ACS Medicinal Chemistry Letters

Cereblon (CRBN) is widely used in targeted protein degradation, but its ligand space has remained dominated by a narrow set of cyclic imide chemotypes. Here, we show that the accessible CRBN ligand space extends substantially beyond this canonical space. A high-throughput screen of > 40,000 compounds, followed by orthogonal biophysical validation, X-ray crystallography and SAR analyses, identified several chemically distinct ligand classes. These include linear acetyl-based motifs, a phthalide-derived scaffold, steroidal compounds, and a range of bicyclic ligands. They engage CRBN through distinct recognition modes, several of which deviate from the canonical hydrogen-bonding pattern. Steroidal scaffolds were particularly notable: cortisone binds the human CRBN thalidomide-binding domain with an affinity comparable to thalidomide, with its A-ring occupying the tri-tryptophan pocket in a glutarimide-like orientation despite lacking the canonical imide NH donor. SAR within this series showed substantial tolerance for chemical modification and scaffold simplification, raising the possibility that endogenous steroidal metabolites may contribute to the physiological ligand landscape of CRBN. Bicyclic lactams additionally provided synthetically accessible scaffolds with tunable affinity and promising sites for linker attachment. Across the identified ligand classes, none of the tested representatives induced detectable degradation of canonical CRBN neosubstrates, and several showed largely clean proteomic profiles. Together, these findings broaden the chemical, mechanistic and potential physiological landscape of CRBN recognition and provide diverse starting points for alternative, potentially neosubstrate-sparing CRBN recruiters.

The binding affinities of a chimeric analog of a riboflavin derivative linked to biotin, (6- (7,8-dimethyl-2,4-dioxo-3,4-dihydrobenzo[g]pteridin-10(2H)-yl)hexyl 5-((3aS,4S,6aR)-2- oxohexahydro-1H-thieno[3,4-d]imidazol-4-yl)pentanoate), referred to as C6-Rf-biotin-tag, to the riboflavin binding retain or streptavidin are in the M range, 1.29 {+/-} 0.277 and 3.00 {+/-} 0.459, respectively. These values suggest that C6-Rf-biotin-tag has potential applications in diagnostic assay and labelling target flavin binding proteins. The C6-Rf-biotin-tag which was characterized with respect to physical and biochemical properties retains UV/Vis spectroscopic and fluorescence behavior similar to riboflavin.

Hybridoma screening in semi-solid medium typically employs antigens labeled with visible fluorophores (e.g., FITC, AF488) to enable single-step identification of antibody-secreting clones. However, conventional chemical conjugation via NHS-esters or isothiocyanate groups frequently modifies lysine residues located within epitopes, potentially abrogating antibody recognition of these critical regions. Here, we describe a SpyTag SpyCatcher-based site-specific labeling strategy that circumvents epitope damage during semi-solid medium screening. A 16-amino-acid SpyTag was genetically fused to the C-terminus of the target antigen, enabling covalent conjugation to an sfGFP SpyCatcher fluorescent probe. In semi-solid medium supplemented with SpyTag-antigen and sfGFPSpyCatcher, positive hybridoma clones were readily identified by distinct fluorescent halos, whereas negative clones showed no detectable signal. Notably, the site-specific method yielded a significantly higher frequency of fluorescence-positive clones compared to the conventional AF488-labeled antigen method, suggesting that epitope preservation enhances screening recovery. Furthermore, this approach did not impair hybridoma growth or final clone positivity, offering a simple, rapid, and epitope-compatible method for monoclonal antibody screening.

