Next-Generation Sequencing of Circulating Tumor DNA Reveals Frequent Alterations in Advanced Hepatocellular Carcinoma.

BACKGROUND: Because imaging has a high sensitivity to diagnose hepatocellular carcinoma (HCC) and tissue biopsies carry risks such as bleeding, the latter are often not performed in HCC. Blood-derived circulating tumor DNA (ctDNA) analysis can identify somatic alterations, but its utility has not been characterized in HCC. MATERIALS AND METHODS: We evaluated 14 patients with advanced HCC (digital ctDNA sequencing [68 genes]). Mutant relative to wild-type allele fraction was calculated. RESULTS: All patients (100%) had somatic alterations (median = 3 alterations/patient [range, 1-8]); median mutant allele fraction, 0.29% (range, 0.1%-37.77%). Mutations were identified in several genes: TP53 (57% of patients), CTNNB1 (29%), PTEN (7%), CDKN2A (7%), ARID1A (7%), and MET (7%); amplifications, in CDK6 (14%), EGFR (14%), MYC (14%), BRAF (7%), RAF1 (7%), FGFR1 (7%), CCNE1 (7%), PIK3CA (7%), and ERBB2/HER2 (7%). Eleven patients (79%) had ≥1 theoretically actionable alteration. No two patients had identical genomic portfolios, suggesting the need for customized treatment. A patient with a CDKN2A-inactivating and a CTNNB1-activating mutation received matched treatment: palbociclib (CDK4/6 inhibitor) and celecoxib (COX-2/Wnt inhibitor); des-gamma-carboxy prothrombin level decreased by 84% at 2 months (1,410 to 242 ng/mL [normal: ≤7.4 ng/mL]; alpha fetoprotein [AFP] low at baseline). A patient with a PTEN-inactivating and a MET-activating mutation (an effect suggested by in silico molecular dynamic simulations) received sirolimus (mechanistic target of rapamycin inhibitor) and cabozantinib (MET inhibitor); AFP declined by 63% (8,320 to 3,045 ng/mL [normal: 0-15 ng/mL]). CONCLUSION: ctDNA derived from noninvasive blood tests can provide exploitable genomic profiles in patients with HCC. IMPLICATIONS FOR PRACTICE: This study reports that blood-derived circulating tumor DNA can provide therapeutically exploitable genomic profiles in hepatocellular cancer, a malignancy that is known to be difficult to biopsy.

Lower than expected hepatitis B virus infection prevalence among first generation Koreans in the U.S.: results of HBV screening in the Southern California Inland Empire.

BACKGROUND: Hepatitis B virus (HBV) infection is prevalent in Asian immigrants in the USA. California's Inland Empire region has a population of approximately four million, including an estimated 19,000 first generation Koreans. Our aim was to screen these adult individuals to establish HBV serological diagnoses, educate, and establish linkage to care. METHODS: A community-based program was conducted in Korean churches from 11/2009 to 2/2010. Subjects were asked to complete a HBV background related questionnaire, provided with HBV education, and tested for serum HBsAg, HBsAb and HBcAb. HBsAg positive subjects were tested for HBV quantitative DNA, HBeAg and HBeAb, counseled and directed to healthcare providers. Subjects unexposed to HBV were invited to attend a HBV vaccination clinic. RESULTS: A total of 973 first generation Koreans were screened, aged 52.3y (18-93y), M/F: 384/589. Most (75%) had a higher than high school education and were from Seoul (62.2%). By questionnaire, 24.7% stated they had been vaccinated against HBV. The serological diagnoses were: HBV infected (3.0%), immune due to natural infection (35.7%), susceptible (20.1%), immune due to vaccination (40.3%), and other (0.9%). Men had a higher infection prevalence (4.9% vs. 1.7%, p = 0.004) and a lower vaccination rate (34.6% vs. 44.0%, p = 0.004) compared to women. Self-reports of immunization status were incorrect for 35.1% of subjects. CONCLUSIONS: This large screening study in first generation Koreans in Southern California demonstrates: 1) a lower than expected HBV prevalence (3%), 2) a continued need for vaccination, and 3) a need for screening despite a reported history of vaccination.

