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Abstract Background Anaplastic lymphoma kinase (ALK) test in advanced non‐small cell lung cancer (NSCLC) can help physicians provide target therapies for patients harboring ALK gene rearrangement. This study aimed to investigate the real‐world test patterns and positive rates of ALK gene rearrangements in advanced NSCLC. Methods In this real‐world study (ChiCTR2000030266), patients with advanced NSCLC who underwent an ALK rearrangement test in 30 medical centers in China between October 1, 2018 and December 31, 2019 were retrospectively analyzed. Interpretation training was conducted before the study was initiated. Quality controls were performed at participating centers using immunohistochemistry (IHC)‐VENTANA‐D5F3. The positive ALK gene rearrangement rate and consistency rate were calculated. The associated clinicopathological characteristics of ALK gene rearrangement were investigated as well. Results The overall ALK gene rearrangement rate was 6.7% in 23,689 patients with advanced NSCLC and 8.2% in 17,436 patients with advanced lung adenocarcinoma. The quality control analysis of IHC‐VENTANA‐D5F3 revealed an intra‐hospital consistency rate of 98.2% (879/895) and an inter‐hospital consistency rate of 99.2% (646/651). IHC‐VENTANA‐D5F3 was used in 53.6%, real‐time polymerase chain reaction (RT‐PCR) in 25.4%, next‐generation sequencing (NGS) in 18.3%, and fluorescence in‐situ hybridization (FISH) in 15.9% in the adenocarcinoma subgroup. For specimens tested with multiple methods, the consistency rates confirmed by IHC‐VENTANA‐D5F3 were 98.0% (822/839) for FISH, 98.7% (1,222/1,238) for NGS, and 91.3% (146/160) for RT‐PCR. The overall ALK gene rearrangement rates were higher in females, patients of ≤ 35 years old, never smokers, tumor cellularity of > 50, and metastatic specimens used for testing in the total NSCLC population and adenocarcinoma subgroup (all P < 0.05). Conclusions This study highlights the real‐world variability and challenges of ALK test in advanced NSCLC, demonstrating a predominant use of IHC‐VENTANA‐D5F3 with high consistency and distinct clinicopathological features in ALK‐positive patients. These findings underscore the need for a consensus on optimal test practices and support the development of refined ALK test strategies to enhance diagnostic accuracy and therapeutic decision‐making in NSCLC.

Abstract Background Accurate lymphoma classification is critical for effective treatment and immunohistochemistry is a cost‐effective and time‐saving approach. Although several machine learning algorithms showed effective results, they focused on a specific task of classification but not the whole classification workflow, thus impractical to be applied in clinical settings. Thus, we aim to develop an effective and economic machine learning‐assisted system that can streamline the lymphoma differential diagnostic workflow using EBER in situ hybridization and immunohistochemical markers. Methods We included pathological reports diagnosed as lymphomas from two cancer centers (Sun Yat‐sen University Cancer Center and Peking University Cancer Hospital & Institute). We proposed a hierarchical approach that mimicked the human diagnostic process and employed simplified panels of markers to perform a series of interpretable classification. The diagnostic accuracy for lymphoma pathological subtypes and the markers saving ratio were investigated in both temporal independent population and external medical center. Results A total of 14,927 patients and corresponding immunohistochemical results from two cancer centers were included. The proposed system had high discriminative ability for differentiating lymphoma pathological subtypes (measured by mean AUC in three validation cohorts, non‐Hodgkin and Hodgkin lymphoma: 0.959; non‐Hodgkin subtypes: 0.983; B‐lymphoma subtypes: 0.868; T‐lymphoma subtypes: 0.962; DLBCL subtypes: 0.957). In addition, the system's well selected characteristics can contribute to the development of agreement on panels of markers for differential diagnosis and help minimize cost of immunohistochemical marker techniques (measured by marker saving ratio compared to real clinical settings, internal primary‐stage cohort: 16.45% saved, p

