Abstract The properties of cancer-associated genetic changes in cell-free DNA (cfDNA) are not fully understood. We performed whole-genome sequencing (WGS) of cfDNA as well as tumor tissue and white blood cells (WBCs) from 1,807 samples of 1,064 patients across eight common cancer types. Characterization of single base substitutions, small insertions and deletions, structural variants (SVs), and phased variants in single cfDNA molecules revealed unique properties of tumor-derived alterations as well as differences in error rates that spanned orders of magnitude. Given the low error rate associated with detection of tumor-specific rearrangement junctions in cfDNA, we hypothesized that these types of changes could enable detection of circulating tumor DNA (ctDNA) without prior knowledge of the alterations in the tumor tissue. As an example of this approach, we scanned each sequenced fragment genome-wide in cfDNA samples from the CheckPAC trial of patients with metastatic pancreatic cancer treated with radiation and immunotherapy to identify putative rearrangement junctions. We identified 22,010,911 such fragments but only 1,572 (0.007%) and 58,339 (0.27%) of these were present in the matched tumor or WBC samples, respectively, with the remaining identified only in cfDNA. We characterized each cfDNA fragment by the SV type, SV size, microhomology and insertion at the breakpoint junction, fragment size, and the location of the breakpoint with respect to the nearest fragment end, identifying differences depending on the origin of the SV. Machine learning analyses of SVs from cfDNA resulted in a high cross-validated performance for detection of tumor-specific SVs with an area under the curve (AUC) of 0.97 (95% CI: 0.97-0.98). After enriching for fragments most likely to be tumor-derived, we found that the number of cfDNA fragments containing SVs was highly correlated with the number obtained using a tumor-informed approach (Pearson correlation coefficient = 0.87, p<0.001), and could recapitulate longitudinal ctDNA levels and clinical outcomes using only low-coverage (∼4x) plasma WGS. The universal nature of tumor-associated sequence and structural alterations in cfDNA may be broadly useful for cancer detection. Citation Format: Daniel C. Bruhm, Carolyn Hruban, Adrianna L. Bartolomucci, Akshaya V. Annapragada, Sarah Short, Shashikant Koul, Kaui P. Lebarbenchon, Julia S. Johansen, Inna M. Chen, Andrei Sorop, Razvan Iacob, Speranta Iacob, Liana Gheorghe, Simona Dima, Katherine A. McGlynn, Manuel Ramírez-Zea, John Groopman, PLCRC-MEDOCC group, Remond J. Fijneman, Gerrit A. Meijer, Zachariah H. Foda, Jillian Phallen, Robert B. Scharpf, Victor E. Velculescu. Sequence and structural DNA alterations in the circulation of patients with cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 2591.

Abstract Hepatocellular carcinoma (HCC) is the third leading cause of cancer death globally and is one of the most rapidly increasing causes of cancer mortality in North America and Europe due to metabolic and emerging risk factors. Here, we show that cell-free DNA (cfDNA) fragmentation profiles reflected underlying disease biology and that a machine learning classifier detected HCC across diverse populations and was enhanced by incorporating patient clinical risk and protein concentrations. Characteristics of the cfDNA fragmentome, including genome-wide chromatin, repeat elements, methylome, and mutational profiles, were altered in patients with HCC, including those with metabolic risk factors and aflatoxin exposure. Novel whole-genome tissue-of-origin deconvolution analyses identified increased representation of cfDNA originating from hepatocytes (p=2.4x10-10) and CD8+ T cells (p=5.8x10-7) and decreased contribution of NK cells (p=3.0x10-6) in patients with cancer. Using a previously locked fragmentome classifier for