Screening for exclusion of high-risk bleeding features of esophageal varices in cirrhosis through CT and MRI.

BACKGROUND & AIM: Esophageal varices (EV) screening guidelines have evolved with improved risk stratification to avoid unnecessary esophagogastroduodenoscopy (EGD) in individuals with low bleeding risks. However, uncertainties persist in the recommendations for certain patient groups, particularly those with hepatocellular carcinoma (HCC) and/or receiving non-selective beta-blockers (NSBB) without prior endoscopy. This study assessed the efficacy of imaging in ruling out EVs and their high-risk features associated with bleeding in patients with cirrhosis and with HCC. We also evaluated the impact of NSBB on the detection of these characteristics. METHODS: A total of 119 patients undergoing EGD with CT and/or MRI within 90 days of the procedure were included. 87 patients had HCC. A new imaging grading system was developed utilizing the size of EVs and the extent of their protrusion into the esophagus lumen. The negative predictive value (NPV) of EVimaging(-) versus EVimaging (+) (grades 1-3) in ruling out the presence of EV and/or high-risk features by EGD was calculated. The predictive performance of imaging was determined by logistic regression. RESULTS: The NPV of imaging for detecting EV and high-risk features was 81 % and 92 %, respectively. Among HCC patients, the NPV for EV and high-risk features was 80 % and 64 %, respectively. Being on NSBB didn't statistically impact the imaging detection of EV. Imaging was a better predictor of high-risk EGD findings than Child-Turcotte-Pugh scores. CONCLUSIONS: Our results suggest that imaging can effectively rule out the presence of EV and high-risk features during EGD, even in patients with HCC and/or receiving NSBB.

Novel urine cell-free DNA methylation markers for hepatocellular carcinoma.

An optimized hepatocellular carcinoma (HCC)-targeted methylation next generation sequencing assay was developed to discover HCC-associated methylation markers directly from urine for HCC screening. Urine cell-free DNA (ucfDNA) isolated from a discovery cohort of 31 non-HCC and 30 HCC was used for biomarker discovery, identifying 29 genes with differentially methylated regions (DMRs). Methylation-specific qPCR (MSqPCR) assays were developed to verify the selected DMRs corresponding to 8 genes (GRASP, CCND2, HOXA9, BMP4, VIM, EMX1, SFRP1, and ECE). Using archived ucfDNA, methylation of GRASP, HOXA9, BMP4, and ECE1, were found to be significantly different (p < 0.05) between HCC and non-HCC patients. The four markers together with previously reported GSTP1 and RASSF1A markers were assessed as a 6-marker panel in an independent training cohort of 87 non-HCC and 78 HCC using logistic regression modeling. AUROC of 0.908 (95% CI, 0.8656-0.9252) was identified for the 6-marker panel with AFP, which was significantly higher than AFP-alone (AUROC 0.841 (95% CI, 0.778-0.904), p = 0.0026). Applying backward selection method, a 4-marker panel was found to exhibit similar performance to the 6-marker panel with AFP having 80% sensitivity compared to 29.5% by AFP-alone at a specificity of 85%. This study supports the potential use of methylated transrenal ucfDNA for HCC screening.

Impact of Cell-Debris and Room-Temperature Storage on Urine Circulating Tumor DNA from Hepatocellular Carcinoma.

This study evaluated the impact of cell debris and 7-day room temperature storage on the quality and yield of transrenal DNA. Archived urine specimens collected from five hepatocellular carcinoma (HCC) patients and two pregnant women carrying male fetuses were used to assess the impact of cell debris on urine cell-free DNA (ucfDNA) isolation as measured by quantitative PCR for Y-chromosome DNA, or HCC-associated mutation and methylation markers, and by capillary electrophoresis. Prospectively collected urine from 21 HCC patients was aliquoted after collection for paired immediate freezing versus 7-day room temperature storage followed by freezing for further analysis. Cell debris contained more Y-chromosome DNA than supernatant in three of the six urine specimens tested from pregnant women, suggesting that cell debris can be associated with 20.6% to 84.9% of transrenal ucfDNA. Ninety-five percent (20 of 21) of frozen and room temperature urine pairs had overlapping DNA size distribution. ucfDNA quantity determined by quantitative PCR for TP53, CTNNB1, TERT, and HCC-associated urine circulating tumor DNA markers were statistically similar between room temperature and frozen samples. This suggests no significant difference in DNA degradation between the groups. The association of transrenal ucfDNA with cell debris and HCC circulating DNA stability at room temperature is significant to further the understanding of transrenal circulating tumor DNA pre-analytical handling for HCC screening.

