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Hepatocellular carcinoma (HCC) is the third most frequent cause of cancer-related death. The immune-rich contexture of the HCC microenvironment makes this tumour an appealing target for immune-based therapies. Here, we discuss how the functional characteristics of the liver microenvironment can potentially be harnessed for the treatment of HCC. We will review the evidence supporting a therapeutic role for vaccines, cell-based therapies and immune-checkpoint inhibitors and discuss the potential for patient stratification in an attempt to overcome the series of failures that has characterised drug development in this disease area.

PURPOSE: To evaluate the potential differences in non-target embolization and vessel microsphere filling of a reflux-control microcatheter (RCM) compared to a standard end-hole microcatheter (SEHM) in a swine model. MATERIALS AND METHODS: Radiopaque microspheres were injected with both RCM and SEHM (2.4-Fr and 2.7-Fr) in the kidneys of a preclinical swine model. Transarterial renal embolization procedures with RCM or SEHM were performed in both kidneys of 14 pigs. Renal arteries were selectively embolized with an automated injection protocol of radio-opaque microspheres. Ex-vivo X-ray microtomography images of the kidneys were utilized to evaluate the embolization by quantification of the deposition of injected microspheres in the target vs. the non-target area of injection. X-ray microtomography images were blindly analyzed by five interventional radiologists. The degree of vessel filling and the non-target embolization were quantified using a scale from 1 to 5 for each parameter. An analysis of variance was used to compare the paired scores. RESULTS: Total volumes of radio-opaque microspheres injected were similar for RCM (11.5±3.6 [SD] mL; range: 6-17mL) and SEHM (10.6±5.2 [SD] mL; range: 4-19mL) (P=0.38). The voxels enhanced ratio in the target (T) vs. non-target (NT) areas was greater with RCM (T=98.3% vs. NT=1.7%) than with SEHM (T=89% vs. NT=11%) but the difference was not significant (P=0.30). The total score blindly given by the five interventional radiologists was significantly different between RCM (12.3±2.1 [SD]; range: 6-15) and the standard catheter (11.3±2.5 [SD]; range: 4-15) (P=0.0073), with a significant decrease of non-target embolization for RCM (3.8±1.3 [SD]; range: 3.5-4.2) compared to SEHM (3.2±1.5 [SD]; range: 2.9-3.5) (P=0.014). CONCLUSION: In an animal model, RCM microcatheters reduce the risk of non-target embolization from 11% to 1.7%, increasing the delivery of microspheres of 98% to the target vessels, compared to SEHM microcatheters. ispartof: DIAGNOSTIC AND INTERVENTIONAL IMAGING vol:102 issue:10 pages:641-648 ispartof: location:France status: published

Hepatocellular carcinoma (HCC) can have viral or non-viral causes1–5. Non-alcoholic steatohepatitis (NASH) is an important driver of HCC. Immunotherapy has been approved for treating HCC, but biomarker-based stratification of patients for optimal response to therapy is an unmet need6,7. Here we report the progressive accumulation of exhausted, unconventionally activated CD8+PD1+ T cells in NASH-affected livers. In preclinical models of NASH-induced HCC, therapeutic immunotherapy targeted at programmed death-1 (PD1) expanded activated CD8+PD1+ T cells within tumours but did not lead to tumour regression, which indicates that tumour immune surveillance was impaired. When given prophylactically, anti-PD1 treatment led to an increase in the incidence of NASH–HCC and in the number and size of tumour nodules, which correlated with increased hepatic CD8+PD1+CXCR6+, TOX+, and TNF+ T cells. The increase in HCC triggered by anti-PD1 treatment was prevented by depletion of CD8+ T cells or TNF neutralization, suggesting that CD8+ T cells help to induce NASH–HCC, rather than invigorating or executing immune surveillance. We found similar phenotypic and functional profiles in hepatic CD8+PD1+ T cells from humans with NAFLD or NASH. A meta-analysis of three randomized phase III clinical trials that tested inhibitors of PDL1 (programmed death-ligand 1) or PD1 in more than 1,600 patients with advanced HCC revealed that immune therapy did not improve survival in patients with non-viral HCC. In two additional cohorts, patients with NASH-driven HCC who received anti-PD1 or anti-PDL1 treatment showed reduced overall survival compared to patients with other aetiologies. Collectively, these data show that non-viral HCC, and particularly NASH–HCC, might be less responsive to immunotherapy, probably owing to NASH-related aberrant T cell activation causing tissue damage that leads to impaired immune surveillance. Our data provide a rationale for stratification of patients with HCC according to underlying aetiology in studies of immunotherapy as a primary or adjuvant treatment., In hepatocellular carcinoma driven by non-alcoholic steatohepatitis, aberrant T cell activation and impaired immune surveillance seem to make hepatocellular carcinoma less responsive to anti-PD1 or anti-PDL1 immunotherapy.

