Susanne U. Mertens-Talcott

The oncogenic microRNA-27a targets genes that regulate specificity protein transcription factors and the G2-M checkpoint in MDA-MB-231 breast cancer cells.

There is evidence that specificity proteins (Sp), such as Sp1, Sp3, and Sp4, are overexpressed in tumors and contribute to the proliferative and angiogenic phenotype associated with cancer cells. Sp1, Sp3, and Sp4 are expressed in a panel of estrogen receptor (ER)-positive and ER-negative breast cancer cell lines, and we hypothesized that regulation of their expression may be due to microRNA-27a (miR-27a), which is also expressed in these cell lines and has been reported to regulate the zinc finger ZBTB10 gene, a putative Sp repressor. Transfection of ER-negative MDA-MB-231 breast cancer cells with antisense miR-27a (as-miR-27a) resulted in increased expression of ZBTB10 mRNA and decreased expression of Sp1, Sp3, and Sp4 at the mRNA and protein levels and also decreased activity in cells transfected with constructs containing Sp1 and Sp3 promoter inserts. In addition, these responses were accompanied by decreased expression of Sp-dependent survival and angiogenic genes, including survivin, vascular endothelial growth factor (VEGF), and VEGF receptor 1 (VEGFR1). Moreover, similar results were observed in MDA-MB-231 cells transfected with ZBTB10 expression plasmid. Both as-miR-27a and ZBTB10 overexpression decreased the percentage of MDA-MB-231 cells in S phase of the cell cycle; however, ZBTB10 increased the percentage of cells in G(0)-G(1), whereas as-miR-27a increased the percentage in G(2)-M. This latter response was associated with induction of Myt-1 (another miR-27a target gene), which inhibits G(2)-M through enhanced phosphorylation and inactivation of cdc2. Thus, the oncogenic activity of miR-27a in MDA-MB-231 cells is due, in part, to suppression of ZBTB10 and Myt-1.

Pharmacokinetics of anthocyanins and antioxidant effects after the consumption of anthocyanin-rich acai juice and pulp (Euterpe oleracea Mart.) in human healthy volunteers.

The acai berry is the fruit of the acai palm and is traditionally consumed in Brazil but has gained popularity abroad as a food and functional ingredient, yet little information exists on its health effect in humans. This study was performed as an acute four-way crossover clinical trial with acai pulp and clarified acai juice compared to applesauce and a non-antioxidant beverage as controls. Healthy volunteers (12) were dosed at 7 mL/kg of body weight after a washout phase and overnight fast, and plasma was repeatedly sampled over 12 h and urine over 24 h after consumption. Noncompartmental pharmacokinetic analysis of total anthocyanins quantified as cyanidin-3-O-glucoside showed Cmax values of 2321 and 1138 ng/L at t max times of 2.2 and 2.0 h, and AUC last values of 8568 and 3314 ng h L(-1) for pulp and juice, respectively. Nonlinear mixed effect modeling identified dose volume as a significant predictor of relative oral bioavailability in a negative nonlinear relationship for acai pulp and juice. Plasma antioxidant capacity was significantly increased by the acai pulp and applesauce. Individual increases in plasma antioxidant capacity of up to 2.3- and 3-fold for acai juice and pulp, respectively were observed. The antioxidant capacity in urine, generation of reactive oxygen species, and uric acid concentrations in plasma were not significantly altered by the treatments. Results demonstrate the absorption and antioxidant effects of anthocyanins in acai in plasma in an acute human consumption trial.

Chemical Composition, Antioxidant Properties, and Thermal Stability of a Phytochemical Enriched Oil from Acai (Euterpe oleracea Mart.)

