use this information to align healthcare with Veteran priorities during their established clinic appointments. Data are collected at baseline and 4-month follow up to assess for changes in treatment burden and use of home and community services. Formative and summative evaluations are also collected to assess for implementation outcomes according to Proctor's implementation framework. CONCLUSIONS: This work has the potential to significantly improve the standard of care by personalizing healthcare and helping patients achieve what is most important to them.

Multinational Drug Survival Study of Omalizumab in Patients With Chronic Urticaria and Potential Predictors for Discontinuation.

Importance: Treating patients with chronic urticaria using omalizumab has been shown to be safe and effective in randomized clinical trials. Multinational studies on long-term omalizumab performance in chronic urticaria in clinical practice settings are lacking, especially on drug survival. Drug survival, which refers to the length of time that patients are treated with a specific drug, is a comprehensive outcome covering effectiveness, safety, and patient and physician preferences. Furthermore, little is known about the reasons and potential predictors for omalizumab discontinuation. Objective: To investigate omalizumab drug survival as well as reasons and potential predictors for discontinuation in a large, diverse population. Design, Setting, and Participants: This international multicenter cohort study was conducted at 14 Urticaria Centers of Reference and Excellence in 10 countries, including all patients with chronic urticaria from these centers who were ever treated with omalizumab. Main Outcomes and Measures: Drug survival analysis was performed to assess time to discontinuation. Patient characteristics and treatment protocols were investigated by Cox regression analysis to identify potential predictors for omalizumab discontinuation. Results: In 2325 patients with chronic urticaria who started omalizumab between June 2009 and July 2022, the mean (SD) age of the cohort was 42 (6) years, and 1650 participants (71%) were female. Overall omalizumab survival rates decreased from 76% to 39% after 1 to 7 years, respectively (median survival time, 3.3 [95 % CI, 2.9-4.0] years), primarily due to discontinuation from well-controlled disease in 576 patients (65%). Ineffectiveness and adverse effects were reasons for discontinuation in a far smaller proportion of patients, totaling 164 patients (18%) and 31 patients (4%), respectively. Fast treatment response was associated with higher rates of omalizumab discontinuation due to well-controlled disease (hazard ratio, 1.45 [95% CI, 1.20-1.75]), and disease duration of more than 2 years was associated with lower rates of discontinuation due to well-controlled disease (HR, 0.81 [95% CI, 0.67-0.98]). Immunosuppressive cotreatment at the start of omalizumab and autoimmune disease was associated with a higher risk for discontinuation due to ineffectiveness (HR, 1.65 [95% CI, 1.12-2.42]). The presence of spontaneous wheals (HR, 0.62 [95% CI, 0.41-0.93]) and access to higher dosages (HR, 0.40 [95% CI, 0.27-0.58) were both associated with a lower risk for discontinuation of omalizumab due to ineffectiveness. Conclusion and Relevance: This multinational omalizumab drug survival cohort study demonstrated that treatment of chronic urticaria with omalizumab in a clinical setting is effective and safe, and well-controlled disease is the main reason for treatment discontinuation. These findings on omalizumab drug survival rates and reasons and potential predictors for discontinuation may guide patients and physicians in clinical decision-making and expectation management. These results may call for the identification of biomarkers for chronic urticaria remission in complete responders to omalizumab treatment.

Guiding Post-Hospital Recovery by 'What Matters:' Implementation of Patient Priorities Identification in a VA Community Living Center.

BACKGROUND: Patient priorities care (PPC) is an effective age-friendly health systems (AFHS) approach to aligning care with goals derived from 'what matters'. The purpose of this quality improvement program was to evaluate the fidelity and feasibility of the health priorities identification (HPI) process in VA Community Living Centers (CLC). METHODS: PPC experts worked with local CLC staff to guide the integration of HPI into the CLC and utilized a Plan-Do-Study-Act (PDSA) model for this quality improvement project. PPC experts reviewed health priorities identification (HPI) encounters and interdisciplinary team (IDT) meetings for fidelity to the HPI process of PPC. Qualitative interviews with local CLC staff determined the appropriateness of the health priorities identification process in the CLC. RESULTS: Over 8 months, nine facilitators completed twenty HPI encounters. Development of a Patient Health Priorities note template, staff education and PPC facilitator training improved fidelity and documentation of HPI encounters in the electronic health record. Facilitator interviews suggested that PPC is appropriate in this setting, not burdensome to staff and fostered a person-centered approach to AFHS. CONCLUSIONS: The HPI process is an acceptable and feasible approach to ask the 'what matters' component of AFHS in a CLC setting.

