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Abstract The escalating drug resistance among microorganisms underscores the urgent need for innovative therapeutic strategies and a comprehensive understanding of bacteria's defense mechanisms against oxidative stress and antibiotics. Among the recently discovered barriers, the endogenous production of hydrogen sulfide (H2S) via the reverse transsulfuration pathway, emerges as a noteworthy factor. In this study, we have explored the catalytic capabilities and crystal structure of cystathionine γ-lyase from Pseudomonas aeruginosa (PaCGL), a multidrug-opportunistic pathogen chiefly responsible for nosocomial infections. In addition to a canonical l-cystathionine hydrolysis, PaCGL efficiently catalyzes the production of H2S using l-cysteine and/or l-homocysteine as alternative substrates. Comparative analysis with the human enzyme and counterparts from other pathogens revealed distinct structural features within the primary enzyme cavities. Specifically, a distinctly folded entrance loop could potentially modulate the access of substrates and/or inhibitors to the catalytic site. Our findings offer significant insights into the structural evolution of CGL enzymes across different pathogens and provide novel opportunities for developing specific inhibitors targeting PaCGL.

Abstract Background Cholangiocarcinoma (CCA) is still a deadly tumour. Histological and molecular aspects of thioacetamide (TAA)-induced intrahepatic CCA (iCCA) in rats mimic those of human iCCA. Carcinogenic changes and therapeutic vulnerabilities in CCA may be captured by molecular investigations in bile, where we performed bile proteomic and metabolomic analyses that help discovery yet unknown pathways relevant to human iCCA. Methods Cholangiocarcinogenesis was induced in rats (TAA) and mice (Jnk Δhepa + CCl4 + DEN model). We performed proteomic and metabolomic analyses in bile from control and CCA-bearing rats. Differential expression was validated in rat and human CCAs. Mechanisms were addressed in human CCA cells, including Huh28-KRASG12D cells. Cell signaling, growth, gene regulation and [U-13C]-D-glucose-serine fluxomics analyses were performed. In vivo studies were performed in the clinically-relevant iCCA mouse model. Results Pathways related to inflammation, oxidative stress and glucose metabolism were identified by proteomic analysis. Oxidative stress and high amounts of the oncogenesis-supporting amino acids serine and glycine were discovered by metabolomic studies. Most relevant hits were confirmed in rat and human CCAs (TCGA). Activation of interleukin-6 (IL6) and epidermal growth factor receptor (EGFR) pathways, and key genes in cancer-related glucose metabolic reprogramming, were validated in TAA-CCAs. In TAA-CCAs, G9a, an epigenetic pro-tumorigenic writer, was also increased. We show that EGFR signaling and mutant KRASG12D can both activate IL6 production in CCA cells. Furthermore, phosphoglycerate dehydrogenase (PHGDH), the rate-limiting enzyme in serine-glycine pathway, was upregulated in human iCCA correlating with G9a expression. In a G9a activity-dependent manner, KRASG12D promoted PHGDH expression, glucose flow towards serine synthesis, and increased CCA cell viability. KRASG12D CAA cells were more sensitive to PHGDH and G9a inhibition than controls. In mouse iCCA, G9a pharmacological targeting reduced PHGDH expression. Conclusions In CCA, we identified new pro-tumorigenic mechanisms: Activation of EGFR signaling or KRAS mutation drives IL6 expression in tumour cells; Glucose metabolism reprogramming in iCCA includes activation of the serine-glycine pathway; Mutant KRAS drives PHGDH expression in a G9a-dependent manner; PHGDH and G9a emerge as therapeutic targets in iCCA.

p53 regulates signalling pathways involved in metabolic homeostasis. Here the authors show that O-GlcNAcylation of p53 in the liver plays a key role in the physiological regulation of glucose homeostasis, potentially via controlling the expression of the gluconeogenic enzyme phosphoenolpyruvate carboxykinase.

