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Background: Colorectal neuroendocrine carcinomas (NECs), including small cell (SCNEC) and large cell (LCNEC) subtypes, are rare but aggressive malignancies with limited population-level data. Moreover, prior studies have limited stratification by histologic subtype and anatomic location, despite evidence of disease heterogeneity in colorectal malignancies. This study aimed to characterize the clinical characteristics, treatment patterns, and outcomes of colorectal NEC by histologic subtype and tumor location, and to compare them with neuroendocrine tumors (NETs) and adenocarcinomas (ACs). Methods: A retrospective cohort review was conducted utilizing the SEER database from 2000 to 2022 to identify patients with colorectal SCNEC, LCNEC, NET, and AC. Demographic, clinical, therapeutic and survival data were analyzed and compared using Kaplan–Meier methods, log-rank tests, and Cox proportional hazards regression. Results: A total of 790 SCNEC, 498 LCNEC, 31,200 NET, and 638,898 AC cases were identified. SCNEC and LCNEC were associated with advanced stage at diagnosis (66% and 55% stage IV, respectively) and inferior survival outcomes compared to NET and AC (median OS (mOS) of 7 and 8 months for SCNEC and LCNEC, respectively, p = 0.0002). Stratified by location, colonic SCNEC had the poorest survival (mOS 5 months), worse than rectal SCNEC (mOS 9 months); this pattern was not observed in LCNEC. Surgical resection was associated with improved OS in both NEC subtypes (HR range 0.25–0.56, all p < 0.012). Radiation therapy was associated with improved survival in NEC overall (SCNEC HR 0.58; LCNEC HR 0.71, both p < 0.05), with the observed benefit appearing greatest in rectal NEC. Demographic factors were associated with survival in NET and AC but had no significant impact in NEC. Conclusions: Colorectal SCNEC and LCNEC are highly aggressive malignancies with poor outcomes, with SCNEC demonstrating modestly inferior outcomes compared to LCNEC. This study demonstrates that both histologic subtype and anatomic site of disease are important determinants of prognosis and treatment response. This supports the concept that colorectal NEC represents a biologically heterogeneous disease, with colonic SCNEC showing the worst outcomes and rectal NEC associated with a potential enhanced response to radiation. These findings support the need for further prospective and molecular studies to better define treatment approaches and targeted therapies for these rare malignancies.

PURPOSE In large B-cell lymphoma (LBCL), use of high-dose methotrexate (HD-MTX) to prevent CNS relapse (so-called CNS prophylaxis) remains controversial. International guidelines continue to recommend HD-MTX in patients deemed ultra high risk (UHR) because of a lack of robust data specific to these subgroups. This study was designed to examine the impact of HD-MTX specifically in UHR patients. METHODS Data from two previous retrospective analyses were combined to produce a cohort of UHR patients (≥1 of the following criteria: CNS international prognostic index score 5-6; testicular, renal/adrenal, or breast involvement; ≥3 extranodal sites), treated with or without HD-MTX. The primary outcome was CNS relapse rate (isolated or concurrent with systemic relapse) measured from diagnosis with additional landmark analysis of all responding patients with no progression event at 6 months. RESULTS Analyses were performed on 1,923 patients meeting UHR criteria. No significant difference in 3-year CNS relapse rate was observed between no HD-MTX (n = 1,051) versus HD-MTX patients (n = 872), including in multivariable analyses adjusting for baseline characteristics (3-year rate, 9.3% v 8.1%; adjusted hazard ratio [HR], 1.13 [95% CI, 0.82 to 1.57]). Analyses restricted to isolated CNS relapse also confirmed no difference (5.9% v 5.7%; adjusted HR, 1.03 [95% CI, 0.69 to 1.53]). In the landmark analysis (no HD-MTX n = 782, HD-MTX n = 773), no difference in 3-year CNS relapse was observed (6.7% v 6.6%, adjusted HR, 0.95 [95% CI, 0.62 to 1.44]). CONCLUSION To our knowledge, this is the largest international comparative data set of patients with LBCL with UHR features showing no significant reduction in CNS relapse with HD-MTX in all UHR or in any individual subgroup. Despite inherent limitations of retrospective analyses, the data strongly support previous studies in suggesting HD-MTX prophylaxis has no meaningful benefit for most patients.