Gabapentin (GBP) and pregabalin (PGB) are widely used gabapentinoids. Previously, we have demonstrated, for the first time, that GBP and PGB modulate the nociceptive response to noxious heat in C. elegans at an optimal concentration. In the current study, we use C. elegans and paired thermal nociception assays with direct internal drug concentration measurements to characterize the pharmacokinetic (PK)/pharmacodynamic (PD) relationship of both compounds. Neither drug altered baseline mobility or quadrant preference, confirming that behavioral effects reflected genuine antinociceptive action. Both GBP and PGB produced dose- and time-dependent reductions in thermal avoidance, with 500 uM exposures generating a biphasic, V-shaped time course in which suppression of thermal sensitivity deepened before partially reversing. This partial reversal occurred later with PGB than with GBP. Internal concentrations confirmed dose-dependent absorption and retention for both drugs, yet at 500 uM, internal drug levels remained elevated through 360 min even as behavioral avoidance recovered, indicating that the recovery limb reflects active counter-regulation rather than passive clearance, consistent with previously reported transcriptional and proteomic signatures. Exposure-response profiles were notably flat, suggesting a saturable pharmacodynamic ceiling. Molecular modeling revealed conserved electronic pharmacophores supporting shared alpha-2-delta engagement, alongside shape-descriptor differences that may contribute to divergent absorption kinetics. These findings position C. elegans as a valuable model for dissecting gabapentinoid PK/PD relationships. Beyond mechanistic insight, these findings support the continued investigation of C. elegans as a screening platform whose validation could help address the 3R (Replacement, Reduction, Refinement) principles guiding animal research.

To combat refractory diseases, such as cancer, multitarget-directed ligands (MTDLs) have become an emerging area of research to exploit synthetic lethality (SL) relationships associated with drug resistance. Herein, we present the in silico design of MTDLs for the polypharmaceutical treatment of endometrial adenocarcinoma (EAC) and our discovery of a novel SL in EAC; PTEN loss of function (LOF) and the inhibition of CDK9. We used high-resolution x-ray crystallographic data to chemically engineer, LCI133, to inhibit CDK9, CDK4/6-and AURKA/B kinases. PTEN LOF in EAC results in augmented deregulated transcription and a massive increase in nascent RNA, a phenotype which encodes a high level of apoptotic sensitivity to LCI133 and CDK9 inhibitors. Treatment with LCI133 results in a rapid decline nose-dive in global nRNA, MYC nRNA levels and TS elongation (TE) in PTEN LOF EAC. PTEN LOF is necessary and sufficient to confer sensitivity of EAC cells to LCI133 and other CDK9 inhibitors.

Amyloid fibrils are highly ordered protein aggregates characterized by a conserved cross-{beta}-sheet architecture despite originating from structurally diverse precursor proteins. Growing evidence suggests that interactions between different amyloidogenic proteins can modulate aggregation pathways through heterologous cross-seeding; however, the influence of seed polymorphism on the structure and biological properties of cross-seeded fibrils remains poorly understood. Here, we investigated the cross-seeding of native human insulin by two structurally distinct polymorphs of hen egg-white lysozyme (HEWL): flexible fibrils (FFs) and rigid fibrils (RFs). Native insulin remained stable under physiological conditions and underwent spontaneous fibrillation only under acidic conditions. In contrast, both HEWL polymorphs efficiently induced insulin aggregation at physiological pH, bypassing the nucleation barrier. Thioflavin T fluorescence, circular dichroism spectroscopy, and transmission electron microscopy revealed that lysozyme FFs templated the formation of insulin flexible fibrils (IFFs), whereas lysozyme RFs produced insulin rigid fibrils (IRFs), demonstrating that the structural characteristics of the parental HEWL polymorphs were propagated during heterologous cross-seeding. The toxicity of the resulting insulin fibrils was evaluated in SH-SY5Y neuronal cells and CCF-STTG1 astrocytes. IFFs exhibited minimal cytotoxicity and only subtle morphological alterations, whereas IRFs caused modest reductions in cell viability accompanied by more pronounced cellular damage. These findings demonstrate that the structural polymorphism of HEWL fibrils governs both the architecture and biological activity of cross-seeded insulin fibrils, highlighting amyloid polymorphism as an important determinant of heterologous amyloid propagation and a potential design principle for engineering functional amyloid-based biomaterials and protein delivery platforms.