Abstract Introduction Gastric cancer (GC) is one of the leading causes of cancer-related death worldwide. Recently, targeted therapies including PD1 (programmed cell death 1) antibodies have been used in advanced GC patients. However, identifying new biomarker for immunotherapy is still urgently needed. The objective of this study is to unveil the immune evasion mechanism of GC cells and identify new biomarkers for immune checkpoint blockade therapy in patients with GC. Methods Coimmunoprecipitation and meRIP were performed to investigate the mechanism of immune evasion of GC cells. Cocuture system was established to evaluate the cytotoxicity of cocultured CD8+ T cells. The clinical significance of HSPA4 upregulation was analyzed by multiplex fluorescent immunohistochemistry staining in GC tumor tissues. Results Histone acetylation causes HSPA4 upregulation in GC tumor tissues. HSPA4 upregulation increases the protein stability of m6A demethylase ALKBH5. ALKBH5 decreases CD58 in GC cells through m6A methylation regulation. The cytotoxicity of CD8+ T cells are impaired and PD1/PDL1 axis is activated when CD8+ T cells are cocultured with HSPA4 overexpressed GC cells. HSPA4 upregulation is associated with worse 5-year overall survival of GC patients receiving only surgery. It is an independent prognosis factor for worse survival of GC patients. In GC patients receiving the combined chemotherapy with anti-PD1 immunotherapy, HSPA4 upregulation is observed in responders compared with non-responders. Conclusion HSPA4 upregulation causes the decrease of CD58 in GC cells via HSPA4/ALKBH5/CD58 axis, followed by PD1/PDL1 activation and impairment of CD8+ T cell’s cytotoxicity, finally induces immune evasion of GC cells. HSPA4 upregulation is associated with worse overall survival of GC patients with only surgery. Meanwhile, HSPA4 upregulation predicts for better response in GC patients receiving the combined immunotherapy.

Summary: Accurate tumor diagnosis by pathologists relies on identifying specific morphological characteristics. However, summarizing these unique morphological features in tumor classifications can be challenging. Although deep learning models have been extensively studied for tumor classification, their indirect and subjective interpretation obstructs pathologists from comprehending the model and discerning the morphological features accountable for classifications. In this study, we introduce a new approach utilizing Style Generative Adversarial Networks, which enables a direct interpretation of deep learning models to detect significant morphological characteristics within datasets representing patients with deficient mismatch repair/microsatellite instability-high gastric cancer. Our approach effectively identifies distinct morphological features crucial for tumor classification, offering valuable insights for pathologists to enhance diagnostic accuracy and foster professional growth.

Abstract Although many studies have compared tumor fibroblasts (T-Fbs) and nontumor fibroblasts (N-Fbs), less is understood about the stromal contribution of metastatic lymph node fibroblasts (LN-Fbs) to the evolving microenvironment. Here, we explored the characteristics of LN-Fbs in esophageal squamous cell carcinoma (ESCC) and the interactions between fibroblasts and ESCC tumor cells in metastatic lymph nodes. Fibroblasts were isolated from tumor, nontumor and metastatic lymph node tissues from different patients with ESCC. Transcriptome sequencing was performed on the fibroblasts. Tumor growth and drug-resistance assays were carried out, and characteristics of T-Fbs, N-Fbs and LN-Fbs were determined. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to assay the culture medium of fibroblasts. The results demonstrated that fibroblasts derived from different tissues had different characteristics. Coculture with LN-Fbs conditioned medium inhibited ESCC tumor cell growth and induced chemoresistance in ESCC cells. LN-Fbs induced chemoresistance to cisplatin in ESCC cells by secreting PI16. Coculture with LN-Fbs conditioned medium decreased cisplatin-induced apoptosis in ESCC cells by regulating the p38 and JNK cell signaling pathways. Survival analyses showed that patients with high PI16 expression in Fbs of lymph nodes exhibited worse overall survival. We also examined PI16 expression in interstitial tissues in ESCC tumor samples of patients receiving platinum-based therapy postsurgery and found that high PI16 expression in tumor interstitial tissues was an independent prognostic factor for ESCC patients. In addition, an in vivo assay demonstrated that PI16 knockdown increased the sensitivity of ESCC cells to cisplatin. Our results suggest that fibroblasts in metastatic lymph nodes decrease apoptosis of ESCC cells via PI16, thereby providing a cisplatin-resistance niche and supporting ESCC tumor cells to survive in metastatic lymph nodes. PI16 is also a potential target for effectively blocking the chemoresistance niche signaling circuit in response to cisplatin.