BackgroundThough the precise criteria for accessing LT are consistently being applied, HCC recurrence (HCC-R_LT) still affects more than 15% of the patients. We analyzed the clinical, histopathological, and biological features of patients with HCC to identify the predictive factors associated with cancer recurrence and survival after LT.MethodsWe retrospectively analyzed 441 patients with HCC who underwent LT in our center. Overall, 70 (15.8%) of them developed HCC-R_LT. We matched them by age at transplant and etiology with 70 non-recurrent patients. A comparable cohort from the Liver Transplant Centre of Bologna served as validation. The clinical and biochemical characteristics and pre-LT criteria (Milan, Metroticket, Metroticket_AFP, and AFP model) were evaluated. Histological analysis and immunohistochemistry for angiopoietin-2 in the tumor and non-tumor tissue of explanted livers were performed. Patients’ follow-up was until death, last clinical evaluation, or 31 December 2021. In patients with HCC-R_LT, the date of diagnosis of recurrence and anatomical site has been reported; if a biopsy of recurrence was available, histologic and immunohistochemical analyses were also performed.ResultsPatients were followed up for a mean period of 62.7 ± 54.7 months (median, 39 months). A higher risk of HCC-R_LT was evident for factors related indirectly (AFP) or directly (endothelial angiopoietin-2, microvascular invasion) to biological HCC aggressiveness. In multivariate analysis, only angiopoietin-2 expression was independently associated with recurrence. Extremely high levels of endothelial angiopoietin-2 expression were also found in hepatic recurrence and all different metastatic locations. In univariate analysis, MELD, Metroticket_AFP Score, Edmondson–Steiner grade, microvascular invasion, and endothelial angiopoietin-2 were significantly related to survival. In multivariate analysis, angiopoietin-2 expression, Metroticket_AFP score, and MELD (in both training and validation cohorts) independently predicted mortality. In time-dependent area under receiver operating characteristic curve analysis, the endothelial angiopoietin-2 expression had the highest specificity and sensitivity for recurrence (AUC 0.922, 95% CI 0.876–0.962, p < 0.0001).ConclusionsEndothelial angiopoietin-2 expression is a powerful independent predictor of post-LT tumor recurrence and mortality, highlighting the fundamental role of tumor biology in defining the patients’ prognosis after liver transplantation. The great advantage of endothelial angiopoietin-2 is that it is evaluable in HCC biopsy before LT and could drive a patient’s priority on the waiting list.

Cysteine plays a major role in the redox homeostasis and antioxidative defense mechanisms of many parasites of the phylum Apicomplexa. Of relevance to human health is Toxoplasma gondii, the causative agent of toxoplasmosis. A major route of cysteine biosynthesis in this parasite is the reverse transsulfuration pathway involving two key enzymes cystathionine β-synthase (CBS) and cystathionine γ-lyase (CGL). CBS from T. gondii (TgCBS) catalyzes the pyridoxal-5́-phosphate-dependent condensation of homocysteine with either serine or O-acetylserine to produce cystathionine. The enzyme can perform alternative reactions that use homocysteine and cysteine as substrates leading to the endogenous biosynthesis of hydrogen sulfide, another key element in maintaining the intracellular redox equilibrium. In contrast with human CBS, TgCBS lacks the N-terminal heme binding domain and is not responsive to S-adenosylmethionine. Herein, we describe the structure of a TgCBS construct that lacks amino acid residues 466-491 and shows the same activity of the native protein. TgCBS Δ466-491 was determined alone and in complex with reaction intermediates. A complementary molecular dynamics analysis revealed a unique domain organization, similar to the pathogenic mutant D444N of human CBS. Our data provides one missing piece in the structural diversity of CBSs by revealing the so far unknown three-dimensional arrangement of the CBS-type of Apicomplexa. This domain distribution is also detected in yeast and bacteria like Pseudomonas aeruginosa. These results pave the way for understanding the mechanisms by which TgCBS regulates the intracellular redox of the parasite, and have far-reaching consequences for the functional understanding of CBSs with similar domain distribution.