Objective: This study examined the prevalence of psychotic experiences (PE) and how early onset and persistence of PE contribute to risk and severity of mental disorders in adolescents at familial high-risk of schizophrenia (FHR-SZ) or bipolar disorder (FHR-BP) and adolescents from a population-based control group (PBC). Methods: This is the second follow-up of a nationwide cohort study including 522 children at FHR-SZ (N=202), FHR-BP (N=120), and PBC (N=200). Participants were assessed at ages 7, 11, and 15 using a semi-structured interview to evaluate PE and mental disorders. Results: At age 15, adolescents at FHR-SZ reported more PE than PBC over the past six months (current) and the past four years, while adolescents at FHR-BP only reported more current PE. PE reported at two or three timepoints (persistent PE) predicted any Axis I disorder in mid-adolescence, corresponding to three- (OR 2.9, 95% CI [1.5-5.7]) and 21-fold (OR 21.4, 95% CI [2.8-162.3]) increased risks, respectively. Persistent PE also predicted multimorbidity, with three- (OR 2.8, 95% CI [1.0-7.6]) and four-fold (OR 4.1, 95% CI [1.2-14.1]) increased risks, respectively. This was after adjustment for sex, early mental disorders, and familial risk. Conclusions: This study demonstrates a strong link between persistent PE and mid-adolescence mental disorders. Our findings emphasize PE as important risk markers for mental disorders during mid-adolescence and highlight the importance of monitoring children with PE before age 7 who develop persistent symptoms.

Study Objective Little is known about where in hospital care for multiple long-term conditions (MLTC) is delivered. We aimed to describe pathways of care (ward transfers) and outcomes for people admitted to hospital for unscheduled care by MLTC status and other key sociodemographic characteristics. Design and setting Analysis of routinely-collected electronic health records from a large acute UK hospital. Participants Adult unscheduled care admissions from 1st July 2018 to 30th June 2019. The presence of two or more of 59 long-term conditions was ascertained using ICD-10 codes from previous hospital discharges. Main outcome measures Markov state transition probabilities were derived for ward moves and compared for MLTC vs no MLTC, age, sex, ethnicity and neighbourhood deprivation. Outcomes (length of stay, death, readmission, move from definitive ward) and time spent in emergency and assessment departments were compared between subgroups. Results A total of 33,252 adults, mean age 56.0 (SD 21.9) years were analysed; 14,834 (42.4%) had MLTC. People with MLTC were more likely to die in hospital (4.2 vs 1.9%, p<0.001), transfer to internal medicine wards or older peoples medicine wards, were less likely to transfer to surgical wards, had longer median length of stay (1.83 vs 0.69 days, p<0.001), stayed longer in acute medical units (15.5 vs 9.6 hours, p<0.001), and were more likely to move from their definitive ward (18.2 vs 16.4%, p=0.002). Conclusion Unscheduled hospital care pathways are complex and differ for people with MLTC, who have worse outcomes and may be less likely to receive optimal care.

Next-generation sequencing (NGS) for the detection of somatic variants has become the method of choice in a variety of molecular oncology fields and in the clinic. Its use ranges from sequencing entire tumor genomes and transcriptomes to targeted clinical diagnostic gene panels. The NYU Langone Genome PACT (Profiling of Actionable Cancer Targets, LG-PACT) assay is a qualitative in vitro diagnostic test that uses targeted next generation sequencing (NGS) of formalin-fixed paraffin-embedded (FFPE) tumor tissue matched with normal specimens from patients to detect gene alterations in a targeted panel covering 606 genes and the TERT promoter. Indications for testing are cancer (solid tumors and hematological malignancies) where a mutational profile from multiple genes would be informative for disease stratification, prognosis, or treatment options including targeted therapies and eligibility for clinical trials. The test is intended to provide information on somatic mutations including point mutations, small insertions/deletions (indels), and copy number aberrations for diagnostic and treatment decisions. LG-PACT is a United States Food and Drug Administration (FDA) cleared diagnostic test (510K: K202304). The clinical interpretation of sequencing data of molecular tumor markers from NGS encompasses automated variant calling tools with human interpretation. This final mostly manual review of data step is intensive, involving highly trained scientists, encompassing literature review, interpretation and clinical tier classification by pathologists, who then provide a complete molecular diagnostic report to the treating oncologists. We provide analysis of 1339 clinical genomic profiles from 31 different cancers and their subtypes, comprising of central nervous system (CNS) 792 (59%) cases (incl. meningioma, glioma and glioblastoma), with 267 (20%) cases predominantly of lung, pancreatic and colorectal and 280 of others (21%). Here, we present the technical challenges of validating an NGS oncological diagnostic targeted assay for clinical grade accuracy and sensitivity for patient care. We show how copy number alterations provide a more comprehensive description of the tumors genomic profile. We then outline the utility of targeted panel sequencing based on certified pathologist selection of reportable variants for our current patient cohort. Where analysis of variant detection has led to 49.4% (661/1339) of our clinical tumor samples containing mutations in known therapy targeted genes, 35.6% (477/1339) with mutation detected in other genes, and 15% (201/1339) cases being negative.