Hepatocellular carcinoma surveillance: a national survey of current practices in the USA.

BACKGROUND: The incidence rate of hepatocellular carcinoma (HCC) in the USA is rapidly rising. Surveillance using biannual ultrasound (US) is recommended, but actual practices are unknown. AIM: To determine current HCC surveillance practices among gastroenterology and hepatology providers in the USA. METHODS: A 21-question electronic survey was emailed to 12,485 gastroenterology and hepatology providers in the USA. The survey contained questions concerning provider background, surveillance practices, and opinions. Pearson chi-square and multivariate logistic regression tests were used to analyze the data. RESULTS: We received 777 responses (6.2% response rate); 656 were eligible for analysis. 92% place cirrhotic patients under surveillance independent of etiology, 79% exclusively use a 6-month interval, and 77% use alpha-fetoprotein. While 93% use US, only 36% use US exclusively and 60% use two or more imaging modalities. Providers from transplant centers favor using additional imaging modalities, instead of only US. Multivariate analysis showed transplant center providers who allocate more time to patient care (OR 1.96, p = 0.004) and see more cirrhotic patients (OR 2.07, p = 0.033) have increased odds of using additional imaging modalities. CONCLUSIONS: Participating providers reported very high rates of surveillance utilization and use of a biannual interval. It is likely that the sample is composed of providers who are very engaged and informed about HCC surveillance. However, their surveillance imaging practices largely deviated from practice guidelines, which all recommend only using US. Providers affiliated with transplant centers tend to use additional imaging modalities such as computed tomography and MRI, instead of US only.

Use of ImPACT to diagnose minimal hepatic encephalopathy: an accurate, practical, user-friendly internet-based neuropsychological test battery.

BACKGROUND AND AIMS: An effective, user-friendly neurocognitive test to diagnose minimal hepatic encephalopathy (MHE) is needed. Immediate Post-concussion Assessment and Cognitive Testing (ImPACT) is a brief, validated, Web-based, neuropsychological test battery resulting in four composite scores [Verbal Memory (VrbM), Visual Memory, Visual Motor Speed (VMS), Reaction Time (RT)]. We compared ImPACT to traditional paper-and-pencil tests in patients at risk for MHE versus controls. METHODS: Ninety cirrhotic patients with no history of overt hepatic encephalopathy were compared with 131 controls on standard psychometric tests (SPT) [Trail Making Test-A, Trail Making Test-B, Digit Symbol Test], 4 ImPACT composite scores, and the Sickness Impact Profile (SIP). MHE+ was defined by a score 2 SD below the normative mean on at least one of the SPT. ImPACT (ImP+) scores of patients were defined as 2 SD from the control mean. RESULTS: Cirrhotic patients scored more poorly than controls on 3/4 of ImPACT scores: VrbM (78.88 vs. 71.37, p<0.001), VMS (26.47 vs. 22.68, p<0.001) and RT (0.89 vs. 1.00, p<0.01), as well as on all 3 SPT. Of the 90 cirrhotics, 16 (18%) were MHE+, who performed more poorly (p<0.001) than patients without MHE on VrbM (58.13 vs. 74.19), VMS (16.77 vs. 23.95) and RT (1.24 vs. 0.95). Of the 90 cirrhotics, 25 (27.8%) were ImP+. MHE+ and ImP+ patients had increased SIP scores versus controls (p<0.001). CONCLUSIONS: Compared to paper-and-pencil testing, ImPACT provides a brief, user-friendly, neuropsychological evaluation of MHE. ImPACT could become a new standard for MHE diagnosis.

CON: Noninvasive Imaging Is the Preferred Strategy for Cardiovascular Risk Stratification in This Patient.

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Rescue liver re-transplantation after graft loss due to severe rejection in the setting of pre-transplant nivolumab therapy.

Immune checkpoint inhibitors (ICI) have been used to treat hepatocellular carcinoma (HCC) since 2017. The safety of ICIs in the setting of solid organ transplantation remains controversial. When used in the post-transplant setting, ICIs have been associated with high allograft rejection rates, but there are few published reports on the use of ICIs prior to transplant. We present the first reported case of rescue liver re-transplantation after loss of the first allograft due to severe acute rejection with extensive hepatic necrosis in the setting of pre-transplant ICI therapy with the PD-1 inhibitor nivolumab. It is likely that the durable immune response triggered by nivolumab contributes to graft rejection, therefore extreme caution should be taken when using ICIs before transplant until further investigation has been conducted on their safety in the pre-transplant setting.

