The effect of biliary obstruction, biliary drainage and bile reinfusion on bile acid metabolism and gut microbiota in mice.

BACKGROUND & AIMS: Preoperative obstructive jaundice is usually associated with higher post-operative mortality. Although external biliary drainage (EBD) has been widely used to relieve obstructive jaundice, the role of bile reinfusion after EBD is still controversial. The aim of our study was to study the effects of biliary obstruction, biliary drainage and bile reinfusion on bile acid metabolism and gut microbiota. METHODS: Firstly, we created a mice bile drainage collection (BDC) model to simulate the process of biliary obstruction, drainage and bile reinfusion. Then, we analysed the faecal, serum, liver and bile samples to investigate the effects of the process on bile acid profiles and gut microbiota. Finally, we evaluated the clinical effects of bile reinfusion. RESULTS: We evaluated the bile acid profiles of faeces, serum, liver and bile of normal mice. During biliary obstruction, secondary bile acids can still be produced, and increased in the liver and serum of mice. Compared with no bile reinfusion, bile reinfusion was beneficial to the recovery of T-ωMCA in the liver and bile, and can restore the colon crypt length shortened by biliary obstruction. Only Ruminococcus_1 proliferated when the biliary obstruction lasted for 12 days. In the clinic, bile reinfusion cannot accelerate the patient's perioperative recovery or prolong long-term survival. CONCLUSION: We have successfully created a mice bile drainage collection model. Short-term bile reinfusion can partially benefit the recovery of the secondary bile acids in the liver and bile, but hardly benefit the patient's perioperative recovery or long-term survival. (247 words).

Comprehensive analysis of genomic alterations of Chinese hilar cholangiocarcinoma patients.

BACKGROUND: Cholangiocarcinoma (CCA) is a rare malignant tumor of the biliary system. The heterogeneity of CCA leads to the lack of effective targeted treatment for CCA subtypes. The molecular characteristic of hilar CCA (hCCA) is still unclear. METHODS: A total of 63 hCCA patients were enrolled from Shanghai Eastern Hepatobiliary Surgery Hospital. Formalin-fixed, paraffin-embedded tumor tissues, and matched blood were collected and deep sequencing targeting 450 cancer genes were performed. Tumor mutation burden (TMB) was measured by an algorithm developed in-house. Correlation analysis was performed by Fisher's exact test. RESULTS: The most commonly mutated genes were TP53 (51.7%), NF1 and KRAS (20%, for both), SMAD4 (16.7%), FAT3 and FRS2 (13.3%, for both), NF1 (11.7%), and KMT2C, MDM2, and ATM (10%, for each) in hCCA. ARID1A, GATA6, and PREX2 mutations commonly occurred in female and KMT2C mutations mainly occurred in patients under 60 years old. Statistical analysis showed the association between ARID1A mutation and tumor stage (P = 0.041) and between NF1 mutation and high TMB (P = 0.0095). Furthermore, ARID1B mutation was identified to associate with the poor prognosis of Chinese hCCA patients (P = 0.004). CONCLUSION: The mutational characterization of hCCA is different from both extrahepatic CCA and intrahepatic CCA. ARID1B is a potential biomarker for prognosis prediction of Chinese hCCA patients.

Transarterial chemoembolization versus percutaneous microwave coagulation therapy for recurrent unresectable intrahepatic cholangiocarcinoma: Development of a prognostic nomogram.

BACKGROUND: Transarterial chemoembolization (TACE) and percutaneous microwave coagulation therapy (PMCT) are commonly used to treat intrahepatic recurrent liver cancers. However, there is no information regarding their effectiveness in patients with recurrent intrahepatic cholangiocarcinoma (ICC) after resection. METHODS: A total of 275 patients with localized recurrent ICC who received either TACE (n = 183) or PMCT (n = 92) were studied. A propensity score matching analysis was performed to compare prognostic impact of TACE and PMCT. Prognostic factors for TACE and PMCT were identified respectively. Predictive nomograms for each TACE and PMCT were developed using the Cox independent prognostic factors and were validated in independent patient groups by receiver operating characteristic curves and area under curve values. RESULTS: Both TACE and PMCT provided curativeness in partial patients (5-year overall survival: 21.4% and 6.1%, respectively), but TACE provided better survival benefit in both overall patients (hazard ratio [HR] = 0.71; 95% confidence interval [CI]: 0.50-0.97; P = 0.034) and propensity score matching analysis (HR = 0.69; 95% CI: 0.47-0.98; P = 0.041). Independent prognostic factors for TACE were tumor size >5 cm, poor differentiation, and major resection, whereas poor differentiation, hepatitis B virus infection, cholelithiasis, and lymph node metastasis were identified for PMCT. Both predictive nomograms for TACE and PMCT were validated to be effective with area under curve values of 0.77 and 0.70, respectively. CONCLUSIONS: TACE provided better survival benefits compared to PMCT. However, there was a disparity in prognostic factors, suggesting evaluation of the two nomograms may be supportive in modality selection. Further prospective validation studies are required for the results to be applied in clinical medicine.