Importance: Patients are increasingly learning Alzheimers disease (AD) genetic and biomarker results through electronic health portals. Evaluation of alternative scalable delivery models for return of AD risk information is needed to best support patient understanding and psychological well-being. Objective: To determine whether a patient-centered digital platform is comparable to clinician-mediated telehealth sessions for returning APOE and plasma pTau-217 results on outcomes of knowledge and psychological well-being. Design: The Evaluation of Self-Mediated Alternatives for Risk Testing Education and Return of Results (eSMARTER) study was a noninferiority trial of a patient-centered digital platform compared to clinician-mediated disclosure of APOE genotype and optional pTau-217 disclosure. Setting: Decentralized, fully remote trial enrolled participants in the contiguous United States (U.S.) between October 2024 and February 2025, with follow-up completed in November 2025. Participants: Eligible participants were aged 60-80 and had previously undergone APOE genotyping (without disclosure) via the GeneMatch program, passed psychological screening, had internet access, and were English-speaking. Interventions: Participants were randomized, 2:1, to the eSMARTER digital platform or clinician-mediated disclosure of APOE genotype. Following the 6-month post-APOE assessment, participants were offered optional pTau-217 disclosure via the same randomized modality. Main Outcomes and Measures: Primary outcomes at 1-7 days following APOE disclosure included changes in anxiety, disease-specific distress, and AD-related knowledge within a priori non-inferiority margins. Results: 674 persons (mean [SD] age 68 [4.7] years; 451 [67%] female; mean [SD] telephone MoCA=19 [2]) were eligible and provided demographic information. 651 participants were randomized to clinician-mediated (n=216) or digital disclosure (n=435) and completed APOE disclosure (66 [10%] APOE4 homozygotes, 377 [58%] heterozygotes, 208 [32%] non-carriers). 604 participants completed the study; 500 completed optional pTau-217 disclosure. Baseline characteristics were balanced across groups. At 1-7 days following APOE disclosure, scores on AD-related knowledge, PROMIS Anxiety, and disease-specific distress measures met non-inferiority. Conclusions and Relevance: Disclosure of APOE genotype by the eSMARTER digital platform is non-inferior to clinician-mediated telehealth disclosure. No significant between group differences were found following disclosure of pTau-217 results. Together, these results suggest that this digital platform may provide an evidence-based scalable approach for returning AD genetic and biomarker results.

Abstract: Objective: Post-traumatic epilepsy (PTE) is a common sequela of traumatic brain injury (TBI). Research indicates that individuals with PTE tend to experience greater cognitive difficulties compared to those with TBI alone. However, it is plausible that a distinct cognitive profile exists that distinguishes between TBI cases with and without PTE. We aimed to identify longitudinal changes in cognitive measures among TBI patients to better assess the changes associated with developing PTE. Setting: Outpatient. Participants: Prospective subjects who had suffered TBI within 6 months post-injury (TBI-6M, n=32), retrospective subjects with pre-existing PTE diagnoses (PTE, n=20), and healthy control subjects (HC, n=41). Design: We examined cognitive performance for TBI patients within 6 months post-injury, then again within 12 months (TBI-12M, n=26), and within 18-months (TBI-18M, n=25), and compared this with cognitive performance among HC and PTE. Main Measures: Cognitive tests administered yielded 15 test components for analysis. We utilized linear mixed effects modeling to examine cohort-level differences cognitive function. Results: 11/15 tests showed a significant performance deficit in the PTE subjects compared to HC. TBI-6M was not significantly different from the PTE subjects; with time, 9/15 tests showed some degree of recovery in TBI subjects. Tests for information processing speed/working memory and executive function showed strong recovery (TBI-6M vs. TBI-18M, SDMT written: p<0.0001, SDMT oral and COWAT: p<0.001). Tests for visual attention/working memory also showed a smaller but significant recovery (TBI-18M vs. PTE, p<0.05). By contrast, tests for verbal memory [HVLT-R Delayed Recall] showed chronic impairment in TBI (TBI-18M vs HC, p<0.0001). TBI subjects generally trend towards recovery in cognitive performance post-TBI. Conclusions: Information processing speed/working memory are strong indicators for TBI recovery, while auditory learning/memory shows chronic impairment. The stagnation of recovery in cognitive domains typically characterized by robust recovery may correlate with an elevated risk of developing PTE.