The escalating drug resistance among microorganisms underscores the urgent need for innovative therapeutic strategies and a comprehensive understanding of bacteria's defense mechanisms against oxidative stress and antibiotics. Among the recently discovered barriers, the endogenous production of hydrogen sulfide (H2S) via the reverse transsulfuration pathway, emerges as a noteworthy factor. In this study, we have explored the catalytic capabilities and crystal structure of cystathionine γ-lyase from Pseudomonas aeruginosa (PaCGL), a multidrug-opportunistic pathogen chiefly responsible for nosocomial infections. In addition to a canonical l-cystathionine hydrolysis, PaCGL efficiently catalyzes the production of H2S using l-cysteine and/or l-homocysteine as alternative substrates. Comparative analysis with the human enzyme and counterparts from other pathogens revealed distinct structural features within the primary enzyme cavities. Specifically, a distinctly folded entrance loop could potentially modulate the access of substrates and/or inhibitors to the catalytic site. Our findings offer significant insights into the structural evolution of CGL enzymes across different pathogens and provide novel opportunities for developing specific inhibitors targeting PaCGL. [ABSTRACT FROM AUTHOR]

Cystathionine γ‐lyase (CGL) is a PLP‐dependent enzyme that catalyzes the last step of the reverse transsulfuration route for endogenous cysteine biosynthesis. The canonical CGL‐catalyzed process consists of an α,γ‐elimination reaction that breaks down cystathionine into cysteine, α‐ketobutyrate, and ammonia. In some species, the enzyme can alternatively use cysteine as a substrate, resulting in the production of hydrogen sulfide (H2S). Importantly, inhibition of the enzyme and consequently of its H2S production activity, makes multiresistant bacteria considerably more susceptible to antibiotics. Other organisms, such as Toxoplasma gondii, the causative agent of toxoplasmosis, encode a CGL enzyme (TgCGL) that almost exclusively catalyzes the canonical process, with only minor reactivity to cysteine. Interestingly, the substitution of N360 by a serine (the equivalent amino acid residue in the human enzyme) at the active site changes the specificity of TgCGL for the catalysis of cystathionine, resulting in an enzyme that can cleave both the CγS and the CβS bond of cystathionine. Based on these findings and to deepen the molecular basis underlying the enzyme‐substrate specificity, we have elucidated the crystal structures of native TgCGL and the variant TgCGL‐N360S from crystals grown in the presence of cystathionine, cysteine, and the inhibitor d,l‐propargylglycine (PPG). Our structures reveal the binding mode of each molecule within the catalytic cavity and help explain the inhibitory behavior of cysteine and PPG. A specific inhibitory mechanism of TgCGL by PPG is proposed. PDB Code(s): 7NL1;8BIS;8BIU;8BIV;8BIW;8BIX;8BIZ [ABSTRACT FROM AUTHOR]

The article presents the highlights of the hybrid XVI International Magnesium Symposium in Glasgow, Scotland in July 2022. Topics include updates on magnesium (Mg2+) research, the evaluation of pathologic hypo- or hyper-magnesemia conditions, and the role of Mg2+ in agriculture. Presenters include Roberta Cazzola of University of Milano, Stefano Iotti of University of Bologna, and Andrea Rosanoff of the Center for Magnesium Education & Research (CMER).

Background: Cholangiocarcinoma (CCA) is still a deadly tumour. Histological and molecular aspects of thioacetamide (TAA)-induced intrahepatic CCA (iCCA) in rats mimic those of human iCCA. Carcinogenic changes and therapeutic vulnerabilities in CCA may be captured by molecular investigations in bile, where we performed bile proteomic and metabolomic analyses that help discovery yet unknown pathways relevant to human iCCA. Methods: Cholangiocarcinogenesis was induced in rats (TAA) and mice (JnkΔhepa + CCl4 + DEN model). We performed proteomic and metabolomic analyses in bile from control and CCA-bearing rats. Differential expression was validated in rat and human CCAs. Mechanisms were addressed in human CCA cells, including Huh28-KRASG12D cells. Cell signaling, growth, gene regulation and [U-13C]-D-glucose-serine fluxomics analyses were performed. In vivo studies were performed in the clinically-relevant iCCA mouse model. Results: Pathways related to inflammation, oxidative stress and glucose metabolism were identified by proteomic analysis. Oxidative stress and high amounts of the oncogenesis-supporting amino acids serine and glycine were discovered by metabolomic studies. Most relevant hits were confirmed in rat and human CCAs (TCGA). Activation of interleukin-6 (IL6) and epidermal growth factor receptor (EGFR) pathways, and key genes in cancer-related glucose metabolic reprogramming, were validated in TAA-CCAs. In TAA-CCAs, G9a, an epigenetic pro-tumorigenic writer, was also increased. We show that EGFR signaling and mutant KRASG12D can both activate IL6 production in CCA cells. Furthermore, phosphoglycerate dehydrogenase (PHGDH), the rate-limiting enzyme in serine-glycine pathway, was upregulated in human iCCA correlating with G9a expression. In a G9a activity-dependent manner, KRASG12D promoted PHGDH expression, glucose flow towards serine synthesis, and increased CCA cell viability. KRASG12D CAA cells were more sensitive to PHGDH and G9a inhibition than controls. In mouse iCCA, G9a pharmacological targeting reduced PHGDH expression. Conclusions: In CCA, we identified new pro-tumorigenic mechanisms: Activation of EGFR signaling or KRAS mutation drives IL6 expression in tumour cells; Glucose metabolism reprogramming in iCCA includes activation of the serine-glycine pathway; Mutant KRAS drives PHGDH expression in a G9a-dependent manner; PHGDH and G9a emerge as therapeutic targets in iCCA. [ABSTRACT FROM AUTHOR]

