Pancoast´s syndrome (PS) includes all signs and symptoms resulting from a tumor affecting the pulmonary apex and adjacent structures. Its main clinical feature is pain in the shoulder, supra-clavicular and arm along with the distribution of the ulnar nerve. The majority of cases are caused by non-small-cell bronchogenic carcinoma. Etiologic diagnosis is mostly achieved by histological examination of a percutaneous transthoracic needle biopsy. PS is a rare manifestation of a hepatocellular carcinoma (HCC). HCC is the most common primary liver tumor having a multifactorial etiology and high mortality rates. It often spreads itself to the lungs and regional lymphatic nodes; bone metastases are uncommon (2-20%); Patients with HCC can experience bone pain with no signs of hepatic dysfunction.

Tisochrysis lutea is a marine haptophyte rich in omega-3 polyunsaturated fatty acids (e.g., docosahexaenoic acid (DHA)) and carotenoids (e.g., fucoxanthin). Because of the nutraceutical applications of these compounds, this microalga is being used in aquaculture to feed oyster and shrimp larvae. In our earlier report, T. lutea organic crude extracts exhibited in vitro cytotoxic activity against human hepatocarcinoma (HepG2) cells. However, so far, the compound(s) accountable for the observed bioactivity have not been identified. Therefore, the aim of this study was to isolate and identify the chemical component(s) responsible for the bioactivity observed. Bioassay-guided fractionation through a combination of silica-gel column chromatography, followed by preparative thin layer chromatography (PTLC), led to the isolation of two diastereomers of a monoterpenoid lactone, namely, loliolide (1) and epi-loliolide (2), isolated for the first time in this species. The structural elucidation of both compounds was carried out by GC-MS and 1D (1 H and13 C APT) and 2D (COSY, HMBC, HSQC-ed, and NOESY) NMR analysis. Both compounds significantly reduced the viability of HepG2 cells and were considerably less toxic towards a non-tumoral murine stromal (S17) cell line, although epi-loliolide was found to be more active than loliolide. © 2020 by the authors.

Purpose: To assess the relevance of diffusion-weighted imaging (DWI) in the hepatocellular carcinoma (HCC) detection as a stand-alone procedure or in combination to conventional magnetic resonance imaging (MRI). Methods: Medical records from nineteen patients with pathological confirmation of HCC, who have been submitted to conventional MRI and DWI in a time frame 3 months prior and 3 months after diagnosis, were retrospectively reviewed. For each patient, apparent diffusion coefficient (ADC) values were measured in the HCC lesion, in the liver parenchyma and in the spleen. Tumor ADC and parenchyma ADC, obtained with and without normalization, were compared through paired-samples t-test. Tumor ADC was analyzed according to lobe, size category and acquisition system using the Mann-Whitney test. For sensitivity assessment of DWI and conventional MRI, individually and combined, a contingency table was made. Results: The calculated DWI sensitivity for HCC detection (47.4%) was lower than conventional MRI sensitivity (68.4%). The highest sensitivity was obtained with the combined reading of both techniques (78.9%). A statistically significant difference between tumor ADC and parenchymal ADC was found, both for normalized and non-normalized values, with lower values for the HCC lesions. Comparison of tumor ADC values according to lobe, size and acquisition system did not show a statistically significant difference. Conclusions: For HCC detection in the setting of liver cirrhosis the use of DWI increases tumor detection compared to conventional MR alone and its use is recommended. This result corroborates other literature reports regarding the added value of DWI in HCC diagnosis, irrespective of the magnetic field strength. Due to the small patient sample of the current series further investigation may be warranted.

