Clinical Epigenetics

BackgroundAgeing is a sex-specific process characterised by a progressive decline in physiological integrity. DNA methylation represents a primary epigenetic hallmark of ageing, yet sex-specific patterns of epigenetic ageing within and across tissues remain poorly understood. This study aims to address these gaps through an integrated analysis of sex-moderated epigenetic ageing across eight human tissues. MethodsA total of 137 DNA methylation datasets comprising over 36,000 individuals aged 10-114 years were analysed using a meta-analytic workflow to identify age-associated differentially methylated positions (aDMPs) and regions (aDMRs), meta-regression to assess sex moderation, and pathway enrichment analyses to interpret functional relevance. FindingsIndividual tissues displayed distinct age-related methylation trajectories, but some DMP sites showed consistent hyper- or hypomethylation across tissues. Across tissues, we identified 68,630 aDMPs (10%) robustly associated with ageing. Age-associated changes at the regional level were less common, with only 80 robust age-associated aDMRs detected across tissues, representing 0.09% of analysed regions. Sex moderation was observed for only 16 aDMPs (0.002%), indicating that sex effects on age-associated DNA methylation are largely tissue-specific rather than shared across tissues. InterpretationOur findings indicate that age-associated DNA methylation changes predominantly occur at isolated CpG sites rather than extended genomic regions and are strongly dependent on tissue and genomic context. The minimal overlap of sex-moderated methylation signals across tissues suggests that age-related sex differences at the epigenetic level are more likely attributable to tissue- and cell-type-specific variation rather than to broadly conserved epigenetic mechanisms shared across tissues. FundingThis study was funded by an Australian Research Council (ARC) Discovery project (DP200101830). Severine Lamon was funded by an ARC Future Fellowship (FT210100278). Nir Eynon was funded by NHMRC Investigator Grant (APP1194159), and a Hevolution/AFAR New Investigator Award in Aging Biology and Geroscience Research. Mandhri D. Abeysooryia was supported by an Australian Government Research Training Program (RTP) Scholarship. Research in context Evidence before this studyDNA methylation is widely recognised as a central epigenetic hallmark of ageing. Previous research has demonstrated that some age-related methylation changes are conserved across tissues, forming the basis of pan-tissue epigenetic clocks. Most studies to date have primarily examined age effects in isolation. Although biological sex influences ageing trajectories and susceptibility to nearly all age-related diseases, sex-moderated epigenetic ageing has received limited investigation. Specifically, pan-tissue clocks, including GrimAge and PhenoAge, are "sex-aware" but were trained and validated in mixed-sex cohorts, limiting their capacity to disentangle tissue-specific sex effects. Consequently, it remains unclear whether sex-moderated epigenetic ageing signals are shared across tissues or are tissue-specific. Added value of this studyThis study provides a large-scale, comprehensive multi-tissue analysis of sex-moderated epigenetic ageing, integrating 137 DNA methylation datasets across eight human tissues and more than 36,000 male and female individuals spanning the lifespan. Our findings show that while age-associated methylation changes are widespread at the CpG level, sex-moderated effects are rare and largely tissue-specific, with minimal overlap across tissues. Implications of all the available evidenceTogether, the available evidence indicates that epigenetic ageing is predominantly driven by shared, conserved age-related methylation changes, whereas sex differences in epigenetic ageing are modest and context dependent. These sex-related effects are more likely to reflect tissue- and cell-type-specific variation rather than widespread, shared mechanisms. This underscores the need to develop sex-specific epigenetic clocks and to conduct longitudinal cohort and intervention studies to more precisely characterise sex-specific dynamics of epigenetic ageing across tissues.

BackgroundX chromosome inactivation (XCI) is the mechanism which randomly silences one X chromosome to equalise gene expression between 46, XX females and 46, XY males. Though XCI is expected to result in a random pattern of mosaicism across tissues, some females display a significantly unbalanced ratio in immune cells, termed XCI-skew, in which [&ge;]75% of cells have the same X inactivated. XCI-skew is associated with adverse health outcomes and its prevalence increases with age - particularly after midlife - yet the specific risk factors have yet to be identified. The menopausal transition, which is driven by profound shifts in sex hormone levels, has significant impact on chronic disease risk yet the molecular and cellular effects are incompletely understood. We hypothesised that the menopausal transition may impact XCI-skew. MethodsUsing XCI data measured in blood-derived DNA from 1,395 females from the TwinsUK population cohort, along with questionnaires, genetic data, and sex hormone measures, we carried out a cross-sectional study to assess the impact of the menopausal transition and sex hormones on XCI-skew. ResultsWe demonstrate that early menopause (<45yrs) is associated with increased risk of XCI-skew. In subset analyses across those who had a surgically induced or natural menopause, we find the association restricted to those who underwent a surgical menopause. We next identify a low polygenic score (PGS) for testosterone levels is significantly associated with XCI-skew, which we replicate in an independent dataset (n=149), while a PGS for age at natural menopause is not associated. Finally, using longitudinal measures across two time points spanning [~]18 years we show XCI-skew is a stable cellular phenotype that typically increases over time. DiscussionThese data represent the first environmental and genetic risk factors of XCI-skew, both of which implicate endogenous sex hormone levels, particularly testosterone. We propose XCI-skew may have clinical relevance in postmenopausal females.