liver cancer detection, we analyzed 377 individuals with and without HCC from two distinct geographic cohorts. We found that the cfDNA fragmentome classifier detected HCC across all stages and diverse etiologies with a sensitivity of 70% (95% CI=65%-75%) and specificity of 94% (95% CI=90%-97%), outperforming the commonly used alpha-fetoprotein (AFP) biomarker which had a sensitivity of 62% (95% CI=57%-67%) and specificity of 93% (95% CI=88%-96%). A combined approach using cfDNA fragmentomes, AFP, and clinical risk achieved a sensitivity of 74% (95% CI=66%-80%) at a specificity of 85% (95% CI=78%-89%) in transplant curable disease (Milan criteria) in both cohorts and was more sensitive than standard-of-care AFP and ultrasound performance for early-stage disease (reported 63% sensitivity at 84% specificity). This study provides insights into the origins of altered cfDNA and circulating proteins for populations at risk of liver cancer and supports the use of a genome-wide fragmentome approach for non-invasive detection of HCC. Citation Format: Hope Orjuela, Carter Norton, Shashikant Koul, Daniel C. Bruhm, Akshaya V. Annapragada, Sarah Short, Keerti Boyapati, Adrianna Bartolomucci, Vilmos Adleff, Nicholas A. Vulpescu, Kauí Lebarbenchon, Jacob Carey, Carter Portwood, Andrei Sorop, Razvan Iacob, Speranta Iacob, Liana Gheorghe, Simoni Dima, Katherine A. McGlynn, Manuel Ramirez-Zea, Jillian Phallen, Robert B. Scharpf, John Groopman, Victor E. Velculescu, Zachariah Foda. Characterizing the cfDNA fragmentome in patients with hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 1125.

Abstract INTRODUCTION: Liver disease occurs on a continuum from steatosis to fibrosis, cirrhosis and ultimately hepatocellular carcinoma (HCC), with a 30% lifetime risk of HCC among those with cirrhosis (LCr). If identified early, steatosis and fibrosis are potentially reversible, and in LCr, surveillance can reduce cancer morbidity and mortality. Despite these benefits, conventional LCr detection modalities are invasive or have limited performance. We previously demonstrated that cost-effective liquid biopsies of genome-wide cell-free DNA (cfDNA) fragmentomes enable early detection of HCC. Here, we use these technologies to detect liver steatosis, fibrosis, and cirrhosis towards improved pre-cancer intervention and HCC surveillance. METHODS: We performed low-coverage, whole genome sequencing of plasma cfDNA from separate Discovery (n=423) and Validation (n=221) cohorts including individuals with no known liver disease (n=397), chronic liver disease and early fibrosis (n=91) including viral hepatitis and metabolic associated steatotic liver disease, or advanced fibrosis/cirrhosis (n=156). We computed genome-wide fragment length, coverage, and repeat element features (DELFI and ARTEMIS), cross-validated a machine learning classifier for fibrosis and LCr detection in the Discovery Cohort and evaluated the locked model in the Validation Cohort. We then performed whole methylome sequencing (n=28) and cell-type deconvolution to reveal mechanisms of change to cfDNA fragmentomes in LCr. RESULTS: Individuals with early liver disease/fibrosis and advanced fibrosis/cirrhosis were detected with high performance (AUC=0.90, 95% CI=0.86-0.95 and AUC=0.95, 95% CI=0.93-0.98, respectively) in the Discovery Cohort. At an 80% specificity locked cutpoint, Validation Cohort sensitivity was 70.8% (90% CI=52.3%-87.5%) for early liver disease/fibrosis and 90.1% (90% CI=84.4%-94.4%) for advanced fibrosis/cirrhosis. The model displayed low cross-reactivity for other fibrotic origin conditions including benign lung nodules or chronic pancreatitis (median scores 0.087 and 0.068 respectively vs. 0.55 for LCr, p<0.0002). The approach outperformed the existing fibrosis index FIB-4, detecting 5.07x (95% CI=3.03-17.35) and 1.2x (95% CI=1.18-1.32) more cases of