Background: Colorectal cancer (CRC) is one of the most common malignancies causing the third highest mortality rate in the world. It is particularly urgent to explore effective therapeutic strategies to overcome this disease. We identified a novel benzothiazole derivative (BTD) that may serve as a potentially effective agent against CRC.Method: MTT assays, cell colony formation assays, EdU staining assays, flow cytometry, RNA-seq, Western blotting, and migration and invasion assays were used to examine the effects of BTD on cell proliferation, apoptosis, metastasis, and the cell cycle. The antitumor activity of BTD in vivo was investigated in a CT26 tumor-bearing mouse model. Immunohistochemistry (IHC) was performed to examine the protein expression in mouse tumors. Hematology, biochemical analysis, and H&E staining were used to assess the biosafety of BTD.Results: We observed that BTD suppressed cell proliferation and metastasis and promoted the apoptosis of tumor cells in vitro. Treatment with BTD at a tolerable dose significantly reduced tumor growth in CT26-tumor-bearing mice and appeared to be safe. Treatment of BTD induced apoptosis by increasing the generation of reactive oxygen species (ROS) and evoking the loss of mitochondrial transmembrane potential. Overall, BTD suppressed cell proliferation and metastasis, and induced apoptosis of colorectal tumor cells through the ROS–mitochondria-mediated apoptotic pathway. The preliminary proof of the antitumor activity and relative safety of BTD were validated in a mouse model.Conclusion: Our findings suggest that BTD could serve as a potentially safe and effective candidate for CRC treatment.

Background: Bladder cancer (BCa) is the leading reason for death among genitourinary malignancies. RNA modifications in tumors closely link to the immune microenvironment. Our study aimed to propose a promising model associated with the “writer” enzymes of five primary RNA adenosine modifications (including m6A, m6Am, m1A, APA, and A-to-I editing), thus characterizing the clinical outcome, immune landscape and therapeutic efficacy of BCa.Methods: Unsupervised clustering was employed to categorize BCa into different RNA modification patterns based on gene expression profiles of 34 RNA modification “writers”. The RNA modification “writers” score (RMS) signature composed of RNA phenotype-associated differentially expressed genes (DEGs) was established using the least absolute shrinkage and selection operator (LASSO), which was evaluated in meta-GEO (including eight independent GEO datasets) training cohort and the TCGA-BLCA validation cohort. The hub genes in the RMS model were determined via weighted gene co-expression network analysis (WGCNA) and were further validated using human specimen. The potential applicability of the RMS model in predicting the therapeutic responsiveness was assessed through the Genomics of Drug Sensitivity in Cancer database and multiple immunotherapy datasets.Results: Two distinct RNA modification patterns were determined among 1,410 BCa samples from a meta-GEO cohort, showing radically varying clinical outcomes and biological characteristics. The RMS model comprising 14 RNA modification phenotype-associated prognostic DEGs positively correlated with the unsatisfactory outcome of BCa patients in meta-GEO training cohort (HR = 3.00, 95% CI = 2.19–4.12) and TCGA-BLCA validation cohort (HR = 1.53, 95% CI = 1.13–2.09). The infiltration of immunosuppressive cells and the activation of EMT, angiogenesis, IL-6/JAK/STAT3 signaling were markedly enriched in RMS-high group. A nomogram exhibited high prognostic prediction accuracy, with a concordance index of 0.785. The therapeutic effect of chemotherapeutic agents and antibody-drug conjugates was significantly different between RMS-low and -high groups. The combination of the RMS model and conventional characteristics (TMB, TNB and PD-L1) achieved an optimal AUC value of 0.828 in differentiating responders from non-responders to immunotherapy.Conclusion: We conferred the first landscape of five forms of RNA modifications in BCa and emphasized the excellent power of an RNA modifications-related model in evaluating BCa prognosis and immune landscape.