Background & Aims Although HBV is a major cause of death in Africa, its genetic variability has been poorly documented. This study aimed to address whether HBV genotype and surface gene variants are associated with HBV-related liver disease in The Gambia. Methods We conducted a case-control study nested in the Prevention of Liver Fibrosis and Cancer in Africa programme. Consecutive treatment-naive patients with chronic HBV infection and detectable viral load were recruited: 211 controls with no significant liver disease and 91 cases (56 cirrhosis and 35 HCC cases). HBV genotypes and surface gene variants were determined by Sanger sequencing or next-generation sequencing (NGS) in serum DNA. Aflatoxin B1 (AFB1)-specific codon 249 TP53 mutation was determined by NGS in circulating cell-free plasma DNA. Results In phylogenetic analysis, 85% of individuals carried HBV genotype E, 14% genotype A, and 1% A/E recombinant viruses. Surface gene variants were more frequently observed in cases (43% and 57% in cirrhosis and HCC cases, respectively) than controls (25%; p 2,000 IU/ml (OR 22.7 [8.0–64.9]), HBsAg levels, Graphical abstract, Highlights • The rarer HBV genotype A (vs. E) is associated with a higher risk of HCC (odds ratio 5.2). • A genotype-independent deletion hotspot was found in the preS2 domain (preS2Δ38–55). • PreS2Δ38–55 was significantly associated with HCC and is an independent risk factor.

Background Turkey is an intermediate hepatitis B virus (HBV) endemic country. However, prevalence among Turkish migrants in Belgium is unknown, especially in those born in Belgium with a foreign-born parent, i.e. second-generation migrants (SGM). Aims To evaluate the prevalence of HBV infection and associated risk factors in Turkish first-generation migrants (FGM), i.e. foreign-born, and SGM. Methods Between September 2017 and May 2019, free outreach testing for hepatitis B surface antigen (HBsAg), hepatitis B core antibodies (anti-HBc), and antibodies against HBsAg was offered to Turkish migrants in Middle-Limburg, Belgium. Face-to-face questionnaire assessed HBV risk factors. HBsAg positive patients were referred and followed up. Turkish SGM were stratified into birth cohort born before and after 1987, since those born after 1987 should be covered by the universal infant vaccination program. Results A total of 1,081/1,113 (97.1%) Turkish did go for HBV testing. Twenty-six (2.4%) were HBsAg positive; 11/26 were unaware of their status and 10/11 were successfully referred. HBsAg prevalence was 3.0% in FGM and 1.5% in SGM,p= .070. Only one out of seven HBsAg positive SGM was born after 1987. In the multiple generalized estimating equations model, the most important risk factors for anti-HBc positivity were male gender (p= .021), older age (p

Cellular senescence is a process that results in irreversible cell-cycle arrest, and is thought to be an autonomous tumor-suppressor mechanism. During senescence, cells develop distinctive metabolic and signaling features, together referred to as the senescence-associated secretory phenotype (SASP). The SASP is implicated in several aging-related pathologies, including various malignancies. Accumulating evidence argues that cellular senescence acts as a double-edged sword in human cancer, and new agents and innovative strategies to tackle senescent cells are in development pipelines to counter the adverse effects of cellular senescence in the clinic. We focus on recent discoveries in senescence research and SASP biology, and highlight the potential of SASP suppression and senescent cell clearance in advancing precision medicine.