early liver disease/fibrosis and advanced fibrosis/cirrhosis in simulations. cfDNA methylome deconvolution revealed increased contributions of liver endothelium (p=0.00016) and blood monocytes (p=5.2x10-5) and decreased contribution of hepatocytes (p=0.00035) with shorter fragment lengths in LCr. CONCLUSIONS: A cfDNA fragmentome biomarker enabled early detection of liver disease including LCr and reflected both liver-derived and immune-cell related changes. These analyses may enable accessible early detection of pre-cancer conditions with potential to improve liver disease management and early detection of HCC. Citation Format: Akshaya Vijaya Annapragada, Zachariah Foda, Hope Orjuela, Carter Norton, Shashi Koul, Noushin Niknafs, Sarah Short, Keerti Boyapati, Adrianna Bartolomucci, Dimitrios Mathios, Michael Noe, Chris Cherry, Jacob Carey, Alessandro Leal, Bryan Chesnick, Nic Dracopoli, Jamie Medina, Nicholas Vulpescu, Daniel Bruhm, Sarah Bacus, Vilmos Adleff, Amy Kim, Steve Baylin, Greg Kirk, Andrei Sorop, Razvan Iacob, Speranta Iacob, Liana Gheorghe, Simona Dima, Katherine McGlynn, Manuel Ramirez-Zea, Claus Feltoft, Julia Johansen, John Groopman, Jillian Phallen, Rob Scharpf, Victor Velculescu. Non-invasive early detection of cancer-predisposing liver diseases using genome-wide cfDNA fragmentomes [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 4074.

BACKGROUND AND AIMS:Inflammatory bowel diseases (IBD), including Crohn's disease (CD), ulcerative colitis (UC), and IBD-unclassified (IBD-U), are increasingly recognized across Eastern Europe, including Romania, where historical data indicated low incidence. Contemporary real-time epidemiological data for our country are scarce. This study evaluated the short-term frequency and epidemiological characteristics of IBD patients presenting to major Romanian gastroenterology centers. METHODS:We conducted a prospective, cross-sectional observational study over a 14-day period in November 2024 across 18 university-affiliated tertiary gastroenterology clinical sites in Romania. All consecutive adult patients with confirmed IBD were enrolled using a centralized online platform. Demographics, disease type and phenotype, severity, and treatment were recorded and analyzed descriptively. RESULTS:A total of 1,045 patients were registered: 52.4% CD, 46.9% UC, and 0.7% IBD-U. Geographical distribution revealed a statistically significant variation, with Crohn's disease being more frequent in Southern Romania, while UC predominated in the Eastern and Central-western regions (p=0.0009). Most patients resided in urban areas, and the majority were in clinical remission at presentation. Phenotypic analysis revealed ileocolonic CD (L3) and left-sided/pancolitis UC (E2/E3) as most frequent. Severe disease history was more common in CD, and prior surgery was significantly higher in CD than UC. Smoking and appendectomy were more frequently associated with CD as previously reported. Therapeutic patterns reflected disease type: anti-TNF use predominated in CD, while other biologics and small molecules were more common in UC. Regional differences in therapy were observed, with southern centers showing higher use of novel therapies, likely reflecting a more mature IBD population, with a higher CD prevalence. Notably, the number of IBD diagnoses increased over time, correlating with Romania's GDP growth (R² = 0.89, p < 0.001), suggesting that socioeconomic factors may influence disease recognition and diagnosis. CONCLUSIONS:This study offers the most recent snapshot of IBD epidemiology in Romania, highlighting a transition toward medium-incidence patterns and growing clinical complexity. These findings provide evidence for the need to establish nationwide population-based surveillance systems and healthcare planning initiatives aimed at mitigating the rising burden of IBD.

