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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 [≤] 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 [≤] 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.

Pancreatic cancer disproportionately affects Black individuals in the United States, but they have limited representation in genetic studies of pancreatic ductal adenocarcinoma (PDAC). To address this gap, we performed admixture mapping and genome-wide association analysis (GWAS) in genetically inferred African ancestry individuals (1,030 cases and 889 controls). Admixture mapping identified three regions with a significantly higher proportion of African ancestry in cases compared to controls (5q33.3, 10p1, 22q12.3). GWAS identified a genome-wide significant association at 5p15.33 (CLPTM1L, rs383009:T>C, T Allele Frequency=0.51, OR:1.45, P value=1.24x10-8), a locus previously associated with PDAC. Known loci at 5p15.33, 7q32.3, 8q24.21 and 7q25.1 also replicated (P value <0.01). Multi-ancestral fine-mapping identified two potential causal SNPs (rs3830069 and rs2735940) at 5p15.33. Collectively these findings identified novel PDAC risk loci and expanded our understanding of this deadly cancer in underrepresented populations, emphasizing the multifactorial nature of PDAC risk including inherited genetic and non-genetic factors. Statement of SignificanceTo understand how genetic variation contributes to PDAC risk in Black people in North American, we studied individuals of genetically-inferred African ancestry. We identified novel risk loci and differences in the contribution of known loci. This demonstrates that ancestry-informed genetic analyses improve our understanding of PDAC risk and enhances discovery.

Background: Acute necrotizing encephalopathy (ANE) is a rare and severe neurologic complication of viral infection in children, thought to result from a hyperacute cytokine storm causing blood-brain barrier disruption and central nervous system injury. Despite characteristic clinical and radiologic features, ANE remains poorly understood at the molecular level, with no validated biomarkers or targeted therapies. We aimed to determine whether genetic predisposition to ANE due to RANBP2 variants is associated with a distinct immunologic signature. Methods: We conducted a prospective biological study of familial ANE (ANE1, NCT06731790). We included 23 heterozygous carriers of the RANBP2 c.1754C>T (p.Thr585Met) variant from 10 families, and 28 noncarriers (median age, 40 years [range, 4-72]). Soluble immune mediators, transcriptomic analyses, multiparameter flow cytometry, and cellular imaging were analysed in peripheral blood mononuclear cells (PBMCs) and monocytes. Baseline and resiquimod stimulated immune responses were analysed within the same statistical model, with genetic status as the primary predictor. Findings: The RANBP2 Thr585Met mutation was associated with a dysregulated inflammatory phenotype characterized by reduced basal mediator production and exaggerated TNF- responses following stimulation (estimated difference, +2,098 pg/mL; 95% CI, 1,121 to 3,076; P=0.0001). Transcriptomic and flow cytometry analyses showed broad reprogramming of myeloid cells with enrichment of CXCR3-high CD14-high subsets. Expansion of these populations was associated with increased long-term disease burden. The RANBP2 variant was the only independent factor associated this inflammatory phenotype. Interpretation: RANBP2-associated ANE is characterised by a distinct immunological signature that can inform disease stratification and support the development of targeted immunotherapeutic approaches.

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 recent approval of KRAS inhibitors supports the therapeutic value of targeting mutant KRAS cancers. However, clinical efficacy is hindered by both primary and treatment-associated acquired resistance. We applied a CRISPR-Cas9 loss-of-function screen and identified loss of KEAP1 as a resistance mechanism to the KRASG12D-selective inhibitor MRTX1133 and the RAS(ON) multi-selective inhibitor RMC-7977 in pancreatic cancer models. RNA-sequencing analyses revealed a KEAP1KO transcriptome that is distinct from the ERK-, MYC-, and YAP/TAZ-TEAD-dependent transcriptional programs that drive KRAS inhibitor resistance, demonstrating a distinct mechanism of resistance. We then established a PDAC KEAP1-deficient (PKD) gene signature that was enriched in patients and preclinical models insensitive to KRAS inhibitor treatment. Finally, we observed that KEAP1-deficient cells exhibited elevated glutamine metabolism, and combination treatment with the glutamine antagonist DRP-104 (sirpiglenastat) enhanced KRAS inhibitor suppression of pancreatic and lung tumors. SIGNIFICANCEKEAP1 loss is associated with reduced response to KRAS inhibitor therapy. We demonstrate that KEAP1 loss-associated resistance can be overcome by pharmacologic inhibition of the KEAP1 loss-induced glutamine dependency, establishing a combination to enhance RAS inhibitor clinical efficacy.