A new highly sensitive point of care screen for spontaneous bacterial peritonitis using the leukocyte esterase method.

BACKGROUND & AIMS: Urine reagent strips measuring leukocyte esterase activity have been studied to screen spontaneous bacterial peritonitis (SBP) but are insensitive. We calibrated a strip specifically for ascitic fluid to achieve high sensitivity in this diagnosis. METHODS: Experiments were conducted on ascitic fluid from patients with cirrhosis. Samples with SBP were diluted with native acellular ascitic fluid to achieve PMN counts below, above, and close to the diagnostic threshold of 250 PMN/microl. A model of SBP was created by spiking negative ascitic fluid samples (<250 PMN/microl) with activated PMN from blood of patients with sepsis, and diluted to achieve a range of PMN. Aliquots were tested at 2, 3, 4, and 10 min with the Periscreen leukocyte esterase strip. PMN/microl was correlated to timings and color scales: white defined negative (PMN <250/microl); and shades of brown, purple, and pink defined positive. Ascitic fluid samples were obtained from 58 patients. Negative ascitic fluid was used from 32 to generate the model SBP. RESULTS: One thousand three hundred and four experiments were performed with a median PMN count of 492/microl (0-7510). After exclusion of uninterpretable colorimetric results, 1089 experiments were analyzed [PMN of 444/microl (0-7510)]. The best result was obtained at 3 min (n=299), with Se: 100%, Sp: 57.9%; NPV: 100%, PPV: 76.5%. The test was not interpretable in bloody, chylous or bilious ascitic fluid, or concurrent imipenem treatment. CONCLUSIONS: This new leukocyte esterase strip calibrated to an ascitic fluid PMN count 250/microl is a robust screening tool when the strip turns any hue of tan/brown at 3 min.

Monitoring peripheral blood CD4+ adenosine triphosphate activity in a liver transplant cohort: insight into the interplay between hepatitis C virus infection and cellular immunity.

Peripheral blood CD4+ adenosine triphosphate [ATP (ng/mL)] release [ImmuKnow Immune Cell Function Assay (ATP)] correlates to immunoreactivity. We hypothesized that ATP levels could provide insight into hepatitis C virus (HCV) infection and recurrent liver disease in liver transplantation (LT). We studied our center's LT cohort, in which ATP levels had been measured off protocol from February 2005 through July 2006. Of the 280 LTs performed since 1993, 114 (58.2%) fit the selection criteria, with a mean age of 49 +/- 10 years. LT (alone/combination) indications included HCV (58%), alcoholic liver disease (41%), hepatocellular carcinoma (16%), and other (33%). Four hundred seventy-seven ATP levels were obtained: 3 (1-17) per patient 25 months (4 days to 19 years) from the time from transplantation. Final diagnoses were normal allograft function (n = 166, 35%), recurrent disease (n = 199, 42%), septic event (n = 34, 7%), other (n = 51, 11%), and undetermined (n = 27, 6%). Two hundred eighty-one ATP levels were obtained [3 (1-18) per patient] in 66 HCV(+) patients. Forty-five (68%) developed biopsy-proven recurrent liver disease [188/281 (67%) ATP levels]. The median ATP level (ng/mL) was 162 (1-761); it was lower in HCV(+) patients (151 +/- 109) versus HCV(-) patients (211 +/- 139; P < 0.0001). ATP ranges in HCV(+) patients were stable from the time from transplantation. In HCV(-) patients, ATP ranges were initially high and eventually decreased to HCV(+) levels (P = 0.01). Immunosuppressant levels were low in 62% of HCV(-) patients versus 38% of HCV(+) patients (P = 0.04). In HCV(+) patients, ATP was lower in disease recurrence (139 +/- 97) versus none (181 +/- 141; P = 0.01) with similar immunosuppression, and ATP decreased with grade (P = 0.05) but not stage. Time from transplantation, aspartate aminotransferase/alanine aminotransferase >1, and low ATP were independently associated with recurrent HCV. In conclusion, after LT, global cellular immune function appears depressed at baseline in HCV(+) patients versus HCV(-) patients and more so in HCV(+) recurrent disease.