Background: Organized population-based colorectal cancer (CRC) screening using fecal immunochemical testing (FIT) followed by diagnostic colonoscopy is the recommended standard across the EU. In 2019, Romania started the ROCCAS pilot screening program in 4 administrative regions in South Muntenia. We evaluated pilot key performance indicators (KPIs) to inform evidence-based decisions for the upcoming national rollout. Methods: From January 2021-December 2023 50,000 average-risk individuals aged 50–74 years were invited to participate in a biennial FIT-based CRC screening program (cutoff for test positivity≥20 µg/g). FIT-positive participants received colonoscopy at the regional screening centre. Prospectively collected registry data were analysed. Multivariable logistic regression analysis was used to identify predictors of adenoma, advanced adenoma (AA), sessile serrated lesion (SSL), and CRC detection. Findings: FIT completion rate was 89%, with 6% positivity (n=2576). Colonoscopy uptake was 40% (n=1038). Caecal intubation (96%) and adenoma detection (55%) rates reached/ exceeded international benchmarks. Advanced adenomas, SSL and CRC were diagnosed inx 29%, 4% and 11% of scoped cases respectively. Multivariable analysis showed that male sex predicted adenoma (OR 2·72, 95%CI 2·11–3·51) and AA detection (OR 1·85, 95%CI 1·41–2·44), and was protective against right-sided SSLs (OR 0·34, 95%CI 0·14–0·81). FIT values correlated with lesion severity and independently predicted CRC (aOR 1·53 per 100 µg/g, p<.001). Interpretation: Our ROCCAS pilot screening program provided promising evidence of high-quality colonoscopy performance and clinically relevant detection rates. Nevertheless, colonoscopy uptake improvement should be prioritized before national rollout. We are currently scaling up awareness efforts and tailored communication strategies to maximize program’s outreach and effectiveness. Funding: ROCCAS was funded by the Romanian Ministry of Health with the support of the European Union Structural Reform Support Program.

Accessible liquid biopsies, including analyses of genome-wide cell-free DNA (cfDNA) fragmentation, are emerging for early detection of cancer but remain largely unexplored in other diseases. Here, we used whole-genome sequencing to examine cfDNA fragmentomes in 1576 individuals, including those with liver disease or with other morbidities such as vascular, autoimmune, and neurodegenerative conditions. As a prototype for disease-specific cfDNA fragmentomic biomarkers, we developed a machine learning classifier that detected early liver disease, advanced fibrosis, and cirrhosis with high sensitivity in separate discovery (n = 423) and validation cohorts (n = 221) and had limited cross-reactivity for other diseases. Genome-wide fragmentome and methylome analyses revealed liver-derived and immune-mediated changes in cfDNA in the circulation of individuals affected with liver disease. Fragmentomic changes were also observed across a range of other human morbidities and reflected disease-specific changes in the circulation. A machine learning model using cfDNA fragmentomes predicted overall survival in separate morbidity discovery (n = 571) and validation cohorts (n = 231). These analyses demonstrate the connection between cfDNA fragmentomes and an individual's physiologic state and provide previously unrecognized possibilities for cfDNA liquid biopsies across human disease.

Background: Hepatocellular carcinoma (HCC) is one of the most common and deadliest cancers worldwide. Alpha-fetoprotein (AFP), a widely used and accessible tumoral marker, has limited performance in the early detection of HCC among high-risk populations. This study aims to evaluate the potential added value of ccfDNA (circulating cell-free DNA) fragment size, alone or in a multiparameter panel, using accessible, feasible ccfDNA analysis. Methods: A prospective cohort of 125 patients with chronic liver disease was analyzed. Patients with incomplete clinical or laboratory data and patients without cirrhosis were excluded from the final analysis. Nonparametric tests, logistic regression and ROC curve analysis were performed. ccfDNA fragment size was measured using on-chip electrophoresis. Results: ccfDNA fragment size was significantly lower in the cirrhosis-HCC subgroup compared to the cirrhosis-only subgroup (p < 0.001). While AFP remains an independent predictor of HCC among cirrhosis patients, ccfDNA fragment size did not prove to be an independent predictor in this cohort. AUROC (area under the receiver operating characteristic curve) analysis revealed that a combined model of AFP, age, liver reserve, and ccfDNA fragment size did not perform better than the corresponding panel without ccfDNA. Moreover, after DeLong comparison, the difference between the two AUROCs proved statistically insignificant. Age and platelet count remain the strongest independent predictors in our exploratory cohort. Conclusions: Although ccfDNA fragment size proved to be lower in the HCC subgroup, its statistical significance fades when included into a multimarker panel. However, all panels should undergo further validation in a larger cohort, in order to better assess the individual contribution of each parameter and to discriminate between added diagnostic value and confounding effect of age and liver reserve parameters.

