Search Results (2,295)

Background/Objectives: Tumor-infiltrating lymphocyte (TIL) therapy is an important option for patients with metastatic melanoma progressing after standard systemic therapy, but real-world data on treatment delivery, toxicity monitoring, and immune recovery remain limited. We evaluated clinical outcomes, treatment tolerance, immune reconstitution, and cardiac biomarker dynamics across three Mayo Clinic sites. Methods: We retrospectively analyzed adults with metastatic melanoma who received lymphodepleting chemotherapy followed by TIL infusion and high-dose interleukin-2 (IL-2) between April 2024 and December 2025. Clinical outcomes, treatment delivery, and adverse events were assessed. Longitudinal immune monitoring included CD4 and CD8 T-cell counts, CD4:CD8 ratio, and immunoglobulin G (IgG) at baseline and follow-up. In a prespecified cardiac sub-cohort, high-sensitivity troponin (hs-Tn) was measured during IL-2 administration to evaluate associations with cardiac events and IL-2 interruption. Results: Thirty-six patients underwent TIL infusion. The objective response rate was 50.0%, including complete responses in 13.9%, and the disease control rate was 72.2%. Median progression-free survival was 3.61 months, and median overall survival was 12.94 months. M1d disease was associated with inferior overall survival on univariable analysis (HR 6.55, 95% CI 2.03–21.17; p = 0.002), with attenuation after multivariable adjustment. Receipt of ≥3 IL-2 doses was associated with longer overall survival on univariable analysis (HR 0.20, 95% CI 0.06–0.64; p = 0.007), but this association also attenuated after adjustment. Longitudinal immune monitoring demonstrated persistent CD4 lymphopenia through 6 months, sustained inversion of the CD4:CD8 ratio, and declining IgG at months 3 and 6. In the cardiac sub-cohort (24 patients; 87 IL-2 doses), post-dose hs-Tn ≥15 ng/L was associated with clinically significant cardiac events (OR 9.6, 95% CI 1.5–60.6; p = 0.016) and IL-2 interruption (OR 3.4, 95% CI 1.1–10.7; p = 0.036). For cardiac events, hs-Tn ≥15 ng/L had 100% sensitivity and 100% negative predictive value. Conclusions: In routine practice, TIL therapy was feasible and active in metastatic melanoma. M1d disease identified a subgroup with poor survival, peri-dose hs-Tn showed promise as a tool to support safer IL-2 delivery, and prolonged CD4 suppression with IgG decline suggests that recovery after TIL therapy extends beyond initial hematologic reconstitution. These findings support prospective validation of biomarker-guided IL-2 monitoring and extended post-treatment immune surveillance. Full article

Background/Objective: During the last two decades, several cases presenting with circulating clonal B-cells with features consistent with possible marginal zone (MZ) derivation have been described under different terminologies. The present study aims to shed more light onto the main disease characteristics and determine prognostic factors for outcome in 98 consecutive cases with clonal B-cell lymphocytosis of marginal zone origin (CBL-MZ). Methods: This is a multicenter retrospective analysis including 98 consecutive CD5(−) CBL cases referred to our Departments between 1999 and 2017. These cases were selected based on the presence of circulating CD5(−) clonal B-cells, irrespectively of their absolute number, without B-symptoms, organomegaly, lymphadenopathy or cytopenias or any other features consistent with a known lymphoproliferative disorder. Clinical, morphologic, biochemical, immunophenotypic, histologic and molecular features of CBL-MZ cases were analyzed. Results: The median absolute lymphocyte counts (ALCs) and circulating CBLs were 6.7 × 10⁹/L and 3.447 × 10⁹/L, respectively. Paraproteinemia was found in 38%. Bone marrow (ΒΜ) was involved in all but one case. MYD-88L265P mutation was positive in 11%. Two subcategories of CBL-MZ were identified: One was characterized by lower ALC/CBL, paraproteinemia, CD38 expression, BM lymphoplasmacytic morphology and more frequent MYD-88L265P mutation. The second category displayed a leukemic picture, higher frequency of CD11c expression, hypogammaglobulinemia and elevated LDH. Treatment-free survival (TFS) was 91%, and median freedom from progression (FFP) was 95.6 months. For TFS, two factors proved significant using multivariate analysis: BM infiltration ≥ 50% and elevated LDH (RR 5.6 and 5.4, respectively). Evolution to splenic marginal zone lymphoma was a rare event (5%). A novel pattern of progression emerged, namely development of cytopenias due to extensive BM infiltration without any other disease localization. Conclusions: CBL-MZ is an indolent lymphoproliferative disorder with excellent outcome and low probability of progression. Based on our findings, CBL-MZ represents a heterogeneous entity where the vast majority of cases remain stable or develop increasing lymphocytosis. Clinically, our data also highlight that the extent of BM infiltration and elevated LDH levels appear to be the most notable predictors for introducing therapy. Full article