Drug-Induced Liver Injury Associated with the Use of Everolimus in a Liver Transplant Patient.

Drug-induced liver injury (DILI) has not been previously reported as a complication of treatment with everolimus. A 56-year-old Caucasian male liver transplant recipient developed DILI after receiving everolimus. Elevations in transaminase levels occurred within a week of starting everolimus and an upward trend in the transaminase levels continued with supporting histopathologic changes confirmed by liver biopsy. Within one week of drug discontinuation, his liver enzymes normalized to baseline. This report includes a brief review of the pharmacokinetic properties of everolimus, a review of the relevant literature, and an analysis using the RUCAM and Naranjo algorithms.

Nonalcoholic fatty liver disease with cirrhosis increases familial risk for advanced fibrosis.

BACKGROUND: The risk of advanced fibrosis in first-degree relatives of patients with nonalcoholic fatty liver disease and cirrhosis (NAFLD-cirrhosis) is unknown and needs to be systematically quantified. We aimed to prospectively assess the risk of advanced fibrosis in first-degree relatives of probands with NAFLD-cirrhosis. METHODS: This is a cross-sectional analysis of a prospective cohort of 26 probands with NAFLD-cirrhosis and 39 first-degree relatives. The control population included 69 community-dwelling twin, sib-sib, or parent-offspring pairs (n = 138), comprising 69 individuals randomly ascertained to be without evidence of NAFLD and 69 of their first-degree relatives. The primary outcome was presence of advanced fibrosis (stage 3 or 4 fibrosis). NAFLD was assessed clinically and quantified by MRI proton density fat fraction (MRI-PDFF). Advanced fibrosis was diagnosed by liver stiffness greater than 3.63 kPa using magnetic resonance elastography (MRE). RESULTS: The prevalence of advanced fibrosis in first-degree relatives of probands with NAFLD-cirrhosis was significantly higher than that in the control population (17.9% vs. 1.4%, P = 0.0032). Compared with controls, the odds of advanced fibrosis among the first-degree relatives of probands with NAFLD-cirrhosis were odds ratio 14.9 (95% CI, 1.8-126.0, P = 0.0133). Even after multivariable adjustment by age, sex, Hispanic ethnicity, BMI, and diabetes status, the risk of advanced fibrosis remained both statistically and clinically significant (multivariable-adjusted odds ratio 12.5; 95% CI, 1.1-146.1, P = 0.0438). CONCLUSION: Using a well-phenotyped familial cohort, we demonstrated that first-degree relatives of probands with NAFLD-cirrhosis have a 12 times higher risk of advanced fibrosis. Advanced fibrosis screening may be considered in first-degree relatives of NAFLD-cirrhosis patients. UCSD IRB: 140084. FUNDING: National Institute of Diabetes and Digestive and Kidney Diseases and National Institute of Environmental Health Sciences, NIH.

A pilot study of an alcoholic liver disease recurrence prevention education program in hospitalized patients with advanced liver disease.

No systematic work has been completed to assess whether or not educational programming might exert lifestyle improvements among alcoholic liver disease (ALD) inpatients. The present pilot study sought to answer this question through the use of a small-scale two-group experiment (five-session education program versus standard care) at a state-of-the art Liver Unit that provided tertiary care of indigent patients with advanced ALD. A total of 44 patients were initially randomly assigned to program conditions, and 25 provided 3-month follow-up data (13 in the program condition, 12 in the control condition). Patients who received the program reported high receptivity to it, and showed greater learning of program material and reported greater lifestyle changes than the control patients. For those ALD inpatients that are able and willing to participate, the program shows promising effects on self-reported lifestyle change.

HFE based re-evaluation of heterozygous hemochromatosis.