Hepatocellular carcinoma (HCC) is a leading cause of cancer death globally with increasing mortality due to emerging metabolic risk factors. Here, we show that in 377 individuals with and without HCC from two geographic cohorts, cell-free DNA (cfDNA) fragmentome characteristics, including chromatin, genomic, methylome, mutational, and repeat element profiles, were altered in patients with cancer of different etiologies, including metabolic risk factors and aflatoxin exposure. A novel methylation-based tissue-of-origin algorithm (MethID) identified cfDNA fragmentome changes originating from liver, vascular, and immune cells in cancer patients. A locked cfDNA fragmentome classifier detected HCC across stages and etiologies, outperforming alpha-fetoprotein (AFP). Combining cfDNA fragmentomes, AFP, and clinical risk achieved high sensitivity in early- and late-stage cancers in both cohorts and was more sensitive than conventional approaches for detecting early-stage disease. This study provides insights into cfDNA origins for populations at risk and validates a genome-wide fragmentome approach for non-invasive detection of HCC.

Background and Objectives: In recent years, frailty has emerged as a prognostic factor in inflammatory bowel diseases (IBD), particularly among patients with active disease. However, evidence regarding its reversibility after treatment optimization remains limited. This study aimed to assess frailty in active IBD and determine whether frailty status improved after 6 months of clinical management and the achievement of clinical remission. Materials and Methods: This prospective, single-center, observational cohort study included adults with active IBD requiring escalation to advanced therapy who achieved clinical remission at the 6-month follow-up. Patients were evaluated at baseline and after 6 months using a modified Fried frailty phenotype. Quality of life was assessed using the Short Inflammatory Bowel Disease Questionnaire (SIBDQ), and depressive symptoms were assessed using the Center for Epidemiologic Studies Depression (CES-D) scale. Univariate and multivariate logistic regressions were utilized to identify independent factors associated with frailty improvement. Results: The analysis included 54 patients (61.1% male; 42.6% with Crohn's disease). At baseline, 20.4% were classified as frail, 72.2% as pre-frail, and 7.4% as robust. Following 6 months of clinical management and the achievement of clinical remission, a 100% resolution of frailty was observed, with the robust cohort expanding to 42.6%. Significant improvements occurred across clinical parameters, including handgrip strength, 400 m walk times, and median SIBDQ scores (increasing from 4.4 to 5.9, p < 0.001) alongside a substantial decline in CES-D scores (p = 0.017). Multivariate logistic regression revealed that severe disease at baseline (aOR = 4.51, 95%CI: 1.26-16.18, p = 0.020), anti-TNF therapy initiation (aOR = 3.69, 95%CI: 1.04-13.18, p = 0.044), and higher baseline CES-D scores (aOR = 1.06, 95%CI: 1.00-1.13, p = 0.038) were independently associated with higher odds of frailty improvement. Conclusions: Among patients who achieved clinical remission, frailty and pre-frailty demonstrate substantial short-term improvement following advanced therapy. Functional and psychological recoveries are associated with successful control of baseline disease severity and systemic inflammation.