BACKGROUND: Turkey is an intermediate hepatitis B virus (HBV) endemic country. However, prevalence among Turkish migrants in Belgium is unknown, especially in those born in Belgium with a foreign-born parent, i.e. second-generation migrants (SGM). AIMS: To evaluate the prevalence of HBV infection and associated risk factors in Turkish first-generation migrants (FGM), i.e. foreign-born, and SGM. METHODS: Between September 2017 and May 2019, free outreach testing for hepatitis B surface antigen (HBsAg), hepatitis B core antibodies (anti-HBc), and antibodies against HBsAg was offered to Turkish migrants in Middle-Limburg, Belgium. Face-to-face questionnaire assessed HBV risk factors. HBsAg positive patients were referred and followed up. Turkish SGM were stratified into birth cohort born before and after 1987, since those born after 1987 should be covered by the universal infant vaccination program. RESULTS: A total of 1,081/1,113 (97.1%) Turkish did go for HBV testing. Twenty-six (2.4%) were HBsAg positive; 11/26 were unaware of their status and 10/11 were successfully referred. HBsAg prevalence was 3.0% in FGM and 1.5% in SGM, p = .070. Only one out of seven HBsAg positive SGM was born after 1987. In the multiple generalized estimating equations model, the most important risk factors for anti-HBc positivity were male gender (p = .021), older age (p < .001), FGM (p < .001), low educational level of the mother (p = .003), HBV infected mother (p = .008), HBV infected siblings (p = .002), HBV infected other family member (p = .004), gynaecological examination in Turkey or unsafe male circumcision (p = .032) and dental treatment in Turkey (p = .049). CONCLUSION: Outreach testing was well-accepted and referral to specialist care was generally successful. National HBV screening should be implemented in the Turkish FGM population and might be considered in SGM not covered by primary prevention strategies. ispartof: PLOS ONE vol:15 issue:7 ispartof: location:United States status: published

Cancer cells need excess energy and essential nutrients/metabolites not only to divide and proliferate but also to migrate and invade distant organs for metastasis. Fatty acid and cholesterol synthesis, considered a hallmark of cancer for anabolism and membrane biogenesis, requires citrate. We review here potential pathways in which citrate is synthesized and/or supplied to cancer cells and the impact of extracellular citrate on cancer cell metabolism and growth. Cancer cells employ different mechanisms to support mitochondrial activity and citrate synthesis when some of the necessary substrates are missing in the extracellular space. We also discuss the different transport mechanisms available for the entry of extracellular citrate into cancer cells and how citrate as a master metabolite enhances ATP production and fuels anabolic pathways. The available literature suggests that cancer cells show an increased metabolic flexibility with which they tackle changing environmental conditions, a phenomenon crucial for cancer cell proliferation and metastasis.

Hepatitis B is endemic in sub-Saharan Africa with ~60 million people chronically infected. While prevention, through vaccination, is central to elimination strategies, only 11 countries have birth dose vaccination and full vaccine coverage remains at suboptimal levels. Furthermore, to fully realize elimination, those chronically infected need to be identified, assessed for therapy and then linked to care. Given current treatment criteria, the precise quantum of people warranting therapy, according to criteria, is essentially unknown. The issue is further complicated by data to suggest differences in the numbers of people requiring treatment when applying WHO as compared to European Association for the Study of the Liver, EASL, criteria. Optimal determination of treatment eligibility is further hindered by the lack of available tools to adequately assess individual patients. It is conceivable that accurately determining the number of those requiring treatment, given the heterogeneity of hepatitis B in Africa, is difficult. Better studies and data are required. More signifcantly, improved access and availability to the diagnostic tools needed to assess patients in additon to access to drugs are as, if not more important, to achieve elimination.