ABSTRACT Metallo-beta-lactamase (MBL)-producing Enterobacterales are a global public health concern. Infections caused by these pathogens are often hard to treat, with few effective agents currently available. While aztreonam-avibactam only recently gained U.S. and European regulatory approval, laboratories have implemented laboratory-developed tests (LDTs) using existing aztreonam and ceftazidime-avibactam reagents. We sought to compare the performance of four phenotypic methods, up to three reagent manufacturers, and two media manufacturers. One hundred two MBL-producing Enterobacterales isolates were evaluated at two sites (Centers for Disease Control and Prevention [CDC] and Johns Hopkins University [JHU] School of Medicine). The performance of four methods was evaluated: broth disk elution (BDE), disk stacking (DS), gradient diffusion strip stacking (SS), and a strip-to-disk proximity (strip-disk) method, which were compared to reference broth microdilution (rBMD) tested in parallel. Respective category agreement for CDC and JHU was 92.2% and 96.1% for BDE, 51%–76.5% and 31%–49% for DS, 71.6%–86.3% and 83.3%–86.3% for SS, and 82.4%–87.3% and 80%–89.2% for the strip-disk method. The SS essential agreement ranged from 46.1% to 62.7% for JHU and 54.9% to 92.2% for CDC. Interlaboratory reproducibility was 93.1% for rBMD, 92.2% for BDE, 47.1%–53.1% for DS, 76.5%–80% for SS, and 87.3%–88.9% for strip-disk. BDE was the only method that exceeded acceptable performance of >90% accuracy and reproducibility. DS had the lowest and most variable performance between sites and among the different disk manufacturers used. LDTs for combination testing can be highly variable, and further work is needed to improve standardization and harmonization to achieve accurate and reliable results for clinical purposes. IMPORTANCE Carbapenem-resistant Enterobacterales are classified as an urgent public health threat in the Centers for Disease Control and Prevention’s 2019 Antibiotic Resistance Threats Report. Among the most concerning resistance mechanisms are carbapenemase enzymes encoded on mobile genetic elements, which can spread rapidly between bacteria. Of the various carbapenemases, metallo-beta-lactamases are particularly challenging, as they confer resistance to nearly all beta-lactams, and current treatment options remain limited. While aztreonam-avibactam has been U.S. Food and Drug Administration (FDA)-approved for treating certain infections with limited or no alternative treatment options, no FDA-cleared susceptibility test is yet available. In this context, laboratory-developed methods, such as combination testing of aztreonam and ceftazidime-avibactam, play a critical role in guiding therapy. Our study evaluates several of the more common laboratory-developed tests compared to reference broth microdilution and found that broth disk elution was the most accurate; in contrast, other methods had variable performance, some demonstrating limited reliability and being prone to overcalling resistance.

Introduction There is a growing interest in therapy applications of cell-free extracellular vesicles (EVs) but systemically administered EVs predominantly accumulate in the liver and thereby limit their applications in extrahepatic tissues. One way of diverting EVs to extrahepatic tissues is by tethering or expressing targeting vectors onto the surface of EVs but this requires chemical or genetic modifications with potential drawbacks. There is an unmet need to direct systemically administered EVs to a desired site without genetic or chemical modifications. Methods Herein, we employed positron emission tomography imaging of 89 Zr-labeled EVs to explore the ability of various substances to alter EV trafficking in the body. Results and discussion Subcutaneous injections of phenothiazine and poly(ethylene) glycol into the hind limb of BALB/c mice significantly increased the accumulation of intravenously administered 89 Zr-labeled EVs to the hind limb. Expanding these findings, we investigated EVs isolated from various sources, the influence of timing of the administration of phenothiazine with respect to labeled EV administration, and the relationship of the trafficking of EVs to the site of phenothiazine administration with overall change in biodistribution. The findings support further exploration of phenothiazine analogs for site specific enhancement of EV trafficking in therapeutic applications.

BACKGROUND: Free fibula flaps have been widely used for bony reconstruction in pediatric patients. While donor site complications have been well studied in adults, data in the pediatric population remains limited. This study exclusively reports donor-site morbidity following fibula flap harvests in the pediatric and adolescent population. METHODS: We reviewed all patients younger than 18 years with open physes who underwent fibula flap harvests at a single academic institution from 1997 to 2024. Demographics, clinical and surgical characteristics, and donor-site complications were recorded. RESULTS: Fifty-seven free fibula flaps were harvested from 55 pediatric patients (mean age 12 years; 29 female) with a mean follow-up of 6 years. Flaps were most commonly used for lower extremity reconstruction (52%) and following tumor resection (74%). Osteoarticular flaps (n=8) were associated with transient deep peroneal nerve palsy in four cases, all of which recovered antigravity dorsiflexion by 6 months. In standard segmental harvests (n=49), donor-site complications occurred in 38%, most frequently valgus ankle deformity (9%) and neuropathic pain (6%). Valgus deformity was more likely in patients with shorter distal residual fibula and an age–residual fibula index (A-RFI) <16. CONCLUSIONS: Vascularized fibula flaps are safe and versatile for pediatric skeletal reconstruction. Donor-site complications are common but generally manageable, with full recovery of nerve function. Osteoarticular harvests carry the highest risk of peroneal nerve injury, and valgus ankle deformity may occur, particularly with an age–residual fibula index <16. Careful planning and age-adjusted fibular preservation are essential to optimize outcomes.

