Sergio Granados-Principal

Hydroxytyrosol: from laboratory investigations to future clinical trials

Mediterranean countries have lower rates of mortality from cardiovascular disease and cancer than Northern European or other Western countries. This has been attributed, at least in part, to the so-called Mediterranean diet, which is composed of specific local foods, including olive oil. Traditionally, many beneficial properties associated with this oil have been ascribed to its high oleic acid content. Today, it is clear that many of the beneficial effects of ingesting virgin olive oil are due to its minor compounds. This review summarizes the existing knowledge concerning the chemistry, pharmacokinetics, and toxicology of hydroxytyrosol, a minor compound of virgin olive oil, as well as this compounds importance for health. The main findings in terms of its beneficial effects in cardiovascular disease and cancer, including its properties against inflammation and platelet aggregation, are emphasized. New evidence and strategies regarding the use of hydroxytyrosol as a natural drug for the prevention and treatment of diseases with high incidences in Western countries are also presented.

New advances in molecular mechanisms and the prevention of adriamycin toxicity by antioxidant nutrients

Anthracyclines (doxorubicin, daunorubicin, epirubicin, and idarubicin) are currently the most effective group of anti-neoplastic drugs used in clinical practice. Of these, doxorubicin (also called adriamycin) is a key chemotherapeutic agent in cancer treatment, although its use is limited as a consequence of the chronic and acute toxicity associated with this drug. The molecular mechanisms of doxorubicin account for both the anti-cancer and the toxic side effects. Many antioxidants have been assayed, with positive or negative results, to prevent the toxicity of doxorubicin. The present review has two main goals: (1) to report the latest findings regarding the molecular mechanisms of doxorubicin toxicity; (2) to update our understanding of the role of natural antioxidants in preventive therapy against doxorubicin-induced toxicity. This review provides new evidence for the chemoprevention of doxorubicin toxicity, making use of natural antioxidants - in particular vitamin E, vitamin C, coenzyme Q, carotenoids, vitamin A, flavonoids, polyphenol, resveratrol, antioxidant from virgin olive oil and selenium - and offers new insights into the molecular mechanisms of doxorubicin toxicity with respect to DNA damage, free radicals and other parameters.

Free radicals in breast carcinogenesis, breast cancer progression and cancer stem cells. Biological bases to develop oxidative-based therapies.

Oxidative stress leads to lipid, carbohydrate, protein and DNA damage in biological systems and affects cell structure and function. Breast cancer cells are subjected to a high level of oxidative stress, both intracellular and extracellular. To survive, cancer cells must acquire adaptive mechanisms that counteract the toxic effects of free radicals exposure. These mechanisms may involve the activation of redox-sensitive transcription factors, increased expression of antioxidant enzymes and antiapoptotic proteins. Moreover, recent data maintain that different breast cancer cell types, show different intracellular antioxidant capacities that may determine their ability to resist radio and chemotherapy. The resistant cell type has been shown to correspond with tumor initiating cells, also known as cancer stem cells (CSCs), which are thought to be responsible for tumor initiation and metastasis. Abrogation of the above-mentioned adaptive mechanisms by redox regulation in cancer cells opens a promising research line that could have significant therapeutic applications.

Hydroxytyrosol ameliorates oxidative stress and mitochondrial dysfunction in doxorubicin-induced cardiotoxicity in rats with breast cancer.

Oxidative stress is involved in several processes including cancer, aging and cardiovascular disease, and has been shown to potentiate the therapeutic effect of drugs such as doxorubicin. Doxorubicin causes significant cardiotoxicity characterized by marked increases in oxidative stress and mitochondrial dysfunction. Herein, we investigate whether doxorubicin-associated chronic cardiac toxicity can be ameliorated with the antioxidant hydroxytyrosol in rats with breast cancer. Thirty-six rats bearing breast tumors induced chemically were divided into 4 groups: control, hydroxytyrosol (0.5mg/kg, 5days/week), doxorubicin (1mg/kg/week), and doxorubicin plus hydroxytyrosol. Cardiac disturbances at the cellular and mitochondrial level, mitochondrial electron transport chain complexes I-IV and apoptosis-inducing factor, and oxidative stress markers have been analyzed. Hydroxytyrosol improved the cardiac disturbances enhanced by doxorubicin by significantly reducing the percentage of altered mitochondria and oxidative damage. These results suggest that hydroxytyrosol improve the mitochondrial electron transport chain. This study demonstrates that hydroxytyrosol protect rat heart damage provoked by doxorubicin decreasing oxidative damage and mitochondrial alterations.

