BACKGROUND AND AIMS: Fontan-associated liver disease (FALD) could lead to liver cirrhosis and hepatocellular carcinoma (HCC). The prognosis of FALD patients is worse when HCC develops. Therefore, a better understanding o...BACKGROUND AND AIMS: Fontan-associated liver disease (FALD) could lead to liver cirrhosis and hepatocellular carcinoma (HCC). The prognosis of FALD patients is worse when HCC develops. Therefore, a better understanding of the mechanisms of FALD hepatocarcinogenesis is critical. We investigated the genomic and transcriptomic signatures of FALD HCC. APPROACH AND RESULTS: Ten patients with FALD HCCs, 2 patients with non-Fontan congenital heart surgery (CHS) HCC, and 1 patient with FALD focal nodular hyperplasia (FNH) were enrolled. Whole-genome sequencing and transcriptome sequencing were performed, and the results were compared with other etiology-related HCCs in common databases. FALD HCCs exhibited comparable tumor mutation burdens to those of HCCs in the databases, as well as mutation signatures related to DNA double-strand break repair deficiency. These features were shared by non-Fontan CHS HCCs and FALD FNH, suggesting that postoperative stress has an impact on genomic alterations. Of note, FALD and non-Fontan CHS HCCs showed a higher amount of copy number alterations and structural variants, including short deletions and translocations, which were rarely detected in FALD FNH. These structural variants were implicated in frequently altered genes of HCC, suggesting a possible trigger of hepatocarcinogenesis. Transcriptome analysis revealed that individual FALD and non-Fontan CHS HCCs were grouped into the reported HCC subclasses, namely progenitor phenotype, rather than an uncommon HCC subclass. CONCLUSIONS: Genomic mutational profiles of FALD and non-Fontan CHS HCCs are distinct from HCCs with other etiologies. Copy number alterations and transcriptional profiles in FALD HCCs classified these into a poor-prognosis group of HCCs.
BACKGROUND AND AIMS: Immune checkpoint inhibitors are now considered part of the standard of care in biliary tract cancer, based on a statistically significant but overall modest survival benefit in recent pivotal trials...BACKGROUND AND AIMS: Immune checkpoint inhibitors are now considered part of the standard of care in biliary tract cancer, based on a statistically significant but overall modest survival benefit in recent pivotal trials. Unlike in other gastrointestinal malignancies, PD-L1-based scores were not significantly associated with survival. Intrahepatic cholangiocarcinomas (iCCA) are morphologically heterogeneous tumors, with a subset of iCCA dominated by stroma-rich areas. We aimed at understanding the potential impact of the intertumoral and intratumoral heterogeneity on PD-L1 expression in iCCA. METHODS: Bulk transcriptome analysis and multiplex immunohistochemistry were performed on 141 clinically annotated resected iCCA. RESULTS: Molecular signatures are critically driven by the relative stroma content, with higher inflammatory gene expression scores in tumor microenvironment (TME)-rich compared with TME-poor tumors. In addition to this intertumoral heterogeneity, we observed a striking intratumoral heterogeneity reflected by an enrichment of PD-L1-expressing cells in stroma-dominant areas. The effect of intratumoral heterogeneity on PD-L1-based scores was confirmed by analyzing "virtual biopsies," that is, randomly selected adjacent tumor regions designed to mimic clinical biopsies, further highlighting the broad challenges associated with biomarker development using needle biopsies. By integrating clinical data, we provide evidence that the prognostic value of PD-L1-expressing tumor cells is overall modest and may depend not only on the magnitude of expression but also on their localization within the tumor. CONCLUSIONS: Our data indicate that stroma content may be a surrogate for inflamed iCCA. Moreover, the intertumoral and intratumoral heterogeneity of iCCA poses a significant challenge to interpreting the PD-L1 signal. To establish tissue-based biomarkers in iCCA, a more granular annotation of their expression in different intratumoral compartments will be critical. IMPACT AND IMPLICATIONS: Intrahepatic cholangiocarcinomas display a striking intertumoral and intratumoral heterogeneity, with broad variation in stroma content. We show that transcriptome-based inflammation-associated scores and signatures segregate with relative stroma content and that stroma areas in individual tumors are enriched in immune cells and PD-L1-positive cells. We argue that because of the extensive intratumor spatial heterogeneity of PD-L1 expression-the assessment of which is exacerbated by the limited amount of tissue routinely obtained in diagnostic biopsies in the locally advanced/metastatic setting-PD-L1-based metrics are likely to be bound to lack robustness as predictive biomarker of response to checkpoint inhibitors in iCCA and may partially account for the lack of correlation between PD-L1-based scores and clinical outcome in the pivotal phase 3 trials. The general implications of the above are: future attempts to identify tissue-based biomarkers should take spatial variation in the tumor microenvironment (TME) into account; TME-rich tumors are inflamed; and the predictive relevance of stroma content should be further investigated.