Over the past decades, cancer became one of the major health problems worldwide. Among the most common types of cancers is liver cancer, which includes hepatocellular carcinoma. Hepatocellular carcinoma is the fourth leading cause of cancer-related death, and its incidence has been growing. The currently available treatment options are poorly effective and severely restricted when the disease is advanced. In these cases, systemic chemotherapy is usually applied. However, it is always accompanied by undesired side effects and a high rate of tumor recurrence. Therefore, it is necessary to develop new strategies, able to overcome these limitations.Gene therapy has been widely studied and has been demonstrating promising results for the treatment of several diseases, including cancer. It consists of the delivery of genetic material to the target cells, inducing a tumor suppressive effect. Even though viral gene delivery systems are most commonly used in the clinical field, non-viral gene delivery systems arise as safer systems and have been widely studied and engineered. Among the various non-viral gene delivery systems, polymeric nanoparticles stand out due to their biocompatibility and versatility. Besides, the techniques used to produce these systems allow a precise control of nanocarriers characteristics and modification of their surface in order to achieve the desired features. Polymeric nanoparticles include, among others, polymeric micelles and polyplexes, that could be synthesized from glycopolymers. These synthetic polymers contain carbohydrate groups in their structure, which promote targeting. Accordingly, glycopolymers containing the monomer LAMA (2-lactobionamidoethyl methacrylate), which has a galactose residue in its structure, were used to produce polymeric micelles and polyplexes. pLAMA is able to bind to the asialoglycoprotein receptor, overexpressed on the surface of HCC cells. Therefore, the main purpose of this work was to develop a polymer-based nanosystem capable of effectively and specifically deliver genetic material into HCC cells.Taking this into account, initially, polymeric micelles were produced using amphiphilic glycopolymers, PAMA103-co-PLAMA19-b-PDEAEMA63 and PAMA50-co-PLAMA10-b-PDEAEMA16. These nanosystems were extensively characterized, presenting a size around 70 nm, and were able to condense and protect genetic material. However, the results obtained after transfection of HepG2 cells with the polymeric micelles showed high levels of toxicity and low levels of biological activity. Additionally, the potential of the polymeric nanoparticles for drug delivery was evaluated. PAMA50-co-PLAMA10-b-PDEAEMA16-based micelles were loaded with sorafenib, a first-line drug approved for HCC treatment, and the values obtained for loading efficiency and loading capacity was 15 and 4 %, respectively, which are above what has been reported in the literature. However, these micelles resulted in high levels of toxicity in HepG2 cells and, for that reason, another approach was evaluated. The alternative consisted of the production of polyplexes using the polymer PAMA73-co-PLAMA21. These nanosystems presented a size around 150 nm and, overall, were able to condense and protect genetic material, maintaining their stability along time. The polyplexes were produced at different N/P charge ratios and resulted in great levels of transgene expression in the different hepatocarcinoma cell lines used. Transfection efficiency of these polyplexes prepared at 40/1 N/P charge ratio was evaluated by flow cytometry, showing an increased percentage of transfected cells comparing to the commercial control, PEI, which was confirmed by fluorescence microscopy. Additionally, the specificity of the developed polyplexes towards the asialoglycoprotein receptor was demonstrated by a competition assay, using the natural ligand for this receptor, asialofetuin. The obtained results were confirmed by flow cytometry and fluorescence microscopy. Cellular uptake and intracellular distribution of the polyplexes were assessed by confocal microscopy, showing that some polyplexes were colocalized with endosomes. As an attempt to promote endosomal escape, a microtubule-stabilizing agent, docetaxel, was used, resulting in a substantial increase of transfection activity. At last, to evaluate the potential of the PAMA73-co-PLAMA21-based polyplexes to mediate an antitumor strategy, a preliminary test was performed, using polyplexes prepared with the therapeutic plasmids encoding p53 or PTEN, having been observed a significant reduction on the cell viability.

Hepatocellular carcinoma (HCC) is one of the main causes of cancer-related death. Sorafenib, which is the first-line therapy for this disease, is associated with reduced therapeutic efficacy that could potentially be overcome by combination with selumetinib. In this context, the main goal of this work was to develop a new nanosystem, composed of a polymeric core coated by a lipid bilayer containing the targeting ligand GalNAc, to specifically and efficiently co-deliver both drugs into HCC cells, in order to significantly increase their therapeutic efficacy. Methods: The physicochemical characterization of hybrid nanosystems (HNP) and their components was performed by dynamic light scattering, zeta potential, matrix-assisted laser desorption ionization – time of flight mass spectroscopy, and transmission electron microscopy. Cellular binding, uptake and specificity of HNP were evaluated through flow cytometry and confocal microscopy. The therapeutic activity was evaluated namely through: cell viability by the Alamar Blue assay; cell death by flow cytometry using FITC-Annexin V; caspases activity by luminescence; mitochondrial membrane potential by flow cytometry; and molecular target levels by Western blot. Results: The obtained data show that these hybrid nanosystems present high stability and loading capacity of both drugs, and suitable physicochemical properties, namely in terms of size and surface charge. Moreover, the generated formulation allows to circumvent drug resistance and presents high specificity, promoting great cell death levels in HCC cells, but not in non-tumor cells. This potentiation of the antitumor effect of co-loaded drugs was carried out by an increased programmed cell death, being associated with a strong reduction in the mitochondrial membrane potential, a significant increase in the activity of caspases 3/7 and caspase 9, and much greater number of annexin V-positive cells. Conclusion: The developed formulation resulted in a high and synergistic antitumor effect, revealing a translational potential to improve therapeutic approaches against HCC.

Half of human cancers harbor TP53 mutations that render p53 inactive as a tumor suppressor. In these cancers, reactivation of mutant p53 (mutp53) through restoration of wild-type-like function constitutes a valuable anticancer therapeutic strategy. In order to search for mutp53 reactivators, a small library of tryptophanol-derived oxazoloisoindolinones was synthesized and the potential of these compounds as mutp53 reactivators and anticancer agents was investigated in human tumor cells and xenograft mouse models. By analysis of their anti-proliferative effect on a panel of p53-null NCI-H1299 tumor cells ectopically expressing highly prevalent mutp53, the compound SLMP53-2 was selected based on its potential reactivation of multiple structural mutp53. In mutp53-Y220C-expressing hepatocellular carcinoma (HCC) cells, SLMP53-2-induced growth inhibition was mediated by cell cycle arrest, apoptosis, and endoplasmic reticulum stress response. In these cells, SLMP53-2 restored wild-type-like conformation and DNA-binding ability of mutp53-Y220C by enhancing its interaction with the heat shock protein 70 (Hsp70), leading to the reestablishment of p53 transcriptional activity. Additionally, SLMP53-2 displayed synergistic effect with sorafenib, the only approved therapy for advanced HCC. Notably, it exhibited potent antitumor activity in human HCC xenograft mouse models with a favorable toxicological profile. Collectively, SLMP53-2 is a new mutp53-targeting agent with promising antitumor activity, particularly against HCC.