Glycosylation is a key structural modification of immunoglobulin G (IgG) that modulates its effector functions and has multiple roles in balancing inflammation. Altered IgG glycosylation has been reported in many diseases, often years before clinical manifestation, suggesting its causal role and biomarker potential. Here, we analyzed IgG glycome composition in 20,405 individuals from 42 different studies processed at the Genos Glycoscience Research Laboratory between 2008 and 2025. Across nearly all diseases, specific IgG glycome profiles reflected accelerated biological aging. Accelerated glycan aging was strongly associated with increased risk of all-cause mortality, independent of established clinical risk factors and potential confounders. Moreover, interventions known to reduce mortality risk, including hormone replacement therapy, therapeutic plasma exchange and caloric restriction, were associated with reversal of glycan aging. Given their role in modulating low-grade systemic inflammation, IgG glycans may represent a functional link between chronic inflammation, aging, disease susceptibility and all-cause mortality.

Background: Elevated lipoprotein(a) [Lp(a)] is a known risk factor for several cardiovascular-related diseases established from multiple genetic and observational studies. However, the underlying mechanisms mediating the effects of Lp(a) levels on cardiovascular disease risk and major adverse cardiovascular events (MACE) are unclear. The aim of this study was to identify proteins downstream of Lp(a) using mendelian randomization (MR) - a genetic causal inference approach. Methods: A two-sample MR was performed by initially identifying Lp(a) genetic instruments based on data from genome wide association studies (GWAS) of Lp(a) blood concentrations. These instruments were then tested for association with proteins from proteomic pQTL data (Olink from UK Biobank, 2940 proteins and SomaScan from deCODE, 4907 proteins). Results: A total of 521 proteins associated with Lp(a) were identified. Using pathway enrichment analysis, the following MACE-relevant pathways were identified comprising a total of 91 Lp(a) downstream proteins: oxidized phospholipid-related, chemotaxis of immune cells and endothelial cell activation, pro-inflammatory monocyte activation, neutrophil activity, coagulation, and lipid metabolism. Conclusion: The results suggest that the influence of Lp(a) treatments is primarily through modifying inflammation rather than lipid-lowering, thus providing insight into the mechanistic framework which mediates the effects of elevated Lp(a) on atherosclerotic cardiovascular disease.

Pregnancy complications such as preeclampsia are associated with circulating cell-free mitochondrial DNA (mtDNA), a damage-associated molecular pattern capable of activating Toll-like receptor 9 (TLR9). We hypothesized that acute mtDNA exposure induces maternal inflammation and endothelial dysfunction during pregnancy via TLR9 activation. Non-pregnant and pregnant rats (gestational days 14-15) were treated intravenously with saline or purified mtDNA and euthanized 4 h after treatment. mtDNA increased cytokine mRNA expression in lung and liver of non-pregnant and pregnant rats, with magnitude varying by pregnancy status and organ. Aortas from pregnant, but not non-pregnant, rats exhibited reduced acetylcholine (ACh)-induced relaxation following mtDNA treatment (Emax, saline: 90.1 {+/-} 3.9 % vs. mtDNA: 62.1 {+/-} 20.7 % KClmax, p<0.05), while uterine artery function was preserved, indicating vascular bed-specific effects. Ex vivo incubation of aortic rings with mtDNA {+/-} white blood cells did not replicate in vivo findings, implicating systemic rather than direct vascular mechanisms. Nuclear DNA did not affect ACh-induced relaxation (p>0.05), confirming that the vascular effects were mtDNA-specific. Pharmacological antagonism of TLR9 with ODN2088 partially attenuated mtDNA-induced maternal endothelial dysfunction. Although overt vascular ROS increases were not detected, aortas from pregnant rats had reduced sod-1 expression (p<0.05) and increased eNOS protein abundance (p<0.05). Acute mtDNA exposure during pregnancy induces maternal organ inflammation and impairs endothelium-dependent vasodilation, with partial TLR9 involvement. In conclusion, aortic transcriptional changes in antioxidant pathways and increased eNOS abundance were also observed, though their functional significance remains to be determined. New & NoteworthyTo our knowledge, this is the first study to demonstrate that acute exposure to circulating mtDNA induces pregnancy-specific maternal endothelial dysfunction and organ-selective inflammatory responses. Our findings reveal pregnancy- and vascular-bed specific responses of the maternal vasculature to mitochondrial danger signals, with partial TLR9 involvement. Aortic transcriptional changes in antioxidant pathways and increased nitric oxide synthase abundance were identified as molecular correlates of this dysfunction.

