Genetic alterations including fusions in fibroblast growth factor receptor 2 (FGFR2) are detected in 10-20% of intrahepatic cholangiocarcinoma (iCCA), and FGFR2 inhibitors are effective for the treatment of iCCA. We examined a prevalence of FGFR2 genetic alterations and their clinicopathological significance in combined hepatocellular-cholangiocarcinoma (cHCC-CCA). FGFR2 expression, which is a surrogate marker for FGFR2 genetic alterations, was immunohistochemically assessed in the liver sections from 75 patients with cHCC-CCA, 35 with small duct-type iCCA, 30 with large duct-type iCCA, and 35 with hepatocellular carcinoma (HCC). FGFR2 genetic alterations were detected by reverse transcription-PCR and direct sequence. An association of FGFR2 expression with clinicopathological features was investigated in cHCC-CCAs. FGFR2 expression was detected in significantly more patients with cHCC-CCA (21.3%) and small duct-type iCCA (25.7%), compared to those with large duct-type iCCA (3.3%) and HCC (0%) (p < 0.05). FGFR2-positive cHCC-CCAs were significantly smaller size (p < 0.05), with more predominant cholangiolocarcinoma component (p < 0.01) and less nestin expression (p < 0.05). Genetic alterations of ARID1A and BAP1 and multiple genes were significantly more frequent in FGFR2-positive cHCC-CCAs (p < 0.05). 5\'/3\' imbalance in FGFR2 genes indicating exon18-truncated FGFR2 was significantly more frequently detected in FGFR2-positive cHCC-CCAs and small duct iCCAs, compared to FGFR2-negative ones (p < 0.05). FGFR2::BICC fusion was detected in a case of cHCC-CCAs. FGFR2 genetic alterations may be prevalent in cHCC-CCAs as well as small duct-type iCCAs, which suggest cHCC-CCAs may also be a possible therapeutic target of FGFR2 inhibitors.

The aim of the study was to comprehensively understand the combined hepatocellular and cholangiocarcinoma (CHC) and develop a nomogram for prognostic prediction of CHC.
Data were collected from the Surveillance, Epidemiology and End Results (SEER) database (year 2004-2014). Propensity-score matching (PSM) was used to match the demographic characteristic of the CHC versus hepatocellular carcinoma (HCC) and intrahepatic cholangiocarcinoma (ICC). A nomogram model was established to predict the prognosis in terms of cancer specific survival (CSS). The established nomogram was externally validated by a multicenter cohort.
A total of 71,756 patients enrolled in our study including 62,877 HCC patients, 566 CHC patients, and 8303 ICC patients. The CHC, HCC, and ICC are not exactly similar in clinical characteristic. After PSM, the CSS of CHC was better than HCC but comparable to ICC. Tumor size, M stage, surgery, chemotherapy, and surgery were independently prognostic factors of CHC and were included in the establishment of novel nomogram. The c-index of the novel nomogram in SEER training set and multicenter validation was 0.779 and 0.780, respectively, which indicated that the model was with better discrimination power. In addition, decision curve analyses proved the favorable potential clinical effect of the predictive model. Lastly, a risk classification based on nomogram also verified the reliability of the model.
CHC had better survival than HCC but was comparable to ICC. The nomogram was established based on tumor size, M stage, chemotherapy, surgery, and radiotherapy and well validated by external multicenter cohort.

OBJECTIVE: The type IV intermediate filament, nestin, may be a candidate diagnostic marker for combined hepatocellular-cholangiocarcinoma (cHCC-CCA). Therefore, the significance of nestin as a diagnostic marker for cHCC-CCA categorized by the World Health Organization (WHO) 2019 classification and its relationship with clinicopathological features were examined in the present study.
RESULTS: Nestin expression was immunohistochemically assessed in the liver sections from 75 patients with cHCC-CCA, 22 with small duct-type intrahepatic cholangiocarcinoma (iCCA), 20 with large duct-type iCCA and 35 with hepatocellular carcinoma (HCC). Nestin expression and its relationship with clinicopathological features and genetic alterations were investigated in cHCC-CCA. Nestin expression was detected in significantly more patients with cHCC-CCA (66.7%) than in those with large duct-type iCCA (5%) (P < 0.01), HCC (2.9%) (P < 0.01) and small duct-type iCCA (40.9%) (P < 0.05). Nestin expression was partly associated with neural cell adhesion molecule (NCAM) and vimentin expression. Nestin expression was also observed in significantly more patients with small duct-type iCCA than in those with large duct-type iCCA and HCC (P < 0.01). Nestin-positive cHCC-CCA was characterized by a smaller tumour size, the more frequent presence of cholangiolocellular carcinoma (CLC) components, a higher rate of p53 overexpression and a higher rate of multiple genetic alterations (P < 0.05). Furthermore, p53 overexpression was associated with a higher histological grade and multiple genetic alterations (P < 0.05) in nestin-positive cHCC-CCA.
CONCLUSIONS: Nestin may be a useful diagnostic marker for a specific subgroup of cHCC-CCA and small duct-type iCCA associated with CLC components, p53 mutations and multiple genetic alterations, which are related to stemness and multipotent differentiation.