This study aimed to investigate the expression of immune checkpoint-associated proteins in phyllodes tumors (PTs) and assess their clinicopathologic and prognostic significance. Surgical resection specimens from 200 patients were included, and the expressions of CD24, Siglec-10, CD47, and SIRPα in both the epithelial and stromal components of the tumor were assessed, with ≥1% positivity considered positive. Of 200 cases, 145 were benign, 44 borderline, and 11 malignant. The expressions of CD24, Siglec-10, CD47, and SIRPα in stromal cells increased with tumor grade: CD24 was associated with stromal cellularity, atypia, and mitosis; Siglec-10 with stromal cellularity and atypia; CD47 with stromal atypia and mitosis; and SIRPα with stromal overgrowth and atypia. While expressions of CD24, CD47, and SIRPα were associated with either shorter disease-free survival (DFS) or shorter overall survival, multivariate analysis identified stromal CD24 expression as an independent predictor of shorter DFS. Immune checkpoint-associated proteins, particularly stromal CD24, CD47, and SIRPα, are associated with adverse features and poor outcomes in PTs, implicating potential prognostic and therapeutic relevance. Full article

The Santiago River is a highly anthropogenically impaired lotic system globally, yet the mechanisms governing its contaminant transport remain poorly understood under static monitoring paradigms. This study evaluates how hydrological forcing dictates the mobilization and bioavailability of trace metals by integrating a 15-year public hydrochemical database from 10 monitoring nodes with SAR-derived discharge estimates and thermodynamic metal modeling (PHREEQC). To validate the structural integrity of the mass load estimates against hydrometric uncertainties, a deterministic boundary-sensitivity analysis was conducted. Results empirically refute the classical dilution paradigm, introducing the “Anthropogenic Pulse” to describe the non-linear acceleration of pollutant export during high-flow events (discharge Q surging from 36.62 to 286.13 m³/s). While climate-driven parameters follow seasonal cycles, industrial stressors (COD, Pb, Cd) remain in a chronic steady state, decoupling from volumetric dilution. Based on coupled × CQ × C (discharge × concentration) estimates, this dynamic induces a synchronized flushing of toxic burdens, exporting monthly peak loads exceeding 51,000 kg of Zinc, 6500 kg of Lead, and 3100 kg of Cadmium. Thermodynamic simulations reveal that this hydrological flushing functions as a chemical activator; the seasonal dilution of natural Alkalinity and Hardness suppresses the river’s theoretical buffered pH (from 8.5 to 7.0), maintaining metals in their uncomplexed free-ion states (Me²⁺). Modeling indicates that nearly 90% of the exported Cadmium remains in this highly labile, toxic form due to a dual coupling with both river Discharge (r_s = 0.87) and pH (r_s = 0.79). The identification of stochastic arsenic peaks 100 times above regulatory limits at Paso de Guadalupe (RS-08) underscores the failure of concentration-based monitoring. Our findings suggest that restoration strategies should shift toward mass-loading-based regulatory frameworks and targeted sediment management at critical nodes to mitigate the chronic export of bioavailable industrial waste. Full article