Yamamoto S, Kodama T, Kuwano A
… +23 more, Maesaka K, Yoshida-Hashidate T, Shindou H, Takeda H, Shirai K, Myojin Y, Murai K, Makino Y, Tahata Y, Saito Y, Hosui A, Nozaki Y, Nakabori T, Ohkawa K, Tanaka S, Nishio A, Miyazaki M, Hikita H, Motomura K, Lujambio A, Taketomi A, Eguchi H, Takehara T
BACKGROUND AND AIMS: Atezolizumab plus bevacizumab (Atez/Bev) is the first-line immunotherapy for advanced hepatocellular carcinoma (HCC), yet many patients show primary or acquired resistance. We aimed to identify tumor...BACKGROUND AND AIMS: Atezolizumab plus bevacizumab (Atez/Bev) is the first-line immunotherapy for advanced hepatocellular carcinoma (HCC), yet many patients show primary or acquired resistance. We aimed to identify tumor-intrinsic mechanisms driving immune cold tumor microenvironments (TMEs) and Atez/Bev resistance. APPROACH AND RESULTS: We used a genetically heterogeneous immunocompetent HCC mouse model generated by hydrodynamic injection of a barcoded oncogene library, integrating single-cell RNA sequencing, spatial transcriptomics, and bulk RNA sequencing of human HCC tissues. Retrospective analyses included a multi-institutional registry of 549 Atez/Bev-treated patients and 199 surgically resected HCC patients. External validation used RNA-seq data from 247 patients registered in the IMbrave150 and GO30140 trials. In mice, Atez/Bev eliminated hot tumors, while pre-existing cold tumors with exhausted effector T cells and Tregs predominated later. Barcode analysis revealed enrichment of NFE2L2 (NRF2) in resistant tumors. NRF2 overexpression suppressed immune infiltration and conferred resistance, which was reversed by NRF2 or COX2 inhibition. In human cohorts, both non-responders and patients with acquired resistance exhibited high NRF2/COX2 expression and immune exclusion, confirmed by spatial profiling. High NRF2 activity predicted shorter progression-free survival (PFS), whereas concomitant COX2 inhibitor use correlated with longer PFS. Plasma prostaglandin E2 (PGE 2 ) independently predicted poor response and survival. In IMbrave150/GO30140, NRF2 and prostaglandin pathway activation correlated with Atez/Bev resistance but not sorafenib response, validating a treatment-specific mechanism. CONCLUSIONS: Tumor-intrinsic activation of the NRF2-COX2-PGE 2 axis drives immune cold TMEs and mediates Atez/Bev resistance in HCC. Targeting this pathway may enhance efficacy, and plasma PGE 2 represents a non-invasive biomarker for stratification.
BACKGROUND AND AIMS: Local tumor progression (LTP) of hepatocellular carcinoma (HCC) after thermal ablation (TA) is associated with tumor invasiveness and poses a significant threat to patient outcomes. We aim to build a...BACKGROUND AND AIMS: Local tumor progression (LTP) of hepatocellular carcinoma (HCC) after thermal ablation (TA) is associated with tumor invasiveness and poses a significant threat to patient outcomes. We aim to build a multimodal model to explicable tumor invasiveness and reduce LTP. APPROACH AND RESULTS: From January 2015 to August 2023, 1208 HCC lesions were collected as training (n=502), validation (n=180), internal test (n=250), and external test (n=276) sets. Contrast-enhanced ultrasound (CEUS), magnetic resonance imaging (MRI), biological, clinical, and prognostic information were collected to build the model. A long short-term memory network and radiomics were used to extract image features. Logistic regression was used to combine image and clinical information. Pathological, immunohistochemical, and RNA sequencing analyses were used to explicable tumor invasiveness. Moderation analysis was applied to determine an appropriate minimum ablation margin (MAM) for high-invasiveness tumors in a safe location to reduce LTP. AUC of the multimodal model was 0.809 and 0.811 in internal and external test sets, respectively. The high-invasiveness group showed lower differentiation, higher microvascular invasion proportion, macrotrabecular-massive HCC proportion, CK-7 and GPC-3 positive rate, and increased expression of VEGFA, MMP-9, and HSPA1A (all p<0.05). KEGG and GSEA analyses revealed the upregulation of pathways related to angiogenesis, tolerance to stress response, and tumor metastasis in high-invasiveness group. The 8-mm MAM ablation strategy can effectively decrease the LTP incidence of high-invasiveness group (from 42.4% to 10.5%, p=0.027) to the level comparable to low-invasiveness group (10.5% vs. 6.1%, p=0.613) in external test set. CONCLUSIONS: The multimodal model achieved satisfactory performance on classifying tumor invasiveness, and provided effective strategy for high-invasiveness tumors to reduce LTP occurrence.