Homozygosity for the C282Y mutation in the HFE gene is strongly associated with hereditary hemochromatosis. More than one subject out of 10 in the general population is a heterozygote for the C282Y mutation. In this study, we address whether or not conclusions drawn from HLA-based family studies regarding the expression of heterozygous hemochromatosis are applicable to C282Y heterozygotes. The correlation between HLA-inferred and HFE genotypes and the variation of serum iron tests according to HFE genotype and other factors were studied in persons from well-characterized hemochromatosis pedigrees. Subjects were tested for both C282Y and H63D mutations. The following factors were studied: age, sex, alcohol consumption, body mass index, liver function tests, serum lipids and glucose, serum iron, transferrin saturation, and ferritin. HLA-inferred heterozygotes were C282Y heterozygotes in only 70% and compound heterozygotes (i.e., heterozygotes for both C282Y and H63D) in 20%. C282Y heterozygotes did not differ from wild type homozygotes in terms of serum iron tests. Only compound heterozygotes presented with slightly increased transferrin saturation. On the other hand, increased serum ferritin was strongly associated with overweight or lipidic or glucose abnormalities. C282Y heterozygotes selected from family studies do not have greater serum iron tests than wild type homozygotes, except for compound heterozygotes, and therefore should not require special followup. The discovery of abnormal iron tests in a C282Y heterozygote should lead to workup for other causes of iron overload.

High-dose tigecycline and colistin for successful treatment of disseminated carbapenem-resistant Klebsiella pneumoniae infection in a liver transplant recipient.

Solid organ transplantation (SOT) is a risk factor for the acquisition of carbapenem-resistant Klebsiella pneumoniae. This infection is associated with a high mortality rate given the limited armamentarium of antibiotics for multidrug-resistant organisms along with continued immunosuppression to prevent graft rejection. We report a case of carbapenem-resistant K. pneumoniae pneumonia, bacteraemia and intra-abdominal infection in a newly transplanted liver recipient. The patient was successfully treated with a long course of high-dose tigecycline and colistin, along with surgical drainage. We discuss SOT-relevant epidemiology, therapeutic options and the rationale for our treatment choice.

Sodium benzoate for treatment of hepatic encephalopathy.

Hepatic encephalopathy (HE) is a serious but usually reversible neuropsychiatric complication of cirrhosis, inborn errors of metabolism involving disorders of the urea cycle, and noncirrhotic portosystemic shunting that most commonly arises from a transjugular intrahepatic portosystemic shunting procedure. Symptoms can include alterations in cognitive function, neuromuscular activity, and consciousness, as well as sleep disorders and mood changes. HE is associated with significant morbidity and mortality and, if not properly treated, will lead to increased hospital admissions and healthcare costs. Although the standard therapies of lactulose and rifaximin (Xifaxan, Salix) are effective for most patients, these drugs may be associated with significant adverse effects and expense and, in some patients, inadequate therapeutic response. A need for adjunctive therapies exists. Drugs that target serum and tissue ammonia metabolism and elimination may be important adjuncts to drugs that reduce ammonia production and absorption from the gastrointestinal tract for patients with severe or persistent overt symptoms of HE. Sodium benzoate is an inexpensive adjunctive agent that can be used in addition to lactulose and rifaximin and may provide an option for some select patients with refractory HE who have failed to respond to standard therapies or who cannot afford them. Although sodium benzoate does not share the same adverse effect profiles of standard therapies for HE, its efficacy has not been well established. Given the significant dose-dependent sodium content of this therapy, it may not be appropriate for patients with significant fluid retention or kidney dysfunction.

Concomitant hepatic encephalopathy and refractory ascites: successful treatment with staged embolization of two large portosystemic shunts and transjugular intrahepatic portosystemic shunt placement.

A patient with cirrhosis, refractory ascites, and two large competitive portosystemic shunts presented with uncontrollable acute exacerbation of chronic hepatic encephalopathy (HE). A staged procedure was performed by first performing embolization of a large mesogonadal shunt to treat the HE. Three months later, a transjugular intrahepatic portosystemic shunt (TIPS) was created to address the ascites. A large paraumbilical vein shunt was embolized at TIPS placement to minimize the risk of recurrent HE. At 9-month follow-up, the ascites was well controlled with medical management with little or no HE.

Prevalence of septic events, type 1 hepatorenal syndrome, and mortality in severe alcoholic hepatitis and utility of discriminant function and MELD score in predicting these adverse events.

