Progranulin mediates immune evasion of pancreatic ductal adenocarcinoma through regulation of MHCI expression.

Immune evasion is indispensable for cancer initiation and progression, although its underlying mechanisms in pancreatic ductal adenocarcinoma (PDAC) are not fully known. Here, we characterize the function of tumor-derived PGRN in promoting immune evasion in primary PDAC. Tumor- but not macrophage-derived PGRN is associated with poor overall survival in PDAC. Multiplex immunohistochemistry shows low MHC class I (MHCI) expression and lack of CD8+ T cell infiltration in PGRN-high tumors. Inhibition of PGRN abrogates autophagy-dependent MHCI degradation and restores MHCI expression on PDAC cells. Antibody-based blockade of PGRN in a PDAC mouse model remarkably decelerates tumor initiation and progression. Notably, tumors expressing LCMV-gp33 as a model antigen are sensitized to gp33-TCR transgenic T cell-mediated cytotoxicity upon PGRN blockade. Overall, our study shows a crucial function of tumor-derived PGRN in regulating immunogenicity of primary PDAC.

Contaminated and misidentified cell lines commonly use in cancer research.

Cell line authentication is important for credibility concern and scientific reproducibility. Authenticated cancer cell lines retain the properties of the cancers of origin and serve valuable resources for medical research. Experimental results commonly will be validated in more than one cell line to avoid specific cell line effect not generalizable to the disease on the whole. The use of appropriate and verified cell lines would therefore be very important in preclinical studies of translational research, bridging basic research to clinics.

Clinical application of Albumin-Bilirubin (ALBI) score: The current status.

Prognosis of patients that have undergone liver resection can be predicted preoperatively. Albumin-Bilirubin (ALBI) is a useful score to evaluate liver function objectively. This score is simple to use, uses objective parameters, and can stratify patients into grades. In combination with other liver cancer scores, the prognosis and survival of patients can be evaluated with more accuracy. Current literatures supporting the use of the ALBI score relating to prognosis of post hepatectomy, radiofrequency ablation (RFA), transarterial chemoembolization (TACE), radiotherapy and systemic therapy are available. Therefore, ALBI score is worthy of clinical application. The following is a review of the current literatures on the ALBI score.

STAT3: An Emerging Therapeutic Target for Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is a major global health problem and its treatment options have been limited. Signal transducer and activator of transcription 3 (STAT3) is a transcription factor important for various cellular processes. Overexpression and constitutive activation of STAT3 have been frequently found in HCC and associated with poor prognosis. Ample evidence has shown that STAT3 plays pivotal roles in the initiation, progression, metastasis and immune suppression of HCC. Thus, STAT3 has attracted attention as a novel therapeutic target in HCC. Clinical trials have investigated STAT3-targeted therapeutics either as monotherapy or in combination with chemotherapeutic agents, immune checkpoint inhibitors and alternative targeted drugs. Some of these studies have yielded encouraging results. Particularly, napabucasin-a cancer stemness inhibitor targeting STAT3-driven gene transcription-has stood out with its promising clinical efficacy and safety profile. Nonetheless, clinical investigations of STAT3-targeted therapies in HCC are limited and more efforts are strongly urged to evaluate their clinical performance in HCC. Here, we provide a comprehensive review of the roles of STAT3 in HCC and follow by comprehensive analysis of STAT3 targeted strategies.

The ATP-binding cassette transporter ABCF1 is a hepatic oncofetal protein that promotes chemoresistance, EMT and cancer stemness in hepatocellular carcinoma.

