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Molecular mechanisms of drug resistance and compensation in SARS-CoV-2 main protease: the interplay between E166 and L50
Full text linkThe SARS-CoV-2 main protease (Mpro) is essential for viral replication and is a primary target for COVID-19 antivirals. Direct-acting antivirals such as nirmatrelvir, the active component of Paxlovid, target the Mpro active site to block viral polyprotein cleavage and thus replication. However, drug resistance mutations at the active site residue Glu166 (E166) have emerged during in vitro selection studies, raising concerns about the durability of current antiviral strategies. Here, we investigate the molecular basis of drug resistance conferred by E166A and E166V mutations against nirmatrelvir and the related PF-00835231, individually and in combination with the distal mutation L50F. We found that E166 mutations reduce nirmatrelvir potency by up to 3,000-fold while preserving substrate cleavage, with catalytic efficiency reduced by only up to twofold. This loss of catalytic efficiency was compensated for by the addition of L50F in the double-mutant variants. We have determined three cocrystal structures of the E166 variants (E166A, E166V, and E166V/L50F) bound to PF-00835231. Comparison of these structures with wild-type enzyme demonstrated that E166 is crucial for dimerization and for shaping the substrate-binding S1 pocket. Our findings highlight the mutability of E166, a prime site for resistance for inhibitors that leverage direct interactions with this position, and the potential emergence of highly resistant and active variants in combination with the compensatory mutation L50F. These insights support the design of inhibitors that target conserved protease features and avoid E166 side-chain interactions to minimize susceptibility to resistance.
Importance: Drug resistance remains a great challenge to modern medicine. This study investigates SARS-CoV-2 main protease variants E166A and E166V which confer nirmatrelvir resistance. These variants can retain considerable enzymatic activity through combination with the compensatory mutation L50F. For single- and double-mutant variant enzymes, we assessed catalytic efficiency, measured loss in potency for nirmatrelvir and its analog PF-00835231, and cocrystallized with inhibitors to investigate drug resistance caused by these mutations. Our results contribute toward understanding of molecular mechanisms of resistance and combinations of mutations, which pushes toward resistance-thwarting inhibitor design. These principles also apply broadly to many quickly evolving drug targets in infectious diseases.No embarg
Prospective SARS-CoV-2 additional vaccination in immunosuppressant-treated individuals with autoimmune diseases in a randomized controlled trial
Full text linkBACKGROUND: Individuals with autoimmune diseases (AD) on immunosuppressants often have suboptimal responses to COVID-19 vaccine. We evaluated the efficacy and safety of additional COVID-19 vaccines in those treated with mycophenolate mofetil/mycophenolic acid (MMF/MPA), methotrexate (MTX), and B cell-depleting therapy (BCDT), including the impact of withholding MMF/MPA and MTX.
METHODS: In this open-label, multicenter, randomized trial, 22 participants taking MMF/MPA, 26 taking MTX, and 93 treated with BCDT who had suboptimal antibody responses to initial COVID-19 vaccines (2 doses of BNT162b2 or mRNA-1273 or 1 dose of AD26.COV2.S) received an additional homologous vaccine. Participants taking MMF/MPA and MTX were randomized (1:1) to continue or withhold treatment around vaccination. The primary outcome was the change in anti-Wuhan-Hu-1 receptor-binding domain (RBD) concentrations at 4 weeks post-additional vaccination. Secondary outcomes included adverse events, COVID-19 , and AD activity through 48 weeks.
RESULTS: Additional vaccination increased anti-RBD concentrations in participants taking MMF/MPA and MTX , irrespective of immunosuppressant withholding. BCDT-treated participants also demonstrated increased anti-RBD concentrations, albeit lower than MMF/MPA- and MTX-treated cohorts. COVID-19 occurred in 33% of participants; infections were predominantly mild and included only three non-fatal hospitalizations. Additional vaccination was well-tolerated, with low frequencies of severe disease flares and adverse events.
CONCLUSION: Additional COVID-19 vaccination is effective and safe in individuals with ADs treated with immunosuppressants, regardless of whether MMF/MPA or MTX is withheld.