Background/Objectives: Mucinous adenocarcinoma (MAC) is a rare and clinically problematic subtype of rectal cancer, tending to present at an advanced stage and to respond poorly to neoadjuvant therapy. The consistently worse prognosis than that of not-otherwise-specified adenocarcinoma (NOS-AC) is not fully understood, potentially owing to intrinsically more aggressive biology or specific immune evasion mechanisms. We used the IMMUNOREACT multicentre cohort, with external validation in TCGA, to investigate the clinical and immunological features of rectal MAC in detail. Methods: Two hundred patients with rectal adenocarcinoma (16 MAC, 184 NOS-AC) from the IMMUNOREACT 1 (NCT04915326) and IMMUNOREACT 2 (NCT04917263) prospective cohorts were included. To account for the imbalance in baseline characteristics, propensity score matching (PSM) was performed on age, sex, neoadjuvant treatment and TNM stage. The immune microenvironment was characterised using immunohistochemistry (CD3, CD4, CD8, CD8β, Tbet, FoxP3, PD-L1, MSH6, PMS2, CD80), flow cytometry and NanoString PanCancer IO 360™ transcriptomics of adjacent healthy mucosa. Findings were externally validated against TCGA rectal and colon adenocarcinoma datasets. Results: MAC presented at significantly more advanced stage than NOS-AC across all TNM parameters: higher T stage (p = 0.006), N stage (p < 0.001), M stage (p = 0.039) and overall TNM stage (p < 0.001). In the unmatched cohort, MAC was associated with worse overall survival (HR 2.53; 95% CI 1.03–6.23; p = 0.043) and disease-free survival (HR 2.86; 95% CI 1.25–6.55; p = 0.013), but both differences became non-significant after PSM. MAC patients had higher haemoglobin after adjusting for confounders (mean difference [MD] 1.26 g/dL, 95% CI 0.30–2.31, p = 0.012), consistent with a hypothesis of reduced chronic rectal bleeding as a possible mechanism for late presentation. Transcriptomically, MAC showed suppression of HLA class II antigen presentation genes (HLA-DQA1, HLA-DQB1, HLA-DRB1) and myeloid activation genes (S100A8/A9/A12) in adjacent healthy mucosa. Loss of MMR proteins MSH6 and PMS2 in histologically normal mucosa was significantly more frequent in MAC. These findings were replicated in the TCGA cohort, which also showed lower tumour mutational burden and a distinct mucin-associated transcriptomic profile in MAC. Conclusions: The worse outcomes of rectal MAC appear to be driven largely by late-stage presentation, possibly owing to later diagnosis. MAC nonetheless carries a distinct immune phenotype, detectable even in histologically normal surrounding mucosa, that likely contributes to its treatment resistance. These observations provide a basis for developing histotype-specific approaches to both early detection and treatment in this uncommon but clinically challenging tumour subtype. Full article

Background/Objectives: Ursodeoxycholic acid (UDCA) is a bile acid widely used for the treatment of cholestatic liver diseases; however, its poor aqueous solubility represents a major limitation for the development of oral liquid formulations, particularly in pediatric patients requiring accurate and flexible dosing. This study aimed to develop and characterize a fully solubilized extemporaneous UDCA oral formulation using the ready-to-use vehicle Wagner, with particular emphasis on the role of hydroxypropyl-β-cyclodextrin (HP-β-CD) as a solubilizing excipient. Methods: Phase-solubility studies, Job’s plot analysis, and ¹H NMR spectroscopy were performed to investigate the host–guest interaction between UDCA and HP-β-CD, confirming the formation of a stable 1:1 inclusion complex responsible for a marked increase in drug solubility. The aqueous solubility of UDCA increased from approximately 0.02 mg/mL in water to 31 ± 1 mg/mL in the Wagner base containing HP-β-CD, compared to ~10 mg/mL in the corresponding cyclodextrin-free vehicle. Chemical stability was evaluated using an HPLC method adapted from the European Pharmacopoeia, employing dual detection (refractive index and photodiode array detector) to ensure specificity and stability-indicating capability. Results: The UDCA solution (20 mg/mL) remained chemically stable for at least 4 months under refrigerated (4–8 °C) and room temperature (25 °C) conditions, with only moderate degradation observed at 40 °C. Physical stability studies confirmed the absence of precipitation, phase separation, or significant pH variations under all storage conditions. Conclusions: Wagner-based formulation enabled the development of a stable and homogeneous UDCA oral solution, providing a complementary formulation strategy to conventional suspension-based preparations. This approach represents a robust and patient-oriented strategy for extemporaneous compounding, particularly suitable for pediatric use. Full article

