Cancer of the biliary confluence also known as hilar cholangiocarcinoma (HC) or Klatskin tumor, is a rare type of neoplastic disease constituting approximately 40%-60% of intrahepatic malignancies, and 2% of all cancers. The prognosis is extremely poor and the majority of Klatskin tumors are deemed unresectable upon diagnosis. Most patients with unresectable bile duct cancer die within the first year after diagnosis, due to hepatic failure, and/or infectious complications secondary to biliary obstruction. Curative treatments include surgical resection and liver transplantation in highly selected patients. Nevertheless, very few patients are eligible for surgery or transplant at the time of diagnosis. For patients with unresectable HC, radiotherapy, chemotherapy, photodynamic therapy, and liver-directed minimally invasive procedures such as percutaneous image-guided ablation and intra-arterial chemoembolization are recommended treatment options. This review focuses on currently available treatment options for unresectable HC and discusses future perspectives that could optimize outcomes.

NUC1031 is a first-in-class ProTide, that is a gemcitabine pro-drug designed to overcome putative mechanisms of resistance, including decreased expression of hENT/hCNT transporters, absence of activating enzymes such as deoxycytidine kinase (dCK) and presence of degrading enzymes such as cytidine deaminase (CDA). We undertook comprehensive pre-clinical evaluation of NUC1031 in biliary tract cancer (BTC) models, given that gemcitabine/cisplatin is a standard first-line therapy in advanced BTC.

Here, we compared the in vitro activity of NUC1031 in comparison to gemcitabine, validate putative mechanism(s) of action, assessed potential biomarkers of sensitivity or resistance, and performed combination studies with cisplatin. We also evaluated the in vivo efficacy of NUC1031 and gemcitabine using a CDA-high cholangiocarcinoma patient-derived xenograft (PDX) model.

In a panel of BTC cell lines (N = 10), NUC1031 had less potency than gemcitabine in multiple cellular assays. NUC1031 did not demonstrate evidence of greater synergy over gemcitabine in combination with cisplatin. Surprisingly, efficacy of both gemcitabine and NUC1031 was not found to be correlated with hENT/hCTN, dCK or CDA transcript levels. Gemcitabine and NUC1031 showed equivalent efficacy in a CDA-high PDX model in vivo contradicting the primary rationale of NUC1031 design.

NUC1031 did not exhibit evidence of superior activity over gemcitabine, as a single-agent, or in combination with cisplatin, in either our in vivo or in vitro BTC models. Given that the largest Phase 3 study (ClinicalTrials.gov: NCT0314666) to date in BTC is underway (N = 828) comparing NUC1031/cisplatin to gemcitabine/cisplatin, our results suggest that a more conservative clinical evaluation path would be more appropriate.

Oxidative stress and their effectors play critical roles in carcinogenesis and chemoresistance. However, the role of oxidative stress-related genes variants in biliary tract cancer (BTC) chemoresistance remains unknown. In this work, we aim to investigate oxidative stress-dependent molecular mechanisms underlying chemoresistance, and find potential biomarkers to predict chemotherapy response for BTC.

Sixty-six SNPs in 21 oxidative stress-related genes were genotyped and analyzed in 367 BTC patients. Immunoblot, immunohistochemical, immunofluorescent, quantitative PCR, chromatin immunoprecipitation analysis and study of animal xenograft models were performed to discover oxidative stress-related susceptibility genes underlying chemoresistance mechanism of BTC.

We found that 3 functional polymorphisms (CAT_rs769217, GPX4_rs4807542, and GSR_rs3779647), which were shown to affect their respective gene expression levels, modified the effect of chemotherapy on overall survival (OS). We then demonstrated that knockdown of GPX4, CAT, or GSR induced chemoresistance through elevation of ROS level and activation of Nrf2-ABCG2 pathway in BTC cell lines. Moreover, the association between Nrf2 expression and BTC prognosis is only found in patients who received chemotherapy. Knockdown of Nrf2 enhanced chemosensitivity or even eliminated postoperative recurrence in BTC xenograft mouse models. Importantly, upon chemotherapy treatment patients harboring high oxidative stress-related score received higher survival benefit from adjuvant chemotherapy compared with patients with low oxidative stress-related score.

The result of our study suggests, for the first time, that the oxidative stress-related score calculated by combining variations in CAT, GPX4, and GSR or Nrf2 expression could be used for predicting the chemosensitivity of BTC patients. FUND: This work was supported by the National Science Foundation of China, Foundation of Shanghai Shen Kang Hospital Development Center, and Shanghai Outstanding Academic Leaders Plan.