ATP-binding cassette (ABC) transporters mediate multidrug resistance and cancer stem cell properties in various model systems. Yet, their biological significance in cancers, especially in hepatocellular carcinoma (HCC), remains unclear. In this study, we investigated the function of ABCF1 in HCC and explored its potential as a therapeutic target. ABCF1 was highly expressed in fetal mouse livers but not in normal adult livers. ABCF1 expression was upregulated in HCCs. These results demonstrate that ABCF1 functions as a hepatic oncofetal protein. We further demonstrated elevated ABCF1 expression in HCC cells upon acquiring chemoresistance. Suppression of ABCF1 by siRNA sensitized both parental cells and chemoresistant cells to chemotherapeutic agents. Reversely, ABCF1 overexpression promoted chemoresistance and drug efflux. In addition, overexpression of ABCF1 enhanced migration, spheroid and colony formation and epithelial-mesenchymal transition (EMT) induction. RNA sequencing analysis revealed EMT inducers HIF1α/IL8 and Sox4 as potential mediators for the oncogenic effect of ABCF1 in HCC progression. Together, this study illustrates that ABCF1 is a novel potential therapeutic target for HCC treatment.

C-terminal truncated HBx protein activates caveolin-1/LRP6/ß-catenin/FRMD5 axis in promoting hepatocarcinogenesis.

Hepatitis B virus X protein mutants, particularly truncated at C-terminal (HBxΔC), generated during random viral integration, are frequently detected in hepatocellular carcinoma (HCC) and exert a more potent oncogenic effect than full-length form (FL). Here, we showed that caveolin-1 (Cav1), a robust metastasis promoter, is transcriptionally upregulated by HBxΔC but not by FL HBx. Promoting effect of HBxΔC in HCC cell aggressiveness is abolished when Cav1 is suppressed. Expression profiling identified FERM domain containing 5 (FRMD5) protein as a downstream target of Cav1. In accordance with the regulation of Cav1, HBxΔC upregulates FRMD5. Knockdown of FRMD5 in HBxΔC cells recapitulated the functional effect of Cav1 knockdown in HBxΔC cells. The regulation of FRMD5 by HBxΔC-induced Cav1 is mediated by the protein stablilization of LRP6 leading to the activation of ß-catenin. Expression of a constitutively active ß-catenin in Cav1 knockdown cells rescued FRMD5 expression and HCC tumorigenesis and metastasis. Clinical relevance of HBxΔC/Cav1/LRP6/FRMD5 pathway is demonstrated by the significant correlation of Cav1, LRP6 and FRMD5 expressions in HCC. The findings of this study uncover a novel HBxΔC-regulated molecular pathway which has profound implications in HCC therapeutics.

Establishment and characterization of new tumor xenografts and cancer cell lines from EBV-positive nasopharyngeal carcinoma.

The lack of representative nasopharyngeal carcinoma (NPC) models has seriously hampered research on EBV carcinogenesis and preclinical studies in NPC. Here we report the successful growth of five NPC patient-derived xenografts (PDXs) from fifty-eight attempts of transplantation of NPC specimens into NOD/SCID mice. The take rates for primary and recurrent NPC are 4.9% and 17.6%, respectively. Successful establishment of a new EBV-positive NPC cell line, NPC43, is achieved directly from patient NPC tissues by including Rho-associated coiled-coil containing kinases inhibitor (Y-27632) in culture medium. Spontaneous lytic reactivation of EBV can be observed in NPC43 upon withdrawal of Y-27632. Whole-exome sequencing (WES) reveals a close similarity in mutational profiles of these NPC PDXs with their corresponding patient NPC. Whole-genome sequencing (WGS) further delineates the genomic landscape and sequences of EBV genomes in these newly established NPC models, which supports their potential use in future studies of NPC.

Novel biomarkers GEP/ABCB5 regulate response to adjuvant transarterial chemoembolization after curative hepatectomy for hepatocellular carcinoma.