Background: Primary liver cancer, around 75%–85% are hepatocellular carcinoma in China, is the fourth most common malignancy and the second leading cause of tumor-related death, thereby posing a significant threat to the life and health of the Chinese people. Summary: Since the publication of Guidelines for Diagnosis and Treatment of Primary Liver Cancer in China in June 2017, which were updated by the National Health Commission in December 2019, additional high-quality evidence has emerged from researchers worldwide regarding the diagnosis, staging, and treatment of liver cancer, that requires the guidelines to be updated again. The new edition (2022 Edition) was written by more than 100 experts in the field of liver cancer in China, which not only reflects the real-world situation in China, but also may re-shape the nationwide diagnosis and treatment of liver cancer. Key Messages: The new guideline aims to encourage the implementation of evidence-based practice, and improve the national average five-year survival rate for patients with liver cancer, as proposed in the "Health China 2030 Blueprint."

Epstein–Barr virus-associated gastric cancer shows a robust response to immune checkpoint inhibitors. Here the authors introduce a deep convolutional neural network and its fusion with pathologists for predicting it from histopathology.

ObjectiveThis study aimed to develop an artificial intelligence model for predicting the pathological complete response (pCR) to neoadjuvant chemoradiotherapy (nCRT) of locally advanced rectal cancer (LARC) using digital pathological images.BackgroundnCRT followed by total mesorectal excision (TME) is a standard treatment strategy for patients with LARC. Predicting the PCR to nCRT of LARC remine difficulty.Methods842 LARC patients treated with standard nCRT from three medical centers were retrospectively recruited and subgrouped into the training, testing and external validation sets. Treatment response was classified as pCR and non-pCR based on the pathological diagnosis after surgery as the ground truth. The hematoxylin & eosin (H&E)-stained biopsy slides were manually annotated and used to develop a deep pathological complete response (DeepPCR) prediction model by deep learning.ResultsThe proposed DeepPCR model achieved an AUC-ROC of 0.710 (95% CI: 0.595, 0.808) in the testing cohort. Similarly, in the external validation cohort, the DeepPCR model achieved an AUC-ROC of 0.723 (95% CI: 0.591, 0.844). The sensitivity and specificity of the DeepPCR model were 72.6% and 46.9% in the testing set and 72.5% and 62.7% in the external validation cohort, respectively. Multivariate logistic regression analysis showed that the DeepPCR model was an independent predictive factor of nCRT (P=0.008 and P=0.004 for the testing set and external validation set, respectively).ConclusionsThe DeepPCR model showed high accuracy in predicting pCR and served as an independent predictive factor for pCR. The model can be used to assist in clinical treatment decision making before surgery.

Abstract The development of deep learning and open access to a substantial collection of imaging data together provide a potential solution for computational image transformation, which is gradually changing the landscape of optical imaging and biomedical research. However, current implementations of deep learning usually operate in a supervised manner, and their reliance on laborious and error-prone data annotation procedures remains a barrier to more general applicability. Here, we propose an unsupervised image transformation to facilitate the utilization of deep learning for optical microscopy, even in some cases in which supervised models cannot be applied. Through the introduction of a saliency constraint, the unsupervised model, named Unsupervised content-preserving Transformation for Optical Microscopy (UTOM), can learn the mapping between two image domains without requiring paired training data while avoiding distortions of the image content. UTOM shows promising performance in a wide range of biomedical image transformation tasks, including in silico histological staining, fluorescence image restoration, and virtual fluorescence labeling. Quantitative evaluations reveal that UTOM achieves stable and high-fidelity image transformations across different imaging conditions and modalities. We anticipate that our framework will encourage a paradigm shift in training neural networks and enable more applications of artificial intelligence in biomedical imaging.