Risk stratification system to predict recurrence of intrahepatic cholangiocarcinoma after hepatic resection.

BACKGROUND: Previous nomograms for intrahepatic cholangiocarcinoma (ICC) were conducted to predict overall survival, which could be influenced by various factors. Herein, we conducted our nomogram to predict recurrence of the tumor only after hepatic resection. METHODS: The nomogram was established with prognostic factors for the relapse-free survival (RFS) analyzed from our single center cohort and was evaluated by comparing with the American Joint Committee on Cancer (AJCC) staging system for the predictive accuracy. RESULTS: Seropositivity of hepatitis B surface antigen (hazard ratio [HR], 0.505; 95% confidence interval [CI], 0.279 to 0.914; P = 0.024), tumor size of larger than 5 cm (HR, 1.947; 95% CI, 1.177 to 3.219; P = 0.009), Child-Pugh score of B (HR, 3.067; 95% CI, 1.293 to 7.275; P = 0.011), and lymph node metastasis (HR, 2.790; 95% CI, 1.628 to 4.781; P < 0.001) were found to be independent prognostic factors that significantly affected RFS. The calibration curve for the prediction revealed excellent agreement between estimation by our stratification system and actual RFS. The concordance C index of the nomogram (0.71; 95% CI, 0.65 to 0.77) revealed to be significantly higher than the AJCC staging system (0.66; 95% CI, 0.60 to 0.72). In the validation cohort, our risk stratification system (C-index 0.65; 95% CI, 0.59 to 0.71) also revealed more precise prediction than the AJCC staging system (C-index, 0.57; 95% CI, 0.50 to 0.64). CONCLUSIONS: Our nomogram could more accurately predict recurrence of ICC after hepatic resection than the AJCC staging system.

Development and validation of a nomogram to predict overall survival in patients with incidental gallbladder cancer: A retrospective cohort study.

Objective: The aim of this study was to develop and validate a nomogram to predict the overall survival of incidental gallbladder cancer. Methods: A total of 383 eligible patients with incidental gallbladder cancer diagnosed in Shanghai Eastern Hepatobiliary Surgery Hospital from 2011 to 2021 were retrospectively included. They were randomly divided into a training cohort (70%) and a validation cohort (30%). Univariate and multivariate analyses and the Akaike information criterion were used to identify variables independently associated with overall survival. A Cox proportional hazards model was used to construct the nomogram. The C-index, area under time-dependent receiver operating characteristic curves and calibration curves were used to evaluate the discrimination and calibration of the nomogram. Results: T stage, N metastasis, peritoneal metastasis, reresection and histology were independent prognostic factors for overall survival. Based on these predictors, a nomogram was successfully established. The C-index of the nomogram in the training cohort and validation cohort was 0.76 and 0.814, respectively. The AUCs of the nomogram in the training cohort were 0.8, 0.819 and 0.815 for predicting OS at 1, 3 and 5 years, respectively, while the AUCs of the nomogram in the validation cohort were 0.846, 0.845 and 0.902 for predicting OS at 1, 3 and 5 years, respectively. Compared with the 8th AJCC staging system, the AUCs of the nomogram in the present study showed a better discriminative ability. Calibration curves for the training and validation cohorts showed excellent agreement between the predicted and observed outcomes at 1, 3 and 5 years. Conclusions: The nomogram in this study showed excellent discrimination and calibration in predicting overall survival in patients with incidental gallbladder cancer. It is useful for physicians to obtain accurate long-term survival information and to help them make optimal treatment and follow-up decisions.

Single-cell atlas of diverse immune populations in the advanced biliary tract cancer microenvironment.

