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1.
Nutrients ; 10(9)2018 Sep 14.
Artigo em Inglês | MEDLINE | ID: mdl-30223482

RESUMO

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-1ß 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-1ß, 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.

2.
Onco Targets Ther ; 11: 2241-2250, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29719406

RESUMO

Purpose: Pertuzumab, a novel anti-epidermal growth factor receptor 2 humanized monoclonal antibody, and trastuzumab-emtansine (TDM1), a novel antibody-drug conjugate made up of trastuzumab covalently linked to the highly potent microtubule inhibitory agent DM1, have been recently approved by the US Food and Drug Administration for increasing the efficiency and safety of breast cancer therapy with trastuzumab. We investigated for the first time the potential cardiotoxic effects of pertuzumab and TDM1, which are not yet fully elucidated, and we tested whether ranolazine could blunt their cardiotoxicity. Methods: The cardiotoxic effects were tested in vitro on rat cardiomyoblasts, human fetal cardiomyocytes, adult-like cardiomyocytes, and in vivo on a mouse model. Results: All the treated cardiac cell lines were significantly affected by treatment with the tested drugs. Surprisingly, TDM1 showed stronger inhibitory effects on cardiac cells with respect to trastuzumab and pertuzumab by more significantly reducing the cell viability and by changing the morphology of these cells. TDM1 also affected the beating phenotype of adult-like cardiomyocytes in vitro and reduced fractional shortening and ejection fraction in vivo in a mouse model. We also found that ranolazine attenuated not only the cardiotoxic side effects of trastuzumab but also those of pertuzumab and TDM1, when used in combinatorial treatments both in vitro and in vivo, as demonstrated by the recovery of fractional shortening and ejection fraction values in mice pretreated with TDM1. Conclusion: We demonstrated that it is possible to predict the eventual cardiotoxic effects of novel approved anticancer drugs early by using in vitro and in vivo approaches, which can also be useful to screen in advance the cardioprotective agents, so as to avoid the onset of unwanted cardiotoxic side effects.

3.
J Gen Virol ; 97(1): 82-94, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26519290

RESUMO

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.


Assuntos
Anticorpos Monoclonais/metabolismo , Antivirais/metabolismo , Claudina-1/antagonistas & inibidores , Hepacivirus/fisiologia , Receptores Depuradores Classe B/antagonistas & inibidores , Anticorpos de Cadeia Única/metabolismo , Internalização do Vírus/efeitos dos fármacos , Anticorpos Monoclonais/isolamento & purificação , Antivirais/isolamento & purificação , Linhagem Celular , Claudina-1/imunologia , Hepatócitos/virologia , Humanos , Modelos Teóricos , Biblioteca de Peptídeos , Receptores Depuradores Classe B/imunologia , Anticorpos de Cadeia Única/isolamento & purificação , Carga Viral , Cultura de Vírus
4.
Biomed Res Int ; 2015: 703213, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26649313

RESUMO

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.


Assuntos
Claudina-1/metabolismo , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Biblioteca de Peptídeos , Anticorpos de Cadeia Única/genética , Claudina-1/química , Células HEK293 , Humanos , Anticorpos de Cadeia Única/química , Anticorpos de Cadeia Única/metabolismo
5.
Protein Eng Des Sel ; 27(3): 83-8, 2014 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-24421342

RESUMO

The inhibition of ErbB2 by the use of human antibodies can be a valuable strategy for the treatment of breast and gastric cancer. Trastuzumab, a humanized anti-ErbB2 antibody in clinical use, is effective but can engender resistance as well as cardiotoxicity. ImmunoRNases, made up of a human anti-ErbB2 scFv and human pancreatic ribonucleases (HP-RNases), have been engineered to overcome the limits of other immunotoxins, such as immunogenicity and nonspecific toxicity. Here, we report that a novel anti-ErbB2 immunoRNase, called Erb-HPDDADD-RNase, obtained by fusing Erbicin, a human ErbB2-directed scFv, with an HP-RNase variant that resists the cytosolic inhibitor protein, binds with high affinity to a panel of ErbB2-positive gastric tumor cells and inhibits their growth more than does the parental immunoRNase, which is not resistant to the inhibitor. Moreover, Erb-HP-DDADD-RNase is endowed with antiproliferative activity for trastuzumab-resistant cancer cells both in vitro and in vivo that is more potent than that of the parental immunoRNase. Importantly, Erb-HP-DDADD-RNase does not show cardiotoxic effects in vitro on human cardiomyocytes and does not impair cardiac function in a mouse model. Thus, Erb-HP-DDADD-RNase could fulfil the therapeutic need of cancer patients ineligible for trastuzumab treatment due to primary or acquired trastuzumab resistance or to cardiac dysfunction.


Assuntos
Antineoplásicos/farmacologia , Cardiotoxinas/toxicidade , Receptor ErbB-2/química , Proteínas Recombinantes de Fusão/farmacologia , Proteínas Recombinantes de Fusão/toxicidade , Ribonuclease Pancreático/química , Animais , Anticorpos Monoclonais Humanizados , Antineoplásicos/química , Cardiotoxinas/química , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Humanos , Camundongos , Miócitos Cardíacos/efeitos dos fármacos , Neoplasias Experimentais/patologia , Engenharia de Proteínas , Receptor ErbB-2/genética , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Ribonuclease Pancreático/genética , Trastuzumab
6.
Gastric Cancer ; 17(1): 107-15, 2014 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-23460348

RESUMO

BACKGROUND: Gastric cancer represents one of the most common causes of cancer deaths worldwide. Overexpression of ErbB2, a tyrosine kinase receptor involved in the pathogenesis of several human cancer types, has been reported also in gastric cancer. Thus, the inhibition of ErbB2 signal transduction pathways by the use of human antibodies could be a valuable strategy for the therapy of this type of cancer. METHODS: We tested for the first time the antitumor effects on gastric cancer cells of Erb-hcAb, a novel fully human compact antibody, prepared in our laboratory, which targets a different epitope of ErbB2 with respect to trastuzumab, the only anti-ErbB2 antibody currently in clinical use for both breast and gastric cancer therapy. RESULTS: Herein we demonstrate that the in vitro and in vivo growth of gastric cancer cells is efficiently inhibited by Erb-hcAb, which shows antitumor effects on the NCI-N87 cell line more potent than those observed for trastuzumab. CONCLUSIONS: Erb-hcAb could be a promising candidate in the immunotherapy of gastric cancer as it combines the antiproliferative effect associated with the inhibition of ErbB2 signaling on tumor target cells with the ability to induce antibody-dependent cellular cytotoxicity.


Assuntos
Antineoplásicos/farmacologia , Imunoconjugados/farmacologia , Terapia de Alvo Molecular/métodos , Receptor ErbB-2/imunologia , Receptor ErbB-2/metabolismo , Neoplasias Gástricas/tratamento farmacológico , Animais , Anticorpos Monoclonais Humanizados/farmacologia , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Feminino , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Transdução de Sinais/efeitos dos fármacos , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologia , Trastuzumab , Ensaios Antitumorais Modelo de Xenoenxerto
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