HCC is one of the leading causes of cancer-related death globally. HCCs are often diagnosed in advanced stages, and most patients are not eligible for curative treatments. Semiannual abdominal ultrasound with serum alpha...HCC is one of the leading causes of cancer-related death globally. HCCs are often diagnosed in advanced stages, and most patients are not eligible for curative treatments. Semiannual abdominal ultrasound with serum alpha-fetoprotein is the current recommended surveillance strategy; however, it has suboptimal sensitivity and implementation, thereby lowering its effectiveness for early-stage HCC detection. Further, post-treatment monitoring of HCC recurrence or progression is not tailored to tumor aggressiveness. These failures highlight critical unmet needs, which can be addressed by novel biomarkers that enable early detection and guide individualized surveillance and post-treatment follow-up for HCC, ultimately leading to improved outcomes in this disease. Cell-free DNA (cfDNA), consisting of small fragments of DNA released from tumor and normal cells into the bloodstream, reflects tumor-specific molecular alterations and has emerged as a promising noninvasive biomarker. This review summarizes recent advances in cfDNA applications in HCC, including risk stratification, early detection, prognostication, and minimal residual disease assessment. Among them, cfDNA has made the most progress in early detection. cfDNA assays, combined with clinical and serum parameters, demonstrate sensitivity noninferior or superior to ultrasound for detecting early HCC, with larger phase 3 validation studies ongoing. In contrast, the use of cfDNA for risk stratification, prognostication, and minimal residual disease detection remains in the early stages of clinical development. Future multicenter studies with standardized methodologies and rigorous validation will be essential to translate cfDNA into routine clinical practice for the management of HCC.
BACKGROUND AND AIMS: Metabolic dysfunction-associated steatotic liver disease (MASLD) is increasing and disproportionately affects Hispanics. The LiverRisk score (LRS), a non-invasive stratification tool, could enhance e...BACKGROUND AND AIMS: Metabolic dysfunction-associated steatotic liver disease (MASLD) is increasing and disproportionately affects Hispanics. The LiverRisk score (LRS), a non-invasive stratification tool, could enhance early identification of patients at risk for liver disease; however, it has not been validated in a large U.S. Veteran cohort with sufficient liver-related outcomes. APPROACH: We conducted a retrospective cohort study of Veterans Health Administration (VHA) patients from 1999 to 2023. Individuals with chronic liver disease, viral hepatitis, or missing LRS values were excluded using laboratory and ICD-10 data. Patients were classified into minimal (LRS <6), low (6 to <10), medium (10 to <15), and high-risk (≥15) categories. Outcomes included liver-related events, mortality, hepatocellular carcinoma (HCC), and C-statistics. Analyses used parametric/non-parametric methods and Cox proportional hazard models with competing risks, adjusting for missing data. RESULTS: Among 170,998 Veterans (93% male; median age 60; 63% White), 71%, 26%, 2%, and 0.5% were classified as minimal-risk, low-risk, medium-risk, and high-risk, respectively. Over 13.5 years median follow-up, 15,463 (9%) developed a liver-related event, 6219 (3.6%) died of liver causes, and 640 (0.5%) developed HCC. C-statistics were 0.61 for events, 0.70 for mortality, and 0.71 for HCC. Accuracy was highest among Hispanic/Latino Veterans (mortality 0.77, 95% CI 0.73-0.81; HCC 0.82, 95% CI 0.75-0.90) versus non-Hispanic Whites (mortality 0.62, 95% CI 0.61-0.62). CONCLUSIONS: LRS effectively predicted liver-related events, mortality, and HCC in Veterans. Its integration into clinical alerts and lab reports could support proactive patient care and liver disease prevention.
Cholangiocarcinoma (CCA) represents a biologically and clinically heterogeneous group of biliary tract malignancies. Growing evidence underscores that the 3 major anatomic subtypes-intrahepatic (iCCA), perihilar (pCCA),...Cholangiocarcinoma (CCA) represents a biologically and clinically heterogeneous group of biliary tract malignancies. Growing evidence underscores that the 3 major anatomic subtypes-intrahepatic (iCCA), perihilar (pCCA), and distal (dCCA)-are distinct tumor entities rather than a single disease. Each subtype differs in embryologic origin, carcinogenesis mechanisms, molecular landscape, and immunogenic profile, all of which contribute to variability in diagnosis, treatment responsiveness, and prognosis. The importance of understanding these distinctions is reinforced by the rising global incidence of CCA, which highlights the need for precision medicine approaches based on these subtypes. Recent advances have deepened our insight into subtype-specific risk factors, including chronic biliary diseases and metabolic conditions, while artificial intelligence-driven tools are improving diagnostic accuracy and risk stratification. Surgical innovations now allow for more tailored resections and perioperative care, and therapeutic strategies are increasingly guided by molecular and immune profiling. Emerging strategies for early diagnosis and chemoprevention in high-risk populations further emphasize the significance of recognizing the unique biology of each subtype. In this review, we provide a comprehensive overview of the distinct anatomic subsets of CCA, integrating their molecular and biological foundations with clinical management to inform practice and guide future research.