Immunotherapies have been explored in treating solid tumors, albeit with disparate clinical effects in distinct cancer types. Systematic interrogation of immune cells in the tumor microenvironment (TME) is vital to the prediction of immunotherapy response and the development of innovative immunotherapeutics. To comprehensively characterize the immune microenvironment in advanced biliary tract cancer (BTC), we utilized single-cell RNA sequencing in unselected viable cells from 16 matched samples, and identified nineteen cell subsets from a total of 45,851 cells, in which exhausted CD8+ T cells, macrophages, and dendritic cells (DCs) in BTC were shown to augment and communicate within the TME. Transcriptional profiles coupled with T cell receptor (TCR) sequences revealed that exhausted CD8+ T cells retained clonal expansion and high proliferation in the TME, and some of them highly expressed the endoplasmic reticulum stress (ER) response gene, XBP1, indicating the role of ER stress in remodeling TME. Functional assays demonstrated that XBP1 and common immune checkpoints (PD1, TIGIT) were significantly upregulated in CD8+ T cells cocultured within the TME of BTC cells (GBC-SD, HCCC-9810). When treating the coculture groups with the specific inhibitor of IRE1α-XBP1 (4µ8C), the downregulation of TIGIT was observed in the treatment group. Collectively, comprehensive transcriptome profiling provides deep insights into the immune atlas in advanced BTC, which might be instrumental in exploring innovative immunotherapy strategies.

Cytoreductive surgery combined with hyperthermic intraperitoneal chemotherapy vs. cytoreductive surgery alone for intrahepatic cholangiocarcinoma with peritoneal metastases: A retrospective cohort study.

BACKGROUND: Cytoreductive surgery (CRS) combined with hyperthermic intraperitoneal chemotherapy (HIPEC) has survival benefits in patients with intraperitoneal malignant lesions, but there is no study specific to intrahepatic cholangiocarcinoma (ICC). PURPOSE: To compare the prognosis of patients with advanced ICC undergoing CRS + HIPEC compared with CRS alone. METHODS: This study was a retrospective cohort study of patients with advanced ICC treated at the Shanghai Eastern Hepatobiliary Surgery Hospital between 01/2014 and 12/2018. The patients were divided into either CRS + HIPEC or CRS group based on the treatment they received. Overall survival (OS), complications, hospital stay, biochemical indicators, tumor markers, and number of HIPEC were examined. RESULTS: There were 51 and 61 patients in the CRS + HIPEC and CRS groups, respectively. There were no differences between the groups regarding preoperative CA19-9 levels (421 ± 381 vs. 523 ± 543 U/mL, P = 0.208). The hospital stay was longer in the CRS + HIPEC group (22.2 ± 10.0 vs. 18.6 ± 7.6 days, P = 0.033). The occurrence of overall complications was similar in the two groups (37.2% vs. 34.4%, P = 0.756). The postoperative CA19-9 levels were lower in the CRS + HIPEC group compared with the CRS group (196 ± 320 vs. 337 ± 396 U/mL, P = 0.044). The median OS was longer in the CRS + HIPEC group than in the CRS group (25.53 vs. 11.17 months, P < 0.001). Compared with the CRS group, the CRS + HIPEC group showed a higher occurrence of leukopenia (7.8% vs. 0, P = 0.040) but a lower occurrence of total bilirubin elevation (15.7% vs. 37.7%, P = 0.032). CONCLUSION: CRS + HIPEC could be a treatment option for patients with advanced ICC, with improved OS and similar complications and adverse events compared with CRS alone.

A combined Cox and logistic model provides accurate predictive performance in estimation of time-dependent probabilities for recurrence of intrahepatic cholangiocarcinoma after resection.

BACKGROUND: Intrahepatic cholangiocarcinoma has heterogeneous outcomes after resection. There remains a need for broadly applicable recurrence-specific tool offering precise evaluation on curativeness of resection. METHODS: A four hospital-based clinical cohort involving 1,655 patients with intrahepatic cholangiocarcinoma who received surgical resection were studied. Cox and logistic models were networked into one system containing risk categories with distinctive probabilities of recurrence. Prediction of time-to-recurrence was performed by formulizing time-dependent risk probabilities. The model was validated in three clinical cohorts (n=332). RESULTS: From the training cohort, 10 and 11 covariates, including diabetes, cholelithiasis, albumin, platelet count, alpha fetoprotein, carbohydrate antigen 19-9, carcinoembryonic antigen, hepatitis B virus infection, tumor size and number, resection type, and lymph node metastasis, from Cox and logistic models were identified significant for recurrence-free survival (RFS). The combined Cox & logistic ranking system (CCLRS)-adjusted time-dependent probabilities were categorized into seven ranks (5-yr RFS for lowest and highest ranks were 75% vs. 0%; hazard ratio 18.5, 95% CI: 14.7-24.9, P<0.0001). The CCLRS was validated with a minimum area under curve value of 0.8086. Prediction of time-to-recurrence was validated to be excellent (Pearson r, 0.8204; P<0.0001). CONCLUSIONS: The CCLRS allows precise estimation on risk of recurrence for intrahepatic cholangiocarcinoma after resection. It could be applicative when estimating time-dependent disease status and stratifying individuals who sole resection of the tumor would not be curative.