BACKGROUND: Transarterial chemoembolization (TACE) is the most commonly used adjuvant therapy for hepatocellular carcinoma (HCC) after curative resection. Responses to TACE are variable due to tumor and patient heterogeneity. We had previously demonstrated that expression of Granulin-epithelin precursor (GEP) and ATP-dependent binding cassette (ABC)B5 in liver cancer stem cells was associated with chemoresistance. The present study aimed to evaluate the association between GEP/ABCB5 expression and response to adjuvant TACE after curative resection for HCC. METHODS: Patients received adjuvant TACE after curative resection for HCC and patients received curative resection alone were identified from a prospectively collected database. Clinical samples were retrieved for biomarker analysis. Patients were categorized into 3 risk groups according to their GEP/ABCB5 status for survival analysis: low (GEP-/ABCB5-), intermediate (either GEP+/ABCB5- or GEP-/ABCB5+) and high (GEP+/ABCB5+). Early recurrence (recurrence within 2 years after resection) and disease-free survival were analyzed. RESULTS: Clinical samples from 44 patients who had followed-up for more than 2 years were retrieved for further biomarker analysis. Among them, 18 received adjuvant TACE and 26 received surgery alone. Patients with adjuvant TACE in the intermediate risk group was associated with significantly better overall survival and 2-year disease-free survival than those who had surgery alone (P = 0.036 and P = 0.011, respectively). Adjuvant TACE did not offer any significant differences in the early recurrence rate, 2-year disease-free survival and overall survival for patients in low and high risk groups. CONCLUSIONS: Adjuvant TACE can only provide survival benefits for patients in the intermediate risk group (either GEP+/ABCB5- or GEP-/ABCB5+). A larger clinical study is warranted to confirm its role in patient selection for adjuvant TACE.

A Brief Overview of Progranulin in Health and Disease.

The purpose of this brief overview of the progranulin protein is to provide a sense of the range and extent of the roles of progranulin in normal physiology and pathology. Progranulin has received attention due to its role in neurodegeneration, where mutation of a single copy of GRN, the gene encoding progranulin, results in frontotemporal dementia, whereas viral delivery of progranulin to the brains of mice exhibiting Parkinson's or Alzheimer's disease phenotypes inhibits the progression of the neurodegenerative phenotypes. Of equal importance, progranulin protects tissues against the harmful effects of poorly controlled inflammation and promotes tissue regeneration after injury at a multitude of sites throughout the body. Progranulin is overexpressed by many types of cancer and contributes to their progression. Given suitable analytical methods and model systems, progranulin offers a wealth of research possibilities.

Methods to Analyze the Role of Progranulin (PGRN/GEP) on Cancer Stem Cell Features.

Emerging evidence suggests that tumors are hierarchically organized with a distinct subpopulation called cancer stem cells (CSCs) lying at the apex of the hierarchy. These cells are not only responsible for tumor initiation and progression but also endowed with stem cell properties, including self-renewal, chemoresistance, and tumor initiation. Although existing therapies can initially eliminate the bulk population of tumor, the stem cell properties of CSCs enable them to survive and repopulate the tumor, resulting in disease relapse. Recently, our group has shown that progranulin (PGRN/GEP) defined a hepatic cancer stem cell subpopulation in hepatocellular carcinoma. This subpopulation demonstrated enhanced ability for colony and spheroid formation, chemoresistance, and tumor initiation. In this chapter, we describe the methods used to isolate the progranulin+ subpopulation and analyze their CSC properties.

Mouse Monoclonal Antibodies Against Progranulin (PGRN/GEP) as Therapeutics in Preclinical Cancer Models.

The use of monoclonal antibody (mAb) has become a unique means of targeted therapy for human cancers. mAb-based therapies have shown survival benefits by applying alone or in combination with chemotherapeutics. Being a humanized biomolecule with exquisite target specificity, mAb demonstrated effects in a relatively lower dose range with limited off-target harm to the patients. Nowadays, novel targets involved in tumorigenic mechanisms and biomarkers expressed exclusively on cancer cell surface are being constantly discovered. The potential effects of their specific mAb could be investigated in the preclinical cancer model. In this chapter, we outlined our experimental procedures in determining the feasibility of novel mAb in the preclinical cancer model, with an example of progranulin (PGRN/GEP) mAb against hepatocellular carcinoma (HCC) tumor in mouse model. This chapter included the establishment of subcutaneous and orthotopic HCC tumor in mouse model, the injection of the mouse monoclonal antibody in combination with cytotoxic chemotherapeutics, the assessment of tumor development, and the analyses of the molecular changes of the tumor cells.