p53 regulates several signaling pathways to maintain the metabolic homeostasis of cells and modulates the cellular response to stress. Deficiency or excess of nutrients causes cellular metabolic stress, and we hypothesized that p53 could be linked to glucose maintenance. We show here that upon starvation hepatic p53 is stabilized by O-GlcNAcylation and plays an essential role in the physiological regulation of glucose homeostasis. More specifically, p53 binds to PCK1 promoter and regulates its transcriptional activation, thereby controlling hepatic glucose production. Mice lacking p53 in the liver show a reduced gluconeogenic response during calorie restriction. Glucagon, adrenaline and glucocorticoids augment protein levels of p53, and administration of these hormones to p53 deficient human hepatocytes and to liver-specific p53 deficient mice fails to increase glucose levels. Moreover, insulin decreases p53 levels, and over-expression of p53 impairs insulin sensitivity. Finally, protein levels of p53, as well as genes responsible of O-GlcNAcylation are elevated in the liver of type 2 diabetic patients and positively correlate with glucose and HOMA-IR. Overall these results indicate that the O-GlcNAcylation of p53 plays an unsuspected key role regulating in vivo glucose homeostasis. p53 regulates signalling pathways involved in metabolic homeostasis. Here the authors show that O-GlcNAcylation of p53 in the liver plays a key role in the physiological regulation of glucose homeostasis, potentially via controlling the expression of the gluconeogenic enzyme phosphoenolpyruvate carboxykinase. [ABSTRACT FROM AUTHOR]

Objective The biological heterogeneity of hepatocellular carcinoma (HCC) makes prognosis difficult. We translate the results of a genome-wide highthroughput analysis into a tool that accurately predicts at presentation tumour growth and survival of patients with HCC. Design Ultrasound surveillance identified HCC in 78 (training set) and 54 (validation set) consecutive patients with cirrhosis. Patients underwent two CT scans 6 weeks apart (no treatment in-between) to determine tumour volumes (V0 and V1) and calculate HCC doubling time. Baseline-paired HCC and surrounding tissue biopsies for microarray study (Agilent Whole Human Genome Oligo Microarrays) were also obtained. Predictors of survival were assessed by multivariate Cox model. Results Calculated tumour doubling times ranged from 30 to 621 days (mean, 107±91 days; median, 83 days) and were divided into quartiles: ⩽53 days (n=19), 54-82 days (n=20), 83-110 days (n=20) and ⩾111 days (n=19). Median survival according to doubling time was significantly lower for the first quartile versus the others (11 vs 41 months, 42, and 47 months, respectively) (p<0.0001). A five-gene transcriptomic hepatic signature including angiopoietin-2 (ANGPT2), delta-like ligand 4 (DLL4), neuropilin (NRP)/tolloid (TLL)-like 2 (NETO2), endothelial cell-specific molecule-1 (ESM1), and nuclear receptor subfamily 4, group A, member 1 (NR4A1) was found to accurately identify rapidly growing HCCs of the first quartile (ROC AUC: 0.961; 95% CI 0.919 to 1.000; p<0.0001) and to be an independent factor for mortality (HR: 3.987; 95% CI 1.941 to 8.193, p<0.0001). Conclusions The hepatic five-gene signature was able to predict HCC growth in individual patient and the consequent risk of death. This implies a role of this molecular tool in the future therapeutic management of patients with HCC. [ABSTRACT FROM AUTHOR]