Establishment and characterization of 38 novel patient-derived primary cancer cell lines using multi-region sampling revealing intra-tumor heterogeneity of gallbladder carcinoma.

Gallbladder carcinoma (GBC) is a lethal biliary tract malignant neoplasm. Patient-derived primary cancer cell lines (PDPCs) are appropriate models to explore biological characteristics and potential therapeutics; however, there is a lack of PDPCs in GBC. In this study, we aimed to establish and characterize the GBC PDPCs, and further investigated the intra-tumor heterogeneity (ITH). Multi-region sampling (3-9 regions) of the operable tumor tissue samples was used to establish PDPCs. Short tandem repeat genotyping for cell authentication and karyotyping was performed, followed by whole-exome sequencing and RNA sequencing to assess the ITH at the genetic and transcriptional levels, respectively. Thirty-eight PDPCs were successfully established from seven GBC patients and characterized. ITH was observed with a median of 38.3% mutations being heterogeneous (range, 26.6-59.4%) across all patients. Similar with other tumor types, TP53 mutations were always truncal. In addition, there were three genes, KMT2C, CDKN2A, and ARID1A, with truncal mutations in at least two patients. A median of 370 differentially expressed genes (DEGs) was identified per patient. Distinct expression patterns were observed between major histocompatibility complex (MHC) class I and II genes. We found the expression of MHC class II genes in the PDPC samples was closely regulated by CIITA, while that of MHC class I genes were not correlated with CIITA expression. The PDPCs established from GBC patients can serve as novel in vitro models to identify the ITH, which may pave a crucial molecular foundation for enhanced understanding of tumorigenesis and progression.

Preoperative portal vein embolization for hilar cholangiocarcinoma--a comparative study.

BACKGROUND/AIMS: Preoperative portal vein embolization (PVE) allows potentially curative hepatic resection to be carried out in patients with hepatobiliary malignancies who are otherwise not candidates for resection because of the small size of the future liver remnant (FLR). However, there have only been a few reports on PVE before hepatectomy for hilar cholangiocarcinoma due to the small number of patients who can be treated with radical surgery. METHODOLOGY: Between January 2007 and March 2009, 49 consecutive patients with hilar cholangiocarcinoma who were planned to have hemi-hepatectomy/extended hemi-hepatectomy plus caudate lobe resection in our tertiary referral center were studied. The change in size of the FLR and the operative outcomes were compared between patients with or without PVE. RESULTS: All patients had liver dysfunction as a result of biliary obstruction due to hilar cholangiocarcinoma although they had all received percutaneous transhepatic biliary drainage. PVE was used in 16 patients with an estimated FLR of <50%, while no PVE was carried out in 33 patients with an estimated FLR of >50%. Complications after PVE occurred in 3 patients (18.8%), which included bile leakage (n=1) and coil displacement (n=2). No complication precluded liver resection. The FLR to total liver volume (TLV) ratio at presentation was significantly smaller in patients who underwent PVE than those who did not undergo PVE (40.3 +/- 7.4% vs. 56.6 +/- 5.0%; p < 0.001). After PVE, the FLR to TLV ratio increased significantly (40.3 +/- 7.4% vs. 43.1 +/- 7.0%; p < 0.001) at a mean time of 14.2 +/- 3.5 days. The mean +/- S.D. increase in FLR was 4.6 +/- 3.0 cm3/day. At surgery, the FLR volume was still significantly smaller in the PVE group than the non-PVE (802 +/- 216 cm3 vs. 979 +/- 202 cm3; p = 0.007). In the PVE group, insufficient hypertrophy of the FRL prevented one patient from having surgery, while local tumor progression and peritoneal dissemination precluded hepatectomy in 2 more patients. Finally, 13 patients (81.3%) underwent radical surgery. The PVE group had similar complication and mortality rates compared with the non-PVE group (complication rate, 69.2% vs. 63.6%; mortality rate, 0.0% vs. 9.1%). The 1- and 2-year overall survivals for the PVE group (with intent-to-treat analysis), PVE group (radical surgery only) and the non-PVE group were 57.3% and 43.0%; 71.3% and 53.5%; 70.4% and 54.4%, respectively. There was no significant difference in the survival outcomes. CONCLUSIONS: The results suggested that PVE is a safe and efficacious procedure in inducing adequate hypertrophy of the FLR before major hepatic resection for hilar cholangiocarcinoma with obstructive jaundice which had been relieved by percutaneous transhepatic biliary drainage.