OBJECTIVE: Precursor lesions of small duct type intrahepatic cholangiocarcinoma (small duct iCCA) have not been clarified so far. We hypothesised that precursor lesions may be frequently distributed in the background liver of small duct iCCA.
RESULTS: We determined by histology the presence of bile duct adenomas and von Meyenburg complexes as candidate precursor lesions in the background liver of small duct iCCA, with other primary liver carcinomas as control. Subjects included 28 patients with small duct iCCA, 29 with large duct iCCAs, 60 with combined hepatocellular-cholangiocarcinoma (Comb) and 40 with hepatocellular carcinoma (HCC). The prevalence of bile duct adenomas in the background liver was significantly higher in small duct iCCA (35.7%) compared to other primary liver carcinomas (Comb, 4.9%; 10%, HCC) (P < 0.01). The prevalence of bile duct adenomas was significantly associated with the presence of von Meyenburg complexes and ductal plate malformation-like patterns in small duct iCCAs and Combs. Von Meyenburg complexes were detected in 11 small duct iCCA (39.3%), five large duct iCCAs (17.2%), 10 Comb (16.4%) and 13 HCC (33.3%), respectively (P > 0.05). Small duct iCCAs showed altered expression of ARID1A (46.4%), p53 (39.3%), PBRM1 (14.3%), IMP3 (85.7%) and EZH2 (82.1%), whereas these markers were negative in bile duct adenomas.
CONCLUSIONS: Bile duct adenomas may be precursor lesions of small duct iCCAs. Alteration of ARID1A, p53 or PBRM1 may be involved in the carcinogenesis of small duct iCCAs.

BACKGROUND: Combined hepatocellular and cholangiocarcinoma (HCC/CC) is a rare primary hepatic malignancy which carries a poor prognosis due to its aggressive nature. Few centers have enough cases to draw definitive conclusions and there is limited understanding of prognosis. Given the rarity of HCC/CC, an analysis of large national cancer database was needed to obtain larger number of HCC/CC cases.
OBJECTIVE: To identify associated factors for 5-year survival of HCC/CC.
METHODS: We conducted a retrospective study of The Surveillance, Epidemiology, and End Results (SEER) database obtained from SEER*Stat 8.3.6 software. Previously defined histology code 8180 for the International Classification of Disease for Oncology, 3rd edition was used to identify HCC/CC cases from 2004 to 2015. We collected demographics, American Joint Committee on Cancer (AJCC) stage, treatment, tumor size, and survival data. These data were converted to categorical variables. The Shapiro-Wilk normality test was used to assess normal distribution. Mann-Whitney U test was used to compare continuous variables without normal distribution, and t-test was used to compare continuous variables with a normal distribution. The Kaplan-Meier survival curve analyzed 5-year survival. Univariate and multivariate logistic regression model was used to analyze factors associated with 5-year survival. Multivariate Cox proportional hazard regression was done on 5-year survival. We defined P < 0.05 was statistically significant.
RESULTS: We identified 497 patients with the following characteristics: Mean age 62.4 years (SD: 11.3), 149 (30.0%) were female, racial distribution was: 276 (55.5%) white, 53 (10.7%) black, 84 (16.9%) Asian and Pacific Islander (API), 77 (15.5%) Hispanic, and 7 (1.4%) others or unknown. Stage I/II disease occurred in 41.5% and tumor size < 50 mm was seen in 35.6% of patients. Twenty-four (4.8%) received locoregional therapy (LRT), 119 (23.9%) underwent resection, and 50 (10.1%) underwent liver transplantation. The overall median survival was 6 mo [Interquartile range (IQR): 1-22]. After multivariate logistic regression, tumor size < 50 mm [Odds ratios (OR): 2.415, P = 0.05], resection (OR: 12.849, P < 0.01), and transplant (OR: 27.129, P < 0.01) showed significance for 5-year survival. Age > 60, sex, race, AJCC stages, metastasis, and LRT were not significant. However, API vs white showed significant OR of 2.793 (CI: 1.120-6.967). Cox proportional hazard regression showed AJCC stages, tumor size < 50 mm, LRT, resection, and transplant showed significant hazard ratio.
CONCLUSIONS: HCC/CC patients with tumor size < 50 mm, resection, and transplant were associated with an increase in 5-year survival. API showed advantageous OR and hazard ratios over white, black.