Human hepatocellular carcinoma (HCC) heterogeneity promotes recurrence and therapeutic resistance. We recently demonstrated that inflammation favors hepatocyte retrodifferentiation into progenitor cells. Here, we identify the molecular effectors that induce metabolic reprogramming, chemoresistance, and invasiveness of retrodifferentiated HCC stem cells. Spheroid cultures of human HepaRG progenitors (HepaRG-Spheres), HBG-BC2, HepG2, and HuH7 cells and isolation of side population (SP) from HepaRG cells (HepaRG-SP) were analyzed by transcriptomics, signaling pathway analysis, and evaluation of chemotherapies. Gene expression profiling of HepaRG-SP and HepaRG-Spheres revealed enriched signatures related to cancer stem cells, metastasis, and recurrence and showed that HepaRG progenitors could retrodifferentiate into an immature state. The transcriptome from these stem cells matched that of proliferative bad outcome HCCs in a cohort of 457 patients. These HCC stem cells expressed high levels of cytokines triggering retrodifferentiation and displayed high migration and invasion potential. They also showed changes in mitochondrial activity with reduced membrane potential, low ATP production, and high lactate production. These changes were, in part, related to angiopoietin-like 4 (ANGPTL4)–induced upregulation of pyruvate dehydrogenase kinase 4 (PDK4), an inhibitor of mitochondrial pyruvate dehydrogenase. Upregulation of ANGPTL4 and PDK4 paralleled that of stem cells markers in human HCC specimens. Moreover, the PDK4 inhibitor dichloroacetate reversed chemoresistance to sorafenib or cisplatin in HCC stem cells derived from four HCC cell lines. In conclusion, retrodifferentiated cancer cells develop enhanced invasion and therapeutic resistance through ANGPTL4 and PDK4. Therefore, restoration of mitochondrial activity in combination with chemotherapy represents an attractive therapeutic approach in HCC. Significance: Restoring mitochondrial function in human hepatocellular carcinomas overcomes cancer resistance.

Liver malignancies, either primary tumours (mainly hepatocellular carcinoma and cholangiocarcinoma) or secondary hepatic metastases, are a major cause of death, with an increasing incidence. Among them, hepatocellular carcinoma (HCC) presents with a dark prognosis because of underlying liver diseases and an often late diagnosis. A curative surgical treatment can therefore only be proposed in 20 to 30% of the patients. However, new treatment options for intermediate to advanced stages, such as internal radionuclide therapy, seem particularly attractive. Transarterial radioembolization (TARE), which consists in the use of intra-arterial injection of a radiolabelled embolising agent, has led to very promising results. TARE with 90Y-loaded microspheres is now becoming an established procedure to treat liver tumours, with two commercially available products (namely, SIR-Sphere® and TheraSphere®). However, this technology remains expensive and is thus not available everywhere. The aim of this review is to describe TARE alternative technologies currently developed and investigated in clinical trials, with special emphasis on HCC.

Late onset of clinical symptoms in hepatocellular carcinoma (HCC) results in late diagnosis and poor disease outcome. Approximately 85% of individuals with HCC have underlying liver cirrhosis. However, not all cirrhotic patients develop cancer. Reliable tools that would distinguish cirrhotic patients who will develop cancer from those who will not are urgently needed. We used the Illumina HumanMethylation450 BeadChip microarray to test whether white blood cell DNA, an easily accessible source of DNA, exhibits site-specific changes in DNA methylation in blood of diagnosed HCC patients (post-diagnostic, 24 cases, 24 controls) and in prospectively collected blood specimens of HCC patients who were cancer-free at blood collection (pre-diagnostic, 21 cases, 21 controls). Out of 22 differentially methylated loci selected for validation by pyrosequencing, 19 loci with neighbouring CpG sites (probes) were confirmed in the pre-diagnostic study group and subjected to verification in a prospective cirrhotic cohort (13 cases, 23 controls). We established for the first time 9 probes that could distinguish HBV-negative cirrhotic patients who subsequently developed HCC from those who stayed cancer-free. These probes were identified within regulatory regions of BARD1, MAGEB3, BRUNOL5, FXYD6, TET1, TSPAN5, DPPA5, KIAA1210, and LSP1. Methylation levels within DPPA5, KIAA1210, and LSP1 were higher in prospective samples from HCC cases versus cirrhotic controls. The remaining probes were hypomethylated in cases compared with controls. Using blood as a minimally invasive material and pyrosequencing as a straightforward quantitative method, the established probes have potential to be developed into a routine clinical test after validation in larger cohorts.