Background and AimsThe rising incidence of Crohns disease (CD) in Westernized countries has been linked to changes in diet and increased consumption of food additives, yet the mechanisms by which these factors fuel intestinal inflammation remain unclear. Adherent-invasive Escherichia coli (AIEC), a pathobiont involved in CD pathogenesis, lacks a clear genetic hallmark but exhibits intestinal colonization and virulence traits, raising questions about the evolutionary forces promoting its emergence among select individuals. Here, we investigated how chronic exposure to two common dietary emulsifiers, carboxymethylcellulose (CMC) and polysorbate 80 (P80), along with host inflammation, drives AIEC genomic evolution and pathogenic potential. MethodsWild-type and Il10-deficient mice were monocolonized with AIEC and chronically exposed to CMC, P80, or water. Bacterial isolates were collected and analyzed for genomic diversification, mutations, and phenotype both in vitro and in vivo. ResultsEmulsifiers accelerated AIEC genomic diversification and selected for mutations linked to increased motility, invasion, and pro-inflammatory activity. Moreover, dietary emulsifier-evolved strains displayed a marked fitness advantage in vivo, outcompeting their counterparts in murine hosts, with the greatest advantage observed when evolution occurred under inflammatory conditions. Notably, evolutionary pathways and phenotypic outcomes were shaped by both emulsifier and the hosts inflammatory status, highlighting synergy between diet and host genetics in fostering pro-inflammatory pathobionts. ConclusionThese findings provide an evolutionary framework connecting modern dietary habits to the emergence of pathogenic AIEC strains, and underscore the importance of dietary interventions in individuals at risk for inflammatory bowel disease.

Mucosa-associated invariant T (MAIT) cells have been paradoxically implicated in both tissue repair and fibrosis. However, when and how they modulate fibrogenesis in the injured liver remain unclear. Here, using the carbon tetrachloride-induced model of liver injury in MR1- and MAIT cell-sufficient and -deficient mice, we identify MAIT cells as an early driver of fibrogenesis. The presence of MAIT cells exacerbated hepatocellular injury, myofibroblast activation, and matrix deposition early in the course of fibrosis development, but not at later stages. This was accompanied by rapid polarization of hepatic MAIT cells toward a MAIT17 phenotype and enrichment of pro-fibrotic transcriptional programs. Concurrently, MAIT cells acquired an exhaustion-associated phenotype while still retaining their effector functions. Mechanistically, we demonstrate that MAIT cells limit hepatic regulatory T (Treg) cell accumulation, accompanied by reduced Ki-67 and CXCR3 levels in the latter population, suggesting their impaired proliferation and tissue recruitment. Furthermore, Treg cell inactivation reversed MAIT cell-dependent differences in the severity of fibrosis, establishing Treg cells as a key downstream mediator. Together, these findings identify MAIT cells as early orchestrators of fibrogenesis and reveal a novel MAIT-Treg axis that can be considered a potential therapeutic target in the early stages of fibrotic diseases.