The male/female ratio of patients with hepatocellular carcinoma (HCC) is often unbalanced towards the male sex, indicating a sex predisposition for HCC development. A possible explanation may be attributed to different hormonal statuses, including the pro-inflammatory action of androgens in men and the protective effects of oestrogen against excessive inflammation in women. Although several studies have studied gene expression in patients with HCC, very few have attempted to identify features that could be distinctive between male and female patients. The present study aimed to identify distinctive signalling mechanisms between men and women that may be associated with HCC progression. The present study analysed a detailed microarray database that was obtained from the prospective study of 78 patients with HCC to study gene expression according to sex. In addition, the present study aimed to evaluate whether the differentially expressed genes were known oestrogen targets. Moreover, RNAs from the HCC cohort were evaluated for microRNA (miRNA/miR) expression, and a relationship between miRNA and gene expression according to sex was investigated. One gene, sineoculis homeobox homolog 1 (SIX1), which is known to be an oestrogen target gene, was revealed to be highly upregulated in hepatitis virus C (HCV)-positive female patients with HCC but not in HCV-positive male patients. In addition, SIX1 upregulation had a significant relationship with tumour growth speed (assessed as tumour doubling time in two CTs performed 6 weeks apart) and survival (P=0.009 and P=0.042, respectively) in female patients only. Furthermore, SIX1 upregulation was related with miR-421 and miR-9-5p only in male patients; however, in female patients, SIX1 upregulation had a direct relationship with miR-181b, miR-503-5p and miR-125b (miRNAs with potential oncogenic capacity), and an inverse correlation with miR139-5p, miR-26b, let7c-3p and let7c-5p (putatively oncosuppressive microRNAs). These data suggested a distinctive model for liver carcinogenesis in HCV-positive women, with downregulation of protective mechanisms against tumour progression and the activation of potential oncogenes, in relation to the oestrogen target gene SIX1. (IRB10/08_CE_UniRer; ClinicalTrials ID: NCT01657695). [ABSTRACT FROM AUTHOR]

We developed an assay for the extraction and simultaneous quantitation of five key metabolites of the methionine metabolic pathway in liver tissue. The metabolites included were 5′-methylthioadenosine, methionine, homocysteine, S-adenosyl- l-homocysteine, and S-adenosyl- l-methionine. The metabolites were extracted using a bead-based homogenization method, and quantitation was carried out using hydrophilic interaction chromatography and time-of-flight mass spectrometry. The extraction procedure was optimized by testing the effect of various solvent combinations. The chromatographic method was optimized for peak shape, signal intensity, and carry-over. With a total chromatographic run time of 5 min, this assay is suitable for the analysis of large sample sets. Time-of-flight mass spectrometry provided high mass accuracy which, combined with isotope pattern matching and use of chemical standards, guarantees high specificity. Moreover, by operating the mass spectrometer in enhanced duty cycle mode the signal strength for the analytes increased three- to tenfold in comparison with the generic full-scan mode. For quantitation, a matrix-spiked calibration method was used. The lowest analyte levels detected and quantified using our method were within the range of concentrations found in the liver. The inter-day coefficients of variance for the analytes were between 5 and 15 % in pooled tissue samples. Interestingly, the CVs between individual liver tissue aliquots were about twice as high. Additional experiments suggested that this higher variability was caused by uneven distribution of the analytes within the liver. In conclusion, an optimized and robust assay is now available for the extraction and quantification of key metabolites in the methionine metabolic pathway. [ABSTRACT FROM AUTHOR]