The distribution of fat-soluble compounds (including antioxidants) in follicular fluid (FF) remains sparsely documented in relation to in vitro fertilization (IVF) outcomes and existing studies have reported diverging associations. This study aimed to describe plasma and FF concentrations of fat-soluble micronutrients in women undergoing IVF and to analyze their adjusted associations with ovarian function, embryo development and pregnancy outcomes. In 2021-2022, plasma and FF samples were collected from 82 women (first IVF cycle) at oocyte puncture, along with lifestyle data covering the three preceding months. Eleven compounds (two tocopherols, three xanthophylls, five carotenes and retinol) were quantified. All compounds were detected in both compartments (lowest in FF) except phytoene, undetectable in FF. Plasma and FF -tocopherol concentrations were positively associated with plasma estradiol levels before oocyte puncture (both p<0.01) while FF -carotene and lycopene were inversely associated with plasma progesterone concentrations (p=0.01 and 0.02, respectively). Plasma phytofluene and phytoene were positively associated with mature oocyte rate (p=0.03 and p=0.01, respectively), while FF retinol was negatively associated (p=0.03). Carotenes, tocopherols and retinol were inversely associated with later IVF outcomes: fertilization rate (p<0.001 for plasma g-tocopherol, 0.02 for FF retinol), top-quality embryo (p=0.02 for plasma phytofluene), biochemical pregnancy at day 7 post-embryo transfer (p=0.05 for plasma -tocopherol, 0.02 for plasma -carotene), clinical pregnancy (p=0.03 for plasma -tocopherol, 0.01 for plasma phytoene) and live birth (p=0.04 for plasma -tocopherol, 0.02 for plasma phytoene). Plasma and FF g-tocopherol were positively associated with embryo fragmentation (both p<0.05). Finally, among xanthophylls, only plasma {beta}-cryptoxanthin was positively associated with plasma progesterone concentrations (p=0.02). Our findings of heterogeneous associations between tocopherols, carotenes, retinol and IVF outcomes across the stages of IVF suggest a beneficial effect limited to early outcomes and support a complex and context-dependent role of these compounds in female reproduction. This manuscript has been submitted to PlosOne on August 19, 2026.

Lysosomes make key contributions to the maintenance of cellular proteostasis, and their functional compromise has been linked to aging and neurodegenerative disease. A defining characteristic of lysosomes is their relative acidity compared to other subcellular compartments, a quality that enables the efficient breakdown of macromolecules. Evidence suggests that neuronal lysosomal pH becomes dysregulated with aging and neurodegenerative disease, yet the mechanisms by which lysosomal pH is maintained remain incompletely understood. To better understand neuronal lysosomal pH regulation, we conducted a genome-wide CRISPRi-based screen in iPSC-derived iNeurons for modifiers of lysosomal pH. We validated several previously known regulators of lysosomal pH and identified novel pathways capable of modifying lysosomal pH, including protein UFMylation and mitochondrial homeostasis. We demonstrate that loss of the lysosomal cationic amino acid exporter, PQLC2, prevents lysosomal acidification in a manner independent of amino acid transport. A novel, tauopathy-associated mutation in PQLC2 impairs lysosomal acidification and drives tau accumulation. Together, this study reveals novel genes that modify lysosomal pH and highlights potential new targets for ameliorating age-related lysosome dysfunction.

