Doxorubicin is a highly active antineoplastic agent, but its clinical use is limited because of its cardiotoxicity. Although nutraceuticals endowed with anti-inflammatory properties exert cardioprotective activity, their bioavailability and stability are inconsistent. In an attempt to address this issue, we evaluated whether bioavailable nanoemulsions loaded with nutraceuticals (curcumin and fresh and dry tomato extracts rich in lycopene) protect cardiomyoblasts (H9C2 cells) from doxorubicin-induced toxicity. Nanoemulsions were produced with a high-pressure homogenizer. H9C2 cells were incubated with nanoemulsions loaded with different nutraceuticals alone or in combination with doxorubicin. Cell viability was evaluated with a modified MTT method. The levels of the lipid peroxidation products malondialdehyde (MDA) and 4-hydroxy-2-butanone (4-HNA), and of the cardiotoxic-related interleukins IL-6, IL-8, IL-1β and IL-10, tumor necrosis factor-alpha (TNF-α), and nitric oxide were analyzed in cardiomyoblasts. The hydrodynamic size of nanoemulsions was around 100 nm. Cell viability enhancement was 35–40% higher in cardiomyoblasts treated with nanoemulsion + doxorubicin than in cardiomyoblasts treated with doxorubicin alone. Nanoemulsions also protected against oxidative stress as witnessed by a reduction of MDA and 4-HNA. Notably, nanoemulsions inhibited the release of IL-6, IL-8, IL-1β, TNF-α and nitric oxide by around 35–40% and increased IL-10 production by 25–27% versus cells not treated with emulsions. Of the nutraceuticals evaluated, lycopene-rich nanoemulsions had the best cardioprotective profile. In conclusion, nanoemulsions loaded with the nutraceuticals described herein protect against cardiotoxicity, by reducing inflammation and lipid oxidative stress. These results set the stage for studies in preclinical models.

Expanding the availability of monoclonal antibodies interfering with hepatitis C virus infection of hepatocytes is an active field of investigation within medical biotechnologies, to prevent graft reinfection in patients subjected to liver transplantation and to overcome resistances elicited by novel antiviral drugs. In this paper, we describe a complete pipeline for screening of phage display libraries of human scFvs against native Claudin-1, a tight-junction protein involved in hepatitis C virus infection, expressed on the cell surface of human hepatocytes. To this aim, we implemented a high-throughput sequencing approach for library screening, followed by a simple and effective strategy to recover active binder clones from enriched sublibraries. The recovered clones were successfully converted to active immunoglobulins, thus demonstrating the effectiveness of the whole procedure. This novel approach can guarantee rapid and cheap isolation of antibodies for virtually any native antigen involved in human diseases, for therapeutic and/or diagnostic applications.

Hepatitis C virus (HCV) is a major cause of chronic hepatitis and liver carcinoma and new therapies based on novel targets are needed. The tight junction protein claudin 1 (CLDN-1) is essential for HCV cell entry and spread, and anti-CLDN-1 rat and mouse mAbs are safe and effective in preventing and treating HCV infection in a human liver chimeric mouse model. To accelerate translation of these observations into a novel approach to treat HCV infection and disease in humans, we screened a phage display library of human single-chain antibody fragments by using a panel of CLDN-1-positive and -negative cell lines and identified phage specifically binding to CLDN-1. The 12 clones showing the highest levels of binding were converted into human IgG4. Some of these mAbs displayed low-nanomolar affinity, and inhibited infection of human hepatoma Huh7.5 cells by different HCV isolates in a dose-dependent manner. Cross-competition experiments identified six inhibitory mAbs that recognized distinct epitopes. Combination of the human anti-SRB1 mAb C-1671 with these anti-CLDN-1 mAbs could either increase or reduce inhibition of cell culture-derived HCV infection in vitro. These novel human anti-CLDN-1 mAbs are potentially useful to develop a new strategy for anti-HCV therapy and lend support to the combined use of antibodies targeting the HCV receptors CLDN-1 and SRB1, but indicate that care must be taken in selecting the proper combination.