CLINICALTRIALS: gov (NCT05000216; registered August 6, 2021: https://clinicaltrials.gov/ct2/show/NCT05000216).No embarg
Connecting research and community: a methodological framework for investigating CMV transmission in childcare settings
Full text linkThe CMV Transmission and Immune Tracking (TransmIT) Study was developed to address critical gaps in understanding of cytomegalovirus (CMV) transmission dynamics in early education and care (EEC) settings. This two-stage, community-engaged study design integrates EEC center partnerships, digital study platforms, and data pipeline infrastructures to enable longitudinal virologic and immunologic surveillance in this high-exposure environment. Stage I focused on establishing foundational components of the study, including a geographically diverse EEC center network, culturally tailored recruitment strategies, a community advisory board, protocols for participant enrollment and saliva sample collection, and optimized laboratory assays to measure viral shedding in saliva. The study approach honed during Stage I is intended to support future longitudinal investigations into viral shedding patterns, immune responses, and co-infections among children and staff in EEC centers. This manuscript presents a methodological framework for conducting community-centered scalable research in early childhood settings with relevance for CMV and other infectious diseases of public health importance.No embarg
Supporting Primary Care for Medically and Socially Complex Patients in Medicaid Managed Care
Full text linkImportance: In 2023, the Massachusetts Medicaid and Children's Health Insurance Program (MassHealth) required accountable care organizations (ACOs) to increase payments to primary care practices and shift to monthly payments, currently calibrated to historical revenues and enhanced practice capabilities, such as being staffed to address behavioral health needs. To prevent rewarding practices for avoiding difficult patients, future payments to primary care practices should reflect their patients' apparent need.
Objective: To describe MassHealth's initiative and a complexity-adjusted payment model.
Design, setting, and participants: This cross-sectional study of payment model development and performance was conducted between February 2022 and November 2024. Participants included all 2019 Massachusetts Medicaid managed-care eligible members who were enrolled for 183 days or longer.
Exposures: Medical and social complexity.
Main outcomes and measures: For each member, the primary care activity level (PCAL) outcome proxies the resources that primary care clinicians need to provide comprehensive, coordinated care. Models were evaluated via R2 and through ratios of observed-to-expected (ie, estimated by the model) outcomes for selected subgroups, which will be approximately 1.0 when payments and expected costs are well matched. The implications of paying practices using PCAL (vs a model based only on age and sex) were explored by examining financial and practice-level characteristics in high and low deciles of practice-level estimated mean.
Results: Among 1 092 742 MassHealth members enrolled in 3602 primary care practices (1 014 252 person-years; mean [SD] age, 25.9 [18.4] years; 538 065 [53.1%] female), the PCAL model achieved R2 = 69.6% and estimates within 10% of observed PCAL spending for high-risk populations (mental health disorders, substance use disorders, complex chronic conditions, and disabilities) and across racial and ethnic groups. Age-adjusted and sex-adjusted payments would overpay practices in the lowest-need decile by 10% and underpay those in the highest-need decile by 34%, while the PCAL model would match payment to estimated need almost exactly in the lowest decile and underpay by just 6% in the highest decile.
Conclusions and relevance: MassHealth's 2023 reform invests in primary care. This cross-sectional study developed a risk model that can adjust primary care payments to patient needs. Neither age and sex adjustments nor inflated historical payments would provide adequate resources to primary care practices caring for the most complex patients.No embarg
Senescent-like microglia limit remyelination through the senescence associated secretory phenotype
Full text linkThe capacity to regenerate myelin in the central nervous system diminishes with age. This decline is particularly evident in multiple sclerosis (MS), a chronic demyelinating disease. Whether cellular senescence, a hallmark of aging, contributes to remyelination impairment remains unknown. Here, we show that senescent cells accumulate within demyelinated lesions after injury, and treatments with senolytics enhances remyelination in young and middle-aged mice but not aged mice. In young mice, we observe the upregulation of senescence-associated transcripts, primarily in microglia and macrophages, after demyelination, followed by a reduction during remyelination. However, in aged mice, senescence-associated factors persist within lesions, correlating with inefficient remyelination. Proteomic analysis of the senescence-associated secretory phenotype (SASP) reveals elevated levels of CCL11/Eotaxin-1 in lesions of aged mice, which is found to inhibit oligodendrocyte maturation. These results suggest therapeutic targeting of SASP components, such as CCL11, may improve remyelination in aging and MS.No embarg
Peer Academic Supports for Success: Pilot Randomised Controlled Feasibility Trial
No full textIntroduction: Mental health conditions are prevalent among university students, putting them at elevated risk for dropout. Universities offer an array of peer programmes, and students often share their concerns with peers before professionals. A well-specified peer intervention to help sustain academic persistence that colleges can directly offer their undergraduates with mental health conditions should benefit this population. The Peer Academic Supports for Success coaching model was developed to address this need.
Objective: This study's goal was to conduct a feasibility and preliminary impact study of the Peer Academic Supports for Success model and feasibility of randomised controlled trial research methods.
Methods: Seventy-two undergraduate students with academically impairing mental health conditions were randomised to receive Peer Academic Supports for Success versus an active control condition. Survey data were collected at baseline and at the end of the next two semesters. Official transcripts were obtained. Intervention implementation data were assessed through coach and participant report.
Results: Peer Academic Supports for Success was delivered with fidelity, successfully attracted and retained students, and was safe. Randomised controlled trial methods proved feasible. Findings revealed significant treatment effects on several of the targeted proximal outcomes.