Desmoid fibromatosis (DF) is a rare mesenchymal neoplasm with an unpredictable clinical course, where spontaneous regression or progression occurs in a significant subset of patients through largely undefined mechanisms. The use of active surveillance (AS) offers the opportunity to investigate whether tumor- or host-driven systemic and local immune features may explain these divergent outcomes, improving patient management. A prospective observational study enrolled 55 patients with primary sporadic DF managed with AS. Clinical evolution was categorized as progression, regression, or stable disease according to RECIST 1.1. Immunomonitoring with multicolor flow cytometry identified distinct systemic T-helper polarization states stratifying clinical trajectories: regressors showed a Th2-skewed profile, while progressors displayed activated T-helper cells and Th1/Th9/Th17 subsets. Higher baseline Th2 levels associated with regression and longer progression-free survival. Plasma proteomic and whole-blood transcriptomic analyses confirmed coordinated IL-4/IL-13-linked pro-resolving programs in regressors and inflammatory, early T-cell activation signatures in progressors. Tumor transcriptomics revealed adaptive, antigen-presentation and restrained immune programs in regressing lesions versus innate inflammatory, interferon and TGF-{beta}-driven fibrotic pathways in progressing tumors. These findings identify systemic T-helper polarization as a biomarker of DF behavior and highlight coordinated systemic-tumoral immune programs underlying clinical outcomes, supporting more precise clinical management.

Genome-wide association studies of idiopathic pulmonary fibrosis (IPF) have identified 35 common genetic risk loci associated with IPF susceptibility. In this study, we evaluated the effects of the reported variants in clinically curated non-European individuals. Despite limited sample sizes, we observed partial replication, limited transferability of some variants and evidence of ancestry-specific effects. The MUC5B promoter variant rs35705950 emerged as the dominant and most consistent signal across ancestries. Our findings highlight the need for larger, well-characterised studies in understudied populations to support robust discovery and translation.

We previously reported that the oxidative stress sensor Kelch-like ECH-associated protein 1 (Keap1) recognizes hepatitis B virus (HBV) X protein (HBx) to activate the NF-E2-related factor 2 (Nrf2) signaling pathway, thereby inhibiting HBV replication, and that HBx promotes K6-linked polyubiquitylation of Nrf2. However, the molecular mechanism remains unclear. Here, we investigated the role of HECT, UBA, and WWE domain-containing E3 ubiquitin ligase 1 (HUWE1) in HBx-mediated K6-linked polyubiquitylation of Nrf2 and its impact on HBV replication. Cell-based ubiquitylation assays demonstrated that HUWE1 knockdown reduced HBx-mediated K6-linked polyubiquitylation of Nrf2, while overexpression of wild-type HUWE1, but not the catalytically inactive HUWE1(C4341A) mutant, enhanced it, indicating that HUWE1 E3 ligase activity is required. Coimmunoprecipitation and proximity ligation assays demonstrated that HUWE1 interacts with HBx in the cytoplasm and binds Nrf2 only in the presence of HBx, suggesting that HBx bridges HUWE1 and Nrf2 into a ternary complex. Cycloheximide chase assays demonstrated that HUWE1 knockdown destabilized Nrf2 in HBx-expressing cells, supporting a role for HUWE1 in Nrf2 stabilization via K6-linked polyubiquitylation. Furthermore, HUWE1 knockdown or treatment with the HUWE1 inhibitor BI8626 significantly increased HBV RNA and pgRNA levels in HBV-infected cells. Collectively, these results demonstrate that HUWE1 promotes K6-linked polyubiquitylation and stabilization of Nrf2 in an HBx-dependent manner to inhibit HBV replication. IMPORTANCEHepatitis B virus (HBV) chronically infects approximately 254 million people worldwide, yet host mechanisms that restrict viral replication remain incompletely understood. The Kelch-like ECH-associated protein 1 (Keap1)/ NF-E2-related factor 2 (Nrf2) signaling pathway is a central defense against oxidative stress. Under basal conditions, Nrf2 is degraded via Keap1/Cullin3-mediated K48-linked polyubiquitylation. We previously demonstrated HBV infection promotes Nrf2 stability through non-canonical K6-linked polyubiquitylation. Here, we identify the E3 ubiquitin ligase HUWE1 as the enzyme responsible for K6-linked polyubiquitylation of Nrf2. HBV X protein (HBx) recruits HUWE1 to Nrf2, forming a HUWE1/HBx/Nrf2 complex that switches Nrf2 ubiquitylation from K48 to K6, stabilizing Nrf2 and suppressing HBV replication. These findings reveal a novel antiviral mechanism exploiting a non-canonical ubiquitin code and highlight HUWE1 as a potential therapeutic target against chronic HBV infection.