Common sex chromosome aneuploidies (SCAs) often present with cognitive and cardiovascular dysfunction in humans. To address SCA effects on gene expression and DNA methylation in relevant cell types, we differentiated neural precursor cells (NPCs) and cardiomyocytes (CMs) from human induced pluripotent stem cells (hiPSCs) with different numbers of sex chromosomes, including isogenic and independent lines. As expected, the expression of genes that escape X inactivation (escapees) mostly increases with the number of inactive X chromosomes (Xi). However, allelic analysis shows dampening of escapees specifically on the Xi in XXY compared to XX in both NPCs and CMs, revealing a novel type of dosage compensation in SCA. In contrast, Y-linked gene expression is higher in XXY versus XY NPCs, but the opposite is observed in CMs. This may explain the greater number of differentially expressed autosomal genes in NPCs versus CMs with an added Y chromosome, while effects of added X chromosomes are similar between cell types. Concordantly, changes in autosomal DNA methylation are mainly driven by the presence of a Y chromosome. These findings highlight the cell-type specificity of sex-linked and autosomal gene regulation in SCA conditions. HighlightsO_LISex chromosome aneuploidy induces cell-type specific changes in gene expression C_LIO_LIDampening of the inactive X chromosome in XXY alleviate X overexpression C_LIO_LIHigh Y-linked gene expression in XXY neuronal precursor cells but not cardiomyocytes C_LIO_LISex chromosome aneuploidy disrupts sex biases in autosomal gene expression C_LI

BackgroundThe retired antigen hypothesis, introduced by Tuohy and colleagues, proposes that tissue-specific proteins expressed conditionally during early life or reproductive stages, then silenced in normal aging tissue, represent safe and effective cancer vaccine targets when re-expressed in tumors. To date, discovery of retired antigens has relied entirely on hypothesis-driven wet lab work, limiting throughput. MethodsHere we present RADAR (Retired Antigen Discovery and Ranking), a multi-omics computational pipeline implemented on a standard server that systematically identifies retired antigen candidates. RADAR comprises four core discovery layers integrating: 1) The Genotype-Tissue Expression Portal (GTEx) normal tissue expression, 2) TCGA tumor re-expression, 3) DNA methylation, and 4) miRNA regulatory networks, each applied sequentially to identify genes exhibiting the epigenetic and post-transcriptional hallmarks of tissue-specific retirement followed by tumor re-activation. Candidate characterization is further supported by three automated modules: 1) protein-level safety screening via the Human Protein Atlas, 2) molecular subtype enrichment analysis, and 3) cross-cancer confirmation, which execute automatically when the relevant data are available for the selected cancer type. ResultsThe pipeline independently validated known targets including alpha-lactalbumin (LALBA, the basis of the Tuohy Phase 1 triple-negative breast cancer vaccine trial) and anti-Mullerian hormone (AMH), consistent with Tuohys ovarian cancer vaccine program targeting AMHR2, and rediscovered multiple known cancer-testis antigens (MAGEA1, MAGEC1, SSX1) as positive controls. Among 4,664 initial candidates derived from GTEx, the pipeline identified 20 high-confidence retired antigen candidates passing all filters. DCAF4L2, COX7B2, TEX19, and CT83 emerge as the highest-priority novel candidates for experimental validation, demonstrating zero expression in critical somatic organs, strong epigenetic silencing, and significant re-expression across multiple cancer types. ConclusionRADAR provides the first systematic computational framework for retired antigen discovery, offering a reproducible and scalable approach to expanding the cancer immunoprevention pipeline beyond individually characterized targets. The pipeline is fully reproducible, requires no specialized hardware, and is immediately extensible to additional TCGA cancer types.

Socioeconomic inequalities in health and well-being are a major public health concern, particularly in ageing populations. Education is a key determinant shaping multiple aspects of health outcomes. We used cross-sectional data from wave 9 of the German sample (n=4,148) of the Survey of Health, Ageing and Retirement in Europe (SHARE) to test whether formal education is associated with well-being in later adulthood, with health literacy, self-rated health, and preventive health behaviours as possible mediators. Our results showed that education was positively associated with greater well-being, but only via indirect pathways. Specifically, self-rated health, health literacy, and fruit and vegetable consumption mediated the relationship between education and well-being accounting for 54.7, 24.7, and 12.6 percent of the total effect, respectively. In addition, there were significant positive correlations between education and health literacy, as well as high-intensity physical activity, daily fruit and vegetable consumption, more preventive health check-ups, and less smoking. In contrast, alcohol consumption was more common among those with higher levels of education. All health behaviours and health literacy were correlated directly or indirectly (i.e., mediated by health) with well-being. These findings highlight the importance of examining indirect pathways linking education to well-being in later life. Interventions aimed at improving health literacy and promoting healthy behaviours may help reduce educational inequalities in quality of life among older adults.