Detecting Liver Cancer Using Cell-Free DNA Fragmentomes.

Liver cancer is a major cause of cancer mortality worldwide. Screening individuals at high risk, including those with cirrhosis and viral hepatitis, provides an avenue for improved survival, but current screening methods are inadequate. In this study, we used whole-genome cell-free DNA (cfDNA) fragmentome analyses to evaluate 724 individuals from the United States, the European Union, or Hong Kong with hepatocellular carcinoma (HCC) or who were at average or high-risk for HCC. Using a machine learning model that incorporated multifeature fragmentome data, the sensitivity for detecting cancer was 88% in an average-risk population at 98% specificity and 85% among high-risk individuals at 80% specificity. We validated these results in an independent population. cfDNA fragmentation changes reflected genomic and chromatin changes in liver cancer, including from transcription factor binding sites. These findings provide a biological basis for changes in cfDNA fragmentation in patients with liver cancer and provide an accessible approach for noninvasive cancer detection. SIGNIFICANCE: There is a great need for accessible and sensitive screening approaches for HCC worldwide. We have developed an approach for examining genome-wide cfDNA fragmentation features to provide a high-performing and cost-effective approach for liver cancer detection. See related commentary Rolfo and Russo, p. 532. This article is highlighted in the In This Issue feature, p. 517.

Patterns and kinetics of hepatocellular carcinoma relapse post-liver transplantation: oligorecurrence and role of local therapies.

Background: Amongst patients with recurrent hepatocellular carcinoma (HCC) post-liver transplantation, systemic therapy options may be limited by immunosuppression or poor performance status. Thus, we aimed to assess the impact of metastasis-directed therapy to all sites of disease (MDT-All) in HCC patients with limited disease recurrence [i.e., oligorecurrence (oligoM1)] post-transplantation and characterize pre-transplant characteristics associated with oligoM1. Methods: In this retrospective cohort study, patients at a single institution with recurrent HCC post-liver transplantation were identified. OligoM1 disease was defined as ≤3 lesions at recurrence, while polyrecurrent (polyM1) disease was defined as >3 lesions. Outcomes were compared in patients with oligoM1 disease by receipt of MDT-All. Regression analyses were used to identify predictors of polyM1 disease and characteristics associated with post-recurrence outcomes. Results: Forty-three patients with recurrent HCC post-liver transplantation from 2005-2022 were identified. Twenty-seven (63%) patients had oligoM1. Microvascular invasion was independently associated with polyM1 [odds ratio (OR): 14.64; 95% confidence interval (CI): 1.48-144.77; P=0.022]. Elevated alpha-fetoprotein (AFP) ≥400 ng/mL [hazard ratio (HR): 2.44; 95% CI: 1.08, 5.52; P=0.033] at recurrence was independently associated with inferior overall survival (OS), while oligoM1 (HR: 0.42; 95% CI: 0.21, 0.87; P=0.018) was independently associated with favorable OS. Amongst patients with oligoM1 who received MDT-All (n=15) median OS was 38.4 vs. 16.1 months for those who did not receive MDT-All (log-rank P=0.021). There was a non-significant improvement in polyprogression-free survival (polyPFS) (median 14.0 vs. 10.7 months, P=0.1) amongst oligoM1 patients who received MDT-All compared to those who did not. Conclusions: Receipt of MDT-All was associated with improved OS amongst patients with limited HCC disease recurrence following liver transplantation.

Magnetic Resonance Imaging Predictors of Hepatocellular Carcinoma Progression and Dropout in Patients in Liver Transplantation Waiting List.

