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Cholangiocarcinomas (CCAs) comprise a mucin-secreting form, intrahepatic or perihilar, and a mixed form located peripherally. We characterized cancer stem cells (CSCs) in CCA subtypes and evaluated their cancerogenic potential. CSC markers were investigated in 25 human CCAs in primary cultures and established cell lines. Tumorigenic potential was evaluated in vitro or in xenografted mice after s.c. or intrahepatic injection in normal and cirrhotic (carbon tetrachloride-induced) mice. CSCs comprised more than 30% of the tumor mass. Although the CSC profile was similar between mucin-intrahepatic and mucin-perihilar subtypes, CD13+ CSCs characterized mixed-intrahepatic, whereas LGR5+ characterized mucin-CCA subtypes. Many neoplastic cells expressed epithelial-mesenchymal transition markers and coexpressed mesenchymal and epithelial markers. In primary cultures, epithelial-mesenchymal transition markers, mesenchymal markers (vimentin, CD90), and CD13 largely predominated over epithelial markers (CD133, EpCAM, and LGR5). In vitro, CSCs expressing epithelial markers formed a higher number of spheroids than CD13+ or CD90+ CSCs. In s.c. tumor xenografts, tumors dominated by stromal markers were formed primarily by CD90+ and CD13+ cells. By contrast, in intrahepatic xenografts in cirrhotic livers, tumors were dominated by epithelial traits reproducing the original human CCAs. In conclusion, CSCs were rich in human CCAs, implicating CCAs as stem cell–based diseases. CSC subpopulations generate different types of cancers depending on the microenvironment. Remarkably, CSCs reproduce the original human CCAs when injected into cirrhotic livers.

Background: The role of liver transplant (LT) for neuroendocrine liver metastasis (NELM) has not been completely defined. While international guidelines included LT as a potential treatment for highly selected patients with advanced NELM, recently, LT has been proposed as an alternative curative treatment for NELM for patients meeting restrictive criteria (Milan criteria). Methods: Using a multi-institutional cohort of patients undergoing liver resection for NELM, the long-term outcomes of patients meeting Milan criteria (resected NET drained by the portal system, stable disease/response to therapies for at least 6 months, metastatic diffusion to < 50% of the total liver volume, a confirmed histology of low-grade, and ≤ 60 years) were investigated. Results: Among the 238 patients included in the study, 28 (12%) patients met the Milan criteria for LT with a 5-year OS of 83%. Furthermore, among patients meeting Milan criteria, subsets of patients with favorable clinic-pathological characteristics had 5-year OS rates greater than 90% including G1 patients (5-year OS, 92%), patients undergoing minor liver resection (5-year OS, 94%), patients with low number of NELM (1–2 NELM), and small tumor size (< 3 cm) (for both groups of patients, 5-year OS, 100%). Conclusions: In our series, only 12% of patients met Milan criteria, and the 5-year OS after liver resection for this small selected group of patients was comparable with that reported in the literature for patients undergoing LT for NELM within Milan criteria. While LT might be the optimal treatment for patients with unresectable NELM, surgical resection should be the first option for patients with resectable NELM.