Conclusions: The findings suggest Peer Academic Supports for Success is a promising university-based intervention to support young adult students with mental health conditions and should be tested in a robust clinical trial
Evaluation of an integrated digital and mobile intervention to improve outcomes for patients with moderate to severe COPD
Full text linkChronic obstructive pulmonary disease (COPD) leads to high rates of emergency department (ED) visits and hospitalizations. This study evaluated a community-based digital health intervention's association with acute care utilization among patients with moderate to severe COPD. In a decentralized, nonrandomized trial, participants received biometric monitoring, symptom tracking, on-demand paramedic services, and digital pulmonary rehabilitation for 6 months. Outcomes were compared to a synthetic control group using weighted optimal matching and multivariable-adjusted regression. The primary outcome was hospitalization; secondary outcomes included readmission rates, ED visits, length of stay, and mortality. Eighty-eight intervention participants (mean age 67 (SD 10)) were compared to a weighted control group of 14,492 (mean age 69 (SD 11)). Intervention participants had lower odds of hospitalization (OR 0.67, 95% CI: 0.46-0.98) and 30-day readmission (OR 0.38, 95% CI: 0.17-0.84). This digital and mobile intervention was associated with reduced acute care use and supports further evaluation of hybrid care models for COPD.No embarg
The Massachusetts Model: A Roadmap for Advancing Serious Illness Care in Nursing Education
No full textThe Massachusetts Nursing Task Force was established to promote the integration of primary palliative nursing competencies in nursing education across the state. A statewide survey revealed that only 23% of baccalaureate nursing programs felt their students were well-prepared in this area. Moreover, there was limited integration of evidence-based resources due to curriculum constraints, lack of faculty time, and financial limitations. To address these challenges, the Massachusetts Nursing Task Force implemented a 2-pronged approach: an End-of-Life Nursing Education Consortium train-the-trainer workshop for faculty and the development of a free, peer-reviewed online nursing resource library aligned with national competencies. The resource library, launched in 2023 and grounded in Kolcaba's Holistic Theory of Comfort, offers diverse materials to facilitate the incorporation of palliative nursing education into existing curricula. Future goals include wider dissemination of these resources, improved assessment of palliative nursing in curricula and practice settings, and expansion of this initiative beyond the state level to advance palliative nursing knowledge and practice.No embarg
The Functional Roles of R-loops in Regulating the Epigenome and Differentiation of Mouse Embryonic Stem Cells
No full textR-loops—RNA:DNA hybrids formed co-transcriptionally—are emerging as important regulators of genome function, yet their influence on chromatin architecture and developmental potential remains poorly understood. Using inducible Rnaseh1 expression in mouse embryonic stem cells (mESCs), we achieved acute, global R-loop depletion to interrogate their role in epigenome regulation and lineage specification. R-loop loss had little effect on steady-state gene expression or self-renewal but caused a marked reduction in H2A.Z occupancy at active and bivalent promoters, coupled with increased nucleosome density. During gastruloid differentiation, R-loop-depleted cells exhibited accelerated ectoderm specification, misregulation of lineage-specific transcription factors, and disrupted cell–cell signaling. These changes coincided with widespread perturbations in gene regulatory networks across multiple early lineages. Together, our findings reveal that R-loops are critical for maintaining promoter architecture via H2A.Z and for preserving balanced lineage trajectories, highlighting their fundamental role in the epigenomic regulation of early development.Interdisciplinary Graduate Program2 years2027-10-1
Skin Ulcer Development and Deterioration of Social Engagement Among Nursing Home Residents
No full textObjectives: Skin ulcers are a critical indicator of quality of care in nursing homes that influence residents' physical, psychological, and social health. The objective of this study is to understand the influence of developing skin ulcers on deterioration in social engagement in nursing home residents.
Design: Observational retrospective cohort study with 1-year follow-up.
Setting and participants: Nursing home residents ≥50 years of age were followed quarterly (2008/2009-2009/2010), the latest years in which the Minimum Data Set 2.0 measured social engagement.
Methods: Cumulative incidence ratios (CIRs) and 95% CIs quantified the association between skin ulcer development and deterioration in residents' social engagement levels.
Results: About 1.25% first developed new skin ulcers at the annual follow-up assessment; 1.22% developed skin ulcers at a quarterly assessment that persisted at the annual assessment, and 4.53% developed skin ulcers at a quarterly assessment that were resolved by annual assessment. Compared with residents who remained ulcer-free, those with new skin ulcer at annual assessment and persistent skin ulcers were at increased risk of experiencing a reduction in social engagement (CIR, 1.26; 95% CI, 1.11-1.44; CIR, 1.32; 95% CI, 1.16-1.50, respectively). Those with resolved skin ulcers were also at increased risk of social engagement deterioration (CIR, 1.12; 95% CI, 1.04-1.21) than those who remained ulcer-free.
Conclusion and implications: Residents who develop skin ulcers are at a higher risk of deteriorating social engagement. Nursing home staff should strive not only to prevent and treat skin ulcers, but also to support social engagement for those with skin ulcers.No embarg