Introduction Continuous Glucose Monitoring (CGM) is considered standard care in high-income countries. There is, however, limited published evidence on CGM use in low- and middle-income countries. The purpose of this study was to assess the usability, acceptability, and feasibility of CGM use among people living with type 1 diabetes (T1D) and caregivers in a low-resource setting. Research Design and Methods This prospective study conducted at the Kenyatta National Hospital purposively enrolled persons aged 4-25 years who had been on management for T1D for at least six months, and caregivers of those under 18 years. Fourty youth living with T1D used CGM for three months in place of self monitoring of blood glucose (SMBG). The System Usability Scale (SUS), a Theoretical Framework of Acceptability-based questionnaire, the Diabetes Distress Scale (DDS), the Glucose Monitoring Satisfaction Survey (GMSS), and a feasibility survey were administered. Outcomes were summarized descriptively, including means, medians, and frequencies using R statistical software. Results The median SUS score was 98.8 (IQR 92.5-100.0). Acceptability was high, and the median total GMSS score improved from 3.73 to 4.73. Among adolescents and adults, the median overall DDS score reduced from 1.54 to 1.36, with reductions in scores in all domains, except for hypoglycemia distress which increased, and physician distress which remained low. Among caregivers, the median overall DDS score declined from 2.05 (moderate distress) to 1.90 (low distress), with modest reductions in teen management and parent-teen relationship distress and a slight increase in personal distress. Median CGM active wear time was 89%. Conclusion This study comprehensively evaluated CGM across usability, acceptability, and feasibility outcomes, with the findings supporting the integration of CGM into routine diabetes management in low-resource settings. The short follow-up period, however, may not capture changing perceptions or long-term adherence.

Introduction: Risk of recurrent opioid use during buprenorphine-naloxone (bup-nx) treatment is dynamic and remains elevated after initiation, with vulnerability shaped in part by treatment intensity and gaps between visits, yet routine outpatient care relies on episodic encounters and retrospective data. This mismatch can delay recognition of emerging instability and limit timely treatment adjustments. This paper reports the development and specification of an intervention strategy to address this mismatch. Methods: We used a structured, multi-phase design process to specify and configure a measurement-based care (MBC) strategy for bup-nx treatment (Bup-MBC) in outpatient addiction clinics through three phases: (1) a systematic review of patient-reported outcome measures (PROMs) for substance use treatment; (2) a qualitative needs assessment using the Theoretical Domains Framework and COM-B (Capability, Opportunity, Motivation-Behavior) model to identify gaps in risk monitoring, agency, and trust; and (3) iterative co-design with multidisciplinary clinicians to refine workflow fit and trust-preserving use of data. Patients informed item and feedback content during the needs assessment but did not participate in the co-design cycles. Results: Bup-MBC integrates (1) brief between-visit PROMs (e.g., withdrawal, craving, adherence); (2) immediate non-punitive patient feedback; (3) clinician-facing summaries and non-directive prompts in the electronic health record (EHR); and (4) an opt-in between-visit outreach pathway with predefined safety triggers, all configured within existing EHR and patient portal infrastructure. It targets patient and clinician capability to recognize changes in risk, opportunity for action through structured monitoring and visit preparation, and trust and agency through non-punitive communication, without adding substantial burden. The full measure set, severity bands, and question-to-action map are provided as supplementary material. Key trade-offs included prioritizing single-item measures for feasibility, balancing opt-in outreach with safety overrides, and assuming routine clinician use of summaries. Conclusion: This development study specifies an EHR-integrated MBC strategy for outpatient bup-nx treatment. As single-center design work with co-design limited to clinicians and delivery contingent on portal or text-message access, its outputs are hypotheses about mechanism and fit rather than demonstrated effects. Feasibility studies are needed to evaluate uptake, acceptability, workflow fit, and effects on treatment.

