Oskar Kornasiewicz

Next generation sequencing reveals microRNA isoforms in liver cirrhosis and hepatocellular carcinoma

Hepatocellular carcinoma (HCC) represents the major histological subtype of liver cancer. Tumorigenic changes in hepatic cells potentially result from aberrant expression of microRNAs (miRNAs). Individual microRNA gene may give rise to miRNAs of different length, named isomiRNAs that proved to be functionally relevant. Since microRNA length heterogeneity in hepatic tissue has not been described before, we employed next-generation sequencing to comprehensively analyze microRNA transcriptome in HCC tumors (n=24) and unaffected tissue adjacent to tumors (n=24), including samples with (n=15) and without cirrhosis (n=9). We detected 374 microRNAs expressed in liver, including miR-122-5p that constituted over 39% of the hepatic miRnome. Among the liver expressed miRs, the levels of 64 significantly differed between tumor and control samples (FDR<0.05, fold change>2). Top deregulated miRNAs included miR-1269a (T/N=22.95), miR-3144-3p (T/N=5.24), miR-183-5p (T/N=4.63), miR-10b-5p (T/N=3.87), miR-490-3p (T/N=0.13), miR-199a-5p (T/N=0.17), miR-199a-3p/miR-199b-3p (T/N=0.19), miR-214-5p (T/N=0.20) and miR-214-3p (T/N=0.21). Almost all miRNA genes produced several mature molecules differing in length (isomiRNAs). The reference sequence was not the most prevalent in 38.6% and completely absent in 10.5% of isomiRNAs. Over 26.1% of miRNAs produced isoforms carrying≥2 alternative seed regions, of which 35.5% constituted novel, previously unknown seeds. This fact sheds new light on the percentage of the human genome regulated by microRNAs and their variants. Among the most deregulated miRNAs, miR-199a-3p/miR-199b-3p (T/N fold change=0.18, FDR=0.005) was expressed in 9 isoforms with 3 different seeds, concertedly leading to upregulation of TGF-beta signaling pathway (OR=1.99; p=0.004). In conclusion, the study reveals the comprehensive miRNome of hepatic tissue and provides new tools for investigation of microRNA-dependent pathways in cirrhotic liver and hepatocellular carcinoma. This article is part of a Directed Issue entitled: Rare Cancers.

Impact of Tumor Characteristic on the Outcome of Liver Transplantation in Patients With Hepatocellular Carcinoma

INTRODUCTION Orthotopic liver transplantation (OLT) is a well-established treatment for cirrhotic patients with hepatocellular carcinoma (HCC) who meet the Milan criteria. The aim of this study was to identify predictors of survival among 65 patients with HCC in cirrhotic livers who underwent liver transplantation (OLT). METHODS From January 2001 to December 2008, we performed 655 OLT in 615 patients. HCC was diagnosed in 58 patients before OLT and in 65 by histological examination of the explanted livers; 74% of the patients met Milan criteria by histological examination. RESULTS The median follow-up was 27 months (range = 1-96). We analyzed patient age and gender, etiology of liver disease, Child score at transplantation, rejection episodes, tumor number/size, vascular invasion, and differentiation grade. There was no significant difference in survival among patients grouped according to the Model for End-stage Liver Disease staging system for HCC. The 5-year survival of patients with low differentiated (G3) HCC was significantly worse than that of those with moderately differentiated (G2) or well-differentiated (G1) HCC: 50%, 81%, and 86% respectively, (P < .01). Patients with microvascular invasion displayed a worse 5-year survival than those without vascular invasion (42% vs 80%; P < .01). CONCLUSIONS The analysis indicated that the histological grade of the tumors and evidences of microscopic vascular invasion were the most useful predictive factors for overall survival among patients with cirrhosis after liver transplantation for HCC.

1000 liver transplantations at the Department of General, Transplant and Liver Surgery, Medical University of Warsaw--analysis of indications and results.