Improving the study of RNA dynamics through advances in RNA-seq with metabolic labeling and nucleotide-recoding chemistry.

RNA metabolic labeling using 4-thiouridine (s4U) captures the dynamics of RNA synthesis and decay. The power of this approach is dependent on appropriate quantification of labeled and unlabeled sequencing reads, which can be compromised by the apparent loss of s4U-labeled reads in a process we refer to as dropout. Here we show that s4U-containing transcripts can be selectively lost when RNA samples are handled under sub-optimal conditions, but that this loss can be minimized using an optimized protocol. We demonstrate a second cause of dropout in nucleotide recoding and RNA sequencing (NR-seq) experiments that is computational and downstream of library preparation. NR-seq experiments involve chemically converting s4U from a uridine analog to a cytidine analog and using the apparent T-to-C mutations to identify the populations of newly synthesized RNA. We show that high levels of T-to-C mutations can prevent read alignment with some computational pipelines, but that this bias can be overcome using improved alignment pipelines. Importantly, kinetic parameter estimates are affected by dropout independent of the NR chemistry employed, and all chemistries are practically indistinguishable in bulk, short-read RNA-seq experiments. Dropout is an avoidable problem that can be identified by including unlabeled controls, and mitigated through improved sample handing and read alignment that together improve the robustness and reproducibility of NR-seq experiments.

Positive Basophil Tests Are Linked to High Disease Activity and Other Features of Autoimmune Chronic Spontaneous Urticaria: A Systematic Review.

BACKGROUND: Chronic spontaneous urticaria (CSU) is believed to be Autoimmune (aiCSU) (type IIb CSU) in at least 8% of patients, associated with mast cell-activating IgG autoantibodies. Basophil tests such as the basophil activation test (BAT) and basophil histamine release assay (BHRA) are considered the best single tests for an aiCSU diagnosis. To date, the strength of associations among a positive BAT and/or BHRA (BAT/BHRA+) and CSU features, patient demographics, and response to treatment remains poorly characterized. OBJECTIVE: To evaluate the strength of current evidence on basophil tests as parameters for CSU characteristics. METHODS: We performed a systematic literature search and review to assess the relationship between BAT/BHRA+ and clinical and laboratory parameters of CSU. Of 1,058 records found in the search, 94 studies were reviewed by experts in urticaria and 42 were included in the analysis. RESULTS: In CSU patients, BAT/BHRA+ showed a strong level of evidence for an association with high disease activity and low levels of total IgE. A weak level of evidence was shown for the association of BAT/BHRA+ and the presence of angioedema, and basopenia. CONCLUSIONS: Our results suggest that aiCSU defined by BAT/BHRA+ is more active or severe and is linked to other aiCSU markers such as low total IgE/basopenia. Basophil tests should be standardized and implemented in routine clinical care to improve the diagnosis and treatment of patients with aiCSU.

WAPL functions as a rheostat of Protocadherin isoform diversity that controls neural wiring.

Neural type-specific expression of clustered Protocadherin (Pcdh) proteins is essential for the establishment of connectivity patterns during brain development. In mammals, deterministic expression of the same Pcdh isoform promotes minimal overlap of tiled projections of serotonergic neuron axons throughout the brain, while stochastic expression of Pcdh genes allows for convergence of tightly packed, overlapping olfactory sensory neuron axons into targeted structures. How can the same gene locus generate opposite transcriptional programs that orchestrate distinct spatial arrangements of axonal patterns? Here, we reveal that cell type-specific Pcdh expression and axonal behavior depend on the activity of cohesin and its unloader, WAPL (wings apart-like protein homolog). While cohesin erases genomic-distance biases in Pcdh choice, WAPL functions as a rheostat of cohesin processivity that determines Pcdh isoform diversity.