Background: Pressure ulcers often develop complex three-dimensional morphologies that extend beyond the visible wound surface. Subsurface extensions such as tunneling and undermining create hidden cavities that complicate clinical assessment and wound management. Despite their clinical relevance, the prevalence and spatial characteristics of these subsurface wound morphologies have not been well characterized at scale. Methods: We performed a registry-based analysis using data from the LIFT-OFF Pressure Ulcer Registry, which captures longitudinal clinical documentation of pressure ulcers treated in routine care. The registry included approximately 18,000 patients with 32,000 documented pressure ulcers. Spatial characteristics of tunneling and undermining were analyzed using measurements recorded during routine wound assessments, including tract length, direction, and circumferential extent. Directional and circumferential distributions of subsurface defects were examined to characterize wound geometry. Results: Tunneling was present in 764 of 14,700 full-thickness pressure ulcers (5.2%), whereas undermining occurred in 2,293 wounds (15.6%). Tunneling tracts were typically short and exhibited directional clustering relative to the wound bed. In contrast, undermining demonstrated broader circumferential distributions and frequently involved larger subsurface separations beneath the wound margin. Both morphologies demonstrated distinct spatial patterns across anatomical locations and wound stages. Conclusion: Tunneling and undermining are common subsurface features of pressure ulcers and exhibit distinct spatial geometries. Whereas tunneling manifests as directional tract-like extensions, undermining more frequently produces circumferential tissue separation beneath wound margins. Improved characterization of subsurface wound architecture may enhance assessment of wound complexity and provide information not captured by surface measurements alone. Future studies should evaluate whether these features contribute to wound severity assessment, prognosis, and risk stratification.

Background: The condition of a patient with acute stroke often changes within hours of ICU admission. Prognostic work here targets fixed endpoints predicted from admission data, and trajectory phenotyping assigns one label per patient. We used longitudinal ICU data to identify interpretable dynamic clinical states, characterize transitions between them, and relate the current state to later events. Methods: Retrospective cohort study of 6368 adults with acute stroke in MIMIC IV v3.1. The first 72 h were divided into twelve 6-hour windows, and a hidden Markov model was fitted to 21 neurological, physiological and organ support variables. State number was chosen against criteria fixed before fitting: statistical fit, restart stability, state occupancy and clinical interpretability. Generalized estimating equations related the current state to new mechanical ventilation and vasopressor use within 12 h, and to ICU death within 72 h. Eleven sensitivity analyses assessed the robustness of the state solution. Results: Four states were selected: neurologically preserved-low support, neurological impairment low support, impairment renal dysfunction and impairment-respiratory support (63.3%, 7.8%, 11.8% and 17.1% of windows). Within 72 h, 40.3% of patients changed state at least once, and transitions ran in both directions rather than along a single severity gradient. States were identified without outcome data, yet ICU mortality by last state ranged from 2.9% to 43.9%. Adjusted for age, sex, subtype and Charlson index, the current state remained associated with organ-support escalation and death. State prevalence differed by at most 1.1 percentage points between training and test sets, and 10 of 11 sensitivity analyses gave a stable four-state solution (ARI 0.754 0.955). Conclusions: The early ICU course of acute stroke can be represented as movement among a small number of clinically interpretable states. The representation was reproducible in a held out set and across admission eras, but requires validation in an independent database before any clinical use.

Background: Polymorphisms in the N-acetyltransferase 2 (NAT2) gene explain much of the interindividual variation in isoniazid (INH) metabolism and determine risk of toxicities. However, there is limited evidence to guide INH dose adjustment according to the NAT2 acetylator profile in weekly rifapentine-INH tuberculosis preventive therapy (TPT). Methods: In a prospective, multicenter, within-subject PK trial (NCT05413551), adults initiating 3HP in Brazil were assigned genotype-guided INH doses (slow: 5 mg/kg <=300 mg; intermediate: 15 mg/kg <=900 mg; rapid: 25 mg/kg <=1,500 mg) alongside a standard 900 mg flat dose on an alternate occasion. AUC0-24 and C24 were estimated from serial blood samples; a two-compartment Michaelis-Menten population PK model characterized NAT2 effects on clearance. Results: Among 228 participants, 47.4% (108/228) were intermediate, 43.4% (99/228) slow, and 9.2% (21/228) rapid acetylators. Genotype-guided dosing reduced AUC0-24 variability approximately two-fold versus standard dosing (CV 58.8% vs 76.8%) and increased exposure uniformity (median AUC0-24 27.2 [IQR 18.8-41.3] vs 43.2 [27.3-71.0] mg h/L). Among slow acetylators, C24 >0.15 ug/mL decreased from 27/42 (64%) with standard dosing to 1/42 (2%) with genotype-guided dosing (P<0.0001). In 104 participants with intensive PK sampling, rapid acetylators receiving guided doses had AUC0-24 similar to standard-dose intermediate acetylators (42.8 vs 39.5 mg h/L; P=.63). Monte Carlo simulations supported doses of 600, 900, and 1,200 mg for slow, intermediate, and rapid acetylators, respectively. Conclusions: NAT2-guided isoniazid dosing reduced variation in drug levels, averting very low and high AUC and C24. These findings inform genotype-stratified dosing of INH for TPT, which might reduce toxicities and improve outcomes.