OBJECTIVES:Idiopathic portal hypertension (IPH) is a rare cause of portal hypertension that lacks a specific diagnostic test. Requiring ruling-out other causes of portal hypertension it is frequently misdiagnosed. This study evaluates whether using high-throughput techniques there is a metabolomic profile allowing a noninvasive diagnosis of IPH.METHODS:Thirty-three IPH patients were included. Matched patients with cirrhosis (CH) and healthy volunteers (HV) were included as controls. Metabolomic analysis of plasma samples was performed using UPLC-time-of-flight-mass spectrometry. We computed Student's P-values, corrected by multiple comparison and VIP score (Variable Importance in the Projection). The metabolites were selected with an adjusted Benjamini Hochberg P value <0.05. We use markers with a greater VIP score, to build partial least squares projection to latent structures regression with discriminant analysis (PLS-DA) representative models to discriminate IPH from CH and from HV. The performance of the PLS-DA model was evaluated using R2 and Q2 parameter. An additional internal cross-validation was done.RESULTS:PLS-DA analysis showed a clear separation of IPH from CH with a model involving 28 metabolites (Q2=0.67, area under the curve (AUC)=0.99) and a clear separation of IPH from healthy subjects with a model including 31 metabolites (Q2=0.75, AUC=0.98). After cross-validation, both models showed high rates of sensitivity (94.8 and 97.5), specificity (89.1 and 89.7), and AUC (0.98 and 0.98), reinforcing the strength of our findings.CONCLUSIONS:A metabolomic profile clearly differentiating patients with IPH from CH and healthy subjects has been identified using subsets of 28 and 31 metabolites, respectively. Therefore, metabolomic analysis appears to be a valuable tool for the noninvasive diagnosis of IPH. [ABSTRACT FROM AUTHOR]

Simple Summary: Thus far, clinical studies have shown that immunotherapy (atezolizumab-bevacizumab) has shown better and favorable overall survival than sorafenib for advanced hepatocellular carcinoma (HCC). However, the treatment outcomes of hepatic arterial infusion chemotherapy (HAIC) with cisplatin compared with sorafenib for intrahepatic advanced HCC remain unclear. We therefore aimed to determine the prognostic factors for HAIC with cisplatin. Our results showed that HAIC with cisplatin could significantly prolong the overall survival for intrahepatic advanced HCC and had a longer prognostic effect than sorafenib. Therefore, our results suggest that HAIC should be used in intrahepatic advanced HCC. Given that the outcome of hepatic arterial infusion chemotherapy (HAIC) with cisplatin for intrahepatic advanced hepatocellular carcinoma (HCC) is unclear, we aimed to compare prognostic factors for overall survival (OS) following HAIC with cisplatin versus sorafenib for intrahepatic advanced HCC using propensity score-matched analysis. We enrolled 331 patients with intrahepatic advanced HCC who received HAIC with cisplatin (n = 88) or sorafenib (n = 243) between June 2006 and March 2020. No significant difference was observed in OS between HAIC with cisplatin and sorafenib cohorts (median survival time [MST]: 14.0 vs. 12.3 months; p = 0.0721). To reduce confounding effects, 166 patients were selected using propensity score-matched analysis (n = 83 for each treatment). HAIC with cisplatin significantly prolonged OS compared with sorafenib (MST: 15.6 vs. 11.0 months; p = 0.0157). Following stratification according to the Child-Pugh classification, for patients with class A (MST: 24.0 vs. 15.0 months; p = 0.0145), HAIC with cisplatin rather than sorafenib significantly prolonged OS. Our findings suggest that HAIC with cisplatin demonstrates longer prognostic effects than sorafenib in intrahepatic advanced HCC. [ABSTRACT FROM AUTHOR]

Simple Summary: The circadian coordination of metabolism is tightly regulated, and its alteration can trigger several diseases, including liver steatohepatitis and cancer. Many factors (such as diet and jet lag) shape both the liver molecular clock and the circadian transcription/translation of genes related to different metabolic pathways. Here, we summarize our current knowledge about the molecular mechanisms that control this circadian regulation of liver metabolism. Circadian clocks control several homeostatic processes in mammals through internal molecular mechanisms. Chronic perturbation of circadian rhythms is associated with metabolic diseases and increased cancer risk, including liver cancer. The hepatic physiology follows a daily rhythm, driven by clock genes that control the expression of several proteins involved in distinct metabolic pathways. Alteration of the liver clock results in metabolic disorders, such as non-alcoholic fatty liver diseases (NAFLD) and impaired glucose metabolism, that can trigger the activation of oncogenic pathways, inducing spontaneous hepatocarcinoma (HCC). In this review, we provide an overview of the role of the liver clock in the metabolic and oncogenic changes that lead to HCC and discuss new potentially useful targets for prevention and management of HCC. [ABSTRACT FROM AUTHOR]