With the rising incidence of hepatocellular carcinoma (HCC), more patients are now eligible for liver transplantation. Consequently, HCC progression and dropout from the waiting list are also anticipated to rise. We developed a predictive model based on radiographic features and alpha-fetoprotein to identify high-risk patients. Methods: This is a case-cohort retrospective study of 76 patients with HCC who were listed for liver transplantation with subsequent liver transplantation or delisting due to HCC progression. We analyzed imaging-based predictive variables including tumor margin (well- versus ill-defined), capsule bulging lesions, volumetric analysis and distance to portal vein, tumor numbers, and tumor diameter. Volumetric analysis of the index lesions was used to quantify index tumor total volume and volumetric enhancement, whereas logistic regression and receiver operating characteristic curve (ROC) analyses were used to predict the main outcome of disease progression. Results: In univariate analyses, the following baseline variables were significantly associated with disease progression: size and number of lesions, sum of lesion diameters, lesions bulging the capsule, and total and venous-enhancing (viable) tumor volumes. Based on multivariable analyses, a risk model including lesion numbers and diameter, capsule bulging, tumor margin (infiltrative versus well-defined), and alpha-fetoprotein was developed to predict HCC progression and dropout. The model has an area under the ROC of 82%, which was significantly higher than Milan criteria that has an area under the ROC of 67%. Conclusions: Our model has a high predictive test for patient dropout due to HCC progression. This model can identify high-risk patients who may benefit from more aggressive HCC treatment early after diagnosis to prevent dropout due to such disease progression.

Long-Term Outcomes of Living Donor Versus Deceased Donor Liver Transplant for Hepatocellular Carcinoma in the United States.

OBJECTIVES: Although living donor liver transplant has become a vital treatment option in hepatocellular carcinoma, controversy remains on whether recurrence and survival rates are different versus deceased donor recipients. Here, we compared clinical characteristics and outcomes between recipients of living and deceased donor liver transplants for hepatocellular carcinoma in the United States. MATERIALS AND METHODS: Our comparisons used data from the United Network of Organ Sharing/Organ Procurement and Transplantation Network. RESULTS: There were 385 living donor and 25 274 deceased donor liver transplant recipients with diagnosis of hepatocellular carcinoma. Transplant list wait times of ≥6 months were more common in deceased donor(55.9%) versus living donor recipients (45.2%; P < .001). Both recipient groups were comparable with regard to alpha-fetoprotein level <200 ng/mL (P = .18). Only a small percentage in both groups had ≥3 total tumors (P = .73); both groups had similar low transplants outside of Milan criteria (P = .45). Overall, 1-, 5-, and 10-year overall survival rates for deceased versus living donor recipients were similar (91.2% vs 92%, 74% vs 76.4%, 58.9% vs 56.5%; P = .69). On multivariate analysis, Black/African American race/ethnicity was associated with worse outcomes than White race/ethnicity as reference (P < .001), whereas Hispanic and Asian race/ethnicity were more protected. Hepatitis C virus as liver disease etiology was associated with worse outcomes than other etiologies. Tumor characteristics, ≥3 lesions, tumor size, and higher alpha-fetoprotein levels were associated with worse outcomes. Living donor transplant was not associated with higher hazard of death. Among living donor recipients only, largest tumor size was associated with higher risk of death (P = .005). CONCLUSIONS: Survival was similarin between the living donor versus deceased donor recipients with hepatocellular carcinoma. With changes in Model for End-Stage Liver Disease exception policies for hepatocellular carcinoma in the United States, living donor transplant for hepatocellular carcinoma could expand the donor pool.

Urine DNA biomarkers for hepatocellular carcinoma screening.

BACKGROUND: Hepatocellular carcinoma (HCC) occurs in a well-defined high-risk patient population, but better screening tests are needed to improve sensitivity and efficacy. Therefore, we investigated the use of urine circulating tumour DNA (ctDNA) as a screening test. METHODS: Candidate markers in urine were selected from HCC and controls. We then enrolled 609 patients from five medical centres to test the selected urine panel. A two-stage model was developed to combine AFP and urine panel as a screening test. RESULTS: Mutated TP53, and methylated RASSF1a, and GSTP1 were selected as the urine panel markers. Serum AFP outperformed the urine panel among all cases of HCC, but the urine panel identified 49% of HCC cases with low AFP < 20 ng/ml. Using the two-stage model, the combined AFP and urine panel identified 148 of the 186 HCC cases (79.6% sensitivity at 90% specificity), which was 30% more than the cases detected with serum AFP alone. It also increased early-stage HCC detection from 62% to 92% (BCLC stage 0), and 40% to 77% (BCLC stage A). CONCLUSION: Urine ctDNA has promising diagnostic utility in patients in HCC, especially in those with low AFP and can be used as a potential non-invasive HCC screening test.

Accuracy of Milan, University of California San Francisco, and Up-To-7 Criteria in Predicting Tumor Recurrence Following Deceased-Donor Liver Transplant in Patients With Hepatocellular Carcinoma.