RNA Sequencing Revealed Signals of Evolution From Gallbladder Stone to Gallbladder Carcinoma.

Gallbladder stone is a major risk factor for gallbladder carcinoma (GBC), while there is still a controversy whether period of follow-up since newly diagnoses of asymptomatic gallstones increases the risk of GBC. In this study, 10 GBC patients and 30 patients with gallstones were admitted to our hospital. Patients with gallstones were divided into 3 groups according to the follow-up time, involving 10 patients with follow-up period of 1-3 years (GS3 group), 10 patients with follow-up period of 5-10 years (GS5 group), and 10 patients with follow-up period of more than 10 years (GS10 group). Tumor and para-tumor tissues of GBC patients, and gallbladder tissues of gallstone patients were collected. RNA sequencing was performed on the 50 samples. Besides, 1,704 differentially expressed genes (DEGs) were identified in tumors compared with para-tumor tissues of 10 GBC patients, which were enriched into some well-known cancer-related pathways, such as PI3K-Akt, mitogen-activated protein kinase (MAPK), Ras, and Wnt signaling pathways, and the most significant pathway was neuroactive ligand-receptor interaction. Patients with gallstones with periods of follow-up equal to 1-3 and > 10 years showed to have higher cancer risk than those with 5-10 years. ALPP and GPR87 are potential biomarkers for predicting cancer risk in patients with gallstones. The in vitro results revealed that GPR-87 can promote the proliferation, migration, and invasion of GBC cells. Herein, we explored the relationship between GBC patients and patients with gallstones with different periods of follow-up in transcriptome level.

Targeted deep sequencing contributes to guiding personalized targeted therapy for advanced biliary tract cancer patients with non­radical resection: A real­world study.

Targeted therapy based on specific genetic alterations has been proven to be an effective treatment for various types of cancer. In the present study, we aimed to explore the efficacy of personalized targeted therapy guided by targeted deep sequencing for patients with advanced biliary tract cancer (BTC) after non­radical resection. Targeted deep sequencing was performed on 49 patients with BTC, to whom biologic agents were recommended. Among 32 patients with stage IV and R2 resection (a non­radical resection), 21 patients underwent conventional chemotherapy (mGEMOX), while the remaining 11 patients received a personalized targeted agent. The genomic landscape of the 49 patients with BTC was determined and the results showed that genetic alterations were enriched in the ERBB family and cell cycle pathway. After a median follow­up of 12 months, the 11 BTC patients with personalized targeted therapy showed a median progression­free survival (PFS) of 4.5 months (2.5­20.5 months), a median overall survival (OS) of 12.9 months (4.7­24.8 months) and a disease control rate (DCR) of 63.6%. In the other 21 BTC patients, who were undergoing conventional chemotherapy, the BTC patients had a median PFS of 1.5 months (0.5­11.6 months), a median OS of 4.1 months (1.3­18.4 months), and a DCR of 33.3%. In addition, 36.4% of the patients in the personalized targeted therapy group experienced grade >2 treatment­related toxicity vs. 19.0% of patients in the conventional chemotherapy group. This real­world study suggests that targeted deep sequencing contributes to the guidance of personalized targeted therapy based on individual actionable mutations, which may benefit advanced BTC patients undergoing non­radical resection.

Latent Risk Intrahepatic Cholangiocarcinoma Susceptible to Adjuvant Treatment After Resection: A Clinical Deep Learning Approach.