Age-related changes in brain mitochondrial DNA deletion and oxidative stress are differentially modulated by dietary fat type and coenzyme Q10

Mitochondria-related oxidative damage is a primary event in aging and age-related neurodegenerative disorders. Some dietary treatments, such as antioxidant supplementation or the enrichment of mitochondrial membranes with less oxidizable fatty acids, reduce lipid peroxidation and lengthen life span in rodents. This study compares life-long feeding on monounsaturated fatty acids (MUFAs), such as virgin olive oil, and n-6 polyunsaturated fatty acids, such as sunflower oil, with or without coenzyme Q₁₀ supplementation, with respect to age-related molecular changes in rat brain mitochondria. The MUFA diet led to diminished age-related phenotypic changes, with lipoxidation-derived protein markers being higher among the older animals, whereas protein carbonyl compounds were lower. It is noteworthy that the MUFA diet prevented the age-related increase in levels of mitochondrial DNA deletions in the brain mitochondria from aged animals. The findings of this study suggest that age-related oxidative stress is related, at the mitochondrial level, to other age-related features such as mitochondrial electron transport and mtDNA alterations, and it can be modulated by selecting an appropriate dietary fat type and/or by suitable supplementation with low levels of the antioxidant/electron carrier molecule coenzyme Q.

Targeting RPL39 and MLF2 reduces tumor initiation and metastasis in breast cancer by inhibiting nitric oxide synthase signaling

Significance This manuscript describes the identification and characterization of two previously unidentified cancer genes, ribosomal protein L39 and myeloid leukemia factor 2, that play an important role in tumor initiation and metastasis. Knockdown of these genes in triple negative breast cancer (TNBC) models significantly reduces primary-tumor growth, as well as metastasis. Mutations in these genes are associated with worse survival in breast-cancer patients. Both genes are regulated by the nitric oxide signaling pathway. Identification of these two genes represents a significant breakthrough in our understanding of treatment resistance in TNBC. Targeting these genes could alter clinical practice for tumor metastasis in future and improve outcomes of patients with breast cancer. We previously described a gene signature for breast cancer stem cells (BCSCs) derived from patient biopsies. Selective shRNA knockdown identified ribosomal protein L39 (RPL39) and myeloid leukemia factor 2 (MLF2) as the top candidates that affect BCSC self-renewal. Knockdown of RPL39 and MLF2 by specific siRNA nanoparticles in patient-derived and human cancer xenografts reduced tumor volume and lung metastases with a concomitant decrease in BCSCs. RNA deep sequencing identified damaging mutations in both genes. These mutations were confirmed in patient lung metastases (n = 53) and were statistically associated with shorter median time to pulmonary metastasis. Both genes affect the nitric oxide synthase pathway and are altered by hypoxia. These findings support that extensive tumor heterogeneity exists within primary cancers; distinct subpopulations associated with stem-like properties have increased metastatic potential.

Hydroxytyrosol inhibits growth and cell proliferation and promotes high expression of sfrp4 in rat mammary tumours

SCOPE Hydroxytyrosol (a phenolic compound derived from virgin olive oil) has demonstrated an anti-tumour effect in several tumour cell lines in addition to other health-related properties. The aim of this study was to investigate, for the first time in an animal model of experimental mammary carcinoma, the anti-cancer ability of hydroxytyrosol and to discover which pathways are modified by hydroxytyrosol. METHODS AND RESULTS Dimethylbenz[α]anthracene-induced mammary tumours were induced in 28 female Sprague-Dawley rats and ten of them were treated with hydroxytyrosol (0.5 mg/kg b.w. 5 days/week for 6 wk). cDNA microarray and quantitative RT-PCR experiments were performed. Hydroxytyrosol was found to inhibit the experimental mammary tumour growth and proliferation rate, with results comparable to those of doxorubicin but better with regard to the histopathological outcome. It also altered the expression of genes related to apoptosis, cell cycle, proliferation, differentiation, survival and transformation pathways. CONCLUSIONS This study shows that hydroxytyrosol exerts anti-cancer effects in Sprague-Dawley rats with experimental mammary tumours, inhibiting growth and cell proliferation in mammary tumours. Moreover, hydroxytyrosol alters several genes associated with cell proliferation, apoptosis and the Wnt signalling pathway, promoting a high expression of Sfrp4. However, further studies are necessary to better understand the mechanisms of hydroxytyrosol.