Abstract / SummarySurvival outcomes for pediatric Burkitt lymphoma (BL) substantially vary depending on geography (50-90%), which also serves as a proxy for the prevalence of Epstein-Barr virus (EBV) within the tumors. Although BL is considered an immunologically "cold" tumor with few tumor-infiltrating lymphocytes (TILs), their functional status has not been fully evaluated, especially for EBV-positive disease. Here, we characterize the exhaustion and activation profiles of T cells in the tumor microenvironment (TME) of EBV-positive BL using orthogonal methods, single-cell gene expression analysis, spectral flow cytometry, and immuno-histochemistry staining (IHC). We found that CD8+ TILs displayed a mosaic of immune inhibitory gene expression encoding, PD1, TIGIT, LAG3 and HAVCR2/TIM3. IHC validated the expression of PD1 and TIGIT on CD8+ TILs, as well as their respective ligands, PDL-1, PVR, and Nectin-2 on malignant B cells. Despite exhaustion-associated signatures, CD8+ TILs retain cytotoxic potential, expressing granules (i.e. Granzyme A, Perforin) and cytokines (i.e. IFN{gamma}) and demonstrate an increased uptake of metabolites such as glucose, arginine, and methionine. In peripheral blood, pediatric BL patients exhibited a significantly higher abundance of PD1+TIGIT+ CD8+ T cells compared to healthy children. Notably, these circulating T cells from BL patients express significantly lower levels of TOX, suggesting they are not irreversibly dysfunctional. Together, our results indicate that CD8+ T cells both in the TME and in circulation of children with BL are not terminally exhausted but remain poised for functional re-invigoration. These findings support the potential integration of immune checkpoint inhibitors into combination chemotherapeutic regimens to improve outcomes for these children. SignificanceEBV-positive BL tumors contain functional, metabolically active CD8+ T cells. Circulating PD1+TIGIT+CD8+ T cells found in BL patients blood are a biomarker for those in the tumor microenvironment.

Gut microbiota are critical determinants of effective immune checkpoint therapy (ICT), yet the microbial mediators and host mechanisms that enhance antitumor immunity remain poorly understood. Here, we identify the microbiota-derived bile acid taurodeoxycholic acid (TDCA) as a metabolite associated with immune checkpoint therapy (ICT) response. TDCA administration alone is sufficient to overcome antibiotic-induced ICT hyporesponsiveness across multiple murine tumor models. Mechanistically, TDCA directly enhances CD8 T cell-mediated antitumor immunity, increasing cytotoxicity. These effects required signaling through the bile acid receptor TGR5. Together, these findings reveal TDCA as a gut microbial metabolite that restores ICT efficacy after antibiotic disruption by directly augmenting CD8 T cell anti-tumor activity. This work supports metabolite replacement as a therapeutic strategy to mitigate antibiotic-associated loss of cancer immunotherapy response. SignificanceTDCA is a microbiota-derived metabolite that restores immune checkpoint therapy efficacy after antibiotic disruption by directly enhancing CD8 T-cell-mediated anti-tumor immunity through bile acid receptor TGR5 signaling. Our findings suggest that supplementation with defined microbial metabolites can mitigate antibiotic-associated loss of immunotherapy response without requiring broader microbiome reconstitution.