Background: Artificial Intelligence (AI) frameworks have emerged as a novel approach in medicine. However, information regarding its applicability and effectiveness in a clinical prognostic factor setting remains unclear. Methods: The AI framework was derived from a pooled dataset of intrahepatic cholangiocarcinoma (ICC) patients from three clinical centers (n = 1,421) by applying the TensorFlow deep learning algorithm to Cox-indicated pathologic (four), serologic (six), and etiologic (two) factors; this algorithm was validated using a dataset of ICC patients from an independent clinical center (n = 234). The model was compared to the commonly used staging system (American Joint Committee on Cancer; AJCC) and methodology (Cox regression) by evaluating the brier score (BS), integrated discrimination improvement (IDI), net reclassification improvement (NRI), and area under curve (AUC) values. Results: The framework (BS, 0.17; AUC, 0.78) was found to be more accurate than the AJCC stage (BS, 0.48; AUC, 0.60; IDI, 0.29; NRI, 11.85; P < 0.001) and the Cox model (BS, 0.49; AUC, 0.70; IDI, 0.46; NRI, 46.11; P < 0.001). Furthermore, hazard ratios greater than three were identified in both overall survival (HR; 3.190; 95% confidence interval [CI], 2.150-4.733; P < 0.001) and disease-free survival (HR, 3.559; 95% CI, 2.500-5.067; P < 0.001) between latent risk and stable groups in validation. In addition, the latent risk subgroup was found to be significantly benefited from adjuvant treatment (HR, 0.459; 95% CI, 0.360-0.586; P < 0.001). Conclusions: The AI framework seems promising in the prognostic estimation and stratification of susceptible individuals for adjuvant treatment in patients with ICC after resection. Future prospective validations are needed for the framework to be applied in clinical practice.

Tumor suppressor LKB1 inhibits the progression of gallbladder carcinoma and predicts the prognosis of patients with this malignancy.

Gallbladder carcinoma (GBC) represents the most common fatal tumors of the biliary tract. The 3-year or 5-year survival rate for patients with this disease are 30 and 5%, respectively. Liver kinase B1 (LKB1), a primary upstream kinase of adenosine monophosphate-activated protein kinase (AMPK) necessary for maintaining cell metabolism and energy homeostasis, has been found to be an important tumor suppressor gene in recent years, and its inactivation has also found to be closely associated with tumor growth, metastasis and cancer stem cell (CSC) proliferation. Nevertheless, the function of LKB1 in GBC remains unclear. In this study, we found that the expression of LKB1 in GBC tissues was decreased compared with that in non-cancerous tissues. LKB1 overexpression suppressed the proliferation, metastasis and expansion of GBC CSCs. Mechanically, LKB1 suppressed GBC cell progression via the JAK/signal transducer and activator of transcription 3 (STAT3) pathway. The use of the JAK2 inhibitor, AZD­1480, attenuated the suppressive effects of LKB1 overexpression on the growth, metastasis and self-renewal ability of the GBC cells, which further demonstrated that JAK/STAT3 was involved in the LKB1-induced suppression of GBC cell growth, metastasis and self-renewal ability. More importantly, the decreased expression of LKB1 was a predictor of a poor prognosis of patients with GBC. On the whole, our data indicate that LKB1 inhibits GBC cell growth, metastasis and self-renewal ability by disrupting JAK/STAT3 signaling, and may thus prove to be a novel prognostic biomarker for patients with GBC.

Phenformin inhibits cell proliferation and induces cell apoptosis and autophagy in cholangiocarcinoma.

Cholangiocarcinoma (CCA) is an aggressive malignant tumor and the prognosis of patients with advanced stage disease remains poor. Therefore, the identification of novel treatment agents for CCA is required. In the present study, the biological effects of the diabetes therapeutic agent, phenformin, in CCA cell lines was investigated. Cell Counting Kit­8 cell viability, cellular clone formation and subcutaneous tumor formation assays were performed, which revealed that phenformin inhibited CCA cell proliferation and growth both in vitro and in vivo. In addition, phenformin induced CCA cell apoptosis and autophagy. Phenformin partly activated the liver kinase B1 (LKB1)/5' AMP­activated protein kinase signaling pathway to exert its biological effects on CCA cell lines, as demonstrated by knockdown of LKB1, which reversed these effects. In conclusion, the present study demonstrated the biological effects of phenformin in CCA and suggested that phenformin may be a potential novel agent for CCA treatment.

Guidance to rational use of pharmaceuticals in gallbladder sarcomatoid carcinoma using patient-derived cancer cells and whole exome sequencing.