OBJECTIVES: We aimed to investigate the accuracy of the Milan, University of California San Francisco, and Up-to-7 criteria in predicting tumor recurrence after liver transplant for hepatocellular carcinoma. MATERIALS AND METHODS: For this study, 165 patients with deceased-donor liver transplant for hepatocellular carcinoma were evaluated. The Milan, University of California San Francisco, and Up-to-7 criteria were calculated based on explant pathology. RESULTS: Tumor recurrence rate after liver transplant was 14.6%. Of 165 patients, 115 (70%) were within Milan, 131 (79%) were within University of California San Francisco, and 135 (82%) were within Up-to-7 criteria. The odds ratio of tumor recurrence in patients outside versus within criteria for Milan, University of California San Francisco, and Up-to-7 was 3.6 (95% confidence interval, 1.5-9.1; P = .005), 7.5 (95% confidence interval, 2.5-19.3; P < .001), and 7.5 (95% confidence interval, 2.9-19.6; P < .001) times higher, respectively. The sensitivity of being outside of Milan in predicting tumor recurrence was comparable to University of California San Francisco and Up-to-7 criteria (56.5%, 56.5%, and 52.2%, respectively). Specificity was highest in Up-to-7 (87.3%) versus 85.2% for University of California San Francisco and 73.9% for Milan criteria. The area under the curve for Milan, University of California San Francisco, and Up-to-7 criteria was 0.63, 0.65, and 0.63. CONCLUSIONS: Application of standard criteria has significantly improved prediction of hepatocellular carcinoma recurrence. However, these criteria are inadequate, supporting the importance of other factors, including tumor biology. Research is ongoing in discovering novel biomarkers as predictors of tumor recurrence.

Detection of Novel Fusion Transcript VTI1A-CFAP46 in Hepatocellular Carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is now the second-highest cause of cancer death worldwide. Recent studies have discovered a wide range of somatic mutations in HCC. These mutations involve various vital signaling pathways such as: Wnt/ß-Catenin, p53, telome-rase reverse transcriptase (TERT), chromatin remodeling, RAS/MAPK signaling, and oxidative stress. However, fusion transcripts have not been broadly explored in HCC. METHODS: To identify novel fusion transcripts in HCC, in the first phase of our study, we performed targeted RNA sequencing (in HCC and paired non-HCC tissues) on 6 patients with a diagnosis of HCC undergoing liver transplantation. RESULTS: As a result of these studies, we discovered the novel fusion transcript, VTI1A-CFAP46. In the second phase of our study, we measured the expression of wild-type VTI1A in 21 HCC specimens, which showed that 10 of 21 exhibited upregulation of wild-type VTI1A in their tumors. VTI1A (Vesicle Transport via Interaction with t-SNARE homolog 1A) is a member of the Soluble N-ethylmaleimide-Sensitive Factor (NSF) attachment protein receptor (SNARE) gene family, which is essential for membrane trafficking and function in endocytosis, autophagy, and Golgi transport. Notably, it is known that autophagy is involved in HCC. CONCLUSIONS: The link between novel fusion transcript VTI1A-CFAP46 and autophagy as a potential therapeutic target in HCC patients deserves further investigation. Moreover, this study shows that fusion transcripts are worthy of additional exploration in HCC.

Role of nucleoside/nucleotide analogues and low-dose hepatitis B immune globülin in prophylaxis of hepatitis B recurrence among cadaveric liver transplant recipients.

BACKGROUND/AIMS: Hepatitis B core antibody (HBcAb) positivity of the donor or the recipient may pose a risk of hepatitis B virus (HBV) reactivation following liver transplantation (LT). We retrospectively investigated patient survival and reactivation among recipients who were given low-dose Hepatitis B Immune Globulin (HBIG) plus antiviral agent (AV) versus AV only. MATERIALS AND METHODS: Records of cadaveric LT recipients, between 2013 and 2016, with positive Hepatitis B surface Antigen (HBsAg) and/or HBcAb and recipients who had received LT from HBcAb-positive donors were reviewed. Patient characteristics and clinical data were extracted. Donor variables were retrieved from the United Network of Organ Sharing (UNOS) database. HBIG (1560 IU/mL) Intravenous (IV) was intraoperatively administered with three daily doses. Entecavir 1 mg daily was also given. STATA was used for statistical analysis. RESULTS: There were 53 recipients; 39 (73.6%) were male with a median age of 59 y. HCV was the major indication in 30 (55.6%) patients. There were 28 recipients (52.8%) who received HBIG plus AV and 25 (47.2%) received AV only. The Model of End Stage Liver Disease (MELD) score between the groups were similar. Survival rates at 6, 12, and 24 months were 100% (n=53), 93.2% (n=44), and 100.0% (n=26), respectively. There was no reactivation; two recipients in the AV group and one in the HBIG plus AV group died within 12 months. CONCLUSION: This study supports the use of low-dose HBIG and AV for post-LT prophylaxis to be as effective as conventionally used high-dose HBIG (9600 IU) plus AV. Future prospective larger studies are warranted to examine the potential benefits of using AV alone without HBIG.