Gallbladder carcinoma (GBC) is the most aggressive gastrointestinal malignancy throughout the world, with wide geographical variance. It is the subtype of biliary tract malignancy that has the poorest prognosis and lower survival among all biliary tract malignancies. Various factors are associated with GBC pathogenesis such as environmental, microbial, metabolic and molecular. Chronic inflammation of gallbladder due to presence of gallstone or microbial infection (eg. Salmonella or H. pylori) results in sustained production of inflammatory mediators in the tissue microenvironment, which can cause genomic changes linked to carcinogenesis. Genetic alterations are one of the major factors, associated with aggressiveness and prognosis. Researches have been done to explore suitable biomarker for early diagnosis and identify altered molecular pathways to develop appropriate biomarkers for early diagnosis, therapy and predicting prognosis. Different agents for targeted therapy against actionable mutations of molecules like EGFR, VEGF, mTOR, HER2, PDL-1, PD-1, MET, PI3K, N-cadherin, VEGFR, MEK1 and MEK2 are being tried. Despite these advancements, there is dismal improvement in the survival of GBC patients. Genetic aberrations other than actionable mutations and epigenetic modification including aberrant expressions of micro-RNAs, are also being studied both as diagnostic biomarker and therapeutic targets. Complex pathogenesis of GBC still needs to be unfolded. In this review we focus on the molecular pathogenesis of GBC elucidated till date along with future directions that can be explored to achieve better management of GBC patients.

Patients with gallbladder carcinoma (GBC) lack effective treatment methods largely due to the inadequacy of both molecular characterisation and potential therapeutic targets. We previously uncovered a spectrum of genomic alterations and identified recurrent mutations in the ErbB pathway in GBC. Here, we aimed to study recurrent mutations of genes and pathways in a larger cohort of patients with GBC and investigate the potential mechanisms and clinical significance of these mutations.

We performed whole-exome sequencing (WES) in 157 patients with GBC. Functional experiments were applied in GBC cell lines to explore the oncogenic roles of ERBB2/ERBB3 hotspot mutations, their correlation with PD-L1 expression and the underlying mechanisms. ERBB inhibitors and a PD-L1 blocker were used to evaluate the anticancer activities in co-culture systems in vitro and in vivo.

WES identified ERBB2 and ERBB3 mutations at a frequency of 7%-8% in the expanded cohort, and patients with ERBB2/ERBB3 mutations exhibited poorer prognoses. A set of in vitro and in vivo experiments revealed increased proliferation/migration on ERBB2/ERBB3 mutation. Ectopic expression of ERBB2/ERBB3 mutants upregulated PD-L1 expression in GBC cells, effectively suppressed normal T-cell-mediated cytotoxicity in vitro through activation of the PI3K/Akt signalling pathway and contributed to the growth and progression of GBC in vivo. Treatment with an ERBB2/ERBB3 inhibitor or a PD-L1 monoclonal antibody reversed these immunosuppressive effects, and combined therapy revealed promising therapeutic activities.

ERBB2/ERBB3 mutations may serve as useful biomarkers in identifying patients who are sensitive to ERBB2/ERBB3 inhibitors and PD-L1 monoclonal antibody treatment.

NCT02442414;Pre-results.

Cholangiocarcinoma (CCA) may arise in the intra- or extrahepatic biliary tract and is associated with a poor prognosis. Despite recent advances, to date there is still no established targeted therapeutic approach available. Non-surgical therapeutic agents are urgently needed, as most patients are non-eligible to surgical resection. Anti-PD-L1 therapy prevents cancer cells from evading the immune system and has emerged as a new treatment option in several cancer entities. Recently, PD-L1 expression has been analyzed in comparably small CCA patient cohorts. However, a systematic validation of different PD-L1 antibodies has not been performed in CCA so far.

We stained a tissue microarray consisting of 170 patients, including 72 intrahepatic cholangiocarcinomas (iCCAs), 57 perihilar cholangiocarcinomas (pCCAs) and 41 distal cholangiocarcinomas (dCCAs) by immunohistochemistry and evaluated PD-L1 positivity in tumor and stromal cells. We analyzed three different PD-L1 antibodies (clones 28-8, SP142, and SP263) that are frequently used and recommended for predictive diagnostic testing in other cancer types.

For PD-L1 antibody clone SP263, 5% of iCCAs, 4% of pCCAs and 3% of dCCAs exhibited PD-L1 expression on tumor cells, thereby showing the highest frequencies of PD-L1 positivity. Accordingly, highest PD-L1 positivity rates of stromal cells with 31% in iCCA, 40% in pCCA and 61% in dCCA were detected for clone SP263. Agreement of PD-L1 positivity in tumor cells was moderate for clone 28-8 and SP263 (κ = 0.44) and poor between 28-8 and SP142 (κ = 0.13), as well as  SP142 and SP263 (κ = 0.11), respectively. Statistical analyses of PD-L1 expression (clone SP263) on tumor cells with clinicopathological data revealed a positive correlation with shortened overall survival in CCA patients.

Selection of appropriate PD-L1 antibodies and careful evaluation of immunohistochemical staining patterns have a significant impact on PD-L1 testing in CCA. Clinical trials are necessary to investigate the putative beneficial effects of PD-L1 targeted immunotherapy in CCA patients.