Background: Primary liver cancer, around 90% are hepatocellular carcinoma in China, is the fourth most common malignancy and the second leading cause of tumor-related death, thereby posing a significant threat to the life and health of the Chinese people. Summary: Since the publication of Guidelines for Diagnosis and Treatment of Primary Liver Cancer (2017 Edition) in 2018, additional high-quality evidence has emerged with relevance to the diagnosis, staging, and treatment of liver cancer in and outside China that requires the guidelines to be updated. The new edition (2019 Edition) was written by more than 70 experts in the field of liver cancer in China. They reflect the real-world situation in China regarding diagnosing and treating liver cancer in recent years. Key Messages: Most importantly, the new guidelines were endorsed and promulgated by the Bureau of Medical Administration of the National Health Commission of the People’s Republic of China in December 2019.

Abstract Background Metaplastic breast carcinoma (MBC) is a rare histological type of breast cancer, which commonly shows resistance to standard therapies and is associated with poor prognosis. The immune microenvironment in MBC and its significance has not been well established due to its low incurrence rate and complex components. We aimed to investigate the diversity of immune parameters including subsets of TILs and PDL1/PD1 expression in MBC, as well as its correlation with prognosis. Methods A total of 60 patients diagnosed with MBC from January 2006 to December 2017 were included in our study. The percentage (%) and quantification (per mm2) of TILs and presence of tertiary lymphoid structures (TLS) were evaluated by hematoxylin and eosin staining (HE). The quantification of CD4+, CD8+ TILs (per mm2), and PD-1/PDL1 expression were evaluated through immunohistochemistry and analyzed in relation to clinicopathological characteristics. A ≥ 1% membranous or cytoplasmatic expression of PD1 and PDL1 was considered a positive expression. Results We found squamous cell carcinoma MBC (33/60, 55%) exhibiting most TILs of all the MBC subtypes (p = 0.043). Thirty-three of 60 (50%) of the patients had coexisting invasive ductal carcinoma of no special type (IDC-NST), and the average percentage of TILs in MBC components was lower compared with NST components (p 300/mm2, 20%), CD4+ TILs (> 250/mm2), and CD8+ TILs (> 70/mm2) in MBC were found associated with longer disease-free survival. Positive expression of PDL1 in tumor cells (≥ 1%) and PD1 in stromal cells (≥ 1%) were also associated with longer survival. Conclusions The immune characteristics differ in various subtypes as well as components of MBC. Immune parameters are key predictive factors of MBC and provide the clinical significance of applying immune checkpoint therapies in patients with MBC.

Background: Hepatocellular carcinoma (HCC) is the most common type of liver cancer and causes many cancer-related deaths worldwide; in China it is the second most prevalent cause of cancer deaths. Most patients are diagnosed clinically with advanced stage disease. Summary: For more than a decade, sorafenib, a small molecular weight tyrosine kinase-inhibitor (SMW TKI) was the only molecular targeted drug available with a survival benefit for the treatment of advanced HCC. With the development of novel TKIs and immune checkpoint inhibitors (ICIs) for advanced HCC, the management of patients has been greatly improved. However, though angiogenic-based targeted therapy remains the backbone for the systemic treatment of HCC, to date no Chinese guidelines for novel molecular targeted therapies to treat advanced HCC have been established. Our interdisciplinary panel on the treatment of advanced HCC comprising hepatologists, hepatobiliary surgeons, oncologists，radiologists，pathologists, orthopedic surgeons, traditional Chinese medicine physicians and interventional radiologists have reviewed the literature in order to develop updated treatment regimens. Key Messages: Panel consensus statements for the appropriate use of new molecular targeted drugs including doses, combination therapies, adverse reaction management as well as efficacy evaluation and predictions for treatment of advanced HCC with evidence levels based on published data are presented, thereby providing an overview of molecular targeted therapies for healthcare professionals.

An amendment to this paper has been published and can be accessed via the original article.