Background: The etiology and nosological status of seasonal affective disorder (SAD) as a specifier of depressive episodes versus a transdiagnostic disorder are the subject of debate. In this study, we investigated the underlying etiology of SAD and dimensional seasonality by examining their association with latitude and genetic risk for a range of traits, and investigated gene-environment interactions. Methods: This study included 12,460 adults aged 18-90 with a history of depression from the Australian Genetics of Depression Study. Regression models included predictors for latitude (distance from equator) and polygenic scores for eight traits; major depressive disorder, bipolar disorder, anxiety disorders, chronotype, sleep duration, body mass index, vitamin D levels, and educational attainment. Outcomes were SAD status and general seasonality score. Results: SAD was positively associated with latitude (OR[95%CI] = 1.05[1.03-1.06], padjusted<0.001), and there was nominal evidence of additive and multiplicative interactions between chronotype genetic risk and latitude (OR = 0.99[0.99-0.99], padjusted=0.381; OR=0.98[0.97-0.99], padjusted=0.489). General seasonality score was associated with latitude (IRR=1.01[1.01-1.01], padjusted 0.001) and genetic risk for major depressive disorder (IRR =1.02[1.01-1.03], padjusted<0.001), bipolar disorder (IRR=1.02[1.01-1.03], padjusted=0.001), anxiety disorders (IRR=1.03[1.01-1.04], padjusted<0.001), vitamin D levels (OR=0.89[0.80-0.95], padjusted=0.048), and educational attainment (IRR=0.97[0.96-0.99], padjusted<0.001). Conclusions: These findings enhance understanding of SAD etiology, highlighting contributions of psychiatric genetic risk and geographic measures on seasonal behavior, and support examining seasonality as a continuous dimension.

Background: Ageing is a systems level biological process underlying the onset and progression of multiple chronic disorders. Rather than arising from a single pathway, age related decline reflects interacting disturbances in metabolic regulation, inflammation, nutrient sensing, cellular stress responses, and tissue repair. Although GLP1 receptor agonists, sodium glucose cotransporter2 inhibitors, metformin, and rapamycin are usually evaluated against disease-specific endpoints. Objective: To develop an SBML compliant quantitative systems pharmacology model in which ageing is the primary pharmacological endpoint and to evaluate which combination therapy provides the greatest benefit for both metabolic and ageing related outcomes. Methods: We developed model comprising four layers: a metabolic/pharmacodynamic layer describing weight loss, HbA1c reduction, and nausea with tolerance; a drug layer capturing class-specific effects of GLP1 agonists, sodium glucose cotransporter2 inhibitors, metformin, and rapamycin; an ageing layer representing damage accumulation, repair capacity, frailty, and biological age gap; and a biomarker layer generating trajectories and estimated glucose disposal rate. Calibration was staged across semaglutide clinical endpoints. Bayesian hierarchical meta analysis, global sensitivity analysis, and practical identifiability analysis were used to assess robustness and interpretability. Results: The model reproduced semaglutide efficacy and tolerability dynamics and supported distinct drug-class profiles across metabolic and ageing axes. Rapamycin showed minimal glycaemic effect but emerged as a dominant driver of repair related ageing outcomes. Combination simulations predicted two distinct optima: one favouring metabolic improvement and one favouring ageing related benefit. Conclusion: The model supports the view that metabolic and ageing optimization are mechanistically distinct objectives and that weight loss and glycaemic improvement alone may be insufficient surrogates for health span benefit.

Background: Prediabetes is highly prevalent in older adults and is characterized by heterogeneous clinical trajectories, including regression to normoglycemia and progression to diabetes. While prediabetes has been associated with impaired physical function and frailty, the longitudinal impact of both a single diagnosis and dynamic glycemic transitions on functional outcomes remains unclear. We aimed to evaluate associations between baseline prediabetes and glycemic transitions over time with trajectories of functional capacity and frailty in older adults. Methods: We conducted a pooled analysis of harmonized data from five nationally representative longitudinal aging cohorts (MHAS, HRS, CHARLS, ELSA, CRELES) within the Gateway to Global Aging Data, including adults aged [&ge;]50 years with [&ge;]1 HbA1c measurements. Prediabetes was defined per ADA criteria (HbA1c 5.7-6.4%). Functional outcomes included activities of daily living (ADL), instrumental ADL (IADL), and frailty assessed using Fried phenotype, FRAIL scale, and a deficit-accumulation Frailty Index (FI). Mixed-effects Poisson models estimated incidence rate ratios (IRRs) for baseline prediabetes, while generalized estimating equations assessed time-varying glycemic status and transition trajectories. Models were adjusted for age, sex, cohort, and time-varying covariates, with sensitivity analyses including BMI, smoking, and alcohol intake. Findings: Among 18,571 participants (median follow-up 13.6 years), baseline prediabetes was associated with increased progression of functional deficits and frailty compared with normoglycemia, including higher FI values and accelerated FI progression. Prediabetes was associated with higher incidence of ADL, IADL, and multimorbidity deficits from early follow-up, although time-dependent changes in incidence rates were not significant. In time-varying analyses (n=7,840), both prediabetes and diabetes were associated with higher incidence of functional deficits compared with normoglycemia, with diabetes showing the strongest effects across all outcomes. Diabetes was associated with greater FI burden and accelerated progression, whereas prediabetes showed a smaller increase, with attenuation over time. Among individuals with baseline prediabetes, regression to normoglycemia occurred in 20.8% and was associated with increased incidence of ADL and frailty deficits. In contrast, progression to diabetes occurred in 24.3%, and was associated with lower risk of incident ADL and Fried frailty deficits compared to stable prediabetes. Interpretation: Prediabetes is associated with increased risk of functional decline, frailty, and deficit accumulation in older adults, independent of progression to diabetes. Regression to normoglycemia was associated with higher risk of functional deterioration. These findings suggest that prediabetes reflects a state of metabolic vulnerability linked to biological aging rather than solely a precursor to diabetes and highlights a need to reframe its clinical significance in older populations. Funding: This research was supported by Instituto Nacional de Geriatria in Mexico. Keywords: Prediabetes; Glycemic transitions; Frailty; Functional decline; Aging; Multimorbidity