We sought to assess prevalence, and utility of discriminant function (DF) and MELD score in predicting septic events (SE), type 1 hepatorenal syndrome (HRS), and short-term mortality in severe alcoholic hepatitis (AH). Charts of patients with AH (group 1) and cirrhosis without AH (group 2) were retrospectively reviewed. Severe AH, discriminant function (DF) >or= 32 was treated with pentoxifylline. One hundred ninety-five patients were enrolled in the study and divided into 2 groups: group 1, n=99, and group 2, n=96. Of those with AH, 82% had a DF >or= 32 at presentation. Group 1 patients had a higher prevalence of SE (38% versus 25%, P=.04), type 1 HRS (30% versus 9%, P=.0003), and short-term mortality (28% versus 7%, P=.0001). In patients with AH, a MELD score >or=20 (but not a DF >or= 32) at presentation was an independent predictor of a SE (odds ratio [OR] 2.8 [1.0-7.9], P=.04), HRS (OR 4.0, 95% confidence interval [CI] 1.0-16.6, P=0.05), and short-term mortality (OR 6.4, 95% CI 1.1-37.6, P=.03). Kaplan-Meier survival curves confirmed that that a MELD >or= 20 but not a DF >or= 32 was associated with a poorer survival (P = .005 and .5, respectively). In conclusion, patients with severe AH have higher prevalence of SE, HRS, and short-term mortality compared to those with cirrhosis without AH. A MELD score >or=20 at presentation is an independent predictor of these adverse events in patients with AH who have been treated with pentoxifylline.

Proposal for a histological scoring and grading system for non-alcoholic fatty liver disease.

AIM/BACKGROUND: The spectrum of histopathological features in non-alcoholic fatty liver disease (NAFLD) has been well described. At least two scoring systems have been established. We propose here a system in which numerical scores are obtained using the different features. METHODS: Twenty-five cases of well-defined NAFLD were identified. Two pathologists blinded to identifiers and clinical data independently scored the liver biopsies twice for portal fibrosis (PF: 0-6), lobular inflammation and necrosis (LIN: 0-3), Mallory bodies (MB: 0-3), hepatocyte ballooning (HB: 0-3), perisinusoidal fibrosis (PSF: 0-3) and fatty change (FC: 1-4). The kappa statistic tested observer concordance. Non-parametric measures of correlation and hierarchical cluster analysis were used to elaborate a grading system. RESULTS: A broad spectrum of NAFLD was observed. Intra- and interobserver concordance was satisfactory. An activity score was created (AS: 0-12) as the sum of LIN, MB, HB and PSF, but not FC. A system for severity of NAFLD was developed: Grade 1 (PF: 0-2 and AS: 0-4), Grade 2 (PF: 3 or AS: 5-7) and Grade 3 (PF: 4-6 or AS: 8-12). Diabetes, elevated alkaline phosphatase and decreased platelets were associated with advanced grade. CONCLUSIONS: This simple, reproducible NAFLD score produces a three-tier severity grade. This numerical system may prove useful in assessing disease severity and interval changes.

Choosing the location for non-image guided abdominal paracentesis.

OBJECTIVES: The optimal location for paracentesis has not been studied scientifically. The evolving obesity epidemic has changed the physique of many patients with cirrhosis and ascites such that needles inserted into the abdominal wall may not reach fluid. We aimed to determine the location for paracentesis that would have the thinnest abdominal wall and the deepest amount of fluid. METHODS: Ultrasound measurements of abdominal wall thickness and depth of ascites were recorded in two locations, the infraumbilical midline (ML) and the left lower quadrant (LLQ), in 52 patients with cirrhosis and ascites admitted to a single inpatient liver unit. RESULTS: The abdominal wall was significantly thinner (1.8 vs. 2.4 cm; P<0.001) and the depth of ascites greater (2.86 vs. 2.29 cm; P=0.017) in the LLQ as compared with the infraumbilical ML position. In the left lateral oblique position, the difference in the depth of ascites was more pronounced when comparing the LLQ with the infraumbilical ML (4.57 vs. 2.78 cm; P<0.0001). CONCLUSIONS: The LLQ is preferable to the ML infraumbilical location for performing paracentesis.