Background: This study aimed to construct a recombinant Pseudomonas aeruginosa outer membrane vesicle (OMV) vector vaccine delivering pcrV and compare the immunological impacts of OMVs as carriers versus as adjuvants. Methods: The recombinant plasmid pBBRMCS5-pcrV was constructed and transformed into P. aeruginosa. Recombinant OMVs (OMV_PcrV) were prepared via ultracentrifugation and characterized in terms of their morphology and particle size by means of transmission electron microscopy (TEM) and nanoparticle tracking analysis (NTA). After a biosafety evaluation, mice were intramuscularly immunized with PcrV or OMV_PcrV, followed by a booster immunization on day 21. On day 42, the mice were challenged subcutaneously and intranasally with PAO1. Bacterial loads in tissues and blood, pulmonary T-cell subsets, and serum antibody levels were assessed. Results: The recombinant plasmid was successfully constructed, and Western blotting confirmed the delivery of PcrV into OMVs. TEM revealed typical spherical nanostructures, and NTA showed a median particle size of 127.4 ± 5.3 nm. Upon subcutaneous challenge, the OMV, OMV_PcrV, and OMV + PcrV groups all achieved 100% protection. Both the OMV_PcrV and OMV + PcrV groups exhibited increased CD4⁺ and CD8⁺ T-cell counts and higher induction levels of specific IgM, IgG1, and IgG2a antibodies. The OMV_PcrV group showed superior clearance of respiratory bacterial colonization and reduced inflammatory injury compared with the PBS control group. Conclusions: The constructed vector successfully delivered the PcrV antigen, and the OMVPcrV vaccine induced effective immune responses. Compared with wild-type outer membrane vesicles (OMVs) and the strategy of directly mixing free PcrV antigen with OMVs (OMV + PcrV), the recombinant OMV_PcrV vaccine exhibited superior immunoprotective efficacy in terms of bacterial clearance and tissue protection, providing experimental evidence for the development of a Pseudomonas aeruginosa vaccine. Full article

Background/Objectives: The gene solute carrier family 52 member 3 (SLC52A3) encodes riboflavin transporter-3, a transmembrane protein essential for riboflavin absorption. Emerging evidence suggests that metabolic transporters may play a role in tumor biology. This study aimed to investigate the expression patterns, prognostic significance, genetic alterations, and functional associations of SLC52A3 in gynecological cancers. Methods: A comprehensive bioinformatic analysis was conducted using multi-omics datasets from The Cancer Genome Atlas (TCGA). Gene expression and survival analyses were performed via GEPIA3. Genetic alterations, including mutations and copy number variations, were assessed using cBioPortal. Immune infiltration correlations were analyzed through TIMER3. Protein–protein interactions and gene enrichment analyses were performed using STRING and GEPIA2, followed by Gene Ontology (GO) and KEGG pathway analyses. Results: SLC52A3 expression was significantly upregulated in ovarian, cervical, and endometrial cancers. Reduced expression of SLC52A3 was associated with poorer overall survival and shorter progression-free interval specifically in endometrial cancer. Genetic alterations in SLC52A3 were not significantly associated with survival outcomes (OS, DFS, and PFS). Functional enrichment analysis indicated that SLC52A3 is involved in biological processes such as cell junction organization and protein localization to the plasma membrane. Additionally, SLC52A3 expression showed positive correlations with genes implicated in tumor progression and metastasis, including NECTIN4, PROM2, TACSTD2, PKP3, SEMA4B, and CD46. Conclusions: These findings suggest that SLC52A3 may serve as a potential prognostic biomarker in endometrial cancer and could play a role in tumor progression pathways. Its functional associations highlight its potential relevance as a therapeutic target, warranting further experimental validation. Full article