Cholangiocarcinoma (CCA) and its subtypes (mucin- and mixed-CCA) arise from the neoplastic transformation of cholangiocytes, the epithelial cells lining the biliary tree. CCA has a high mortality rate owing to its aggressiveness, late diagnosis and high resistance to radiotherapy and chemotherapeutics. We have demonstrated that CCA is enriched for cancer stem cells which express epithelial to mesenchymal transition (EMT) traits, with these features being associated with aggressiveness and drug resistance. TGF-β signaling is upregu-lated in CCA and involved in EMT. We have recently established primary cell cultures from human mucin- and mixed-intrahepatic CCA. In human CCA primary cultures with different levels of EMT trait expression, we evaluated the anticancer effects of: (i) CX-4945, a casein kinase-2 (CK2) inhibitor that blocks TGF-β1-induced EMT; and (ii) LY2157299, a TGF-β receptor I kinase inhibitor. We tested primary cell lines expressing EMT trait markers (vimentin, N-cadherin and nuclear catenin) but negative for epithelial markers, and cell lines expressing epithelial markers (CK19-positive) in association with EMT traits. Cell viability was evaluated by MTS assays, apoptosis by Annexin V FITC and cell migration by wound-healing assay. Results: at a dose of 10 μM, CX4945 significantly decreased cell viability of primary human cell cultures from both mucin and mixed CCA, whereas in CK19-positive cell cultures, the effect of CX4945 on cell viability required higher concentrations (>30μM). At the same concentrations, CX4945 also induced apoptosis (3- fold increase vs controls) which correlated with the expression level of CK2 in the different CCA cell lines (mucin- and mixed-CCA). Indeed, no apoptotic effects were observed in CK19-positive cells expressing lower CK2 levels. The effects of CX4945 on viability and apoptosis were associated with an increased number of γ-H2ax (biomarker for DNA double-strand breaks) foci, suggesting the active role of CK2 as a repair mechanism in CCAs. LY2157299 failed to influence cell proliferation or apoptosis but significantly inhibited cell migration. At a 50 μM concentration, in fact, LY2157299 significantly impaired (at 24, 48 and 120 hrs) the wound-healing of primary cell cultures from both mucin-and mixed-CCA. In conclusion, we demonstrated that CX4945 and LY2157299 exert relevant but distinct anticancer effects against human CCA cells, with CX4945 acting on cell viability and apoptosis, and LY2157299 impairing cell migration. These results suggest that targeting the TGF-β signaling with a combination of CX-4945 and LY2157299 could have potential benefits in the treatment of human CCA.

Even though surgery remains the only potentially curative option for patients with neuroendocrine liver metastases, the factors determining a patient's prognosis following hepatectomy are poorly understood. Using a multicentric database including patients who underwent hepatectomy for NELMs at seven tertiary referral hepato-biliary-pancreatic centers between January 1990 and December 2014, we sought to identify the predictors of survival and develop a clinical tool to predict patient's prognosis after liver resection for NELMs. The median age of the 238 patients included in the study was 61.9 years (interquartile range 51.5-70.1) and 55.9 % (n = 133) of patients were men. The number of NELMs (hazard ratio = 1.05), tumor size (HR = 1.01), and Ki-67 index (HR = 1.07) were the predictors of overall survival. These variables were used to develop a nomogram able to predict survival. According to the predicted 5-year OS, patients were divided into three different risk classes: 19.3, 55.5, and 25.2 % of patients were in low (> 80 % predicted 5-year OS), medium (40-80 % predicted 5-year OS), and high (< 40 % predicted 5-year OS) risk classes. The 10-year OS was 97.0, 55.9, and 20.0 % in the low, medium, and high-risk classes, respectively (p < 0.001). We developed a novel nomogram that accurately (c-index > 70 %) staged and predicted the prognosis of patients undergoing liver resection for NELMs.

Aim To compare clinical presentation, operative management and short- and long-term outcomes of congenital bile duct cysts (BDC) in adults with children. Methods Retrospective multi-institutional Association Francaise de Chirurgie study of Todani types I+IVB and IVA BDC. Results During the 37-year period to 2011, 33 centers included 314 patients (98 children; 216 adults). The adult population included more high-risk patients, with more active, more frequent prior treatment (47.7% vs 11.2%; p < 0.0001), more complicated presentation (50.5% vs 35.7%; p = 0.015), more synchronous biliary cancer (11.6% vs 0%; p = 0.0118) and more major surgery (23.6% vs 2%; p < 0.0001), but this latter feature was only true for type I+IVB BDC. Compared to children, the postoperative morbidity (48.1% vs 20.4%; p < 0.0001), the need for repeat procedures and the status at follow-up were worse in adults (27% vs 8.8%; p = 0.0009). However, severe postoperative morbidity and fair or poor status at follow-up were not statistically different for type IVA BDC, irrespective of patients' age. Synchronous cancer, prior HBP surgery and Todani type IVA BDC were independent predictive factors of poor or fair long-term outcome. Conclusion BDC is a more indolent disease in children compared to adults, except for Todani type IV-A BDC.