Background & Aims: Effectiveness of surveillance for hepatocellular carcinoma is controversial. We here explore its effects in "real life'' clinical practice. Methods: Patients with hepatocellular carcinoma diagnosed in the period 2005-2012 in five Dutch academic centers were evaluated. Surveillance was defined as >= 2 screening tests during three preceding years and at least one radiologic imaging test within 18 months before diagnosis. Results: 295 (27%) of 1074 cases underwent surveillance. Median time interval between last negative radiologic imaging and hepatocellular carcinoma diagnosis was 7.5 months. In the surveillance group, cirrhosis (97% vs. 60%, p < 0.001) and viral hepatitis were more frequent, and non-alcoholic fatty liver disease or absence of risk factors less frequent. In case of surveillance, tumor size was significantly smaller (2.7 vs. 6.0 cm), with lower alpha-fetoprotein levels (16 vs. 44 mu g/L), earlier tumor stage (BCLC 0 and A combined: 61% vs. 21%) and resection/transplantation (34% vs. 25%) or radiofrequency ablation (23% vs. 7%) more often applied, with significantly higher 1-, 3-, and 5-year survival rates. Survival benefit by surveillance remained significant after adjustment for lead-time bias based on assumed tumor doubling time of 90 days, but not with doubling time of >= 120 days. In multivariate analysis, surveillance was an independent predictor for mortality (for interval 9 months: adjusted HRs 0.51 and 0.50, 95% confidence intervals: 0.39-0.67 and 0.37-0.69). Conclusions: Surveillance for hepatocellular carcinoma was associated with smaller tumor size, earlier tumor stage, with an impact on therapeutic strategy and was an independent predictor of survival. (C) 2015 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Latinos born in the US, 36 million, comprise 65% of all US Latinos. Yet their cancer experience is nearly always analyzed together with their foreign-born counterparts, 19 million, who constitute a steady influx of truly lower-risk populations from abroad. To highlight specific cancer vulnerabilities for US-born Latinos, we compare their cancer mortality to the majority non-Latino white (NLW) population, foreign-born Latinos, and non-Latino blacks. Methods We analyzed 465,751 cancer deaths from 2008 to 2012 occurring among residents of California and Texas, the two most populous states, accounting for 47% of US Latinos. This cross-sectional analysis, based on granular data obtained from death certificates on cause of death, age, race, ethnicity and birthplace, makes use of normal standardization techniques and negative binomial regression models. Results While Latinos overall have lower all-cancers-combined mortality rates than NLWs, these numbers were largely driven by low rates among the foreign born while mortality rates for US-born Latinos approach those of NLWs. Among Texas males, rates were 210 per 100,000 for NLWs and 166 for Latinos combined, but 201 per 100,000 for US-born Latinos and 125 for foreign-born Latinos. Compared to NLWs, US-born Latino males in California had mortality rate ratios of 2.83 (95% CI: 2.52–3.18) for liver cancer, 1.44 (95% CI: 1.30–1.61) for kidney cancer, and 1.25 (95% CI: 1.17–1.34) for colorectal cancer (CRC). Texas results showed a similar site-specific pattern. Conclusions Specific cancer patterns for US-born Latinos, who have relatively high cancer mortality, similar overall to NLWs, are masked by aggregation of all Latinos, US-born and foreign-born. While NLWs had high mortality for lung cancer, US-born Latinos had high mortality for liver, kidney and male colorectal cancers. HCV testing and reinforcement of the need for CRC screening should be a priority in this specific and understudied population. The unprecedented proximity of overall rates between NLWs and US-born Latino populations runs counter to the prevailing narrative of Latinos having significantly lower cancer risk and mortality. Birthplace data are critical in detecting meaningful differences among Latinos; these findings merit not only clinical but also public health attention

Hepatocellular carcinoma (HCC) represents the second cause of cancer-related mortality worldwide and is associated with poor prognosis, especially in patients not amenable for curative treatments. The multi-kinase inhibitor sorafenib represents the first-line treatment option for advanced HCC; nevertheless, its effectiveness is limited due to tumor heterogeneity as well as innate or acquired drug resistance, raising the need for new therapeutic strategies. MicroRNAs (miRNAs) involvement in treatment response as well as their safety and efficacy in preclinical models and clinical trials have been widely documented in the oncologic field, including HCC. Here, we identified miR-494 upregulation in a subgroup of human and rat HCCs with stem cell-like characteristics, as well as multiple epigenetic mechanisms involved in its aberrant expression in HCC cell lines and patients. Moreover, we identified p27, puma and pten among miR-494 targets, contributing to speed up cell cycle progression, enhance survival potential in stressful conditions and increase invasive and clonogenic capabilities. MiR-494 overexpression increased sorafenib resistance via mTOR pathway activation in HCC cell lines and, in line, high miR-494 levels associated with decreased sorafenib response in two HCC animal models. A sorafenib-combined anti-miR-494-based strategy revealed an enhanced anti-tumor potential with respect to sorafenib-only treatment in our HCC rat model. In conclusion, our findings suggested miR-494 as a possible therapeutic target as well as a candidate biomarker for patient stratification in advanced HCC.