Background: Tumor-infiltrating lymphocytes (TILs) are clinically significant in triple-negative breast cancer (TNBC). Although a standardized methodology for visual TILs assessment (VTA) exists, it has several inherent limitations. We established a deep learning-based computational TIL assessment (CTA) method broadly following VTA guideline and compared it with VTA for TNBC to determine the prognostic value of the CTA and a reasonable CTA workflow for clinical practice. Methods: We trained three deep neural networks for nuclei segmentation, nuclei classification and necrosis classification to establish a CTA workflow. The automatic TIL (aTIL) score generated was compared with manual TIL (mTIL) scores provided by three pathologists in an Asian (n = 184) and a Caucasian (n = 117) TNBC cohort to evaluate scoring concordance and prognostic value. Findings: The intraclass correlations (ICCs) between aTILs and mTILs varied from 0.40 to 0.70 in two cohorts. Multivariate Cox proportional hazards analysis revealed that the aTIL score was associated with disease free survival (DFS) in both cohorts, as either a continuous [hazard ratio (HR)=0.96, 95% CI 0.94–0.99] or dichotomous variable (HR=0.29, 95% CI 0.12–0.72). A higher C-index was observed in a composite mTIL/aTIL three-tier stratification model than in the dichotomous model, using either mTILs or aTILs alone. Interpretation: The current study provides a useful tool for stromal TIL assessment and prognosis evaluation for patients with TNBC. A workflow integrating both VTA and CTA may aid pathologists in performing risk management and decision-making tasks.

Abstract Background Kinesins play important roles in the development and progression of many human cancers. The functions and underlying mechanisms of kinesin family member C1 (KIFC1), a member of the kinesin-14 family, in the pathogenesis of hepatocellular carcinoma (HCC) have not been fully elucidated. Methods In this study, 168 HCC samples were first analyzed to examine the association between KIFC1 expression and patient clinicopathological features and prognosis. The role of KIFC1 in HCC cell proliferation and metastasis was investigated both in vivo and in vitro. The upstream regulation and downstream targets of KIFC1 were studied by qRT-PCR, western blotting, coimmunoprecipitation, chromatin immunoprecipitation (ChIP) and dual-luciferase reporter assays. Results KIFC1 was highly expressed in HCC tissues and positively associated with advanced stages and poor prognosis. KIFC1 knockdown suppressed HCC cell proliferation and invasion both in vitro and in vivo. Furthermore, KIFC1 knockdown decreased invadopodia formation and reduced epithelial-mesenchymal transition (EMT). HMGA1, an architectural transcriptional factor, was identified to interact with KIFC1. HMGA1 could bind to the promoters of Stat3, MMP2 and EMT-related genes and promote gene transcription. KIFC1 enhanced HMGA1 transcriptional activity and facilitated HCC proliferation and invasion. Moreover, KIFC1 was activated by TCF-4, and KIFC1 inhibition enhanced HCC cell sensitivity to paclitaxel. Conclusions Our findings suggest that KIFC1, activated by TCF-4, functions as an oncogene and promotes HCC pathogenesis through regulating HMGA1 transcriptional activity and that KIFC1 is a potential therapeutic target to enhance the paclitaxel sensitivity of HCC.

Abstract Whether the association between body size or shape and nasopharyngeal carcinoma (NPC) risk exists or varies by age‐specific body size indicators is unclear. In a population‐based case–control study conducted in Southern China between 2010 and 2014, self‐reported height, weight, and body shape at age 20 and 10 years before interview were collected from 2448 histopathologically confirmed NPC cases and 2534 population‐based controls. Body mass index (BMI) was categorized according to the World Health Organization guidelines for Asian populations: underweight (

The Epstein–Barr virus (EBV) can cause different types of cancer in human beings when the virus infects different cell types with various latent patterns. EBV shapes a distinct and immunosuppressive tumor microenvironment (TME) to its benefit by influencing and interacting with different components in the TME. Different EBV-associated malignancies adopt similar but slightly specific immunosuppressive mechanisms by encoding different EBV products to escape both innate and adaptive immune responses. Strategies reversing the immunosuppressive TME of EBV-associated malignancies have been under evaluation in clinical practice. As the interactions among EBV, tumor cells, and TME are intricate, in this review, we mainly discuss the epidemiology of EBV, the life cycle of EBV, the cellular and molecular composition of TME, and a landscape of different EBV-associated malignancies and immunotherapy by targeting the TME.

In the original publication of this article [1], the indicated stages (I-II III-IV) were missing in the clinical stage part at both Table 1 and Table 2.