Hepatocellular carcinoma (HCC) is the third most common cause of death related to cancer diseases worldwide. The current treatment options have many limitations and reduced success rates. In this regard, advances in gene therapy have shown promising results in novel therapeutic strategies. However, the success of gene therapy depends on the efficient and specific delivery of genetic material into target cells. In this regard, the main goal of this work was to develop a new lipid-based nanosystem formulation containing the lipid lactosyl-PE for specific and efficient gene delivery into HCC cells. The obtained results showed that incorporation of 15% of lactosyl-PE into liposomes induces a strong potentiation of lipoplex biological activity in HepG2 cells, not only in terms of transgene expression levels but also in terms of percentage of transfected cells. In the presence of galactose, which competes with lactosyl-PE for the binding to the asialoglycoprotein receptor (ASGP-R), a significant reduction in biological activity was observed, showing that the potentiation of transfection induced by the presence of lactosyl-PE could be due to its specific interaction with ASGP-R, which is overexpressed in HCC. In addition, it was found that the incorporation of lactosyl-PE in the nanosystems promotes an increase in their cell binding and uptake. Regarding the physicochemical properties of lipoplexes, the presence of lactosyl-PE resulted in a significant increase in DNA protection and in a substantial decrease in their mean diameter and zeta potential, conferring them suitable characteristics for in vivo application. Overall, the results obtained in this study suggest that the potentiation of the biological activity induced by the presence of lactosyl-PE is due to its specific binding to the ASGP-R, showing that this novel formulation could constitute a new gene delivery nanosystem for application in therapeutic strategies in HCC.

Abstract Long-chain non-encoded RNAs (lncRNAs) are important in many life activities and can participate in the occurrence of hepatocellular carcinoma (HCC). Moreover, lncRNAs can be used as basis for developing new strategies to hinder liver cancer. To investigate the utility of lncRNAs in HCC as potential biomarkers for early detection and diagnosis, we mined genomic data from the Cancer Genome Atlas (TCGA), and analyzed the gene expressions from 374 tumor patients and 50 normal patients. The abnormal expressions of 387 differentially expressed lncRNAs (DElncRNAs) were identified from a total of 3099 lncRNAs. Moreover, 18 modules were divided based on WGCNA, and 2 of the 18 modules were positively correlated with stage and grade, and negatively correlated with survival time. Finally, 10 lncRNAs were found and their main functions are the enhancement of cellular metabolic capacity and cell proliferation. These 10 lncRNAs may serve as novel prognostic markers and therapeutic targets, and may help guide subsequent studies on HCC.

Astrocyte elevated gene-1 (AEG-1) is a positive regulator of tumorigenesis in human cancer cells. Human AEG-1 gene is located in chromosome 8q22 having 12 exons/11 introns. Chromosome 8q22 is known to be a hot spot for genomic alterations in several cancerous cells involving HCC. The aim of the study was assess association between the negative regulatory region of AEG-1 promoter mutations and genetic susceptibility to hepatocellular carcinoma. The negative regulatory region of the human AEG-1 promoter was evaluated in a total of 50 Iranian hepatocellular carcinomas (HCC) patients. For investigating AEG-1 promoter polymorphisms the PCR-sequencing method was used. In this study was found two new mutation C>T (-633) and G>C (-660) in the patient group. But it was not revealed the statistically significant association between any mutations in this region of the AEG-1 promoter with HCC susceptibility. According to presented data, we can say that the negative regulatory region of the AEG-1 promoter mutations did not exihibit significant relevance with hepatocellular carcinoma. We recommend further studies on the efficacy of the AEG-1 promoter in therapeutic targeting of the HCC.

Abstract Heparanase activity is involved in cancer growth and development in humans and single nucleotide polymorphisms (SNPs) in the heparanase gene (HPSE) have been shown to be associated with tumors. In this study, we investigated whether SNPs in HPSE were a risk factor for hepatocellular carcinoma (HCC) by undertaking a comprehensive haplotype-tagging, case-control study. For this, six haplotype-tagging SNPs (htSNPs) in HPSE were genotyped in 400 HCC patients and 480 controls by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis. A log-additive model revealed significant correlations between the HPSE polymorphisms rs12331678 and rs12503843 and the risk of HCC in the overall samples (p = 0.0046 and p = 0.0055). When the analysis was stratified based on hepatitis B virus (HBV) carrier status, significant interactions between rs12331678 and rs12503843 and HBV were observed. Conditional logistic regression analysis for the independent effect of one significant SNP suggested that rs12331678 or rs12503843 contributed an independent effect to the significant association with the risk of HCC, respectively. Our findings suggest that the SNPs rs12331678 and rs12503843 are HCC risk factors, although the potential functional roles of these two SNPs remain to be fully elucidated.