Diabetes mellitus is characterized by chronic hyperglycemia and loss of pancreatic {beta}-cell function and mass. Current therapies focus on {beta}-cell protection and regeneration, led by GLP-1 receptor agonists. The G protein -subunit (Gs) acts as a key signaling node downstream of numerous GPCRs, integrating diverse signals that impact {beta}-cell mass and function. Elucidating the integrative role of pancreatic Gs signaling is thus crucial for understanding {beta}-cell biology. Our map of the pancreatic Gs-coupled GPCR landscape reveals sophisticated, cell-type-specific networks, positioning Gs as a central hub for intra-pancreatic communication. Previous studies in mice with {beta}-cell-specific or whole-pancreatic Gs deletion demonstrated reduced {beta}-cell mass, impaired insulin secretion, and glucose intolerance. The stronger phenotype in the whole-pancreas model--marked by -cell expansion and abnormal distribution--points to a crucial role for Gs in differential control of postnatal - and {beta}-cell proliferation. Here, we analyze the organ-wide consequences of Gs deletion using pancreas-specific Gs knockout mice (PGsKO). Consistent with prior findings, PGsKO mice exhibit reduced weight gain from four weeks and severe diabetes due to decreased {beta}-cell mass and concomitant -cell expansion. Furthermore, Gs loss induces profound architectural and functional defects in the exocrine pancreas, linked to YAP reactivation in acinar cells. Importantly, we observed attempted {beta}-cell regeneration in PGsKO mice. Although insufficient to reverse diabetes, our results delineate the full pancreatic phenotype that may facilitate these regenerative efforts and suggest that strategically biasing GPCR signaling network away from Gs could be a viable strategy to promote {beta}-cell regeneration from other pancreatic cell types. ARTICLE HIGHLIGHTSO_LIGs is a central signaling hub that integrates diverse GPCR inputs across pancreatic cell types, yet its organ-wide role remained poorly defined. C_LIO_LIWe addressed how pancreas-wide Gs deletion disrupts both endocrine and exocrine compartments, and whether regenerative programs are engaged. C_LIO_LIGs loss caused severe diabetes through {beta}-cell loss and -cell expansion, induced profound exocrine defects with YAP reactivation, and triggered attempted {beta}-cell regeneration from ducts and potentially other cell types. C_LIO_LIOur findings suggest that strategically biasing GPCR signaling away from Gs could promote regeneration from non-{beta}-cell sources, offering new therapeutic avenues for diabetes. C_LI

Genome-guided dietary advice is a goal of precision nutrition. However, the contribution of gene-diet interactions (GxDs) to disease risk remains unclear, hindering the identification of diet-outcome pairs more likely amenable to genetic-based recommendations. We thus implemented a two-step approach: first, we comprehensively assessed the contributions of genome-wide GxDs to cardiometabolic outcomes across a broad array of dietary exposures in UK Biobank participants (N = 141,144 to 325,989). Second, we selected the 20 significant diet-outcome pairs from the 713 pairs tested (p < 7.0 x 10-5) and derived GxD polygenic scores. In an independent sample, all scores were nominally associated with their corresponding outcomes, with 12 of 20 polygenic scores Bonferroni significant (p < 0.0025). Further analyses revealed GxD polygenic scores were associated with clinical outcomes such as incident gout, suggesting translational potential. Altogether, these results showcase the promise of GxD scores to inform precision nutrition.

Although the retinoblastoma protein (pRB) is functionally inactivated by hyperphosphorylation in the majority of colorectal cancers (CRC) - with RB1 rarely mutated and even amplified at the genomic level - three critical gaps remain unaddressed: no study has systematically compared which first-line chemotherapeutic agent best synergizes with CDK4/6 inhibition using head-to-head quantitative analysis; functional differences between palbociclib and abemaciclib in chemotherapy combinations have not been characterized in CRC; and direct genetic evidence of RB dependency in this combinatorial context is lacking. Here, we addressed these gaps by evaluating palbociclib and abemaciclib combined with oxaliplatin, 5-fluorouracil, and SN-38 in HCT116 CRC cells, with validation in SW480 cells, RB1-silenced HCT116 cells (shRNA-RB), and non-tumoral intestinal epithelial cells (IEC-6), using quantitative drug interaction analysis (Chou-Talalay), cell cycle profiling, apoptosis assessment, and pRB phosphorylation measurement. Oxaliplatin was the most consistently synergistic partner for both CDK4/6 inhibitors (CI < 1 across all tested concentrations), while combinations with SN-38 yielded variable results and 5-FU combinations approached additivity. The oxaliplatin combination reinforced G1 arrest and enhanced cell death, with abemaciclib producing more pronounced apoptotic induction than palbociclib - an effect not explained by differential pRB target engagement (both inhibitors reduced pRB Ser807/811 phosphorylation by [~]50%), likely reflecting abemaciclibs broader kinase inhibitory profile. shRNA-mediated RB1 silencing partially attenuated the combinatorial effect, providing direct genetic evidence that the synergy is RB-dependent. Importantly, the combination did not significantly potentiate oxaliplatin cytotoxicity in non-tumoral IEC-6 intestinal epithelial cells, in contrast to the pronounced enhancement observed in tumor cells, and synergistic benefit was preserved at sub-cytotoxic inhibitor concentrations. These findings identify oxaliplatin as the optimal chemotherapeutic partner for CDK4/6 inhibition in CRC, with a mechanism involving RB-dependent potentiation of apoptosis that is preferentially active against tumor cells and maintained at clinically relevant inhibitor doses.

