M. Colledan

A prospective randomised, open-labeled, trial comparing sirolimus-containing versus mTOR-inhibitor-free immunosuppression in patients undergoing liver transplantation for hepatocellular carcinoma

BackgroundThe potential anti-cancer effects of mammalian target of rapamycin (mTOR) inhibitors are being intensively studied. To date, however, few randomised clinical trials (RCT) have been performed to demonstrate anti-neoplastic effects in the pure oncology setting, and at present, no oncology endpoint-directed RCT has been reported in the high-malignancy risk population of immunosuppressed transplant recipients. Interestingly, since mTOR inhibitors have both immunosuppressive and anti-cancer effects, they have the potential to simultaneously protect against immunologic graft loss and tumour development. Therefore, we designed a prospective RCT to determine if the mTOR inhibitor sirolimus can improve hepatocellular carcinoma (HCC)-free patient survival in liver transplant (LT) recipients with a pre-transplant diagnosis of HCC.Methods/DesignThe study is an open-labelled, randomised, RCT comparing sirolimus-containing versus mTOR-inhibitor-free immunosuppression in patients undergoing LT for HCC. Patients with a histologically confirmed HCC diagnosis are randomised into 2 groups within 4-6 weeks after LT; one arm is maintained on a centre-specific mTOR-inhibitor-free immunosuppressive protocol and the second arm is maintained on a centre-specific mTOR-inhibitor-free immunosuppressive protocol for the first 4-6 weeks, at which time sirolimus is initiated. A 21/2 -year recruitment phase is planned with a 5-year follow-up, testing HCC-free survival as the primary endpoint. Our hypothesis is that sirolimus use in the second arm of the study will improve HCC-free survival. The study is a non-commercial investigator-initiated trial (IIT) sponsored by the University Hospital Regensburg and is endorsed by the European Liver and Intestine Transplant Association; 13 countries within Europe, Canada and Australia are participating.DiscussionIf our hypothesis is correct that mTOR inhibition can reduce HCC tumour growth while simultaneously providing immunosuppression to protect the liver allograft from rejection, patients should experience less post-transplant problems with HCC recurrence, and therefore could expect a longer and better quality of life. A positive outcome will likely change the standard of posttransplant immunosuppressive care for LT patients with HCC.Trial RegisterTrial registered at http://www.clinicaltrials.gov: NCT00355862(EudraCT Number: 2005-005362-36)

Hemolytic uremic syndrome: a fatal outcome after kidney and liver transplantation performed to correct factor h gene mutation.

Factor H‐associated hemolytic uremic syndrome (HUS) is a genetic form of thrombotic microangiopathy characterized by deficient factor H (HF‐1) levels/activity and uncontrolled complement activation. The disorder mostly leads to end‐stage renal disease and often recurs after kidney transplantation. We previously demonstrated that in a child with HF‐1‐associated HUS a simultaneous kidney and liver transplantation restored the defective HF‐1 with no recurrence of the disease in the transplanted kidney.

Patterns of hepatitis delta virus reinfection and disease in liver transplantation.

Twenty-seven carriers of the hepatitis B surface antigen who underwent liver transplantation in Italy and Belgium for terminal Hepatitis delta virus (HDV) cirrhosis were investigated. In 22 of the patients, HDV infection recurred. Two patients died of coexisting HDV and hepatitis B virus (HBV) reactivation. Four patients who died of unrelated causes were found to have HDV without signs of HBV reactivation. Five patients (18%) cleared both HBV and HDV after transplantation with no evidence of hepatitis (mean follow-up, 29 months). In many surviving patients. HDV infection recurred early without signs of HBV reactivation. Disease returned in the 11 HDV-infected patients in whom HBV also recurred. Histological hepatitis did not recur during an interim of 12-33 months in the 5 HDV-infected patients in whom HBV did not return. The overall medium-term survival in patients with HDV who underwent transplantation was 77.7%. Liver transplantation offers patients with HDV a hope of cure from disease despite a high risk of reinfection. In the transplantation setting. HDV can cause subclinical infections without any apparent assistance from HBV; these infections become symptomatic only if and when HBV reactivates. Thus, HDV may not be in itself pathogenic but requires cooperation from HBV to cause the appearance of the disease.

Split-liver transplantation eliminates the need for living-donor liver transplantation in children with end-stage cholestatic liver disease.