Pre- Versus Posttransplant Treatment of Hepatitis C Virus With Direct-Acting Antivirals in Liver Transplant Recipients: More Issues to be Solved.

OBJECTIVES: Our goal was to investigate wait times related to hepatitis C virus treatment with direct acting antivirals before versus after liver transplant at a single center as well as wait times for insurance approval for preemptive treatment with these agents after liver transplant. MATERIALS AND METHODS: We retrospectively evaluated hepatitis C virus infections in transplant recipients of deceased liver donations in 2014 and 2015. Demographics, hepatocellular carcinoma incidence, Model for End-Stage Liver Disease scores, and transplant wait times were compared between patients treated before or after liver transplant. Wait times to approval of direct-acting antiviral treatment were evaluated in those untreated before transplant. RESULTS: During our study period, of 67 deceased-donor liver transplants, 21 patients received hepatitis C virus treatment pretransplant (treated group) and 46 patients were not treated pretransplant (untreated group). Twenty-five patients in the untreated group received hepatitis C virus-positive donations, with all in this group treated with direct-acting antivirals. We found no statistically significant differences regarding age, sex, race, donation after cardiac death, or incidence of hepatocellular carcinoma between groups. The treated group had a longer median wait time (287 vs 172 days; P = .02). Twelve of the 46 untreated patients (26.1%) developed biopsy-proven hepatitis C virus-related relapse (median 87 days; range, 55-383 days). Preemptive direct-acting antiviral therapy was initiated at a median of 81 days in the untreated group. CONCLUSIONS: Although treatment of hepatitis C virus before liver transplant is an attractive option to eliminate the risk of complications, it can limit the donor pool for recipients to uninfected donors, significantly increasing wait times in regions with large hepatitis C virus-positive donor pools. Allocation of Model for End-Stage Liver Disease score was not different between the treated and untreated groups. Insurance companies should revise their policies for rapid approval of preemptive direct-acting antiviral treatment after liver transplant.

Microvascular Invasion in Hepatocellular Carcinoma and Liver Transplant.

OBJECTIVES: Curative therapy for hepatocellular carcinoma is liver transplant. To date, the Milan Criteria remain the best pretransplant clinical surrogate for tumor behavior and overall prognosis. Microvascular invasion portends a poor prognosis; however, it is often undetectable before transplant. Furthermore, its pretransplant indicators are not well established. In this study, we investigated the presurgical and pathologic predictors of microvascular invasion in patients with hepatocellular carcinoma. MATERIALS AND METHODS: Between August 2000 and August 2013, 156 liver transplants were performed for hepatocellular carcinoma at the Johns Hopkins Medical Center. Information on clinical characteristics and pathology data, including microvascular invasion, were available for 107 patients on liver explants. Logistic regression was used to assess the effects of Milan Criteria, alpha-fetoprotein, tumor differentiation, and multilobar involvement on the presence of microvascular invasion on explant pathology. RESULTS: In 107 patients, 24 (22%) had microvascular invasion on pathology. In patients with microvascular invasion, 41% were outside of Milan Criteria versus 19.3% of patients within but without microvascular invasion. In patients with microvascular invasion, the rate of poor differentiation and alpha-fetoprotein level > 1000 ng/mL were more common than in patients without microvascular invasion; however, on univariate and multivariable analyses, Milan Criteria, alphafetoprotein level, multilobar involvement, and differentiation did not reach statistical significance in predicting microvascular invasion on pathology. CONCLUSIONS: In this study, potential predictors of microvascular invasion, including Milan Criteria, alphafetoprotein level, tumor differentiation, and multilobar involvement, were not predictive. Preoperative prediction of microvascular invasion remains a challenge, suggesting the need for future studies.