Although cholangiolocellular carcinoma is considered a combined hepatocellular and cholangiocarcinoma, we feel that this classification is not appropriate. Therefore, we compared the diagnostic imaging findings, surgical prognosis, and pathological features of cholangiolocellular carcinoma with those of other combined hepatocellular and cholangiocarcinoma subtypes, hepatocellular carcinoma, and cholangiocarcinoma.
The study patients included 7 with classical type combined hepatocellular and cholangiocarcinoma; 8 with stem cell feature, intermediate type combined hepatocellular and cholangiocarcinoma; 13 with cholangiolocellular carcinoma; 58 with cholangiocarcinoma; and 359 with hepatocellular carcinoma. All patients underwent hepatectomy or living-related donor liver transplantation from 2001 to 2014.
cholangiolocellular carcinoma could be distinguished from hepatocellular carcinom, other combined hepatocellular and cholangiocarcinoma subtypes, and cholangiocarcinoma by the presence of intratumoral Glisson\'s pedicle, hepatic vein penetration, and tumor-staining pattern on angiography-assisted CT. Cholangiolocellular carcinoma was associated with a significantly lower SUV-max than that of cholangiocarcinoma on FDG-PET. Hepatocellular carcinoma, classical type, and cholangiolocellular carcinoma had significantly better prognoses than stem cell feature, intermediate type and cholangiocarcinoma. A cholangiocarcinoma component was detected in cholangiolocellular carcinoma that progressed to the hepatic hilum, and the cholangiocarcinoma component was found in perineural invasion and lymph node metastases.
From the viewpoint of surgeon, cholangiolocellular carcinoma should be classified as a good-prognosis subtype of biliary tract carcinoma because of its tendency to differentiate into cholangiocarcinoma during its progression, and its distinctive imaging and few recurrence rates different from other combined hepatocellular and cholangiocarcinoma subtypes.

Background: The clinical significance of Albumin-to-Alkaline Phosphatase Ratio (AAPR) has been discussed in hepatocellular carcinoma (HCC) and cholangiocarcinoma (CC). The aim of this study is to clarify the prognostic value of AAPR in patients with combined hepatocellular and cholangiocarcinoma (cHCC-CCA). Methods: A total of 267 patients pathologically diagnosed as Allen type C cHCC-CCA in our institution were retrospectively enrolled and randomly divided into the training (N=187) cohort and validation (N=80) cohort. The prognostic value of AAPR was evaluated and validated. An AAPR-based nomogram was constructed and its prediction performance was assessed. Results: We identified 0.43 as the optimal threshold value of AAPR by the X-tile software. In the training cohort, the median overall survival (OS) of patients with AAPR < 0.43 was significant shorter than that of those with AAPR ≥ 0.43(15.8 months vs 35 months, respectively, P < 0.001). Univariate and multivariate analyses demonstrated that AAPR was a strong indicator of OS. The concordance index (C-index), receiver operating characteristic (ROC) curves, likelihood ratio tests (LAT), Akaike information criteria (AIC) and decision curve analysis (DCA) demonstrated that AAPR outperformed the Child-Pugh (CP) grade and albumin-bilirubin (ALBI) grade in predicting OS. These findings were further verified in the validation cohort. The AAPR-based nomogram achieved C-index values of 0.76 (95%CI: 0.71-0.81) in the training cohort and 0.69 (95%CI: 0.60-0.78) in the validation cohort, which presented significant superiority to TNM stage. Conclusions: Preoperative AAPR is an independent prognostic predictor in cHCC-CCA. The AAPR-based nomogram contributes to personalized prognosis prediction and clinical decision making for cHCC-CCA.