Phenolic compounds present in crude oil extracts from acai fruit ( Euterpe oleracea) were identified for the first time. The stability of acai oil that contained three concentrations of phenolics was evaluated under short- and long-term storage for lipid oxidation and phenolic retention impacting antioxidant capacity. Similar to acai fruit itself, acai oil isolates contained phenolic acids such as vanillic acid (1,616 +/- 94 mg/kg), syringic acid (1,073 +/- 62 mg/kg), p-hydroxybenzoic acid (892 +/- 52 mg/kg), protocatechuic acid (630 +/- 36 mg/kg), and ferulic acid (101 +/- 5.9 mg/kg) at highly enriched concentrations in relation to acai pulp as well as (+)-catechin (66.7 +/- 4.8 mg/kg) and numerous procyanidin oligomers (3,102 +/- 130 mg/kg). Phenolic acids experienced up to 16% loss after 10 weeks of storage at 20 or 30 degrees C and up to 33% loss at 40 degrees C. Procyanidin oligomers degraded more extensively (23% at 20 degrees C, 39% at 30 degrees C, and 74% at 40 degrees C), in both high- and low-phenolic acai oils. The hydrophilic antioxidant capacity of acai oil isolates with the highest phenolic concentration was 21.5 +/- 1.7 micromol Trolox equivalents/g, and the total soluble phenolic content was 1252 +/- 11 mg gallic acid equivalents/kg, and each decreased by up to 30 and 40%, respectively, during long-term storage. The short-term heating stability at 150 and 170 degrees C for up to 20 min exhibited only minor losses (<10%) in phenolics and antioxidant capacity. Because of its high phenolic content, the phytochemical-enriched acai oil from acai fruit offers a promising alternative to traditional tropical oils for food, supplements, and cosmetic applications.

Grapefruit‐Drug Interactions: Can Interactions With Drugs Be Avoided?

Grapefruit is rich in flavonoids, which have been demonstrated to have a preventive influence on many chronic diseases, such as cancer and cardiovascular disease. However, since the early 1990s, the potential health benefits of grapefruit have been overshadowed by the possible risk of interactions between drugs and grapefruit and grapefruit juice. Several drugs interacting with grapefruit are known in different drug classes, such as HMG‐CoA reductase inhibitors, calcium antagonists, and immunosu‐pressives. Currently known mechanisms of interaction include the inhibition of cytochrome P450 as a major mechanism, but potential interactions with P‐glycoprotein and organic anion transporters have also been reported. This review is designed to provide a comprehensive summary of underlying mechanisms of interaction and human clinical trials performed in the area of grapefruit drug interactions and to point out possible replacements for drugs with a high potential for interactions.

Oncogenic microRNA-27a is a target for anticancer agent methyl 2-cyano-3,11-dioxo-18β-olean-1,12-dien-30-oate in colon cancer cells

Methyl 2‐cyano‐3,11‐dioxo‐18β‐olean‐1,12‐dien‐30‐oate (CDODA‐Me) is a synthetic derivative of glycyrrhetinic acid, a triterpenoid phytochemical found in licorice extracts. CDODA‐Me inhibited growth of RKO and SW480 colon cancer cells and this was accompanied by decreased expression of Sp1, Sp3 and Sp4 protein and mRNA and several Sp‐dependent genes including survivin, vascular endothelial growth factor (VEGF), and VEGF receptor 1 (VEGFR1 or Flt‐1). CDODA‐Me also induced apoptosis, arrested RKO and SW480 cells at G2/M, and inhibited tumor growth in athymic nude mice bearing RKO cells as xenografts. CDODA‐Me decreased expression of microRNA‐27a (miR‐27a), and this was accompanied by increased expression of 2 miR‐27a‐regulated mRNAs, namely ZBTB10 (an Sp repressor) and Myt‐1 which catalyzes phosphorylation of cdc2 to inhibit progression of cells through G2/M. Both CDODA‐Me and antisense miR‐27a induced comparable responses in RKO and SW480 cells, suggesting that the potent anticarcinogenic activity of CDODA‐Me is due to repression of oncogenic miR‐27a.

Anticarcinogenic effects of polyphenolics from mango (Mangifera indica) varieties.