Hepatocellular carcinoma (HCC) is rare in the Netherlands, even though the incidence has increased quite sharply in recent years. Standard treatment options consist of surgery, orthotopic liver transplantation, radiofrequency ablation, transarterial chemoembolisation (TACE) and systemic therapy with sorafenib. The consensus-based Dutch HCC guideline, established in 2013, serves to guide surveillance, diagnosis and treatment options: Surveillance should be performed by ultrasound at six-month intervals in well-defined cirrhotic patients and in selected high-risk hepatitis B carriers; A nodule > 1 cm in cirrhotic patients with arterial hypervascularity and venous or delayed phase washout at four-phase CT or MRI scan establishes the diagnosis of HCC; In patients with HCC without underlying cirrhosis, resection should be considered regardless of tumour size; In cirrhotic HCC patients, tumour stage, severity of underlying cirrhosis, and performance status determine treatment options. The algorithm of the Barcelona Clinic Liver Cancer (BCLC) staging system should be followed; Patients with Child-Pugh A-B cirrhosis (CP

Inflammation is recognized as one of the drivers of cancer. Yet, the individual immune components that possess pro- and anti-tumorigenic functions in individual cancers remain largely unknown. NKG2D is a potent activating immunoreceptor that has emerged as an important player in inflammatory disorders besides its well-established function as tumour suppressor. Here, we provide genetic evidence of an unexpected tumour-promoting effect of NKG2D in a model of inflammation-driven liver cancer. Compared to NKG2D-deficient mice, NKG2D-sufficient mice display accelerated tumour growth associated with, an increased recruitment of memory CD8+T cells to the liver and exacerbated pro-inflammatory milieu. In addition, we show that NKG2D contributes to liver damage and consequent hepatocyte proliferation known to favour tumorigenesis. Thus, the NKG2D/NKG2D-ligand pathway provides an additional mechanism linking chronic inflammation to tumour development in hepatocellular carcinoma. Our findings expose the need to selectively target the types of cancer that could benefit from NKG2D-based immunotherapy., Expression of NKG2D immunoreceptor ligands on tumour cells is believed to inhibit tumour growth through engaging NKG2D-expressing immune cells. Here, the authors show that in a model of liver cancer the NKG2D/NKG2D-ligand pathway can also promote tumour formation by sustaining an inflammatory environment.

Background:Vegetable and/or fruit intakes in association with hepatocellular carcinoma (HCC) risk have been investigated in case-control studies conducted in specific European countries and cohort studies conducted in Asia, with inconclusive results. No multi-centre European cohort has investigated the indicated associations.Methods:In 486 799 men/women from the European Prospective Investigation into Cancer and nutrition, we identified 201 HCC cases after 11 years median follow-up. We calculated adjusted hazard ratios (HRs) for HCC incidence for sex-specific quintiles and per 100 g d(-1) increments of vegetable/fruit intakes.Results:Higher vegetable intake was associated with a statistically significant, monotonic reduction of HCC risk: HR (100 g d(-1) increment): 0.83; 95% CI: 0.71-0.98. This association was consistent in sensitivity analyses with no apparent heterogeneity across strata of HCC risk factors. Fruit intake was not associated with HCC incidence: HR (100 g d(-1) increment): 1.01; 95% CI: 0.92-1.11.Conclusions:Vegetable, but not fruit, intake is associated with lower HCC risk with no evidence for heterogeneity of this association in strata of important HCC risk factors. Mechanistic studies should clarify pathways underlying this association. Given that HCC prognosis is poor and that vegetables are practically universally accessible, our results may be important, especially for those at high risk for the disease.British Journal of Cancer advance online publication, 5 March 2015; doi:10.1038/bjc.2014.654 www.bjcancer.com.