Background. End-stage cholestatic liver disease (ESCLD) is the main indication for liver replacement in children. Pediatric cadaver–organ-donor shortage has prompted the most important evolutions in the technique of liver transplantation, in particular living-donor liver transplantation (LDLT) and split-liver transplantation (SLT). Methods. Between November 1997 and June 2001, 127 children with ESCLD were evaluated for liver transplantation, and 124 underwent 138 liver transplantations after a median time of 40 days. Causes of liver disease were congenital biliary atresia (n=96), Alagille’s syndrome (n=12), Byler’s disease (n=8), and other cholestatic diseases (n=8). Results. Ninety (73%) patients received a split-liver graft, 28 (23%) a whole liver, and 6 (4%) a reduced-size liver. Overall 2- and 4-year patient survival rates were 93% and 91%, respectively; the 2- and 4-year graft-survival rates were 84% and 80%, respectively. In split-liver recipients, 4-year patient and graft-survival rates were 91% and 83%, respectively; these were 93% and 78%, respectively, in whole-liver recipients and 67% and 63%, respectively, in reduced-size liver recipients. Retransplantation rate was 11%, whereas mortality rate was 8%. Overall incidence of vascular and biliary complication were 16% and 27%, respectively. Conclusions. SLT can provide liver grafts for children with ESCLD with an outcome similar to the one reported following LDLT, eliminating mortality while they are on a transplantation wait list. The need for pediatric LDLT should be reevaluated and programs of SLT strongly encouraged and supported at a national and international level.

Sirolimus Use in Liver Transplant Recipients With Hepatocellular Carcinoma: A Randomized, Multicenter, Open-Label Phase 3 Trial.

Background We investigated whether sirolimus-based immunosuppression improves outcomes in liver transplantation (LTx) candidates with hepatocellular carcinoma (HCC). Methods In a prospective-randomized open-label international trial, 525 LTx recipients with HCC initially receiving mammalian target of rapamycin inhibitor–free immunosuppression were randomized 4 to 6 weeks after transplantation into a group on mammalian target of rapamycin inhibitor–free immunosuppression (group A: 264 patients) or a group incorporating sirolimus (group B: 261). The primary endpoint was recurrence-free survival (RFS); intention-to-treat (ITT) analysis was conducted after 8 years. Overall survival (OS) was a secondary endpoint. Results Recurrence-free survival was 64.5% in group A and 70.2% in group B at study end, this difference was not significant (P = 0.28; hazard ratio [HR], 0.84; 95% confidence interval [95% CI], 0.62; 1.15). In a planned analysis of RFS rates at yearly intervals, group B showed better outcomes 3 years after transplantation (HR, 0.7; 95% CI, 0.48-1.00). Similarly, OS (P = 0.21; HR, 0.81; 95% CI, 0.58-1.13) was not statistically better in group B at study end, but yearly analyses showed improvement out to 5 years (HR, 0.7; 95% CI, 0.49-1.00). Interestingly, subgroup (Milan Criteria-based) analyses revealed that low-risk, rather than high-risk, patients benefited most from sirolimus; furthermore, younger recipients (age ⩽60) also benefited, as well sirolimus monotherapy patients. Serious adverse event numbers were alike in groups A (860) and B (874). Conclusions Sirolimus in LTx recipients with HCC does not improve long-term RFS beyond 5 years. However, a RFS and OS benefit is evident in the first 3 to 5 years, especially in low-risk patients. This trial provides the first high-level evidence base for selecting immunosuppression in LTx recipients with HCC.

Split and whole liver transplantation outcomes: A comparative cohort study

A specific split liver transplantation (SLT) program has been pursued in the North Italian Transplant program (NITp) since November 1997. After 5 yr, 1,449 liver transplants were performed in 7 transplant centers, using 1,304 cadaveric donors. Whole liver transplantation (WLT) and SLT were performed in 1,126 and 323 cases, respectively. SLTs were performed in situ as 147 left lateral segments (LLS), 154 right trisegment liver (RTL) grafts, and 22 modified split livers (MSL), used for couples of adult recipients. After a median posttransplant follow‐up of 22 months, SLTs achieved a 3‐yr patient and graft survival not significantly different from the entire series of transplants (79.4 and 72.2% vs. 80.6 and 74.9%, respectively). Recipients receiving a WLT or a LLS showed significantly better outcomes than patients receiving RTL and MSL (P < 0.03 for patients and P < 0.04 for graft survival). At the multivariate analysis, donor age of >60 yr, RTL transplant, <50 annual transplants volume, urgent transplantation (United Network for Organ Sharing (UNOS) status I and IIA), ischemia time of >7 hours, and retransplantation were factors independently related to graft failure and to significantly worst patient survival. Right grafts procured from RTL and either split procured as MSL had a similar outcome of marginal whole livers. In conclusion, in 5 yr, the increased number of pediatric transplants due to split liver donation reduced to 3% the in‐list children mortality, and a decrease in the adult patient dropout rate from 27.2 to 16.2% was observed. Such results justify a more widespread adoption of SLT protocols, organizational difficulties not being a limit for the application of such technique. Liver Transpl 12:402–410, 2006.