THE AIM OF THE STUDY was to analyze indications and results of the first one thousand liver transplantations at Chair and Clinic of General, Transplantation and Liver Surgery, Medical University of Warsaw. MATERIAL AND METHODS Data from 1000 transplantations (944 patients) performed at Chair and Clinic of General, Transplantation and Liver Surgery between 1994 and 2011 were analyzed retrospectively. These included 943 first transplantations and 55 retransplantations and 2 re-retransplantations. Frequency of particular indications for first transplantation and retransplantations was established. Perioperative mortality was defined as death within 30 days after the transplantation. Kaplan-Meier survival analysis was used to estimate 5-year patient and graft survival. RESULTS The most common indications for first transplantation included: liver failure caused by hepatitis C infection (27.8%) and hepatitis B infection (18%) and alcoholic liver disease (17.7%). Early (< 6 months) and late (> 6 months) retransplantations were dominated by hepatic artery thrombosis (54.3%) and recurrence of the underlying disease (45%). Perioperative mortality rate was 8.9% for first transplantations and 34.5% for retransplantations. Five-year patient and graft survival rate was 74.3% and 71%, respectively, after first transplantations and 54.7% and 52.9%, respectively, after retransplantations. CONCLUSIONS Development of liver transplantation program provided more than 1000 transplantations and excellent long-term results. Liver failure caused by hepatitis C and B infections remains the most common cause of liver transplantation and structure of other indications is consistent with European data.

Incidence, pattern and clinical relevance of microbial contamination of preservation fluid in liver transplantation

BACKGROUND Transmission of pathogens via preservation fluid (PF) is a potential cause of infection among liver transplant recipients. Here, we evaluated the incidence and pattern of microbial contamination of PF and its impact on postoperative graft function after liver transplantation. MATERIAL/METHODS This longitudinal study included data from 41 primary liver transplantations and 5 re-transplantations performed between December 2010 and September 2011. Results of microbiological analyses of 92 PF samples collected before and after the back-table procedure were evaluated in order to establish the incidence and pattern of contamination. The impact of positive PF cultures on early graft function and rate of pathogen transmission was assessed. Post-transplant antibiotic protocol was based on piperacillin/tazobactam administration for a minimum of 10 days. RESULTS The incidence of contamination was 84.8% (39/46), both for samples collected before and after the back-table procedure. Gram-positive low-virulence organisms typical for superficial saprophytic flora, mainly coagulase-negative staphylococci, were predominant. There were no cases of pathogen transmission from PF to the recipient. Positive cultures of PF samples obtained after the back-table procedure were associated with significant elevation of aspartate (p=0.034) and alanine aminotransferase (p=0.048) on the first 5 postoperative days. No significant differences were found regarding serum bilirubin concentration (p=0.335) and international normalized ratio (p=0.137). CONCLUSIONS Despite high incidence of PF contamination, infections caused by pathogens isolated from PF were not observed. However, presence of pathogens in PF might lead to temporary impairment of graft function.

Choice of Transplantation Techniques and Indications for Liver Transplantation in Polycystic Liver Disease in Patients With No Signs of End-Stage Liver Disease

OBJECTIVE Since the initiation of the Liver Transplant Program, 500 liver procedures have been performed. Polycystic liver disease (PLD) and polycystic kidney-liver disease (PKLD) have been rare indications for orthotopic liver transplantation (OLT). Only 7 patients (1.4%) underwent transplantation due to PLD and PKLD. MATERIALS AND METHODS The group consisted of 4 patients who underwent OLT (0.8%) and 3 patients who received simultaneous liver kidney transplantation (LKT; 0.6%). Our objective was to analyze the indications for either OLT or combined LKT as well as indications for surgical techniques during OLT among patients with PLD or PKLD. RESULTS The main indication for OLT was massive hepatomegaly causing severe physical handicaps, fatigue, and clinically advanced malnutrition. All 3 patients with indications for combined LKT were dialysis-dependent. None of the patients had symptoms of end-stage liver disease and/or hepatic failure. In 4 cases, a portal bypass was applied, and the piggy-back method used in the other 3 cases. The hepatectomy caused no uncommon difficulty. In cases of simultaneous transplantations, the kidney was implanted separately after OLT. All patients are alive following the transplantation; major surgical complications have occurred. CONCLUSIONS Patients with PLD can undergo OLT safely with good results. They benefit from the relief of abdominal distension and anorexia. Patients with PKLD who are dialysis-dependent should undergo simultaneous LKT. The surgical technique was solely dependent on the intraoperative conditions determined during the dissection phase.