Many polyphenolics contained in mango have shown anticancer activity. The objective of this study was to compare the anticancer properties of polyphenolic extracts from several mango varieties (Francis, Kent, Ataulfo, Tommy Atkins, and Haden) in cancer cell lines, including Molt-4 leukemia, A-549 lung, MDA-MB-231 breast, LnCap prostate, and SW-480 colon cancer cells and the noncancer colon cell line CCD-18Co. Cell lines were incubated with Ataulfo and Haden extracts, selected on the basis of their superior antioxidant capacity compared to the other varieties, where SW-480 and MOLT-4 were statistically equally most sensitive to both cultivars followed by MDA-MB-231, A-549, and LnCap in order of decreasing efficacy as determined by cell counting. The efficacy of extracts from all mango varieties in the inhibition of cell growth was tested in SW-480 colon carcinoma cells, where Ataulfo and Haden demonstrated superior efficacy, followed by Kent, Francis, and Tommy Atkins. At 5 mg of GAE/L, Ataulfo inhibited the growth of colon SW-480 cancer cells by approximately 72% while the growth of noncancer colonic myofibroblast CCD-18Co cells was not inhibited. The growth inhibition exerted by Ataulfo and Haden polyphenolics in SW-480 was associated with an increased mRNA expression of pro-apoptotic biomarkers and cell cycle regulators, cell cycle arrest, and a decrease in the generation of reactive oxygen species. Overall, polyphenolics from several mango varieties exerted anticancer effects, where compounds from Haden and Ataulfo mango varieties possessed superior chemopreventive activity.

MicroRNA-27a Indirectly Regulates Estrogen Receptor α Expression and Hormone Responsiveness in MCF-7 Breast Cancer Cells

MicroRNA-27a (miR-27a) is expressed in MCF-7 breast cancer cells, and antisense miR-27a (as-miR-27a) induces ZBTB10, a specificity protein (Sp) repressor. Both as-miR-27a and overexpression of ZBTB10 decreased Sp1, Sp3, and Sp4 mRNA and protein expression in MCF-7 cells, and this was also accompanied by decreased levels of estrogen receptor alpha (ERalpha) mRNA and protein. RNA interference studies confirmed that basal expression of ERalpha was dependent on Sp1 but not Sp3 or Sp4 in MCF-7 cells. as-miR-27a and overexpression of ZBTB10 inhibited 17beta-estradiol (E2)-induced transactivation in MCF-7 cells, and this was accompanied by decreased binding of Sp and ER proteins in cell lysates to oligonucleotides containing GC-rich motifs or estrogen-responsive elements, respectively. as-miR-27a and overexpression of ZBTB10 arrested MCF-7 cells in G(0)/G(1) and inhibited E2-induced G(0)/G(1) to S phase progression. as-miR-27a induced only a minimal increase in Myt-1, another miR-27a regulated gene, and this was not accompanied by Myt-1-dependent G(2)/M arrest as observed previously in ER-negative MDA-MB-231 breast cancer cells. Thus, miR-27a indirectly regulates E2-responsiveness in MCF-7 cells through suppression of ZBTB10, thereby enhancing expression of ERalpha.

Protective Effects of Standardized Pomegranate (Punica granatum L.) Polyphenolic Extract in Ultraviolet-Irradiated Human Skin Fibroblasts

Exposure to ultraviolet (UV) radiation has been associated with several acute and chronic conditions, including sunburn, edema, hyperplasia, immunosuppression, photoaging, and skin cancer. The role of naturally occurring phytochemicals in the prevention of such UV-related conditions has captured increased interest. Pomegranate ( Punica granatum L.) is a rich source of polyphenolics, which have been shown to exert anti-inflammatory, antioxidant, and anticarcinogenic activity in numerous in vivo and in vitro studies. This work investigated potential protective effects of a pomegranate fruit extract standardized to punicalagins against UVA- and UVB-induced damage in SKU-1064 human skin fibroblast cells. Pomegranate extract (PE), in a range from 5 to 60 mg/L, was effective at protecting human skin fibroblasts from cell death following UV exposure, likely related to a reduced activation of the pro-inflammatory transcription factor NF-kappaB, a downregulation of proapoptotic caspase-3, and an increased G0/G1 phase, associated with DNA repair. Higher polyphenolic concentrations (500-10000 mg/L) were needed to achieve a significant reduction in UV-induced reactive oxygen species levels and increased intracellular antioxidant capacity (from 1.9 to 8.6 muM Trolox equivalents/mL). Results from this study demonstrate the protective effects of PE against UVA- and UVB-induced cell damage and the potential use of pomegranate polyphenolics in topical applications.