A new splitting technique for liver grafts

The in-situ split-liver (ISSL) technique allows the division of the liver of a cadaver donor in two parts that can be transplanted in two different patients. The two grafts obtained are different in size, the left one being generally suitable for transplantation only in small children. We describe here the successful use of an alternative technique, generating two grafts more similar in size, both of which are transplantable into adults or large children. The donor was a brain-dead 33-year-old man 164 cm tall and weighing 60 kg. The multiple organ harvesting was done with standard technique apart from division of the liver. After a cholecistectomy the right and left portal and hepatic arterial branches were identified. The left hepatic duct was sectioned. The right lobe was then mobilised and freed from the inferior vena cava (IVC); the right vein was encircled at its confluence with the IVC. Parenchyma transection was then done along a plane directed from the right border of the gallbladder fossa, to the division of the portal vein and to the right margin of the median hepatic vein, leaving the right lobe connected only with its vascular attachments (figure). After flushing of the organs the heart was retrieved, then came the right lobe of the liver, including the five to eight segments, the right branches of the hepatic artery and of the portal vein, the common bile duct, and the right hepatic vein with a patch of IVC. The left lobe of the liver, including the segments one to four was then removed, with the coeliac axis and the hepatic artery, the portal trunk, the left hepatic duct, and the median and left hepatic veins in continuity with the IVC. Finally, the kidneys were removed. During the 4 h operation, seven units of packed red blood cells were transfused. The right and left grafts weighed 685 g and 480 g, respectively, and were transplanted in a 53-year-old man, 178 cm tall and weighing 79 kg, with alcoholic cirrhosis, and in a 13-year-old girl 155 cm tall and weighed 48 kg, with autoimmune cirrhosis and hepatitis C virus infection. Both the grafts had immediate good function and the patients had rapid recovery. They were discharged on the 15th and 24th postoperative day, respectively, and are alive and well with normal liver function at 5 months. This alternative technique of ISSL provided two grafts of similar size, showing excellent function in two recipients of adult size. The blood loss during the split was similar to that reported from harvesting a right-lobe graft from a living donor, with a similar technique. With increasing experience this figure can be reduced. ISSL has proved a safe and effective way to increase the number of liver transplants with the available donor pool, dividing the liver mass between the large right lobe and the small left lateral segment. The minimum amount of liver tissue needed to achieve sufficient immediate function in a given patient is not well known, but should probably not be lower then 25–50% of his predicted liver volume. The left lateral segment seldon exceeds 300 g in weight, being therefore inadequate for most adults and large children. The net benefit from standard ISSL is therefore only for small children, representing a limited percentage of candidates for liver transplantation. The extension of ISSL to increase the number of adult transplants can be a further step in optimising the use of cadaver donors. Our technique increases the flexibility of the procedure and can virtually double the pool of liver grafts for adult recipients.

Balancing Donor and Recipient Risk Factors in Liver Transplantation: The Value of D‐MELD With Particular Reference to HCV Recipients

Donor–recipient match is a matter of debate in liver transplantation. D‐MELD (donor age × recipient biochemical model for end‐stage liver disease [MELD]) and other factors were analyzed on a national Italian database recording 5946 liver transplants. Primary endpoint was to determine factors predictive of 3‐year patient survival. D‐MELD cutoff predictive of 5‐year patient survival <50% (5yrsPS<50%) was investigated. A prognosis calculator was implemented (http://www.D‐MELD.com). Differences among D‐MELD deciles allowed their regrouping into three D‐MELD classes (A < 338, B 338–1628, C >1628). At 3 years, the odds ratio (OR) for death was 2.03 (95% confidence interval [CI], 1.44–2.85) in D‐MELD class C versus B. The OR was 0.40 (95% CI, 0.24–0.66) in class A versus class B. Other predictors were hepatitis C virus (HCV; OR = 1.42; 95% CI, 1.11–1.81), hepatitis B virus (HBV; OR = 0.69; 95% CI, 0.51–0.93), retransplant (OR = 1.82; 95% CI, 1.16–2.87) and low‐volume center (OR = 1.48; 95% CI, 1.11–1.99). Cox regressions up to 90 months confirmed results. The hazard ratio was 1.97 (95% CI, 1.59–2.43) for D‐MELD class C versus class B and 0.42 (95% CI, 0.29–0.60) for D‐MELD class A versus class B. Recipient age, HCV, HBV and retransplant were also significant. The 5yrsPS<50% cutoff was identified only in HCV patients (D‐MELD ≥ 1750). The innovative approach offered by D‐MELD and covariates is helpful in predicting outcome after liver transplantation, especially in HCV recipients.