Combined hepatocellular and cholangiocarcinoma (cHCC-CCA) is a rare but aggressive primary liver cancer with dismal prognosis. We aim to develop a new scoring method for personalized prognostic prediction in patients with cHCC-CCA undergoing surgical resection.
Between January 1993 and December 2015, a total of 296 Allen type C cHCC-CCA patients who had received surgical resection in Liver Cancer Institute, Zhongshan Hospital were retrospectively enrolled. A novel prognostic scoring method for cHCC-CCA (PSM-CHCC model) was established and validated. The predictive value of the new model was compared with current prognostic staging systems.
The scoring model was developed based on the independent prognostic variables identified by Cox regression model. Based on the PSM-CHCC model, patients were stratified into three prognostic subgroups according to their individual score: A (scoring 0-2), B (scoring 3-5), and C (scoring > 5). The prediction performance of the PSM-CHCC model outperformed the widely accepted TNM staging system and other staging systems in both training and validation cohorts. Subgroup analysis also verified the discrimination efficacy of the PSM-CHCC model.
The newly established PSM-CHCC model may facilitate prognostic stratification and clinical decision-making in patients with cHCC-CCA.

OBJECTIVE: Combined hepatocellular-cholangiocarcinoma (HCC-CCA) is a rare liver malignancy distinct from either hepatocellular carcinoma (HCC) or cholangiocarcinoma. Liver transplantation (LT) is not recommended for HCC-CCA because of suboptimal outcomes. Non-invasive diagnosis of HCC-CCA is extremely challenging; thus, some HCC-CCAs are presumed as HCC on imaging and listed for LT with the correct diagnosis ultimately made on explant pathology. We compared HCC-CCA with HCC to determine the utility of response to pre-transplant loco-regional therapy (LRT) in predicting outcomes for HCC-CCA after LT as a potential means of identifying appropriate HCC-CCA patients for LT.
METHODS: Retrospective review of 19 patients with pathologically confirmed HCC-CCA were individually matched to 38 HCC patients (1:2) based on age, sex, and Milan criteria at listing was performed. The modified response evaluation criteria in solid tumors was used to categorize patients as responders or non-responders to pre-transplant LRT based on imaging performed before and after LRT. Overall survival (OS) and recurrence-free survival (RFS) were examined.
RESULTS: OS at 3 years post-transplant was 74% for HCC-CCA and 87% for HCC. RFS at 3 years was 74% for HCC-CCA, and 87% for HCC. Among responders to LRT, the 3-year OS was 92% for HCC-CCA and 88% for HCC; among non-responders, 3-year OS was 43% for HCC-CCA and 83% for HCC. Higher 3-year OS was observed among HCC-CCA responders (77%) compared with HCC-CCA non-responders (23%).
CONCLUSIONS: OS was similarly high among.

BACKGROUND: The Hippo pathway plays a vital role in liver regeneration and development by determining cellular lineage and regulating cell proliferation and apoptosis. In this study, we aimed to assess the role of the Hippo pathway in hepatic carcinogenesis and morphogenesis by examining Yes-associated protein 1 (YAP1) expression in the spectrum of hepatic carcinomas based on cellular lineage.
METHODS: We examined 913 primary hepatic carcinomas, including hepatocellular carcinomas (HCCs), combined hepatocellular and cholangiocarcinomas (cHC-CCAs), intrahepatic cholangiocarcinomas (IHCCAs) and perihilar extrahepatic bile duct carcinomas (EHBCAs). Our study group was categorized into 8 disease groups, based on histological diagnosis and cytokeratin 19 (CK19) expression, and immunohistochemistry was used to detect and compare YAP1 expression levels between the groups. The eight disease groups we identified were: 1) CK19(-) HCC, 2) CK19(-) scirrhous HCC, 3) CK19(+) HCC, 4) stem cell feature of cHC-CCA, 5) classical cHC-CCA, 6) cholangiolocellular IHCCA, 7) non-cholangiolocellular IHCCA, and 8) EHBCA.
RESULTS: Positive rates of YAP1 were the highest in the EHBCA group (21%). CK19(+) HCC and non-cholangiolocellular IHCCA groups also showed high expression levels (10% -11%), while the CK19 (-) HCC, CK19 (-) scirrhous HCC, cHC-CCA, and cholangiolocellular IHCCA groups showed low expression levels, ranging between 0% and 5%. Survival analysis, restricted to pT1 stage HCCs and IHCCAs, showed poor overall survival for YAP1(+) IHCCA patients (39 ± 17 vs. 109 ± 10 months, mean ± SD, log rank p-value 0.005). For HCCs, a trend of poor progression-free survival for YAP1(+) HCCs was observed (39 ± 18 vs. 81 ± 5 months, mean ± SD, log rank p-value 0.205) CONCLUSIONS: YAP1 activation was more commonly found in CCAs than in pure HCCs. However, a differing pattern of YAP1 expression between cHC-CCAs and CK19(+) HCCs and the poor prognosis of YAP1 positive hepatic carcinomas suggests that YAP1 may have a preferential role in aggressive tumor behavior, rather than in the determination of cellular lineage in hepatic carcinomas.