Increased literature support the pathogenetic role of dysfunctional energetic metabolism in the setup and progression of organ damage and failure. Genetic diseases often offer the possibility to investigate pathogenetic mechanisms. In particular, excessive cardiac damage is the most frequent cause of mortality in Fabry disease (FD), a genetic condition caused by deficient -galactosidase A (GLA) activity, leading to globotriaosylceramide (Gb3) accumulation. Beyond Gb3 storage, metabolic alterations and mitochondrial dysfunction, supported by in vitro evidence or studies in other tissues, may contribute to FD cardiomyopathy. This study investigated, for the first time, the mechanisms of mitochondrial involvement in FD, its role in determining cardiac manifestations, and its potential as a therapeutic target. We used a humanized FD mouse model (R301Q-Tg/GLA knockout), along with derived embryonic fibroblasts and neonatal and adult cardiomyocytes, to assess mitochondrial function across the lifespan. FD cells showed impaired mitophagy, reduced mitochondrial respiration, and increased reactive oxygen species production. Importantly, this mitochondrial dysfunction exacerbated the lysosomal deficit in FD cells, forming a vicious cycle. In cardiomyocytes, these alterations progressed with age, leading to the accumulation of dysfunctional mitochondria, energetic failure, and, in adult hearts, terminal mitochondrial damage and apoptosis. These events ultimately result in cardiac remodeling and dysfunction, including hypertrophy and diastolic impairment. Indeed, L-arginine supplementation, which promotes NO/PGC-1-dependent mitochondrial rescue, prevented the development of cardiac abnormalities in FD mice. Our findings identify early mitochondrial dysfunction as a key driver of FD cardiomyopathy and support mitochondrial targeting, including L-arginine supplementation, as a promising adjuvant therapeutic strategy. The mechanistic link between lysosomal dysfunction, altered mitochondrial turnover, and energetic collapse emerges as a key targetable pathway in organ damage, extending beyond FD. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=134 SRC="FIGDIR/small/718770v1_ufig1.gif" ALT="Figure 1"> View larger version (62K): org.highwire.dtl.DTLVardef@2a5c4borg.highwire.dtl.DTLVardef@1117767org.highwire.dtl.DTLVardef@1b634c5org.highwire.dtl.DTLVardef@1429b6c_HPS_FORMAT_FIGEXP M_FIG C_FIG Cardiac manifestations vs mitochondrial alterations in Fabry disease: the visible tip and the hidden base of the icebergCardiac manifestations in hR301Q Tg/KO mice become evident from 9 months of age. However, mitochondrial homeostasis is perturbed much earlier (neonatal to young stages), with impaired mitophagy, reduced mitochondrial respiration and membrane potential, increased ROS production and PGC-1 downregulation. At later stages, from 6 months of age, mitochondrial dysfunction progresses and begins to impact cellular energetics, as indicated by reduced ETC expression and the onset of energetic deficit (ATP reduction). The resulting energetic collapse, together with progressive mitochondrial leakage, leads to cardiomyocyte hypertrophy, apoptosis, and dysfunction, which become detectable from 9 months of age, when clinical signs emerge. These findings support a mechanistic model in which 1) lysosomal incompetence due to GLA deficit is the initiating event inducing impairment of mitophagy; 2) Unsuccessful mitophagy, induces downregulation of PGC-1a-dependent mitogenesis; 3) exhausted mitochondria accumulate, inducing energetic collapse (able to exacerbate lysosomal dysfunction and further perturb mitophagy in a vitious cycle); 4) ultimate mitochondrial leakage induces Cytochrome C release and apoptosis activation. This cascade of molecular events is responsible for clinical manifestations, and mitochondrial targeting prevents cardiac organ damage. Significance statementFabry disease is a rare genetic disorder in which cardiac complications are a major cause of death, yet underlying mechanisms remain unclear. Here, we identify mitochondrial dysfunction as an early pathogenic event associated with impaired mitophagy, whereby defective mitochondrial quality control both results from and exacerbates lysosomal dysfunction, creating a self-reinforcing cycle that drives disease progression. Using a humanized model, we demonstrate that mitochondrial dysfunction is a key determinant of cardiac phenotype in vivo, driving energetic failure, oxidative stress, and cardiac damage. Importantly, L-arginine treatment restores mitochondrial function and prevents cardiac abnormalities. Our findings define a broadly relevant pathogenic axis linking lysosomal dysfunction, mitophagy failure, and mitochondrial impairment, that lead to impaired energetic metabolism and consequent cardiac hypertrophy, independently from GB3 accumulation. The implications of our study go beyond Fabry disease and support the therapeutic targeting of cellular energy homeostasis to prevent and treat organ damage and failure in chronic diseases. IMPORTANTO_LIManuscripts submitted to Review Commons are peer reviewed in a journal-agnostic way. C_LIO_LIUpon transfer of the peer reviewed preprint to a journal, the referee reports will be available in full to the handling editor. C_LIO_LIThe identity of the referees will NOT be communicated to the authors unless the reviewers choose to sign their report. C_LIO_LIThe identity of the referee will be confidentially disclosed to any affiliate journals to which the manuscript is transferred. C_LI GUIDELINESO_LIFor reviewers: https://www.reviewcommons.org/reviewers C_LIO_LIFor authors: https://www.reviewcommons.org/authors C_LI CONTACTThe Review Commons office can be contacted directly at: office@reviewcommons.org