Early post-operative prediction of morbidity and mortality after a major liver resection for colorectal metastases

BACKGROUND An early prediction of poor outcomes is essential in the management of patients after a liver resection. The aim of this study was to evaluate the role of selected biochemical parameters on post-operative day 1 (POD 1) in the prediction of morbidity and mortality after a liver resection for colorectal metastases. METHOD This retrospective study was based on 236 major liver resections for colorectal metastases performed between 2006 and 2011. Results of biochemical tests of blood samples obtained on POD 1 were assessed as predictors of primary outcome measures (hepatic and overall morbidity, 90-day mortality) using multiple regression and receiver-operating characteristics (ROC). RESULTS Hepatic morbidity, overall morbidity and 90-day mortality rates were 18.6%, 28.0% and 4.7%, respectively. On the basis of multiple regression analysis and comparisons of the prediction models, serum bilirubin was selected for the prediction of hepatic (>2.05 mg/dl, sensitivity 69.2%, specificity 71.2%) and overall (>2.05 mg/dl, sensitivity 61.1% and specificity 71.2%) morbidity, and aspartate aminotransferase (AST) was selected for the prediction of 90-day mortality (>798 U/l, sensitivity 62.5% and specificity 90.4%). DISCUSSION Biochemical analyses of blood on POD1 enables stratification of patients into low- and high-risk groups for negative outcomes, with serum bilirubin associated with overall and hepatic morbidity and AST associated with mortality.

Post-transplant outcomes of patients with and without hepatitis C virus infection according to donor age and gender matching

BACKGROUND The purpose of this study was to evaluate the impact of donor age and donor-recipient gender matching on liver transplantation outcomes, focusing on differences between patients with and without hepatitis C virus (HCV) infection. MATERIAL AND METHODS This retrospective cohort study evaluated 622 liver transplantation recipients. HCV (n=164) and non-HCV (n=458) patients were subdivided by donor age (≤ 30, 31-50, and >50 years) and donor-recipient gender configurations. Five-year patient survival (PS) and graft survival (GS) were set as outcome measures. RESULTS Five-year PS was 83.1% for HCV-positive and 81.6% for HCV-negative patients (p=0.614), with the corresponding GS rates of 81.2% and 79.3% (p=0.538), respectively. In HCV patients, transplantations from donors older than 50 years were associated with lower PS (p=0.035) and GS (p=0.006) than those from donors aged 31-50 years. This difference was not observed among non-HCV recipients (PS, p=0.994; GS, p=0.878). Regarding donor-recipient gender configurations, outcomes were similar in HCV (PS, p=0.751; GS, p=0.592) and non-HCV patients (PS, p=0.217; GS, p=0.249), except for a tendency toward lower PS for male-to-female transplantations than female-to-female transplantations in non-HCV patients (p=0.064). Outcomes of HCV patients were superior to those of non-HCV patients after transplantation from donors aged 31-50 years (PS, p=0.080; GS, p=0.026). CONCLUSIONS Avoiding the transplantation of grafts from donors aged over 50 years to patients with HCV infection might improve the general outcomes of liver transplantation programs. There is no specific rationale for gender matching with respect to HCV status.

The impact of surgical technique on the results of liver transplantation in patients with hepatocellular carcinoma

BACKGROUND Although piggyback technique has gained widespread acceptance for liver transplantation in general, there is an exceptional lack of data on the choice of appropriate surgical technique for patients with hepatocellular carcinoma (HCC). The purpose of this study was to evaluate the impact of surgical technique on outcomes after liver transplantation for HCC. MATERIAL AND METHODS We conducted a retrospective cohort study on 90 HCC patients who underwent liver transplantation with the conventional (n=19) or piggyback (n=71) technique. Both techniques were compared with respect to intraoperative variables and long-term outcomes, determined by 3-year overall (OS) and recurrence-free (RFS) survival. The potential role of confounding factors was excluded in a series of Cox proportional regression models. RESULTS The piggyback technique was associated with shorter procedure duration (p=0.0005), shorter anatomical anhepatic phase (p<0.0001), shorter duration of total (p=0.018) and warm ischemia (p<0.0001), and fewer blood transfusions (p=0.006). Three-year OS was 89.1% after piggyback and 49.9% after conventional transplantation (p=0.0008), with 3-year RFS of 89.4% and 56.0% (p=0.0006), respectively. Piggyback transplantations provided outcomes superior to conventional procedures both in patients within (p=0.019 for OS; p=0.003 for RFS) and beyond (p=0.023 for OS; p=0.031 for RFS) Milan criteria. Multivariate analysis of the risks of death and recurrence confirmed the benefits of piggyback technique. CONCLUSIONS Given its superior long-term outcome, piggyback transplantation might be considered primarily for HCC patients.