Introduction: Circulating cell-free DNA (cfDNA) has emerged as a promising minimally invasive biomarker in hepatocellular carcinoma (HCC), with potential applications in disease detection and prognostic stratification. This exploratory study aimed to evaluate the relationship between circulating cell-free DNA (cfDNA) concentration, liver dysfunction parameters, and hepatocellular carcinoma stage in an exploratory cohort: we sought to explore the extent to which cfDNA variability may be explained by the underlying liver disease environment and whether cfDNA concentration provides incremental information beyond routinely available markers of liver reserve for the discrimination between early- and late-stage hepatocellular carcinoma. Methods: Sixty-four newly diagnosed HCC patients were included. Clinical, laboratory, and staging data were collected. cfDNA was isolated from plasma, confirmed by on-chip electrophoresis, and quantified by fluorimetry. Logistic regression and ROC curve analyses were performed to assess the ability of several biomarker panels to discriminate early-stage HCC (BCLC 0-A) from intermediate/advanced-stage disease (BCLC B-D). Bootstrap resampling (2000 iterations) evaluated model robustness and coefficient stability. Additional linear regression analyses explored associations between cfDNA concentration and liver dysfunction parameters. Results: Linear regression demonstrated that liver dysfunction parameters explained approximately 53% of cfDNA variability, while HCC stage contributed minimally after adjustment. Models incorporating albumin, bilirubin, and platelet count as individual parameters demonstrated the best discriminatory performance after adjustment for liver disease etiology, achieving AUROCs up to 0.857. The only incremental value that cfDNA concentration added to the panel was an increase to its specificity (from 79.4% to 94.1%), while reducing sensitivity by 13.4%. Bilirubin and platelet count remained the most stable predictors after bootstrap, whereas cfDNA concentration was unstable. Limitations: The study was limited by its small sample size, cross-sectional design, and lack of longitudinal outcome assessment. External validation in larger prospective cohorts is necessary. Conclusion: A substantial proportion of cfDNA concentration variability (approximately 53%) was explained by routinely available liver dysfunction parameters, whereas HCC stage had minimal contribution. These findings suggest that circulating cfDNA concentration in patients with HCC may be influenced to a greater extent by the underlying cirrhotic liver environment and hepatocyte injury than by tumor burden alone. Integrated multimarker panels combining liver reserve parameters and liver disease etiology may be of interest for minimally invasive stratification of HCC. Although cfDNA concentration increased specificity for early- versus advanced-stage disease discrimination, its incremental value was low. Further validation in larger prospective cohorts is required.

Abstract Background Patients diagnosed with inflammatory bowel disease (IBD) are at increased risk of developing mental health issues, such as depression, especially when experiencing disability and poor quality of life (QoL). Recent studies showed that the relationship between depression and IBD appears to be bidirectional, through the brain-gut axis. Methods We aimed to evaluate the risk for depression and the QoL in patients with active IBD hospitalized between January 2021-April 2024 in our clinic, at the moment of biologic initiation and at 6 months follow-up, for those achieving clinical remission. We screened for symptoms of depression using the self-administered Center for Epidemiologic Studies Depression Scale (CES-D) – ranging score 0-60 (0-15 no to mild; 16-23 moderate; 24-60 severe symptomatology). The self-administered Short Inflammatory Bowel Disease Questionnaire (SIBDQ) was used to evaluate the health-related QoL through social, bowel, emotional and systemic dimension, with a score ranging from 10 (poor) to 70 (optimal). Results We included 47 patients (53.19% ulcerative colitis, 57.44% males, age 38.51±12.26 years old), with a disease duration of 6.43±6.28 years. 21 patients were reevaluated at 6 months. In patients with active IBD, the CES-D score was 14.15±13.04 and the SIBDQ score was 42.2±15.8 (42.6±19.2 social; 44.3±16 bowel; 43.6±16.9 emotional; 36.5±18 systemic). 18 patients (38.29%) were at risk for clinical depression, of whom 11 patients (23.40%) had severe depressive symptomatology. Patients being at risk for depression had lower SIBDQ scores (p<0.001), lower levels of serum albumin (p=0.007) and lower body mass index (p=0.004). In the subgroup analysis, both CES-D (17.23±15.63 vs. 8.09±7.19, p=0.005) and SIBDQ (40.81±16.45 vs. 56.09±10.68, p=0.002) were significantly improved after achieving clinical remission. Conclusion Considering the high rate of patients prone for depressive symptomatology, screening tools for depression and assessing QoL should be routinely used, and specialized psychiatric treatment should be integrated in the comprehensive care for IBD patients.