Background: Plasticizers, including phthalate esters and phthalate-free alternatives, are widely detected environmental chemicals. Although increasing evidence suggests that plasticizers may disrupt gastrointestinal homeostasis, their potential molecular links with inflammatory gastrointestinal disorders (IGDs) remain unclear. Methods: This study aimed to systematically identify potential molecular targets and pathways linking representative plasticizers with IGDs. An integrative network toxicology framework was applied to investigate four plasticizers, including dimethyl phthalate (DMP), diethyl phthalate (DEP), dioctyl phthalate/di(2-ethylhexyl) phthalate (DOP/DEHP), and acetyl tributyl citrate (ATBC), in relation to Crohn’s disease (CD), ulcerative colitis (UC), esophagitis, and gastritis. Plasticizer- and disease-related targets were collected from public databases, followed by overlapping target screening, protein–protein interaction network analysis, functional enrichment analysis, GEO-based transcriptomic validation, molecular docking, molecular dynamics simulation, and single-cell RNA-seq analysis. Results: Disease-specific candidate targets were identified, including CXCL8 and FN1 for CD, IL1B for UC, MAPK3, FASN, FN1, PPARG, CXCL8, FOS, and HIF1A for esophagitis, and MMP9, TNF, TLR4, IL6, CCR2, IFNG, and PTGS2 for gastritis. Cross-disease analysis further identified plasticizer-associated signature targets, including MMP7 for DMP, HMOX1 and NOS2 for DEP, and LTF and CCL11 for ATBC. Enrichment analysis indicated that these targets were mainly involved in inflammatory, chemokine, MAPK-related, and xenobiotic response pathways. Molecular docking and dynamics simulations suggested stable interactions between selected plasticizers and candidate targets, while single-cell analysis revealed their cell-type-specific expression patterns in epithelial, immune, and stromal compartments. Conclusions: This study provides an exploratory network toxicology framework for identifying potential molecular associations between plasticizer exposure and IGDs. The findings highlight disease-specific and plasticizer-associated candidate targets that may guide future experimental validation and environmental risk assessment. Full article

Pediatric acute myeloid leukemia (pedAML) is a childhood malignancy with relapse rates of approximately 30%. CD68, a macrophage marker involved in phagocytosis and macrophage recruitment, may contribute to AML biology. We analyzed CD68 expression using the TARGET database and performed survival analyses, mRNA/protein profiling, and functional assays in AML cell lines, pedAML samples, and cord blood samples. High CD68 transcript levels correlated with KMT2A-rearrangements and inversion 16. Survival analysis showed that high CD68 predicted worse event-free survival, though not independently in a multivariate analysis. Flow cytometry confirmed higher CD68 expression in 7/8 pedAML samples compared to cord blood samples. Functionally, CD68 knockdown reduced proliferation and increased drug sensitivity, while overexpression promoted growth and resistance. Gene set enrichment analysis (GSEA) indicated enrichment of MAPK signaling, AP-1–mediated stress response, and epithelial–mesenchymal transition (EMT)/migration-associated pathways in CD68-high models. Together, these findings suggest that CD68 contributes to a pro-tumorigenic and stress-adaptive phenotype in pedAML and may represent a biologically relevant therapeutic target. Full article