INTRODUCTIONHealthful dietary patterns may attenuate dementia risk by preserving cerebrovascular health. Prior work has focused on systemic arterial stiffness, but cerebrovascular measures may be more sensitive to neuroprotective effects of diet. We examined associations between Mediterranean diet adherence, prefrontal cortex (PFC) arterial elasticity, and cognition in older adults. METHODSParticipants were 198 older adults (58% female; mean age 65.6 years) from the Newcastle ACTIVate cohort. Mediterranean Diet (MedDiet) scores were derived from the Australian Eating Survey food frequency questionnaire. Pulse Relaxation Function (PReFx), an index of PFC arterial elasticity, was measured using pulse Diffuse Optical Tomography. Cognition was assessed with CANTAB and a cued task-switching paradigm. RESULTSHigher MedDiet was associated with higher PFC arterial elasticity. MedDiet was not associated with cognition, and PReFx did not mediate diet-cognition associations. DISCUSSIONGreater Mediterranean diet alignment was cross-sectionally associated with PFC arterial elasticity, suggesting a pathway through which diet may influence brain health in ageing.

In the tumor microenvironment (TME), dynamic interactions between cells and soluble factors promote tumor progression. We previously demonstrated that parathyroid hormone-related peptide (PTHrP), a TME-associated cytokine, enhances the aggressive phenotype of HCT116 colorectal cancer (CRC) cells, and that conditioned medium from PTHrP-treated HMEC-1 endothelial stromal cells (CM) induces epithelial-to-mesenchymal transition (EMT) in CRC cells. Here, Western blot analysis showed that CM modulates Met receptor expression and activation and promotes cancer stem cell (CSC) traits in HCT116 cells. Since PTHrP induces CPT-11 chemoresistance through Met signaling, we investigated the involvement of the CM-Met axis in this process. Viability assays revealed that CM increases cell number and confers CPT11 resistance through Met activation. Transforming growth factor beta 1 (TGF{beta}1), upregulated in PTHrP-treated HMEC-1 cells, was evaluated as a potential mediator. Its neutralization reversed the CM-induced increase in cell number but did not affect chemoresistance. In silico analyses revealed differences between CRC and normal tissues related to TGF{beta}1 signaling and Met activation, along with positive correlations among the analyzed markers. Immunohistochemical observation of human samples is consistent with our previous findings. Overall, these findings support a role for PTHrP in promoting CRC aggressiveness through coordinated effects on tumor and stromal compartments

Background: Glioblastoma (GBM) is the most aggressive primary brain tumor in adults. Radioresistance, partly mediated by glioma stem-like cells, represents a major clinical challenge which could be overcome by the identification of the modulators of radioresistance. Existing CRISPR screens in human GBM models have largely used two-dimensional cultures with short-term viability readouts, failing to capture the long-term clonogenic behaviour underlying tumour recurrence after radiotherapy. Method: We developed ClonoScreen3D-CRISPRi, combining CRISPRi-mediated gene knockdown with three-dimensional clonogenic survival assays. Two GBM cell lines (G7 and GBML20), differing in MGMT promoter methylation status, were engineered to express the KRAB-dCas9 editor. Nine candidate radiosensitivity modifiers, selected through transcriptomic analysis, pharmacological studies, and literature review, were examined in both lines. Target validation was performed using full radiation dose-response assays and a pharmacological inhibitor. Results: The majority of candidate genes significantly altered survival fraction following irradiation in both cell lines. Knockdown of NFKB2, RELB, and CDK9 produced the most potent radiosensitization, with sensitizer enhancement ratios of 1.39-1.70 in validation studies, exceeding those of established radiosensitizers including PARP and ATM inhibitors. Notably, knockdown of these genes induced no significant cytotoxicity in the absence of radiation. Pharmacological validation using an IKK inhibitor confirmed these findings, implicating non-canonical NF-{kappa}{beta} signalling and CDK9-dependent transcriptional elongation as critical adaptive mechanisms in GBM radioresistance. Conclusions: ClonoScreen3D-CRISPRi is a scalable, physiologically relevant platform for identifying genetic modifiers of radioresistance. The non-canonical NF-{kappa}{beta} pathway and CDK9 represent promising radiosensitizing targets, and larger screens could enable systematic prioritisation of candidates for clinical translation.