80% of adults in the US have metabolic risk factors for Liver Cirrhosis (LCr), but LCr diagnosis is challenging. Elastography and blood-based fibrosis indices have limited performance, and biopsies are invasive. The lifetime risk of hepatocellular carcinoma (HCC) in individuals with LCr is ∼30%, yet <20% of individuals undergo any HCC surveillance. We previously demonstrated that genome-wide cell-free DNA (cfDNA) fragmentomes can detect HCC in the blood. Here, we expand these approaches to pre-neoplasia, for LCr detection to facilitate management and HCC surveillance. We evaluated cfDNA fragmentomes in separate Discovery (n=465) and External Validation (n=279) Cohorts. These cohorts comprised individuals with LCr (n=132), at high-risk for LCr with viral hepatitis (n=26), metabolic associated steatotic liver disease (MASLD) and/or non-cirrhotic fibrosis (n=44), or aflatoxin exposure (n=10), or from healthy screening populations (n=532, including 126 with metabolic risk factors). For all individuals, we extracted cfDNA from plasma, performed low coverage (1-2x) whole genome sequencing, and computed genome-wide fragment length, coverage and repeat element features (DELFI and ARTEMIS). We cross-validated a machine learning model with these features for detection of LCr in the Discovery Cohort and evaluated the locked model in the Validation Cohort. In the Discovery Cohort, individuals with LCr were detected with high performance (AUC=0.97, 95% CI 0.94-1.0 and AUC=0.95, 95% CI=0.92-0.98, for individuals with and without metabolic risk factors). Scores were higher in LCr than in healthy populations and increased with cirrhosis severity (p<3.2x10-6 for Child-Pugh A, B and C). In the Validation Cohort, the locked model achieved 78% sensitivity and 92% specificity when locked at a threshold of 90% specificity and 90% sensitivity in the Discovery Cohort (AUC=0.95, 95% CI=0.91-0.99, and AUC=0.93, 95% CI=0.89-0.97, for individuals with and without metabolic risk factors), outperforming common fibrosis indices APRI and FIB-4. Among high-risk individuals without LCr but with aflatoxin exposure, MASLD, or fibrosis, scores were higher than in healthy individuals (p<6.0x10-16), but remained lower than for individuals with LCr (p<2.2x10-16). Fragmentomic analyses of transcription factor binding sites and single nucleotide variants revealed molecular alterations linked to both liver-tissue derived and inflammatory changes of cirrhosis. cfDNA fragmentomes enable detection of LCr, a pre-cancer condition that increases HCC risk. HCC surveillance in high-risk populations is critical, but accessibility and adherence remain low. A facile, effective screening approach for LCr may enable early identification towards improved management and initiation of HCC surveillance. Akshaya V. Annapragada, Zachariah H. Foda, Noushin Niknafs, Sarah Short, Dimitrios Mathios, Shashikant Koul, Keerti Boyapati, Adrianna Bartolomucci, Jamie E. Medina, Nicholas A. Vulpescu, Chris Cherry, Daniel C. Bruhm, Vilmos Adleff, Amy Kim, Andrei Sorop, Razvan Iacob, Speranta Iacob, Liana Gheorghe, Simona Dima, Katherine A. McGlynn, Manuel Ramirez-Zea, John Groopman, Jillian Phallen, Robert B. Scharpf, Victor E. Velculescu. Cell-free DNA fragmentomes enable early identification of liver cirrhosis to facilitate cancer surveillance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6426.