Granulin epithelin precursor promotes colorectal carcinogenesis by activating MARK/ERK pathway.

BACKGROUND: Granulin epithelin precursor (GEP) is reported to function as a growth factor stimulating proliferation and migration, and conferring chemoresistance in many cancer types. However, the expression and functional roles of GEP in colorectal cancer (CRC) remain elusive. The aim of this study was thus to investigate the clinical significance of GEP in CRC and reveal the molecular mechanism of GEP in CRC initiation and progression. METHODS: The mRNA expression of GEP in CRC cell lines were detected by qRT-PCR. The GEP protein expression was validated by immunohistochemistry in tissue microarray (TMA) including 190 CRC patient samples. The clinicopathological correlation analysis were achieved by GEP expression on TMA. Functional roles of GEP were determined by MTT proliferation, monolayer colony formation, cell invasion and migration and in vivo studies through siRNA/shRNA mediated knockdown assays. The cancer signaling pathway identification was acquired by flow cytometry, western blot and luciferase activity assays. RESULTS: The mRNA expression of GEP in CRC was significantly higher than it in normal colon tissues. GEP protein was predominantly localized in the cytoplasm and most of the CRC cases demonstrated abundant GEP protein compared with non-tumorous tissues. GEP overexpression was associated with non-rectal location, advanced AJCC stage, regional lymph node and distant metastasis. By Kaplan-Meier survival analysis, GEP abundance served as a prognostic marker for worse survival in CRC patients. GEP knockdown exhibited anti-cancer effect such as inhibiting cell proliferation, monolayer colony formation, cell invasion and migration in DLD-1 and HCT 116 cells and decelerating xenograft formation in nude mice. siGEP also induced G1 cell cycle arrest and apoptosis. Luciferase activity assays further demonstrated GEP activation was involved in MAPK/ERK signaling pathway. CONCLUSION: In summary, we compressively delineate the oncogenic role of GEP in colorectal tumorigenesis by activating MAPK/ERK signaling pathway. GEP might serve as a useful prognostic biomarker and therapeutic target for CRC.

Treatment of advanced hepatocellular carcinoma: immunotherapy from checkpoint blockade to potential of cellular treatment.

The absence of potent therapeutic option accounts for the dismal prognosis of advanced hepatocellular carcinoma (HCC) with high mortality and recurrence rate. For a decade, sorafenib is the only approved systemic drug in the first-line setting and warrants as the standard-of-care for HCC in the advanced stage. Given the common failures of chemotherapies and targeted therapies in the field of HCC treatment, promising breakthroughs were eagerly needed and until recently, immunotherapies have opened a new era of anticancer treatment. The liver organ is perceived as "immunotolerant" owing to its functional role, and the hepatic immune balance is found to be deregulated during chronic liver inflammation and HCC tumorigenesis. Restoring a competent immunity by mitigation of immunosuppression signals is a contemporary approach. In this regard, novel immune checkpoint inhibitors have revolutionized cancer pharmacological treatment options with remarkable clinical outcomes in hematologic malignancy and multiple solid tumors including advanced HCC. Nivolumab, an immunotherapeutic agent to block programmed cell death protein 1 (PD-1), showed high efficacy potential for patients progressed with sorafenib and granted accelerated approval by the US Food and Drug Administration (FDA) recently. The development of this class of immunotherapeutic drug is currently based on myriad studies established on the role of T-cell mediated immunosuppression through immune checkpoints. Heterogeneous results have led to further explorations to the profile of oncogenic processes and signaling pathways associated with PD1/PD-L1 axis. Emerging evidence from preclinical studies implicate natural killer (NK) cells as a mediator to the PD-1 checkpoint signaling immunoevasion. The strategy of adopting immunomodulating ability of NK cells by immune checkpoints inhibitors is potential to additive effects in stimulating anticancer immunity. This idea is not entirely newfound but has recently gained prominence because of advances in defining phenotypic heterogeneity of NK cell populations. The physiological significance and synergistic value of NK cells await further investigation in clinical trials. In this review, an overview of the treatment paradigm shift of HCC management is presented. Current knowledge concerning immunological mechanisms of immune checkpoints attributed to T cell is further discussed and relevant ongoing clinical trials are summarized. We proposed that NK cells should be viewed as part of the network of checkpoint immunoevasion and delineate current evidence of translational clinical research in this area. It is conceivable that immune checkpoint inhibitors in combination with NK cell-based therapeutic strategies will be great promise for treatment of advanced HCC.