Advances in genomic sequencing and bioinformatics have led to the prospect of precision medicine where therapeutics can be advised by the genetic background of individuals. For example, mapping cancer genomics has revealed numerous genes that affect the therapeutic outcome of a drug. Through materials and cell engineering, many opportunities exist for engineers to contribute to precision medicine, such as engineering biosensors for diagnosis and health status monitoring, developing smart formulations for the controlled release of drugs, programming immune cells for targeted cancer therapy, differentiating pluripotent stem cells into desired lineages, fabricating bioscaffolds that support cell growth, or constructing "organs-on-chips" that can screen the effects of drugs. Collective engineering efforts will help transform precision medicine into a more personalized and effective healthcare approach. As continuous progress is made in engineering techniques, more tools will be available to fully realize precision medicine's potential.

Biliary tracts cancers (BTCs) are a diverse group of aggressive malignancies with an overall poor prognosis. Genomic characterization has uncovered many putative clinically actionable aberrations that can also facilitate the prognostication of patients. As such, comprehensive genomic profiling is playing a growing role in the clinical management of BTCs. Currently however, there is only one precision medicine approved by the US Food and Drug Administration (FDA) for the treatment of BTCs. Herein, we highlight the prevalence and prognostic, diagnostic, and predictive significance of recurrent mutations and other genomic aberrations with current clinical implications or emerging relevance to clinical practice. Some ongoing clinical trials, as well as future areas of exploration for precision oncology in BTCs are highlighted.

Gallbladder carcinoma is highly aggressive and resistant to chemotherapy, with no consistent strategy to guide first line chemotherapy. However, patient-derived xenograft (PDX) model has been increasingly used as an effective model for in preclinical study of chemosensitivity.

Mini-PDX model was established using freshly resected primary lesions from 12 patients with gallbladder to examine the sensitivity with five of the most commonly used chemotherapeutic agents, namely gemcitabine, oxaliplatin, 5-fluorouracil, nanoparticle albumin-bound (nab)-paclitaxel, and irinotecan. The results were used to guide the selection of chemotherapeutic agents for adjunctive treatment after the surgery. Kaplan-Meier method was used to compare overall survival (OS) and disease free survival (DFS) with 45 patients who received conventional chemotherapy with gemcitabine and oxaliplatin.

Cell viability assays based on mini-PDX model revealed significant heterogeneities in drug responsiveness. Kaplan-Meier analysis showed that patients in the PDX-guided chemotherapy group had significantly longer median OS (18.6 months; 95% CI 15.9-21.3 months) than patients in the conventional chemotherapy group (13.9 months; 95% CI 11.7-16.2 months) (P = 0.030; HR 3.18; 95% CI 1.47-6.91). Patients in the PDX-guided chemotherapy group also had significantly longer median DFS (17.6 months; 95% CI 14.5-20.6 months) than patients in the conventional chemotherapy group (12.0 months; 95% CI 9.7-14.4 months) (P = 0.014; HR 3.37; 95% CI 1.67-6.79).

The use of mini-PDX model to guide selection of chemotherapeutic regimens could improve the outcome in patients with gallbladder carcinoma.

Cholangiocarcinoma represents 10% of primary liver malignancies and accounts for less than 3% of all gastrointestinal malignant tumors, with an enormous geographical variation. This neoplasia can arise from the biliary tract epithelium or hepatic progenitor cells. Depending on the anatomic localization, it is classified into three subtypes: intrahepatic, perihilar and distal. This fact is one of the main difficulties, because there are many studies that indistinctly include the results in the management of these different types of cholangiocarcinoma, without differentiating its location and even including gallbladder cancer. There are many controversial points in epidemiology, liver transplantation as a treatment, limitations of different results by group and type of treatment, histological testing and chemotherapy. This is a narrative review about topics in cholangiocarcinoma. This article is part of a Special Issue entitled: Cholangiocytes in Health and Disease edited by Jesus Banales, Marco Marzioni, Nicholas LaRusso and Peter Jansen.

The utility of molecules derived from cancer cells as biomarkers of the pathological status in biliary tract and pancreatic cancers is still limited. Soluble LDL receptor relative with 11 ligand-binding repeats (sLR11), a molecule released from immature cells, has been shown to be a circulating biomarker for early stage hematological malignancies.

We have evaluated the pathological significance of bile sLR11 levels in 147 samples from 72 patients with biliary tract cancer (BTC), pancreatic cancer (PC), or benign diseases.

The bile sLR11 levels in the cancer patients were significantly increased compared with those in patients without cancer, independent of cytological detection of cancer cells in bile. The average bile sLR11 levels in cancer patients were significantly higher than in those with benign diseases, while levels of bile carbohydrate antigen 19-9 (CA19-9) and carcinoembryonic antigen (CEA) were not different. LR11 protein was found to be highly expressed in the BTC and PC cells. The LR11 transcript levels in cholangiocarcinoma and pancreatic cancer cell lines were sharply induced during proliferation and significantly increased under hypoxic conditions.

Therefore, sLR11 levels in bile may be indicative of cancer cell conditions and may serve as potential novel biomarker in patients with BTC and PC.