Background Female sex hormones used in feminizing hormone therapy (FHT) may influence activation and persistence of HIV pre‐exposure prophylaxis (PrEP) medications. The clinical relevance of this interaction remains unclear, particularly for transgender and cisgender women (TGW and CGW). To address this gap, we examined drug metabolites in HIV susceptible cells isolated from blood and rectal tissues of TGW and CGW. Materials and Methods We enrolled 23 TGW, 13 on FHT and 10 CGW. Participants received five directly observed daily PrEP doses: emtricitabine 200 mg with either tenofovir disoproxil fumarate 300 mg (FTC/TDF) or tenofovir alafenamide 25 mg (FTC/TAF). Blood and rectal tissue samples were obtained. Mononuclear and CD4 + cells were isolated and analysed for intracellular triphosphorylated nucleotides (TFVdp and FTCtp) and total nucleotide pools (pTFV and pFTC) using HPLC‐MS/MS. Serum hormone concentrations were quantified via enzyme immunoassay. Results and Conclusion Total nucleotide pools (pTFV and pFTC) correlated with active metabolites (TFVdp and FTCtp) in blood and rectal CD4 + cells ( r 2 > 0.75, p < .001). No significant differences in pTFV or pFTC concentrations were observed between high and low oestradiol groups in either CGW or TGW ( p > .1). No correlation was observed between serum oestradiol concentrations and nucleotide concentrations. TAF resulted in 6.8‐fold higher median pTFV concentrations in blood CD4 + cells ( p < .001) but eightfold lower median pTFV concentrations in rectal CD4 + cells compared to TDF ( p = .008). These findings suggest that neither endogenous hormonal fluctuations in CGW nor exogenous FHT in TGW adversely impact intracellular PrEP pharmacology in key HIV target cells.

Introduction Angina with no obstructive coronary artery disease (ANOCA) affects millions and is frequently under-recognised because diagnostic pathways and risk tools predominantly target obstructive coronary artery disease (CAD). This protocol describes shared methods for two machine-learning (ML) studies: (1) differentiating ANOCA from stable angina with obstructive CAD and (2) predicting long-term mortality among patients with ANOCA and obstructive CAD. Methods and analysis We will develop and cross-site validate ML classification models using a multicentre retrospective cohort drawn from the Alberta Provincial Project for Outcome Assessment in Coronary Heart Disease registry and institutional datasets from the University of Ottawa Heart Institute and the University Health Network. Eligible participants are adults (≥18 years) undergoing initial cardiac catheterisation for chest pain/anginal equivalents since 1995, excluding prior revascularisation, major structural heart disease and predefined non-anginal indications. Outcomes are (1) ANOCA (0% to <50% stenosis) versus obstructive CAD (≥50% stenosis) and (2) 1, 3 and 5-year mortality, modelled separately for ANOCA and obstructive CAD. Model development will use nested cross-validation with stratified k-fold inner-loop tuning and leave-one-site-out cross-validation for repeated external validation. Candidate predictors will be harmonised across sites, filtered for missingness and refined using expert/directed acyclic graph-guided selection plus Boruta and Least Absolute Shrinkage and Selection Operator. Preprocessing includes appropriate encoding, missing-data imputation (multivariate imputation by chained equations) and feature scaling. Algorithms will include elastic-net logistic regression, random forest, LightGBM and multilayer perceptron models; hyperparameters will be optimised via Bayesian optimisation. Performance and threshold tuning will be reported. Explainability and subgroup fairness will be assessed using SHapley Additive exPlanations. Final models will be deployed as a web-based clinical risk calculator. Ethics and dissemination Ethics approval has been obtained from the University of Calgary and the University Health Network (#24-5916). Analyses will use deidentified data in secure environments; only aggregate results will be reported. Findings will be disseminated via peer-reviewed publications, conferences and a web-based calculator.