Hepatocellular carcinoma (HCC), a common consequence of chronic liver disease, ranks among the most prevalent cancers globally and contributes significantly to cancer-related mortality. Liver fibrosis is intimately associated with hepatic function and the likelihood of future HCC occurrence. Despite the fact that liver biopsy continues to be the gold standard for diagnosing fibrosis, its utility is hindered by cost and invasiveness, along with patient unease, procedural rejection, and potential adverse effects. Liver elastography has become a leading noninvasive means of assessing tissue stiffness with considerable diagnostic precision. Malignant tumors generally exhibit higher cellularity in comparison to benign ones, resulting in increased stiffness. Elastography techniques capitalize on alterations in tissue elasticity stemming from specific pathological or physiological processes. Technological innovations, such as advanced ultrasound imaging and artificial intelligence (AI)-integrated systems, are paving the way for enhanced diagnostic accuracy and risk prediction. Recent research underscores the potential of elastography in managing HCC patients, presenting novel clinical applications, including prediction of HCC development, differentiation between malignant and benign liver lesions, evaluating treatment response, and forecasting recurrence post-treatment, though certain findings remain contentious. Therefore, this review aims to sum up the latest advancements in liver elastography for HCC patients, outlining its applications while addressing existing limitations and avenues for future progress.

Background: Hepatitis B (HBV) and Delta (HDV) virus infections pose critical public health challenges, particularly in Romania, where HDV co-infection is underdiagnosed. Methods: This study investigates the epidemiology, risk factors, and clinical outcomes of HBV/HDV co-infection in vulnerable populations, leveraging data from the LIVE(RO2) program. Conducted between July 2021 and November 2023, the program screened 320,000 individuals across 24 counties, targeting socially disadvantaged groups such as rural residents, the Roma community, and those lacking health insurance. Results: Among 6813 hepatitis B surface antigen (HBsAg)-positive individuals, HDV antibody prevalence was 4.87%, with active replication confirmed in 75.6% of HDV-positive cases. Regional disparities emerged, with higher HDV prevalence and replication rates in the Eastern region compared to the South. HDV-positive individuals were more likely to be younger, male, and from rural or socioeconomically disadvantaged backgrounds. Clinically, HDV co-infection correlated with increased liver stiffness, advanced fibrosis stages, and lower steatosis levels compared to HBV mono-infection. Psychiatric comorbidities were more prevalent among HDV-positive patients, highlighting the need for integrated care. Conclusions: This study underscores the urgent need for targeted public health interventions, including enhanced screening, education, and access to novel antiviral therapies like bulevirtide to address the significant burden of HBV/HDV co-infection in Romania.

Given that delta hepatitis is associated with a 2-6 times higher risk for hepatocellular carcinoma (HCC) compared to HBV monoinfection, we aimed to identify the negative prognostic factors for complications associated with HDV infection (particularly HCC) and to validate BEA score as a screening tool for HCC in HDV. Our retrospective single centre study included all consecutive admissions of adult patients with chronic HDV infection in the period 01.01.2021-31.12.2022. The negative prognostic factors identified were higher MELD (p < 0.0001) and higher BEA score on admission (p < 0.0001), older age on HBV diagnosis (p < 0.0001) and advanced fibrosis when PegINF was administered (p = 0.01). Good prognostic factors were: Class A-BEA score (p = 0.001), normal platelet count (p = 0.00001), normal albumin level (p = 0.001) and prior treatment with PegInf (p = 0.01). ROC curve showed 78.5% sensitivity for BEA score > 2, validating it as a potential screening tool for HCC. Hence, for patients with BEA score > 2 imaging screening should be intensified in order to early diagnose HCC and prompt access to curative treatment. Additionally, the negative prognostic factors identified (MELD > 15, advanced fibrosis when treated with PegINF or diagnosis with HBV infection at an older age) should encourage more frequent monitoring for HCC compared to local guidelines recommendations.