This study aimed to investigate the expression pattern and prognostic value of friend leukemia virus integration 1 (FLI-1) in nasopharyngeal carcinoma (NPC). Immunohistochemistry (IHC) staining of FLI-1 was performed in specimens from 198 untreated NPC patients. Ninety-nine patients were randomly assigned to the training set to analyze the prognostic value of FLI-1 and other clinicopathological characteristics, while the others were assigned to the testing set for validation. Clinicopathological data were compared using the Pearson chi-square test. Univariate and multivariate analyses were performed using the Cox proportional hazards model to test independent prognostic factors and calculate the hazard ratio (HR) and 95% confidence interval (CI). Cytoplasmic FLI-1 expression positively correlated with N stage, distant metastasis and death (P

BackgroundThe metastasis-associated in colon cancer 1 gene (MACC1) has been found to be associated with cancer development and progression. The aim of this study was to investigate the prognostic value of MACC1 in early-stage and AFP-normal hepatocellular carcinoma (HCC).MethodsmRNA and protein levels of MACC1 expression in one normal liver epithelial cells THLE3 and 15 HCC cell lines were examined using reverse transcription-PCR and Western blot. MACC1 expression was also comparatively studied in 6 paired HCC lesions and the adjacent non-cancerous tissue samples. Immunohistochemistry was employed to analyze MACC1 expression in 308 clinicopathologically characterized HCC cases. Statistical analyses were applied to derive association between MACC1 expression scores and clinical staging as well as patient survival.ResultsLevels of MACC1 mRNA and protein were higher in HCC cell lines and HCC lesions than in normal liver epithelial cells and the paired adjacent noncancerous tissues. Significant difference in MACC1 expression was found in patients of different TNM stages (PConclusionsMACC1 protein may represent a promising biomarker for predicting the prognosis of HCC, including in early-stage and AFP-normal patients.

Fibulin-3, originally identified in senescent and Werner syndrome fibroblasts, has been implicated in cell morphology, growth, adhesion and motility. Fibulin-3 exhibits both antitumor and oncogenic activities towards human cancers; however, the role of Fibulin-3 in hepatocellular carcinoma (HCC) remains elusive. In this study, we showed that both the mRNA and protein levels of Fibulin-3 were remarkably downregulated in HCC cell lines and fresh tissues. Immunohistochemical data revealed that Fibulin-3 was decreased in tumorous tissues in 67.1% (171/255) of cases compared to the corresponding adjacent nontumorous tissues. The results of statistical analysis indicated that low Fibulin-3 expression, defined by the receiver operating characteristic curve (ROC), was significantly associated with tumor differentiation (P=0.008), clinical stage (P=0.014) and serum AFP levels (P

Liver cancer ranks in prevalence and mortality among top five cancers worldwide. Accumulating interests have been focused in developing new strategies for liver cancer treatment. We have previously showed that dihydroartemisinin (DHA) exhibited antitumor activity towards liver cancer. In this study, we demonstrated that histone deacetylase inhibitors (HDACi) significantly augmented the antineoplastic effect of DHA via increasing apoptosis in vitro and in vivo. Inhibition of ERK phosphorylation contributed to DHA-induced apoptosis, due to the fact that inhibitor of ERK phosphorylation (PD98059) increased DHA-induced apoptosis. Compared with DHA alone, the combined treatment with DHA and HDACi reduced mitochondria membrane potential, released cytochrome c into cytoplasm, increased p53 and Bak, decreased Mcl-1 and p-ERK, activated caspase 3 and PARP, and induced apoptotic cells. Furthermore, we showed that HDACi pretreatment facilitated DHA-induced apoptosis. In Hep G2-xenograft carrying nude mice, the intraperitoneal injection of DHA and SAHA resulted in significant inhibition of xenograft tumors. Results of TUNEL and H&E staining showed more apoptosis induced by combined treatment. Immunohistochemistry data revealed the activation of PARP, and the decrease of Ki-67, p-ERK and Mcl-1. Taken together, our data suggest that the combination of HDACi and DHA offers an antitumor effect on liver cancer, and this combination treatment should be considered as a promising strategy for chemotherapy.