Objective The objective of this study was to develop and characterize alginate microspheres suitable for embolization with on-demand triggered doxorubicin (DOX) release and whereby the microspheres as well as the drug releasing process can be visualized in vivo using MRI. Methods and Findings For this purpose, barium crosslinked alginate microspheres were loaded with temperature sensitive liposomes (TSL/TSL-Ba-ms), which release their payload upon mild hyperthermia. These TSL contained DOX and [Gd(HPDO3A)(H2O)], a T-1 MRI contrast agent, for real time visualization of the release. Empty alginate microspheres crosslinked with holmium ions (T-2* MRI contrast agent, Ho-ms) were mixed with TSL-Ba-ms to allow microsphere visualization. TSL-Ba-ms and Ho-ms were prepared with a homemade spray device and sized by sieving. Encapsulation of TSL in barium crosslinked microspheres changed the triggered release properties only slightly: 95% of the loaded DOX was released from free TSL vs. 86% release for TSL-Ba-ms within 30 seconds in 50% FBS at 42 degrees C. TSL-Ba-ms (76 +/- 41 mu m) and Ho-ms (64 +/- 29 mu m) had a comparable size, which most likely will result in a similar in vivo tissue distribution after an i.v. co-injection and therefore Ho-ms can be used as tracer for the TSL-Ba-ms. MR imaging of a TSL-Ba-ms and Ho-ms mixture (ratio 95: 5) before and after hyperthermia allowed in vitro and in vivo visualization of microsphere deposition (T-2*-weighted images) as well as temperature-triggered release (T-1-weighted images). The [Gd(HPDO3A)(H2O)] release and clusters of microspheres containing holmium ions were visualized in a VX2 tumor model in a rabbit using MRI. Conclusions In conclusion, these TSL-Ba-ms and Ho-ms are promising systems for real-time, MRguided embolization and triggered release of drugs in vivo.

Hepatocellular carcinoma usually afflicts individuals in their later years following longstanding liver disease. In Peru, hepatocellular carcinoma exists in a unique clinical presentation, which affects patients around age 25 with a normal, healthy liver. In order to deepen our understanding of the molecular processes ongoing in Peruvian liver tumors, mutation spectrum analysis was carried out on hepatocellular carcinomas from 80 Peruvian patients. Sequencing analysis focused on nine genes typically altered during liver carcinogenesis, i.e. ARID2, AXIN1, BRAF, CTNNB1, NFE2L2, H/K/N-RAS, and TP53. We also assessed the transcription level of factors involved in the control of the alpha-fetoprotein expression and the Hippo signaling pathway that controls contact inhibition in metazoans. The mutation spectrum of Peruvian patients was unique with a major class of alterations represented by Insertions/Deletions. There were no changes at hepatocellular carcinoma-associated mutation hotspots in more than half of the specimens analyzed. Furthermore, our findings support the theory of a consistent collapse in the Hippo axis, as well as an expression of the stemness factor NANOG in high alpha-fetoprotein-expressing hepatocellular carcinomas. These results confirm the specificity of Peruvian hepatocellular carcinoma at the molecular genetic level. The present study emphasizes the necessity to widen cancer research to include historically neglected patients from South America, and more broadly the Global South, where cancer genetics and tumor presentation are divergent from canonical neoplasms.

Two human leukemia cells K562 and THP-1, the breast cancer lines MCF-7 and ZR-75-1, and the melanoma line MDA-MB-435S were compared by flowcytometry for their behaviour at increasing levels of 3BP. K562 and THP-1 responded to 3BP by membrane depolarization and increased ROS; MCF-7 and ZR-75-1 showed decreased polarization and low ROS increase; MDA-MB-435S had limited depolarization and no ROS increase. THP-1 cells exposed to a range of 3BP concentrations in combination with DCA showed increase of polarization, slight ROS increase, and weakened nuclear integrity. 3BP and DCA show no synergism, but have complementary destructive effects on THP-1 cells. The data led to the conclusion that the THP-1 cells do not carry a functional membrane monocarboxylate transporter (MCT) or that 3BP circumvents MCT binding and can enter these cells independently.