Epcoritamab, a subcutaneous CD3×CD20 bispecific antibody, has shown substantial activity in relapsed or refractory (R/R) B-cell lymphomas, but the immunological correlates of durable remission and treatment discontinuation remain unclear. We retrospectively analyzed 21 consecutive patients who initiated epcoritamab at our institution between 1 December 2023 and 31 December 2025, including 17 with R/R large B-cell lymphoma (LBCL) and 4 with R/R follicular lymphoma (FL). Clinical follow-up was updated through 18 May 2026. Serial cytotoxic T lymphocyte (CTL) subset and T-cell receptor (TCR) Vβ repertoire analyses were performed in selected cases. Among response-evaluable patients, the overall response rate was 9/14 in LBCL and 4/4 in FL. Median overall survival was 431 days in LBCL and 431.5 days in FL. Progression-free survival was analyzed descriptively because of the small sample size and substantial censoring. A patient with clinically and radiologically suspected central nervous system relapse of LBCL achieved radiological complete remission after epcoritamab treatment. In two LBCL and one FL case in whom epcoritamab was electively discontinued after complete remission, Vβ-skewed CTL populations were observed, and total memory CTLs exceeded total effector CTLs at discontinuation. These exploratory findings suggest that epcoritamab treatment may be associated with longitudinal remodeling of CTL subsets and Vβ-skewed CTL populations in selected responders. The potential relevance of these immunological patterns to durable response and treatment discontinuation should be validated in larger prospective cohorts with functional and sequence-based T-cell analyses. Full article

Human metapneumovirus (HMPV) is a major cause of respiratory infections, but its impact on macrophage antibacterial functions remains poorly understood. Macrophages play a crucial role in host defense through phagocytosis, and impairment of this function may increase susceptibility to secondary infections. Here, we show that HMPV infection of THP-1-derived macrophages significantly reduces bacterial uptake in a replication-dependent manner. This effect was restricted to infected cells and was not recapitulated by cell-free supernatants, indicating a cell-intrinsic mechanism. HMPV infection was also associated with reduced expression of the scavenger receptor CD36. Viral gene knockdown studies further implicated the HMPV G protein in this phenotype, as silencing the G protein restored phagocytic function. Analysis of single-cell RNA-sequencing datasets from HMPV-infected mouse lungs revealed reduced CD36 expression and broader alterations in phagocytosis-associated gene programs across lung macrophage subsets. Supporting these observations, expression of Cd36 and Marco was reduced in lung tissue from HMPV-infected mice. Overall, these findings demonstrate that HMPV impairs macrophage-mediated bacterial uptake through a replication-dependent, cell-intrinsic mechanism and identify reduced scavenger receptor expression and the viral G protein as factors associated with this phenotype. These alterations may contribute to increase susceptibility to secondary bacterial infections during HMPV infection. Full article

Background and Aim: The gluten-free diet (GFD) can have a huge impact on the quality of life of people with celiac disease (CD), especially on a social level. The objective of this work is to evaluate a structured nutrition education program focused on CD and GFD that aims to increase knowledge and improve inclusion attitudes about the disease in children. Methods: This is a one-month intervention for school children aged 10–12 years called Zeliakide (8 sessions). It was carried out through a STEAM methodology, using inquiry-based learning. The participants responses were evaluated through questionnaires before and after the intervention, and participants were also followed up one month later. The control group was a similar group of students who followed their regular school curriculum. Results: 299 children from one school of Vitoria-Gasteiz took part in the study (155 intervention group; 144 control group). Zeliakide significantly improved knowledge about CD and GFD in children, and this knowledge was retained for one month. Concretely, students increased their ability to explain what CD is, to assess gluten, and to classify food groups according to gluten content. The intervention contributed to augmenting the selection of behaviors to overcome differences between individuals, assessed one month after the intervention. In addition, the program allowed students to understand the work of scientists. Conclusions: Zeliakide can contribute to nutrition education initiatives that aim to improve knowledge of CD and GFD in the general population, while promoting empathetic behavior towards people with CD. Registration: clinicaltrials.gov, NCT05467865 on 21 July 2022. Full article