PURPOSE: Gallbladder sarcomatoid carcinoma is a rare cancer with no clinical standard treatment. With the rapid development of next generation sequencing, it has been able to provide reasonable treatment options for patients based on genetic variations. However, most cancer drugs are not approval for gallbladder sarcomatoid carcinoma indications. The correlation between drug response and a genetic variation needs to be further elucidated. EXPERIMENTAL DESIGN: Three patient-derived cells-JXQ-3D-001, JXQ-3D-002, and JXQ-3D-003, were derived from biopsy samples of one gallbladder sarcomatoid carcinoma patient with progression and have been characterized. In order to study the relationship between drug sensitivity and gene alteration, genetic mutations of three patient-derived cells were discovered by whole exome sequencing, and drug screening has been performed based on the gene alterations and related signaling pathways that are associated with drug targets. RESULTS: It has been found that there are differences in biological characteristics such as morphology, cell proliferation, cell migration and colony formation activity among these three patient-derived cells although they are derived from the same patient. Their sensitivities to the chemotherapy drugs-Fluorouracil, Doxorubicin, and Cisplatin are distinct. Moreover, none of common chemotherapy drugs could inhibit the proliferations of all three patient-derived cells. Comprehensive analysis of their whole exome sequencing demonstrated that tumor-associated genes TP53, AKT2, FGFR3, FGF10, SDHA, and PI3KCA were mutated or amplified. Part of these alterations are actionable. By screening a set of compounds that are associated with the genetic alteration, it has been found that GDC-0941 and PF-04691502 for PI3K-AKT-mTOR pathway inhibitors could dramatically decrease the proliferation of three patient-derived cells. Importantly, expression of phosphorylated AKT and phosphorylated S6 were markedly decreased after treatments with PI3K-AKT-mTOR pathway inhibitors GDC-0941 (0.5 µM) and PF-04691502 (0.1 µM) in all three patient-derived cells. These data suggested that inhibition of the PI3K-AKT-mTOR pathway that was activated by PIK3CA amplification in all three patient-derived cells could reduce the cell proliferation. CONCLUSIONS: A patient-derived cell model combined with whole exome sequencing is a powerful tool to elucidate relationship between drug sensitivities and genetic alternations. In these gallbladder sarcomatoid carcinoma patient-derived cells, it is found that PIK3CA amplification could be used as a biomarker to indicate PI3K-AKT-mTOR pathway activation. Block of the pathway may benefit the gallbladder sarcomatoid carcinoma patient with this alternation in hypothesis. The real efficacy needs to be confirmed in vivo or in a clinical trial.

Surgical therapy for hiliar cholangiocarcinoma: analysis of 198 cases.

BACKGROUND: Carcinoma of the hepatic duct confluence is the most common site of bile duct malignancies. Although hilar cholangiocarcinoma has been characterized as a slow-growing and late metastasizing tumor, post-therapeutic prognosis has remained poor. The study was undertaken to analyze factors influencing the surgical curative effect of hilar cholangiocarcinoma. METHODS: A retrospective clinical analysis was made of 198 patients with hilar cholangiocarcinoma who had been surgically treated at our hospital from 1997 to 2002. Jaundice (94.5%, 187 patients), pruritus (56.6%, 112) and abdominal pain (33.8%, 67) were the main symptoms. According to the Bismuth-Corlette classification, there were 14 type I patients, 19 type II patients, 12 type IIIa patients, 15 type IIIb patients, 112 type IV patients, and 26 unclassified patients. 144 patients received laparotomy, and 120 tumor resection including radical resection (59 patients) and palliative resection (61). Fifty-four patients were treated by endoscopic surgery and 16 patients by postoperative adjuvant radiation. RESULTS: Occupation, preoperative level of total serum bilirubin, operative procedure and postoperative adjuvant radiation affected postoperative survival of the patients. The postoperative survivals of endoscopic nose-biliary drainage (ENBD) group, endoscopic retrograde biliary drainage (ERBD) or endoscopic metal biliary endoprosthesis (EMBE) group, biliary exploration and drainage group, palliative resection group and radical resection group differed (chi2=87.0489, P<0.01). CONCLUSION: Early diagnosis and radical resection are important to improve the prognosis of hilar cholangiocarcinoma.

Dynamic Change of Total Bilirubin after Portal Vein Embolization is Predictive of Major Complications and Posthepatectomy Mortality in Patients with Hilar Cholangiocarcinoma.