BACKGROUND AND AIMS: The mechanisms underlying metabolic dysfunction-associated steatohepatitis (MASH)-associated hepatocellular carcinoma (HCC) are poorly understood, and effective treatments are lacking. This study exp...BACKGROUND AND AIMS: The mechanisms underlying metabolic dysfunction-associated steatohepatitis (MASH)-associated hepatocellular carcinoma (HCC) are poorly understood, and effective treatments are lacking. This study explored the translational potential of Resmetirom, a clinically approved thyroid hormone receptor beta (Thrb) agonist, and investigated the mechanistic basis of MASH-associated hepatocarcinogenesis. APPROACH AND RESULTS: Repurposing Resmetirom for MASH-HCC treatment demonstrated significant tumor-suppressive effects across multiple preclinical models. To further investigate its mechanisms, we employed a Western diet plus CCl 4 -induced murine MASH-HCC model, complemented by single-cell RNA sequencing (scRNA-seq) analyses of liver and tumor tissues. These analyses revealed active cell-cell communication within the tumor microenvironment, particularly involving hepatic stellate cells (HSCs) and dysplastic hepatocytes (dys-Heps). These cells showed marked upregulation of midkine (MDK), which in human HCC correlated with shorter relapse-free survival, specifically in non-viral, non-alcohol-associated cases. In mouse models, MDK facilitated M2-like macrophage polarization via interaction with the receptor LRP1, contributing to disease progression from MASH to fibrosis and eventual HCC. Silencing LRP1 in macrophages abolished MDK-driven M2 polarization and increased cytotoxic cytokine secretion, while LRP1-positive macrophages contributed to T cell exhaustion through the CXCL16-CXCR6 axis. MDK expression negatively correlated with Thrb and lipolytic genes, but positively with lipogenesis genes. Resmetirom treatment not only significantly suppressed tumor growth and reduced steatosis but also decreased MDK expression and increased Thrb levels. Combining Resmetirom and an MDK inhibitor (iMDK) synergistically suppressed tumorigenesis in vivo. CONCLUSIONS: Targeting the MDK/LRP1 axis with Resmetirom offers a promising therapeutic strategy for MASH-associated HCC, addressing both metabolic dysfunction and tumor progression.
BACKGROUND AND AIMS: Accurate N-staging of intrahepatic cholangiocarcinoma (iCCA) remains challenging using noninvasive approaches. We aimed to develop a model to refine lymph node (LN) involvement stratification and inf...BACKGROUND AND AIMS: Accurate N-staging of intrahepatic cholangiocarcinoma (iCCA) remains challenging using noninvasive approaches. We aimed to develop a model to refine lymph node (LN) involvement stratification and inform therapeutic consideration. APPROACH AND RESULTS: This study enrolled a discovery cohort (n=682), an internal test cohort from the FU-iCCA (n=204), and an external multicenter cohort (n=88) for model development, and a neoadjuvant therapy (NAT) cohort (n=145) for therapeutic evaluation of the model. A SwinU-CliRad framework was constructed by integrating Swin UNEt TRansformers (Swin UNETR)-based magnetic resonance imaging-derived outputs of LN involvement with clinicoradiological features. Correlations between SwinU outputs and tumor multi-omics profiles were explored. The SwinU-CliRad model achieved areas under the curve (AUCs) of 0.932, 0.867, and 0.888 in LN risk stratification, and outperformed radiologist-based assessments by correcting more misclassifications than it introduced across the discovery, internal, and external test cohorts (18.8% vs. 7.3%, 18.1% vs. 4.9%, and 17.0% vs. 5.7%), respectively. In the NAT cohort, patients classified as high LN-involved risk by the SwinU-CliRad exhibited lower residual viable tumor rates than those with low LN-involved risk, with higher rates of pathological complete response (12.0% vs. 4.2%) and major pathological response (14.0% vs. 8.4%). SwinU outputs were associated with KRAS mutations, MUC5AC overexpression, and the large-duct histological subtype. Single-cell RNA sequencing analysis linked LN involvement to an immune-suppressive stroma tumor microenvironment. CONCLUSIONS: The SwinU-CliRad model can serve as a biologically interpretable tool for LN risk stratification in iCCA surgical candidates, with high-risk patients identified by the model potentially deriving benefit from NAT.