Granulin-epithelin precursor interacts with 78-kDa glucose-regulated protein in hepatocellular carcinoma.

BACKGROUND: Granulin-epithelin precursor (GEP) is a secretory growth factor, which has been demonstrated to control cancer growth, invasion, drug resistance and immune escape. Our previous studies and others also demonstrated its potential in targeted therapy. Comprehensive characterization of GEP partner on cancer cells are warranted. We have previously shown that GEP interacted with heparan sulfate on the surface of liver cancer cells and the interaction is crucial for GEP-mediated signaling transduction. This study aims to characterize GEP protein partner at the cell membrane with the co-immunoprecipitation and mass spectrometry approach. METHODS: The membrane fraction from liver cancer model Hep3B was used for capturing binding partner with the specific monoclonal antibody against GEP. The precipitated proteins were analyzed by mass spectrometry. After identifying the GEP binding partner, this specific interaction was validated in additional liver cancer cell line HepG2 by co-immunoprecipitation using GRP78 and GEP antibodies, respectively, as the bait. GRP78 transcript levels in hepatocellular carcinoma (HCC) clinical samples (n = 77 pairs) were examined by real-time quantitative RT-PCR. GEP and GRP78 protein expressions were investigated by immunohistochemistry on paraffin sections. RESULTS: We identified the GEP-binding protein as 78-kDa glucose-regulated protein (GRP78, also named heat shock 70-kDa protein 5, HSPA5). This interaction was validated in independent HCC cell lines. Increased GRP78 mRNA levels were demonstrated in liver cancer tissues compared with the paralleled liver tissues (t-test, P = 0.002). GRP78 and GEP transcript levels were significantly correlated (Spearman's correlation, P = 0.001), and the proteins were also detectable in the cytoplasm of liver cancer cells by immunohistochemical staining. CONCLUSIONS: GRP78 and GEP are interacting protein partners in liver cancer cells and may play a role in GEP-mediated cancer progression in HCC.

Octamer 4/microRNA-1246 signaling axis drives Wnt/ß-catenin activation in liver cancer stem cells.

Wnt/ß-catenin signaling is activated in CD133 liver cancer stem cells (CSCs), a subset of cells known to be a root of tumor recurrence and therapy resistance in hepatocellular carcinoma (HCC). However, the regulatory mechanism of this pathway in CSCs remains unclear. Here, we show that human microRNA (miRNA), miR-1246, promotes cancer stemness, including self-renewal, drug resistance, tumorigencity, and metastasis, by activation of the Wnt/ß-catenin pathway through suppressing the expression of AXIN2 and glycogen synthase kinase 3ß (GSK3ß), two key members of the ß-catenin destruction complex. Clinically, high endogenous and circulating miR-1246 was identified in HCC clinical samples and correlated with a worse prognosis. Further functional analysis identified octamer 4 (Oct4) to be the direct upstream regulator of miR-1246, which cooperatively drive ß-catenin activation in liver CSCs. CONCLUSION: These findings uncover the noncanonical regulation of Wnt/ß-catenin in liver CSCs by the Oct4/miR-1246 signaling axis, and also provide a novel diagnostic marker as well as therapeutic intervention for HCC. (Hepatology 2016;64:2062-2076).