Predicting disease risk from prior diagnoses is fundamental to clinical decision-making, particularly during health emergencies such as the COVID-19 pandemic, when individuals with long-term conditions may be disproportionately vulnerable to adverse outcomes. Despite intense interest in developing models to predict disease risk from prior diagnoses (1-3), most prediction models do not estimate effects of each prior diagnosis on disease risk conditional on other diagnoses, limiting interpretability and clinical utility. We developed the Comorbidity Risk Score (CRS), trained on 13 million individuals (age 40-69) from linked electronic health record (EHR) datasets of the entire population of England, to predict COVID-19 hospitalisation and 87 other disease outcomes. CRS was trained at close to saturated sample size and precisely estimated the effects of 212 prior diagnoses on the 88 disease outcomes, conditional on all other prior diagnoses. Correlations of CRS effect sizes across outcomes (e.g. 0.76 for myocardial infarction vs. hyperlipidaemia) matched the corresponding genetic correlations (e.g. 0.79 for myocardial infarction vs. hyperlipidaemia), confirming that comorbidity architectures capture disease aetiology. On average, CRS identified 5% of the population with 3.4-fold higher disease risk, including myocardial infarction (4.4-fold), lung cancer (6.5-fold), and COVID-19 hospitalisation (6.3-fold). Using prior diagnoses alone, CRS outperformed state-of-the-art clinical COVID-19 models (4). Furthermore, CRS (N=13 million) substantially outperformed state-of-the-art AI (1) (N=0.5 million) and linear (3) (N=0.5 million) models in predicting disease risk, suggesting that training sample size outweighs model complexity. CRS attained near-perfect transferability across self-reported ethnicities (e.g., Black vs. White: AUROC ratio = 97.3%). Finally, CRS distinguished independently predictive comorbidities from indirect associations, e.g., lipid metabolism disorder was a strong predictor of myocardial infarction risk but not ischaemic stroke, after conditioning on other prior diagnoses. In conclusion, CRS provides a comprehensive resource for understanding the impact of comorbidities on COVID-19 and other future diseases, revealing disease aetiology while enabling powerful prediction of disease risk.

Improvements in data availability, sharing, and integration, together with the development of explainable artificial intelligence (XAI) techniques, are advancing precision medicine for pediatric cancer by facilitating diagnosis, biomarker discovery, and drug development. Data sharing commons and initiatives like the Childhood Cancer Data Initiative (CCDI) provide access to pediatric-specific genomic and clinical data cohorts and improve data availability for pediatric cancer research. Based on CCDI, a scalable AI platform, Graph Artificial Intelligence for Pediatric Oncology (GAIPO), integrates various data modalities from bulk and single-cell omics data to clinical information. Such multi-modal data facilitates the training and development of advanced XAI models for pediatric cancers. We then developed an end-to-end multi-modality framework, PCGS, for pediatric cancer by incorporating omics-specific representation learning via GNN models with cross-attention fusion and multi-objective learning for downstream tasks such as classification, clustering, and survival analysis. This framework outperforms previous supervised multi-omics integration baseline approaches based on glioma and Wilms tumor cohorts and enables GNN model explainability via Shapley value-based feature attribution approaches to explain the contributions of gene-level features across various biomedical tasks, including classification and survival. Given specific background samples (e.g., age groups, sex, grades) as baselines, this explainable GNN model estimates and ranks the importance scores for input features from each omics modality. It identifies background-specific key features for biomarker discovery, risk group identification, and survival analysis in glioma and Wilms tumor, with potential applicability to other pediatric cancers.