Metabolic dysfunction-associated steatotic liver disease (MASLD) is the most prevalent chronic liver disease and strongly linked to obesity and insulin resistance. We previously reported that the common nuclear envelope variant rs6461378 (g.842031C>T; SUN1 H118Y) associated with MASLD and related traits including insulin resistance. To gain insight into how wild-type (WT) and H118Y SUN1 might differentially impact insulin signaling, we performed affinity purification-mass spectrometry (AP-MS) in human liver-derived cells stably expressing WT or H118Y SUN1. Unbiased AP-MS revealed a novel SUN1-CUL3 interaction, with comparative analysis showing that WT SUN1 interacted robustly with CUL3, while CUL3 interaction was markedly diminished with H118Y SUN1. Cells in which SUN1 was silenced via siRNA, or in which H118Y SUN1 was ectopically expressed, showed increased CUL3 neddylation, which is required for cullin RING ligase (CRL)-mediated ubiquitination of insulin receptor substrate (IRS) proteins. Inhibition of neddylation restored IRS-1 levels and insulin signaling in H118Y SUN1-expressing cells. Together, our findings provide a potential mechanism of H118Y SUN1-driven insulin resistance and a viable therapeutic approach for its reversal.

Allogeneic hematopoietic stem cell transplantation (HSCT) is a life-saving treatment for hematologic malignancies, but long-term survivors present with lower muscle mass and functional capacity. In adult HSCT survivors 10-20 years after treatment, single nucleus RNA sequencing uncovered elevated XRRA1 expression levels in all muscle nuclei populations, which was retained in primary muscle stem cell cultures. HSCT survivors were characterized in vivo by impaired neuromuscular innervation that associated with muscle weakness, and lower muscle stem cell neurotrophic action. Despite these impairments, the molecular and physiological responses to heavy resistance training (HReT) were preserved in HSCT survivors, as demonstrated in a pre-registered clinical trial (ClinicalTrials.gov: NCT04922970). After 12 weeks of HReT, gains in muscle mass and strength were similar in HSCT survivors and healthy controls. In addition, we observed that [~]9% of muscle-resident immune cells persist into adulthood and that bone marrow derived cells do not adopt alternative cell fates in muscle tissue, resolving long-standing questions in human muscle biology. Together, these findings uncover molecular mechanisms of HSCT sequelae in muscle nuclei and muscle stem cells, which, importantly, can at least partly be overcome by mechanical loading. Given the growing population of HSCT survivors and the multitude of benefits of HReT for all organ systems, our findings support the importance of HReT in this population to promote healthspan.