Liver Match, a prospective observational cohort study on liver transplantation in Italy: Study design and current practice of donor-recipient matching

BACKGROUND The Liver Match is an observational cohort study that prospectively enrolled liver transplantations performed at 20 out of 21 Italian Transplant Centres between June 2007 and May 2009. Aim of the study is to investigate the impact of donor/recipient matching on outcomes. In this report we describe the study methodology and provide a cross-sectional description of donor and recipient characteristics and of graft allocation. METHODS Adult primary transplants performed with deceased heart-beating donors were included. Relevant information on donors and recipients, organ procurement and allocation were prospectively entered in an ad hoc database within the National Transplant Centre web-based Network. Data were blindly analysed by an independent Biostatistical Board. RESULTS The study enrolled 1530 donor/recipient matches. Median donor age was 56 years. Female donors (n = 681, median 58, range 12-92 years) were older than males (n = 849, median 53, range 2-97 years, p < 0.0001). Donors older than 60 years were 42.2%, including 4.2% octogenarians. Brain death was due to non-traumatic causes in 1126 (73.6%) cases. Half of the donor population was overweight, 10.1% was obese and 7.6% diabetic. Hepatitis B core antibody (HBcAb) was present in 245 (16.0%) donors. The median Donor Risk Index (DRI) was 1.57 (>1.7 in 35.8%). The median cold ischaemia time was 7.3h (≥ 10 in 10.6%). Median age of recipients was 54 years, and 77.7% were males. Hepatocellular carcinoma (HCC) was the most frequent indication overall (44.4%), being a coindication in roughly 1/3 of cases, followed by viral cirrhosis without HCC (28.2%) and alcoholic cirrhosis without HCC (10.2%). Hepatitis C virus infection (with or without HCC) was the most frequent etiologic factor (45.9% of the whole population and 71.4% of viral-related cirrhosis), yet hepatitis B virus infection accounted for 28.6% of viral-related cirrhosis, and HBcAb positivity was found in 49.7% of recipients. The median Model for End Stage Liver Disease (MELD) at transplant was 12 in patients with HCC and 18 in those without. Multivariate analysis showed a slight but significant inverse association between DRI and MELD at transplant. CONCLUSIONS The deceased donor population in Italy has a high-risk profile compared to other countries, mainly due to older donor age. Almost half of the grafts are transplanted in recipients with HCC. Higher risk donors tend to be preferentially allocated to recipients with HCC, who are usually less ill and older. No other relevant allocation strategy is currently adopted at national level.

Platelet‐derived growth factor‐D and Rho GTPases regulate recruitment of cancer‐associated fibroblasts in cholangiocarcinoma

Cholangiocarcinoma (CCA) is characterized by an abundant stromal reaction. Cancer‐associated fibroblasts (CAFs) are pivotal in tumor growth and invasiveness and represent a potential therapeutic target. To understand the mechanisms leading to CAF recruitment in CCA, we studied (1) expression of epithelial‐mesenchymal transition (EMT) in surgical CCA specimens and CCA cells, (2) lineage tracking of an enhanced green fluorescent protein (EGFP)‐expressing human male CCA cell line (EGI‐1) after xenotransplantation into severe‐combined‐immunodeficient mice, (3) expression of platelet‐derived growth factors (PDGFs) and their receptors in vivo and in vitro, (4) secretion of PDGFs by CCA cells, (5) the role of PDGF‐D in fibroblast recruitment in vitro, and (6) downstream effectors of PDGF‐D signaling. CCA cells expressed several EMT biomarkers, but not alpha smooth muscle actin (α‐SMA). Xenotransplanted CCA masses were surrounded and infiltrated by α‐SMA‐expressing CAFs, which were negative for EGFP and the human Y‐probe, but positive for the murine Y‐probe. CCA cells were strongly immunoreactive for PDGF‐A and ‐D, whereas CAFs expressed PDGF receptor (PDGFR)β. PDGF‐D, a PDGFRβ agonist, was exclusively secreted by cultured CCA cells. Fibroblast migration was potently induced by PDGF‐D and CCA conditioned medium and was significantly inhibited by PDGFRβ blockade with Imatinib and by silencing PDGF‐D expression in CCA cells. In fibroblasts, PDGF‐D activated the Rac1 and Cdc42 Rho GTPases and c‐Jun N‐terminal kinase (JNK). Selective inhibition of Rho GTPases (particularly Rac1) and of JNK strongly reduced PDGF‐D‐induced fibroblast migration. Conclusion: CCA cells express several mesenchymal markers, but do not transdifferentiate into CAFs. Instead, CCA cells recruit CAFs by secreting PDGF‐D, which stimulates fibroblast migration through PDGFRβ and Rho GTPase and JNK activation. Targeting tumor or stroma interactions with inhibitors of the PDGF‐D pathway may offer a novel therapeutic approach. (Hepatology 2013;53:1042–1053)