Inactivation of KLF6 is common in hepatocellular carcinoma (HCC) associated with hepatitis C virus (HCV) infection, thereby abrogating its normal antiproliferative activity in liver cells. The aim of the study was to evaluate the impact of KLF6 depletion on human HCC and experimental hepatocarcinogenesis in vivo. In patients with surgically resected HCC, reduced tumor expression of KLF6 was associated with decreased survival. Consistent with its role as a tumor suppressor, KLF6+/+ mice developed significantly more tumors in response to the chemical carcinogen diethyl nitrosamine (DEN) than wild-type animals. Gene expression signatures in both surrounding tissue and tumors of KLF6+/- mice closely recapitulated those associated with aggressive human HCCs. Expression microarray profiling also revealed an increase in Mdm2 mRNA in tumors from KLF6+/- compared with KLF6+/+ mice, which was validated by way of quantitative real-time polymerase chain reaction and western blot analysis in both human HCC and DEN-induced murine tumors. Moreover, chromatin immunoprecipitation and cotransfection assays established the P2 intronic promoter of Mdm2 as a bona fide transcriptional target repressed by KLF6. Whereas KLF6 overexpression in HCC cell lines and primary hepatocytes led to reduced MDM2 levels and increased p53 protein and transcriptional activity, reduction in KLF6 by small interfering RNA led to increased MDM2 and reduced p53. Conclusion: Our findings indicate that KLF6 deficiency contributes significantly to the carcinogenic milieu in human and murine HCC and uncover a novel tumor suppressor activity of KLF6 in HCC by linking its transcriptional repression of Mdm2 to stabilizing p53. (HEPATOLOGY 2011;54:522-531)

International audience; Hepatitis C virus (HCV) core, known to be involved in liver carcinogenesis, is processed in the endoplasmic reticulum (ER). We thus investigated the impact of three HCV core isolates on ER stress, ER calcium signalling and apoptosis. We show that HCV core constructs trigger hyperexpression of Grp78/BiP, Grp 94, calreticulin and sarco/endoplasmic reticulum calcium ATPase, inducing ER stress. By using the ER-targeted aequorin calcium probe, we found that ER calcium depletion follows ER stress in core-expressing cells. HCV core induces apoptosis through overexpression of the CHOP/GADD153 proapoptotic factor, Bax translocation to mitochondria, mitochondrial membrane depolarization, cytochrome c release, caspase-3 and PARP cleavage. Furthermore, reversion of HCV core-induced ER calcium depletion (by transfection of SERCA2) completely abolished mitochondrial membrane depolarization, suggesting that both ER stress (through CHOP overexpression) and calcium signalling play a major role in the HCV core-mediated control of apoptosis. ER stress and apoptosis were also found in a proportion of HCV-full-length replicon-expressing cells and in the liver of HCV core transgenic mice. In conclusion, our data demonstrate that HCV core deregulates the control of apoptosis by inducing ER stress and ER calcium depletion providing new elements to understand the mechanisms involved in HCV-related liver chronic diseases.

Objectives: Objectives: Long–Evans Cinnamon (LEC) rats are characterized by an abnormal hepatic deposition of copper (Cu) due to a lack of the Cu-transporter P-type adenosine triphosphatase: accordingly, the strain is a good animal model of Wilson's disease. The effect of oral zinc (Zn) acetate treatment on the development of acute hepatitis and the biochemical parameters of Cu-induced liver damage was studied in 5-week-old LEC rats (n=52). Methods: Rats receiving 50 or 80 mg/ml/day Zn acetate by gavage and control rats receiving a daily dose of glucose solution 0.02 g/ml by gastric intubation were killed at 1, 2 or 8 weeks after the start of treatment. Results: Treatment with Zn acetate resulted in the prevention of acute hepatitis: 10 of the 13 untreated rats developed signs and symptoms compatible with acute hepatitis between the 6th and 7th week of treatment. Tissue metallothionein (MT) significantly increased in the treated rats and positively correlated with Zn concentrations within the liver. Control rats had a significantly higher iron concentration in the liver and kidneys compared with supplemented rats, after both short- and long-term experiments. 8-hydroxy-2′-deoxyguanosine amounts were significantly lower in untreated rats. Conclusions: Zn acetate prevents acute hepatitis, by increasing tissue MT concentrations, reducing Cu absorption and interfering with Fe metabolism.