Background: Heart failure (HF) is a major and growing public health problem, and prior studies support a meaningful genetic contribution to HF susceptibility. Clinically, HF is commonly categorized into the major clinical sub-types of HF with reduced ejection fraction (HFrEF) and HF with preserved ejection fraction (HFpEF), which differ in pathophysiology and clinical profiles. However, previous genome-wide association studies have focused on autosomal variation and have routinely excluded the X chromosome, leaving X-linked genetic contributions to HF and its subtypes under-characterized. Methods: We performed X-chromosome wide association study (XWAS) utilizing directly genotyped data from 590,568 Million Veteran Program participants, including 90,694 HF cases across European, African, Hispanic, and Asian Americans. Sex- and ancestry-stratified logistic regression was used with XWAS quality control measures, adjusting for age and population structure, followed by fixed-effects multi-ancestry meta-analysis. Functional annotation, gene-based testing, fine-mapping, and colocalization were performed. We replicated genetic associations with all-cause HF in the UK Biobank. Results: In the multi-ancestry meta-analysis, we identified five X-chromosome-wide significant loci for all-cause HF, five for HFrEF, and one locus for HFpEF in males. No loci reached significance in female-specific analyses. In sex-combined analyses, we identified six loci for all-cause HF and four for HFrEF. The strongest and most emphasized signals mapped to genes were BRWD3, FHL1, and CHRDL1. Ancestry-specific analyses revealed additional loci, including NDP and WDR44 in African ancestry and PHF8 in Hispanic ancestry. One locus, BRWD3, was replicated in UK Biobank HF cohort. Integrated post-GWAS analyses (fine-mapping, colocalization and pleiotropy trait association studies) reinforced the biological plausibility of the X-linked signals. Conclusions: This multi-ancestry, sex-stratified XWAS identifies X-linked genetic contributions to HF and its subtypes and highlights the role of X-chromosome in heart failure pathogenesis.

L-type amino acid transporter (LAT1) drives the uptake of essential amino acids (EAA) and activation of mTORC1 signaling in cancer cells. Current models propose that glutamine exchange is required for LAT1-dependent EAA transport; however, in many tumours, including MYC-driven cancers, glutamine is simultaneously consumed for bioenergetic and biosynthetic processes. How LAT1 activity is maintained under these conditions remains unclear. Here, we identify histidine as an efficient bidirectional LAT1 substrate that is preferentially utilised under glutamine-limitation in Gln-dependent tumour cells. Histidine uptake is modulated by glutamine availability, revealing an unexpected role for histidine in maintaining amino acid homeostasis. We demonstrate that histidine availability supports LAT1-mediated transport, promotes mTORC1/4E-BP1 signaling, and enhances protein synthesis and supports tumour cell proliferation. Under histidine limitation, MYC and ATF4 induce amino acid transporter expression, including LAT1, to preserve intracellular EAA levels. Importantly, histidine restriction sensitizes tumour cells to LAT1 inhibition, enhancing sensitivity to LAT1 inhibition and reducing tumour burden. Together, our findings establish histidine as a key regulator of LAT1 function and mTORC1 activity, suggesting a potential metabolic vulnerability in glutamine-dependent tumours.