OBJECTIVES: This study aims to evaluate the role of dynamic change in total bilirubin after portal vein embolization (PVE) in predicting major complications and 30-day mortality in patients with hilar cholangiocarcinoma (HCCA). METHODS: Retrospective analysis of prospectively maintained data of 64 HCCA patients who underwent PVE before hepatectomy in our institution was used. Total bilirubin and other parameters were measured daily in peri-PVE period. The difference between them and the baseline value from days 0-5 to day -1 (∆D1) and days 5-14 to day -1 (∆D2) were calculated. The relationship between ∆D1 and ∆D2 of total bilirubin and major complications as well as 30-day mortality was analyzed. RESULTS: Out of 64 patients, 10 developed major complications (15.6 %) and 6 patients (9.3 %) had died within 30 days after surgery. The ∆D2 of total bilirubin after PVE was most significantly associated with major complications (P < 0.001) and 30-day mortality (P = 0.002). In addition, it was found to be an independent predictor of major complications after PVE (odds ratio (OR) = 1.050; 95 % CI 1.017-1.084). ASA >3 (OR = 12.048; 95 % CI 1.019-143.321), ∆D2 of total bilirubin (OR = 1.058; 95 % CI 1.007-1.112), and ∆D2 of prealbumin (OR = 0.975; 95 % CI 0.952-0.999) were associated with higher risk of 30-day mortality after PVE. Receiver operating characteristic curves showed that ∆D2 of total bilirubin were better predictors than ∆D1 for major complications (AUC (∆D2) 0.817; P = 0.002 vs. AUC (∆D1) 0.769; P = 0.007) and 30-day mortality (ACU(∆D2) 0.868; P = 0.003 vs. AUC(∆D1) 0.721;P = 0.076). CONCLUSION: Patients with increased total bilirubin in 5-14 days after PVE may indicate a higher risk of major complications and 30-day mortality if the major hepatectomy were performed.

[Analysis of the relation between surgery and prognosis of hilar cholangiocarcinoma].

OBJECTIVE: To explore the prognosis factors of hilar cholangiocarcinoma, and investigate the relation between operative procedure and prognosis of it. METHODS: A retrospective cohort study was investigated in 198 patients with hilar cholangiocarcinoma, who were treated in our hospital from December 1997 to December 2002. There were 117 males and 81 females. The age ranged from 27 to 81 years old with a mean of 56. Jaundice (94.5%), pruritus (56.6%) and abdominal pain (33.8%) were the main present symptoms. According to Bismuth-Corlette classification, there were 14 type I cases, 19 type II cases, 12 type IIIa cases, 15 type IIIb cases, 112 type IV cases and 26 unclassifiable cases. One hundred and forty four cases received open operative treatment, and the others only were treated with endoscopic approach (including ERBD or EMBE 21 cases, ENBD 31 cases) or percutaneous transhepatic cholangiodrainage (2 cases). Tumor resection was performed on 120 cases with a resection rate of 83.3%, included radical resection 59 cases (41.0%). Twenty-four cases underwent paunched biliary exploration and drainage. RESULTS: The Cox's regression model analysis showed that occupation, preoperative maximum total serum bilirubin level, operative procedure and postoperative adjuvant radiation affected postoperative survival significantly, but gender, age, choledocholithiasis, hepatitis, preoperative serum CA19-9 level, Bismuth-Corlette type, histopathologic grading and postoperative chemotherapy were not significant prognostic factors. The postoperative survival of biliary drainage group, palliative resection group and radical resection group, which statistically differed pairwise. Between ERBD or EMBE group and palliative resection group, there was no statistical difference. So was between ERBD or EMBE group and biliary drainage group, or between ENBD group and biliary drainage group. The survival differed statistically between ERBD or EMBE group and ENBD group. CONCLUSIONS: Operative procedure was the most important prognosic factor of hilar cholangiocarcinoma, radical resection still was the primary measure to cure and long term survival. For irresectable hilar cholangiocarcinoma, the effect of ERBD or EMBE could not be considered to be worse than that of open operative treatment.

Circulating miR-106a is a Novel Prognostic and Lymph Node Metastasis Indicator for Cholangiocarcinoma.

Cholangiocarcinoma (CCA) is a common biliary malignancy. Despite continuing advances, novel indicators are urgently needed to identify patients with a poor prognosis. Several microRNAs (miRNAs) have been reported to be dysregulated in CCA tissues. The purpose of the current study was to explore the potential use of certain miRNAs as serum indicators. A total of 157 individuals, including103 CCA patients, were recruited into this study. We first used qRT-PCR to evaluate 5 CCA-related miRNAs in the serum of 95 individuals to identify significantly deregulated miRNAs. A logistic regression was used to analyse the potential variables influencing lymph node metastasis. Cox proportional hazards regression models were applied to determine the association between possible prognostic variables and overall survival (OS). We observed that decreased serum miR-106a confers a higher likelihood of lymph node metastasis [hazard ratio (HR) 18.3, 95% confidence interval (CI) 5.9-56.4, p < 0.01]. Additionally, lower circulating miR-106a levels (HR 5.1; 95% CI 2.2-11.8; p < 0.01) and non-radical surgery (HR 4.2; 95% CI 2.3-7.7; p < 0.01) were independent predictors for poor prognosis. Together, reduced expression of serum miR-106a is a powerful prognostic indicator for CCA patients. The dismal outcome of these CCA patients might correlate with a higher risk of lymph node metastasis.