Background: Bronchoscopic cryotherapy may enhance anti-tumor immunity and improve the effect of immune checkpoint blockade, but CD8+ T cell dynamics after cryotherapy combined with pembrolizumab-based therapy in metastatic non-small cell lung cancer (NSCLC) remain insufficiently characterized. Methods: In this prospective, exploratory, randomized, controlled, single-center study, metastatic NSCLC patients were assigned to bronchoscopic cryotherapy performed 7 ± 1 days before first-line pembrolizumab-based therapy or to standard treatment alone. Peripheral blood mononuclear cells were analyzed by flow cytometry at baseline, week 3, and week 6. CD8+ T cell subsets defined by CD45RO, CD28, granzyme B (GzB), IFNγ, Ki-67, and PD-1 were evaluated in relation to treatment group, radiologic response, progression-free survival (PFS), and overall survival (OS). Results: Flow cytometry was performed in 76 patients, including 34 in the cryotherapy group and 42 in the control group. Cryotherapy was associated with a treatment-specific increase in circulating GzB+ CD8+ T cells by week 6. In contrast, Ki-67+ CD8+ and GzB+Ki-67+ cells increased in both treatment groups, suggesting that early peripheral CD8+ proliferation was largely shared across pembrolizumab-based therapy. Radiologic responders demonstrated more sustained proliferative CD8+ dynamics, most consistently reflected by increased CD28+ Ki-67+ cells. In exploratory landmark survival analyses, a higher week-3-to-baseline GzB+ Ki-67+ CD8+ ratio showed a hypothesis-generating association with longer PFS and OS. Conclusions: In metastatic NSCLC patients receiving first-line pembrolizumab-based therapy, the addition of bronchoscopic cryotherapy was associated with peripheral CD8+ T cell remodeling toward enhanced cytotoxic activity. Proliferative CD8+ T cell changes were associated with radiologic response and better survival. These findings support bronchoscopic cryotherapy as a potential immune-modulating adjunct and warrant validation in larger studies. Full article

Background/Objectives: Celiac disease (CD) is a T-cell-mediated autoimmune condition, triggered by gluten ingestion. Duodenal biopsy is the gold-standard diagnosis for CD, which is often limited by interobserver variability between pathologists. Immunohistochemistry (IHC) is a powerful technique for detecting biomarkers with potential diagnostic significance. This study aims to investigate five candidate biomarkers, BTNL8, NKp46, TdT, THEMIS, and TCRδ, that might improve the reproducibility of the diagnosis of CD. Methods: Formalin-fixed paraffin-embedded material, surplus to diagnostic requirements, was obtained from 46 subjects (untreated CD: n = 21, CD treated with gluten-free diet: n = 5; controls: n = 20) and immunostained for BTNL8, NKp46, TdT, THEMIS and TCRδ. BTNL8 staining was scored on a 0–3 semi-quantitative scale. NKp46, TdT, THEMIS, and TCR delta-positive intra-epithelial lymphocytes (IELs) were quantified as mean counts per 100 epithelial cells (ECs). Results: TCRδ-positive IELs were markedly elevated in CD biopsies (median 9.4 IELs/100 ECs) compared to healthy controls (median 0.5 IELs/100 ECs; p < 0.001), with a threshold of >2.1 TCRδ-positive IELs per 100 ECs yielding an AUC of 0.94 and interobserver agreement of 0.82. NKp46 expression was also increased in CD (median 13.8 IELs/100 ECs) versus controls (median 9.6; p < 0.001), with >12.8 NKp46-positive IELs per 100 ECs achieving an AUC of 0.86 and interobserver agreement of 0.82. Immunostaining for the other biomarkers demonstrated less clear differences between CD and healthy controls. Conclusions: Corroborating several recent publications, TCRδ immunostaining provides high diagnostic accuracy and good interobserver agreement in the diagnosis of CD on duodenal biopsy, even for patients on a gluten-free diet. Full article