Simple Summary: Racial/ethnic disparities in the incidence and outcomes of hepatocellular carcinoma (HCC) is previously described. Yet, due to the challenges for ascertainment of the underlying etiology (e.g., hepatitis B and hepatitis C) in registry-based studies, the contribution of the underlying etiology to the racial disparities is poorly described. Utilizing comprehensive data on tumor characteristics, lifestyle, and outcomes in the Multiethnic Cohort Study, we explored racial disparities in HCC. We show significant racial disparities in the underlying etiology, mortality, and treatment patterns. We further show that underlying etiology is a significant contributor to racial disparities in mortality by race and should be considered in future research. Backgrounds: HCC incidence varies by race/ethnicity. We characterized racial differences in underlying etiology, presentation, and survival in the linkage of Multiethnic Cohort Study with SEER and Medicare claims. Methods: HCC characteristics, treatment, and underlying etiology in participants were obtained. Deaths were ascertained using state death certificates and the National Death Index. Risk factors were collected via questionnaires. Cox models were used to calculate hazard ratios (HR) and 95% confidence intervals (CI) for death. Results: Among 359 cases, the average age at diagnosis was 75.1. The most common etiology was hepatitis C (HCV) (33%), followed by nonalcoholic fatty liver disease (NAFLD) (31%), and different by ethnicity (p < 0.0001). African Americans (AA) (59.5%) and Latinos (40.6%) were more likely to be diagnosed with HCV-related HCC. In Japanese Americans (33.1%), Native Hawaiians (39.1%), and whites (34.8%), NAFLD was the most common etiology. Receipt of treatment varied across ethnic groups (p = 0.0005); AA had the highest proportion of no treatment (50.0%), followed by Latinos (45.3%), vs. whites (15.2%). HCC (72.2%) was the most common cause of death. In a multivariate analysis, AA (HR = 1.87; 95% CI: 1.06–3.28) had significantly higher mortality compared to whites. Conclusions: We found significant ethnic differences in HCC underlying etiology, receipt of treatment, and outcome. The findings are important for reducing disparities. [ABSTRACT FROM AUTHOR]

Simple Summary: The α-fetoprotein (AFP) model officially replaced the Milan criteria in France for liver transplantation (LT) for hepatocellular carcinoma (HCC) in January 2013. The aim of our retrospective study was to analyze the agreement of the criteria and the results of LT with an intention-to-treat design since the adoption of the AFP model and to compare them to the practice and results of LT before the adoption of the AFP model. We did not observe significant changes in practices in 523 consecutively listed patients, with a good agreement (88%) to AFP criteria on the explants before and after the adoption of the AFP model. However, the prognosis of patients listed in the most recent period was worse, maybe because of a significant increase in bridging treatments and in the waiting time. This observational study provides an insight into the real-life course of LT for HCC. Purpose: To compare the agreement for the criteria on the explant and the results of liver transplantation (LT) before and after adoption of the AFP (α-fetoprotein) model. Methods: 523 patients consecutively listed in five French centers were reviewed to compare results of the Milan criteria period (MilanCP, n = 199) (before 2013) and the AFP score period (AFPscP, n = 324) (after 2013). (NCT03156582). Results: During AFPscP, there was a significantly longer waiting time on the list (12.3 vs. 7.7 months, p < 0.001) and higher rate of bridging therapies (84 vs. 75%, p = 0.012) compared to the MilanCP. Dropout rate was slightly higher in the AFPscP (31 vs. 24%, p = 0.073). No difference was found in the histological AFP score between groups (p = 0.838) with a global agreement in 88% of patients. Post-LT recurrence was 9.2% in MilanCP vs. 13.2% in AFPscP (p = 0.239) and predictive factors were AFP > 2 on the last imaging, downstaging policy and salvage transplantation. Post-LT survival was similar (83 vs. 87% after 2 years, p = 0.100), but after propensity score analysis, the post-listing overall survival (OS) was worse in the AFPscP (HR 1.45, p = 0.045). Conclusions: Agreement for the AFP model on explant analysis (≤2) did not significantly change. AFP score > 2 was the major prognostic factor for recurrence. Graft allocation policy has a major impact on prognosis, with a post-listing OS significantly decreased, probably due to the increase in waiting time, increase in bridging therapies, downstaging policy and salvage transplantation. [ABSTRACT FROM AUTHOR]