Comprehensive characterization of the patient-derived xenograft and the paralleled primary hepatocellular carcinoma cell line.

BACKGROUND: Hepatocellular carcinoma (HCC) is an aggressive cancer with high mortality and morbidity worldwide. The limited clinically relevant model has impeded the development of effective HCC treatment strategy. Patient-derived xenograft (PDX) models retain most of the characteristics of original tumors and were shown to be highly predictive for clinical outcomes. Notably, primary cell line models allow in-depth molecular characterization and high-throughput analysis. Combined usage of the two models would provide an excellent tool for systematic study of therapeutic strategies. Here, we comprehensively characterized the novel PDX and the paralleled primary HCC cell line model. METHODS: Tumor tissues were collected from HCC surgical specimens. HCC cells were sorted for in vivo PDX and in vitro cell line establishment by the expression of hepatic cancer stem cell marker to enhance cell viability and the rate of success on subsequent culture. The PDX and its matching primary cell line were authenticated and characterized in vitro and in vivo. RESULTS: Among the successful cases for generating PDXs and primary cells, HCC40 is capable for both PDX and primary cell line establishment, which were then further characterized. The novel HCC40-PDX and HCC40-CL exhibited consistent phenotypic characteristics as the original tumor in terms of HBV protein and AFP expressions. In common with HCC40-PDX, HCC40-CL was tumorigenic in immunocompromised mice. The migration ability in vitro and metastatic properties in vivo echoed the clinical feature of venous infiltration. Genetic profiling by short tandem repeat analysis and p53 mutation pattern consolidated that both the HCC40-PDX and HCC40-CL models were derived from the HCC40 clinical specimen. CONCLUSIONS: The paralleled establishment of PDX and primary cell line would serve as useful models in comprehensive studies for HCC pathogenesis and therapeutics development for personalized treatment.

Overexpression of PIN1 Enhances Cancer Growth and Aggressiveness with Cyclin D1 Induction in EBV-Associated Nasopharyngeal Carcinoma.

BACKGROUND: Nasopharyngeal carcinoma (NPC) is a peculiar Epstein Barr virus (EBV)-associated malignancy that is prevalent in South-East Asia. Peptidyl-prolyl cis-trans isomerase NIMA-interacting 1 (PIN1) isomerizes specific phosphorylated amino acid residues, which makes it an important regulator in cell survival and apoptosis. In this study, we investigated the contribution made by PIN1 in NPC tumorigenesis and PIN1's potential role as a therapeutic target. METHODS: The expression of PIN1 was examined in a panel of NPC cell lines, xenografts and primary tumors. The functional roles of PIN1 in NPC cells were elucidated by the knockdown and overexpression of PIN1 in in vitro and in vivo nude mice models by siRNA and lenti-viral transfection, respectively. The antitumor effects of the PIN1 inhibitor Juglone in NPC cells were also evaluated. RESULTS: We revealed the consistent overexpression of PIN1 in almost all EBV-associated NPC cell lines, xenografts and primary tumors. PIN1 suppression was capable of inhibiting cyclin D1 expression and activating caspase-3 in NPC cells. It positively regulated NPC cell proliferation, colony formation and anchorage-independent growth. The inhibition of PIN1 suppressed tumor growth in vitro and in vivo. CONCLUSIONS: This study demonstrates the oncogenic role of PIN1 in NPC tumorigenesis, and shows that its overexpression can enhance tumor cell growth via the upregulation of cyclinD1. Our findings inform the development of novel treatments targeting PIN1 for NPC patients.