Resveratrol and quercetin in combination have anticancer activity in colon cancer cells and repress oncogenic microRNA-27a.

Resveratrol and quercetin (RQ) in combination (1:1 ratio) previously inhibited growth in human leukemia cells. This study investigated the anticancer activity of the same mixture in HT-29 colon cancer cells. RQ decreased the generation of reactive oxygen species (ROS) by up to 2.25-fold and increased the antioxidant capacity by up to 3-fold in HT-29 cells (3.8–60 μg/mL), whereas IC50 values for viability were 18.13, 18.73, and 11.85 μg/mL, respectively. RQ also induced caspase-3-cleavage (2-fold) and increased PARP cleavage. Specificity protein (Sp) transcription factors are overexpressed in colon and other cancers and regulate genes required for cell proliferation survival and angiogenesis. RQ treatment decreased the expression of Sp1, Sp3, and Sp4 mRNA and this was accompanied by decreased protein expression. Moreover, the Sp-dependent antiapoptotic survival gene survivin was also significantly reduced, both at mRNA and protein levels. RQ decreased microRNA-27a (miR-27a) and induced zinc finger protein ZBTB10, an Sp-repressor, suggesting that interactions of RQ with the miR-27a-ZBTB10-axis play a role in Sp downregulation. This was confirmed by transfection of cells with the specific mimic for miR-27a, which partially reversed the effects of RQ. These findings are consistent with previous studies on botanical anticancer agents in colon cancer cells.

Betulinic acid decreases ER‐negative breast cancer cell growth in vitro and in vivo: Role of Sp transcription factors and microRNA‐27a:ZBTB10

Betulinic acid (BA), a pentacyclic triterpenoid isolated from tree bark is cytotoxic to cancer cells. There is evidence that specificity proteins (Sps), such as Sp1, Sp3, and Sp4, are overexpressed in tumors and contribute to the proliferative and angiogenic phenotype associated with cancer cells. The objective of this study was to determine the efficacy of BA in decreasing the Sps expression and underlying mechanisms. Results show that BA decreased proliferation and induced apoptosis of estrogen‐receptor‐negative breast cancer MDA‐MB‐231 cells. The BA‐induced Sp1, Sp3, and Sp4 downregulation was accompanied by increased zinc finger ZBTB10 expression, a putative Sp‐repressor and decreased microRNA‐27a levels, a microRNA involved in the regulation of ZBTB10. Similar results were observed in MDA‐MB‐231 cells transfected with ZBTB10 expression plasmid. BA induced cell cycle arrest in the G2/M phase and increased Myt‐1 mRNA (a microRNA‐27a target gene), which causes inhibition in G2/M by phosphorylation of cdc2. The effects of BA were reversed by transient transfection with a mimic of microRNA‐27a. In nude mice with xenografted MDA‐MB‐231 cells, tumor size and weight were significantly decreased by BA treatment. In tumor tissue, ZBTB10 mRNA was increased while mRNA and protein of Sp1, Sp3 and Sp4, as well as mRNA of vascular endothelial growth factor receptor (VEGFR), survivin and microRNA‐27a were decreased by BA. In lungs of xenografted mice, human β2‐microglobulin mRNA was decreased in BA‐treated animals. These results show that the anticancer effects of BA are at least in part based on interactions with the microRNA‐27a‐ZBTB10‐Sp‐axis causing increased cell death.