Polo-like kinase 4 (PLK4), belonging to serine/threonine kinase family, is critical for centriole replication and cell cycle progression. PLK4 has been proposed as a tumor suppressor in hepatocellular carcinoma (HCC). However, its expression and significance in HCC have not been well studied. In the present study, we found that PLK4 was markedly downregulated in both HCC cell lines and fresh cancer tissues, using quantitative real-time-PCR and western blot. Immunohistochemistry data also revealed that decreased expression of PLK4 was present in 72.4% (178/246) of HCC tissues, compared with the corresponding adjacent nontumorous tissues. Furthermore, PLK4 expression significantly correlated with clinicopathological parameters, including clinical stage (P=0.034), serum α-fetoprotein (AFP) (P=0.019) and tumor size (P=0.032). Moreover, HCC patients with low PLK4 expression survived shorter than those with high PLK4 expression, as indicated by overall survival (P=0.002) and disease-free survival (P=0.012) assessed by the Kaplan-Meier method. In addition, multivariate analysis suggested PLK4 as an independent predictor of overall survival (HR, 0.556; 95%CI, 0.376-0.822; P=0.003) and disease-free survival (HR, 0.547; 95%CI, 0.382-0.783; P=0.001). Collectively, our study demonstrated that PLK4 was remarkably downregulated in HCC and could be served as a potential prognostic marker for patients with this deadly disease.

SIRT3, a mitochondrial sirtuin belonging to nicotinamide adenine nucleotide (NAD) dependent deacetylases, is implicated in metabolism, longevity and carcinogenesis. SIRT3 expression and its significance in hepatocellular carcinoma (HCC) remain largely unclear. In this study, we demonstrated that SIRT3 expression in HCC tissue was much lower than that in paracarcinoma tissue, at both mRNA and protein levels. The cutoff value for low SIRT3 expression in HCC was defined according to receiver operating characteristic curve (ROC) analysis. As disclosed by immunohistochemistry (IHC) results, low SIRT3 expression was present in 67.3% (167/248) of HCC cases. Furthermore, low expression of SIRT3 was significantly correlated to differentiation (P = 0.013), clinical stage (P = 0.005), serum AFP level (P

The endocannabinoid system (ECS) is dysregulated in various liver diseases. Previously, we had shown that the major endocannabinoid 2-arachidonoyl glycerol (2-AG) promoted tumorigenesis of intrahepatic cholangiocarcinoma (ICC). However, biosynthesis regulation and clinical significance of 2-AG remain elusive. In the present study, we quantified 2-AG by gas chromatography/mass spectrometry (GC/MS) and showed that 2-AG was enriched in patients with ICC samples as well as in thioacetamide-induced orthotopic rat ICC model. Moreover, we found that diacylglycerol lipase β (DAGLβ) was the principal synthesizing enzyme of 2-AG that significantly upregulated in ICC. DAGLβ promoted tumorigenesis and metastasis of ICC in vitro and in vivo and positively correlated with clinical stage and poor survival in patients with ICC. Functional studies showed that activator protein-1 (AP-1; heterodimers of c-Jun and FRA1) directly bound to the promoter and regulated transcription of DAGLβ, which can be enhanced by lipopolysaccharide (LPS). miR-4516 was identified as the tumor-suppressing miRNA of ICC that can be significantly suppressed by LPS, 2-AG, or ectopic DAGLβ overexpression. FRA1 and STAT3 were targets of miR-4516 and overexpression of miRNA-4516 significantly suppressed expression of FRA1, SATA3, and DAGLβ. Expression of miRNA-4516 was negatively correlated with FRA1, SATA3, and DAGLβ in patients with ICC samples. Our findings identify DAGLβ as the principal synthesizing enzyme of 2-AG in ICC. DAGLβ promotes oncogenesis and metastasis of ICC and is transcriptionally regulated by a novel AP-1/DAGLβ/miR4516 feedforward circuitry. [ABSTRACT FROM AUTHOR]