Pancreatic cancer risk variant in LINC00673 creates a miR-1231 binding site and interferes with PTPN11 degradation.

Genome-wide association studies have identified several loci associated with pancreatic cancer risk; however, the mechanisms by which genetic factors influence the development of sporadic pancreatic cancer remain largely unknown. Here, by using genome-wide association analysis and functional characterization, we identify a long intergenic noncoding RNA (lincRNA), LINC00673, as a potential tumor suppressor whose germline variation is associated with pancreatic cancer risk. LINC00673 is able to reinforce the interaction of PTPN11 with PRPF19, an E3 ubiquitin ligase, and promote PTPN11 degradation through ubiquitination, which causes diminished SRC-ERK oncogenic signaling and enhanced activation of the STAT1-dependent antitumor response. A G>A change at rs11655237 in exon 4 of LINC00673 creates a target site for miR-1231 binding, which diminishes the effect of LINC00673 in an allele-specific manner and thus confers susceptibility to tumorigenesis. These findings shed new light on the important role of LINC00673 in maintaining cell homeostasis and how its germline variation might confer susceptibility to pancreatic cancer.

Hepatic cancer stem cell marker granulin-epithelin precursor and ß-catenin expression associate with recurrence in hepatocellular carcinoma.

Granulin-epithelin precursor (GEP) has been demonstrated to confer enhanced cancer stem-like cell properties in hepatocellular carcinoma (HCC) cell line models in our previous studies. Here, we aimed to examine the GEP-expressing cells in relation to the stem cell related molecules and stem-like cell properties in the prospective HCC clinical cohort. GEP protein levels were significantly higher in HCCs than the paralleled non-tumor liver tissues, and associated with venous infiltration. GEPhigh cells isolated from clinical HCC samples exhibited higher levels of stem cell marker CD133, pluripotency-associated signaling molecules ß-catenin, Oct4, SOX2, Nanog, and chemodrug transporter ABCB5. In addition, GEPhigh cells possessed preferential ability to form colonies and spheroids, and enhanced in vivo tumor-initiating ability while their xenografts were able to be serially subpassaged into secondary mouse recipients. Expression levels of GEP and pluripotency-associated genes were further examined in the retrospective HCC cohort and demonstrated significant correlation of GEP with ß-catenin. Notably, HCC patients with high GEP and ß-catenin levels demonstrated poor recurrence-free survival. In summary, GEP-positive HCC cells directly isolated from clinical specimens showed ß-catenin elevation and cancer stem-like cell properties.

Restoration of natural killer activity in hepatocellular carcinoma by treatment with antibody against granulin-epithelin precursor.

Impairment of natural killer (NK) cell activity is an important mechanism of tumor immunoevasion. We have previously shown that expression of granulin-epithelin precursor (GEP) in hepatocellular carcinoma (HCC) cells rendered the cells resistant to NK cell immunosurveillance. Here, we examined whether targeting GEP could rescue NK activity in HCC patients. The current study demonstrated that quantities and activities of NK cells were significantly lower in HCC patients compared with healthy individuals, and were negatively correlated with GEP levels in HCC cells. NK cells demonstrated enhanced expression of the stimulatory receptors natural-killer group 2, member D (NKG2D) and CD69, increased secretion of IFN-γ and perforin, and cytotoxicity against HCC cells upon GEP suppression. Opposite phenotypes of NK cells were observed when GEP was overexpressed in HCC cells. Importantly, GEP blockage by monoclonal antibody A23 restored NK activity in HCC patients and sensitized HCC cells to NK cytotoxicity. Furthermore, A23 induced NK-mediated antibody-dependent cell-mediated cytotoxicity against HCC. In summary, the activity of NK cells in HCC was impaired by GEP expression, which could be rescued by GEP antibody. This study provides new insight for treatments targeting GEP to boost NK activity in HCC patients.