BackgroundLynch syndrome (LS) is a cancer susceptibility syndrome caused by germline pathogenic variants in DNA mismatch repair (MMR) genes. Due to increased risk of colorectal cancer (CRC), enhanced colonoscopic surveillance is recommended for heterozygote MMR-carriers. ObjectiveUsing a registry of English LS patients linked to digital National Health Service records, we aimed to assess adherence of MMR-carriers to national surveillance guidelines, and to determine the impact of surveillance on CRC incidence and mortality. DesignWe described the frequency of colonoscopies in 4,732 MMR-carriers and used logistic regression to determine predictors of surveillance adherence. For MMR-carriers with a record of surveillance and those without, we: estimated age-specific annual CRC incidence rates (AS-AIRs) and cumulative lifetime risks, assessed for stage-shift by comparing CRC stage distributions and stage-specific AS-AIRs, and estimated risks of death from CRC and any cause using Kaplan-Meier methods and Cox Proportional Hazards regression. ResultsSurveillance at a mean interval of [&le;] 3 years (n=3028) was associated with a decrease in CRC-specific and all-cause mortality, without an associated change in total CRC incidence, even after multivariate adjustment. No strong evidence of stage-shift was observed. Colonoscopic surveillance at a mean interval of [&le;] 2 years (n=1569) was associated with an increase in total CRC incidence. Incidence of early-stage cancers was also higher, with no corresponding decrease in late-stage cancers, which may reflect the short follow-up period or the impact of overdiagnosis. ConclusionThe observed reduction in all-cause mortality amongst regularly-surveilled MMR-carriers may indicate an impact of surveillance on CRC-specific mortality, though in the context of a non-randomised study likely reflects the influence of selection bias. KEY MESSAGES OF ARTICLEO_ST_ABSWhat is already known on this topicC_ST_ABSRegular surveillance colonoscopy is recommended in Lynch syndrome, though evidence to support this remains mixed. We searched PubMed for articles published from inception to 01/05/2024 using the terms "Lynch syndrome", "HNPCC", "colonoscopy", "sigmoidoscopy", "surveillance", and "screening". We found one controlled trial and several small analytical studies dating from the early 2000s which compared surveilled and non-surveilled populations and found surveillance to be associated with reduced colorectal cancer (CRC) incidence and improved survival. More recent longitudinal observational studies, most without comparator groups, found a high incidence of CRC in LS populations despite being resident in countries where surveillance was recommended. A small number of studies directly assessed time since last colonoscopy against CRC incidence and stage with mixed findings. Finally, cross-sectional comparisons between countries of CRC incidence rates and surveillance interval recommendations found no relationship between the two1,2. What this study addsHere, we conduct an observational cohort study on a large national cohort of MMR germline pathogenic variant (GPV) carriers (MMR-carriers) in England (n=4,732), comparing CRC incidence and mortality in individuals with a record of regular surveillance to those without. Through linkage of the English National Lynch Syndrome Registry to Hospital Episodes Statistics data, we are uniquely able to study a comprehensive national population of MMR-carriers and identify the dates on which colonoscopies were undertaken over time, allowing assessment of adherence to national surveillance guidelines and the impact this has on CRC outcomes. Notably, receipt of regular colonoscopy was strongly associated with deprivation as well as ethnicity. The results show that regular surveillance at an average interval of 3 years (or less) is not associated with a reduction in CRC incidence when compared to less frequent surveillance, but an apparent decrease in both CRC-specific and overall mortality is observed, even after adjustment for confounding variables. Conversely, regular surveillance at an average interval of 2 years (or less) is associated with an increase in CRC incidence when compared to less frequent surveillance, which may suggest increased diagnosis of early-stage cancers or, due to the absence of a reduction in late-stage cancers, overdiagnosis. The observed impact of surveillance on overall mortality may demonstrate the impact of surveillance on CRC-specific mortality, or, in the context of an observational (non-randomised) study, indicate that the results are subject to selection bias. How this study might affect research, practice, or policyEvidence for the benefit of surveillance colonoscopy remains mixed. Whilst polypectomy would be anticipated to prevent CRC development (thus reducing CRC incidence), several studies have observed increased frequency of CRCs in MMR-carriers undergoing frequent surveillance colonoscopy, which may reflect overdiagnosis. The selection bias inherent to observational studies of surveillance renders mortality outcomes challenging to interpret. Randomised controlled trials of colonoscopic surveillance in MMR-carriers are required for effectiveness of this intervention to be accurately assessed. Given ethical and feasibility challenges, randomised controlled trials might be complemented by quasi-experimental designs using advanced observational methods for assessing effectiveness.