Colocalized Delivery of Rapamycin and Paclitaxel to Tumors Enhances Synergistic Targeting of the PI3K/Akt/mTOR Pathway

Ongoing clinical trials target the aberrant PI3K/Akt/mammalian target of rapamycin (mTOR) pathway in breast cancer through administration of rapamycin, an allosteric mTOR inhibitor, in combination with paclitaxel. However, synergy may not be fully exploited clinically because of distinct pharmacokinetic parameters of drugs. This study explores the synergistic potential of site-specific, colocalized delivery of rapamycin and paclitaxel through nanoparticle incorporation. Nanoparticle drug loading was accurately controlled, and synergistic drug ratios established in vitro. Precise drug ratios were maintained in tumors 48 hours after nanoparticle administration to mice, at levels twofold greater than liver and spleen, yielding superior antitumor activity compared to controls. Simultaneous and preferential in vivo delivery of rapamycin and paclitaxel to tumors yielded mechanistic insights into synergy involving suppression of feedback loop Akt phosphorylation and its downstream targets. Findings demonstrate that a same time, same place, and specific amount approach to combination chemotherapy by means of nanoparticle delivery has the potential to successfully translate in vitro synergistic findings in vivo. Predictive in vitro models can be used to determine optimum drug ratios for antitumor efficacy, while nanoparticle delivery of combination chemotherapies in preclinical animal models may lead to enhanced understanding of mechanisms of synergy, ultimately opening several avenues for personalized therapy.

Diets Based on Virgin Olive Oil or Fish Oil but Not on Sunflower Oil Prevent Age-Related Alveolar Bone Resorption by Mitochondrial-Related Mechanisms

Background/Objectives Aging enhances frequency of chronic diseases like cardiovascular diseases or periodontitis. Here we reproduced an age-dependent model of the periodontium, a fully physiological approach to periodontal conditions, to evaluate the impact of dietary fat type on gingival tissue of young (6 months old) and old (24 months old) rats. Methods/Findings Animals were fed life-long on diets based on monounsaturated fatty acids (MUFA) as virgin olive oil, n-6 polyunsaturated fatty acids (n-6PUFA), as sunflower oil, or n-3PUFA, as fish oil. Age-related alveolar bone loss was higher in n-6PUFA fed rats, probably as a consequence of the ablation of the cell capacity to adapt to aging. Gene expression analysis suggests that MUFA or n-3PUFA allowed mitochondria to maintain an adequate turnover through induction of biogenesis, autophagy and the antioxidant systems, and avoiding mitochondrial electron transport system alterations. Conclusions The main finding is that the enhanced alveolar bone loss associated to age may be targeted by an appropriate dietary treatment. The mechanisms involved in this phenomenon are related with an ablation of the cell capacity to adapt to aging. Thus, MUFA or n-3PUFA might allow mitochondrial maintaining turnover through biogenesis or autophagy. They might also be able to induce the corresponding antioxidant systems to counteract age-related oxidative stress, and do not inhibit mitochondrial electron transport chain. From the nutritional and clinical point of view, it is noteworthy that the potential treatments to attenuate alveolar bone loss (a feature of periodontal disease) associated to age could be similar to some of the proposed for the prevention and treatment of cardiovascular diseases, a group of pathologies recently associated with age-related periodontitis.

Doxorubicin-induced oxidative stress in rats is efficiently counteracted by dietary anthocyanin differently enriched strawberry (Fragaria × ananassa Duch.).

This study investigated the effects of two different strawberry cultivars, Adria and Sveva, against doxorubicin (DOX)-induced toxicity in rats. A controlled dietary intervention was conducted over 16 weeks with four groups: (i) normal diet; (ii) normal diet + DOX injection; (iii) Adria supplementation + DOX injection; and (iv) Sveva supplementation + DOX injection. Sveva presented higher total antioxidant capacity value and phenol and and vitamin C levels than Adria, which in turn presented higher anthocyanin contents. DOX drastically increased lymphocyte DNA damage, liver biomarkers of protein and lipid oxidation, and mitochondrial ROS content and markedly decreased plasma retinol level, liver antioxidant enzymes, and mitochondrial functionality. After 2 months of strawberry supplementation, rats presented a significant reduction of DNA damage and ROS concentration and a significant improvement of oxidative stress biomarkers, antioxidant enzyme activities, and mitochondrial performance. These results suggest that strawberry supplementation can counteract DOX toxicity, confirming the potential health benefit of strawberry in vivo against oxidative stress.