Evaluation of Total Tumor Volume and Pretransplantation α-Fetoprotein Level as Selection Criteria for Liver Transplantation in Patients With Hepatocellular Cancer

INTRODUCTION Appropriate selection of hepatocellular cancer (HCC) patients for liver transplantation is crucial to minimize the risk of recurrence and provide long-term outcomes comparable with those for other indications. Selection criteria based on total tumor volume (TTV) and α-fetoprotein (AFP) concentrations were proposed in a recent large study. The aim of this study was to evaluate the results of liver transplantation for HCC within and beyond these criteria. MATERIAL AND METHODS This retrospective study included 104 patients with HCC who underwent liver transplantation. Risk factors for overall survival and tumor recurrence were evaluated. Overall survival and cumulative tumor recurrence rate for patients with TTV <115 cm(3), AFP concentration <400 ng/mL, and no macrovascular invasion (76/104; 73.1%) were evaluated and compared with those for the remaining patients (28/104; 26.9%). RESULTS Pretransplantation AFP concentration >400 ng/mL (P = .016; hazard ratio [HR], 3.36; 95% confidence intervals [CI], 1.25-9.03) was the only risk factor for overall survival. TTV >115 cm(3) (P = .021; HR 4.29; 95% CI, 1.24-14.81) and AFP concentration >400 ng/mL (P = .002; HR 6.97; 95% CI, 2.02-24.03) were independent risk factors for recurrence. The estimated 3-year tumor recurrence rate was 4.2% for patients with TTV <115 cm(3), AFP concentration <400 ng/mL, and no macrovascular invasion compared with 57.2% for the remaining patients (P < .00001). The 3-year overall survival rate of patients within and beyond this criteria was 81.7% and 64.6%, respectively (P = .0628). CONCLUSIONS In contrast to other criteria, selection of HCC patients for liver transplantation on the basis of TTV and AFP concentration relates to both morphological features and tumor biology. Although fulfillment of these criteria was more than 1.5-fold higher than that of the Milan criteria, the rate of tumor recurrence was exceptionally low.

Poor Outcomes After Liver Transplantation in Patients With Incidental Cholangiocarcinoma Irrespective of Tumor Localization

INTRODUCTION After liver transplantation for cholangiocarcinoma (CCC), patients have a poor prognosis without use of specific therapeutic strategies. Accordingly, recipients with incidental CCC might have the highest risk of recurrent disease; however, sparse data on the long-term outcome of unselected patients with incidental CCC have been published. The aim of this study was to evaluate the post-transplantation outcomes of patients with incidental CCC with special focus on tumor localization. MATERIAL AND METHODS There were 11 primary liver transplantations in patients with incidental CCC of 1310 liver transplantation procedures performed between December 1994 and August 2013. All patients with incidental CCC received a chemotherapy regiment including gemcitabine/5 fluorouracil, doxorubicin, and mitomycin. The patients were switched from calcineurin inhibitors to mammalian target of rapamycin inhibitor-based immunosuppression shortly after CCC diagnosis. RESULTS Intra- and extrahepatic tumors were found in 6 and 5 patients, respectively. At median follow-up examination of 26.3 months there were 8 CCC recurrences and 7 patient deaths. Overall survival after liver transplantation for incidental CCC was 88.9% at 1 year, 44.4% at 2 years, and 14.8% at 3 years. The corresponding rates of recurrence-free survival were 45.7%, 45.7%, and 0.0%, respectively. Post-transplantation CCC recurrences were universal with 0% 3-year recurrence-free survival both in patients with intra- and extrahepatic tumors (P = .475). CONCLUSIONS Incidental CCC in liver transplantation is associated with poor outcomes irrespective of tumor localization. Introduction of new adjuvant multimodal treatment concepts is necessary to improve the prognosis for this subgroup of patients.