BackgroundPancreatic adenocarcinoma (PDAC) is an abysmal disease, with a poor clinical outcome, largely due to limited life-extending treatments for patients. Notoriously, PDAC displays a T cell-suppressive tumor microenvironment where underlying molecular mechanisms that lead to this phenotype remain poorly understood. To unravel specific mechanisms, we utilized bioinformatic analyses with functional studies and revealed the cytolinker protein plectin (PLEC) as a novel player in regulating the T cell-suppressive tumor microenvironment of PDAC. MethodsUtilizing the TCGA-PAAD dataset, tumor samples were separated by PLEC expression to evaluate patient survival, and pathway analyses associated with increased tumorigenesis. Evaluation of immune infiltration and subsequent immune deconvolution was performed using tidyestimate and CIBERSORTx R packages. Single-cell RNA-seq (scRNA-seq) analysis from 229 PDAC patients was analyzed to investigate signaling dynamics and immune cell infiltration in PLECHigh patients. Functional validation was provided using a monoclonal antibody (mAb) against cell surface plectin (CSP) in two murine PDAC models to examine changes in tumor growth and immune cell subset abundance. ResultsOur studies revealed that high plectin expression results in an overall worse survival associated with activation of pro-tumorigenic pathways and decreased anti-tumor immune signature in PDAC patients. Analysis via GSEA indicates PLECHigh patients display an aggressive phenotype and suppressed pro-inflammatory signaling pathways. Immune ESTIMATE scores were significantly decreased in PLECHigh patients, and scRNA-seq analysis revealed that PLECHigh tumors display a decrease in anti-tumor CD8+ T cells. In vivo analyses using an anti-CSP mAb revealed a reduction in tumor growth kinetics compared to IgG control corresponding with a significant increase in proliferating and activated cytotoxic CD8+ T cells. Anti-CSP-mediated tumor suppression was inhibited when CD8+ T cells were depleted, indicating that anti-CSP treatment is contingent on cytotoxic T cell functionality. ConclusionOur findings identify plectin as a biomarker of aggressive disease in PDAC, with high plectin expression associated with decreased T cell infiltration, and that treatment with anti-CSP mAb reinstates anti-tumor immunity and decreases tumor volume in vivo. These findings position plectin as a high-priority therapeutic target, with the potential to fundamentally reshape immune responses in PDAC and improve outcomes for patients with few remaining options.

The Million Veteran Program (MVP) represents the largest and one of the most diverse single cohorts associated with longitudinal Electronic Health Record data (EHR) data. We profiled a subset of samples from MVP using the Illumina Infinium MethylationEPIC Beadchip (EPIC array) to generate one of the largest single cohort methylation dataset to-date. Methylation profiles were analyzed for 45,460 total individuals, with the most populous ancestries composed of 27,455 Europeans, 11,798 African Americans, and 4,859 Admixed Americans. We detail the strict quality control standards implemented to ensure the most robust method of methylation profiling of the MVP cohort. This dataset was then applied to evaluate the effects of smoking exposure on DNA methylation in MVP participants. Ancestry-stratified epigenome-wide association studies (EWAS) of smoking status (ever/never) were performed using over 750,000 probes with certifiable signal. Our multi-ancestry meta-analysis demonstrates replicability with existing EWAS and identifies 3,207 novel probe-smoking associations unlocked via the depth and breadth of data in this cohort.

Background: Substance use disorders (SUDs), including alcohol and drug dependence, and smoking, pose a public health threat with their high prevalence and comorbidity with other diseases, and contribution to mortality. SUDs are highly correlated, and their genetic background is shared to some degree. Objectives: We aimed to investigate the genetic associations of previously reported loci for a wide range of SUDs in an unstudied Ukrainian population. Methods: We collected data from 595 individuals (339 women, 253 men), including 321 participants from two rehab centres. Based on clinical review and questionnaire data we defined drug dependence, alcohol dependence, alcohol abuse, binge drinking, smoking, opiate, amphetamine, cannabis, and hallucinogen use, along with several intermediary alcohol use and smoking variables considering the amount of use and the level of dependence. We genotyped COMT-rs4680, ADH1B-ADH1C-rs1789891, and HTR2A-rs6313, and applied logistic and ordered logistic regression assuming an additive inheritance model, controlling for the recruitment group, other substance uses, age, and sex, in the association analyses. Results: We replicate (P<0.05) the associations at COMT-rs4680 with smoking status (OR[95% CI]=1.56[1.01-2.41], P=0.047) and heaviness (1.37[1.04-1.80], P=0.026), and at ADH1B-ADH1C-rs1789891 and HTR2A-rs6313 with alcohol dependence (1.69[1.03-2.76], P=0.038 and 0.66[0.47-0.92, P=0.016], respectively). Furthermore, we provide evidence for an association at HTR2A-rs6313 with hallucinogen use (0.58[0.35-0.98], P=0.040). Conclusion: In this study on multiple SUDs we shed light on the genetic background of SUDs in Ukrainians and provide further evidence that variation at COMT is mainly associated with smoking, at ADH1B-ADH1C with alcohol-related variables, whereas HTR2A is a more general SUD-associated locus.