Simple Summary: Liver transplantation and surgical resection are potentially curative options in patients with liver cirrhosis and hepatocellular carcinoma. However, tumor recurrence is frequent, and it is associated with a poor prognosis. The selection of candidates is paramount to maximize survival while decreasing tumor recurrence rates, particularly regarding liver transplantation in a context of worldwide organ shortage. Circulating tumor cells are an attractive method of liquid biopsy that could represent a novel strategy to delineate the optimal therapeutic approach in hepatocellular carcinoma. This observational, prospective study aims to assess the role of circulating tumor cells in patients undergoing surgical resection or liver transplantation, as well as their potential association with other widely known surrogate markers of poor prognosis. Background: In patients with hepatocellular carcinoma (HCC), a complete clearance of circulating tumor cells (CTCs) early after liver transplantation (LT) or surgical resection (LR) could prevent tumor recurrence. Methods: prospective pilot study including patients with HCC who underwent LR or LT from September 2017 to May 2020. Enumeration of CTCs was performed in peripheral blood samples (7 mL) using the Isoflux® system (Fluxion Biosciences) immediately before surgery, at post-operative day 5 and at day 30. A clinically relevant number of CTCs was defined as >30 CTCs/sample. Results: 41 HCC patients were included (mean age 58.7 ± 6.3; 82.9% male). LR was performed in 10 patients (24.4%) and 31 patients (75.6%) underwent LT. The main etiology of liver disease was chronic hepatitis C (31.7%). Patients undergoing LR and LT were similar in terms of preoperative CTC count (p = 0.99), but clearance of CTCs within the first month was more pronounced in the LT group. Clusters of CTCs at baseline were associated with incomplete clearance of CTCs at day 30 (54.2% vs. 11.8%, p = 0.005), which in turn impacted negatively on survival (p = 0.038). Conclusion: Incomplete clearance of CTCs after surgery could be a surrogate marker of HCC aggressiveness. [ABSTRACT FROM AUTHOR]

Simple Summary: Metabolic reprogramming is a hallmark of malignancy. Hepatocellular carcinoma (HCC) cancer cells alterations in metabolism are due to the adaption to hypoxia and hypo-nutrient conditions. Several proteins and metabolites associated with glycolysis, tricarboxylic acid cycle and pyrimidine synthesis were found to be differentially regulated in serum, tumor and peritumoral tissues with increased tumor size, suggesting that microenvironment and tumor cell cooperate to regulate metabolism. In this study, the metabolomic characterization of HCC using paired tumor and adjacent liver tissues indicated tumor size-dependent metabolic reprogramming. Targeting cancer metabolism provides potential diagnostic and prognostic metabolic biomarkers. Our study brings new insight into the potential therapeutic use of metabolic targets and a methodological framework and diagnostic and prognostic metabolic markers that may be used in a clinical setting. The stratification of future clinical trials based on these metabolic subsets should improve the development of effective therapies and more intensive surveillance. Hepatocellular carcinoma (HCC) is a common malignancy with poor prognosis, high morbidity and mortality concerning with lack of effective diagnosis and high postoperative recurrence. Similar with other cancers, HCC cancer cells have to alter their metabolism to adapt to the changing requirements imposed by the environment of the growing tumor. In less vascularized regions of tumor, cancer cells experience hypoxia and nutrient starvation. Here, we show that HCC undergoes a global metabolic reprogramming during tumor growth. A combined proteomics and metabolomics analysis of paired peritumoral and tumor tissues from 200 HCC patients revealed liver-specific metabolic reprogramming and metabolic alterations with increasing tumor sizes. Several proteins and metabolites associated with glycolysis, the tricarboxylic acid cycle and pyrimidine synthesis were found to be differentially regulated in serum, tumor and peritumoral tissue with increased tumor sizes. Several prognostic metabolite biomarkers involved in HCC metabolic reprogramming were identified and integrated with clinical and pathological data. We built and validated this combined model to discriminate against patients with different recurrence risks. An integrated and comprehensive metabolomic analysis of HCC is provided by our present work. Metabolomic alterations associated with the advanced stage of the disease and poor clinical outcomes, were revealed. Targeting cancer metabolism may deliver effective therapies for HCC. [ABSTRACT FROM AUTHOR]