ImportanceEmerging data show that B-cell depleting chemotherapies, which are increasingly used to treat autoimmune disorders and multiple sclerosis, can be associated with mucosal side effects such as inflammatory vaginitis. ObjectiveEvaluate the impact of rituximab treatment on vaginal mucosal immune markers, endocervical immune cell populations and vaginal microbiome. DesignCross-sectional observational study conducted between 2022 - 2024. SettingAcademic medical center, Boston Massachusetts. ParticipantsWe enrolled women aged >18 years who were either 1) receiving rituximab for autoimmune renal disease or were 2) healthy controls ExposureTreatment with rituximab, an anti CD20 monoclonal antibody. Main outcome and measureWe compared endocervical immune cell populations, vaginal fluid immune markers, vaginal fluid immunoglobulins and vaginal microbiome composition between individuals being treated with rituximab and healthy controls. ResultsWe enrolled 26 women treated with rituximab for autoimmune renal disease and 26 healthy controls. Median circulating and endocervical B-cell and plasma cell proportions were significantly lower in treated participants compared to controls. Median vaginal fluid IgA concentrations were significantly lower in participants treated with rituximab, while ILE, IgM, IgG1, IgG2, IgG3 and IgG4 were not different between groups. Total T cell frequencies were similar between groups, but the proportion of activated T cells (CD4+CD38+HLADR+) was significantly lower in people treated with rituximab. Concentrations of IL10, IL13, IL17, IL21, IL23, IL4, ITAC and TNFa were elevated in vaginal fluid from the rituximab group, while IL-8 was lower. A CST-IV-C, low-Lactobacillus pattern of vaginal microbiota was more common in the rituximab group. Conclusions and RelevanceSystemic B-cell depletion is associated with reduced vaginal fluid IgA, a more diverse microbiome composition, and increases in many vaginal fluid immune markers compared to healthy controls. The reduction in vaginal fluid IgA may provide opportunities for vaginal bacteria to induce inflammation. Key pointsO_ST_ABSQuestionC_ST_ABSHow does circulating B-cell depletion impact the vaginal microenvironment? FindingsIn this cross-sectional study of 52 women, B cell and plasma cell proportions were significantly lower in both blood and vaginal mucosa among rituximab-treated participants compared to healthy controls. Vaginal IgA concentrations, but not other immunoglobulins, were significantly lower in rituximab treated participants. In treated participants, vaginal cytokine concentrations were elevated, and microbiome composition shifted toward non-Lactobacillus-dominant communities. In six people with inflammatory vaginitis, both circulating and endocervical B cells were lowest in people with the most severe symptoms. MeaningSystemic B cell depletion is associated with alterations in vaginal mucosal immune markers and microbiome composition which increase local inflammation.

Objectives: Non-suicidal self-injury (NSSI) and self-harming sexual behaviours share functional and behavioural overlaps. However, the relationship between NSSI and problematic sexual behaviour (PSB) remains underexplored. This study aimed to investigate the association between NSSI and PSB in two cohorts - a non-clinical university cohort and a clinical PSB patient cohort. Methods: Data were collected from 2,189 university participants and 477 clinical PSB patients. NSSI was assessed via self-report, and PSB was measured with the Sexual Addiction Screening Test-Revised (SAST-R) Core. The four core addictive dimensions of PSB: relationship disturbance, loss of control, preoccupation, and affect disturbance, were also evaluated. Logistic regression analyses were conducted to examine the association between PSB (presence/absence and severity) and NSSI, looking at effects of gender and contributions of addictive dimensions of PSB. Results: Rates of NSSI were similar in the university (7.1%) and patient (5.7%) cohorts; stratified by gender, a higher proportion of women PSB patients had NSSI compared to in the university cohort (29.3% vs 9.3%). In the university group, who had milder PSB than patients, PSB was associated with NSSI (OR=2.11, p<0.001); a significant gender by PSB interaction was found showing that women with PSB were over four times more likely to have NSSI than men without PSB (OR=4.44, p=0.037). In contrast, PSB severity was not associated with NSSI in PSB patients (OR=1.10, p=0.25). Associations of the addictive dimensions of PSB with NSSI were observed only in the subgroup of university women, in the 'preoccupation' dimension (p<0.001). Conclusions: Our findings highlight gender-specific patterns in the association between PSB and NSSI, suggesting the need for further research and possibly targeted prevention and intervention strategies in women.