Phytoestrogens are a group of plant-derived compounds with an estrogen-like activity. In mammalians, phytoestrogens bind to the estrogen receptor (ER) and participate in the regulation of cell growth and gene transcription. There are several reports of the cytotoxic effects of phytoestrogens in different cancer cell lines. The aim of this study was to measure the phytoestrogen activity against breast cancer cells with different levels of ER expression and to elucidate the molecular pathways regulated by the leader compound. Methods used in the study include immunoblotting, transfection with a luciferase reporter vector, and a MTT test. We demonstrated the absence of a significant difference between ER+ and ER- breast cancer cell lines in their response to cytotoxic stimuli: treatment with high doses of phytoestrogens (apigenin, genistein, quercetin, naringenin) had the same efficiency in ER-positive and ER-negative cells. Incubation of breast cancer cells with apigenin revealed the highest cytotoxicity of this compound; on the contrary, naringenin treatment resulted in a low cytotoxic activity. It was shown that high doses of apigenin (50 μM) do not display estrogen-like activity and can suppress ER activation by 17β-estradiol. Cultivation of HER2-positive breast cancer SKBR3 cells in the presence of apigenin resulted in a decrease in HER2/neu expression, accompanied by cleavage of an apoptosis substrate PARP. Therefore, the cytotoxic effects of phytoestrogens are not associated with the steroid receptors of breast cancer cells. Apigenin was found to be the most effective phytoestrogen that strongly inhibits the growth of breast cancer cells, including HER2-positive ones.

Programmed death ligand 1 (PD-L1) is expressed by many cancer cell types, as well as by activated T cells and antigen-presenting cells. Constitutive and inducible PD-L1 expression contributes to immune evasion by breast cancer (BC) cells. We show here that the dietary phytochemical apigenin inhibited interferon (IFN)-γ-induced PD-L1 upregulation by triple-negative MDA-MB-468 BC cells, HER2(+) SK-BR-3 BC cells, and 4T1 mouse mammary carcinoma cells, as well as human mammary epithelial cells, but did not affect constitutive PD-L1 expression by triple-negative MDA-MB-231 BC cells. IFN-β-induced expression of PD-L1 by MDA-MB-468 cells was also inhibited by apigenin. In addition, luteolin, the major metabolite of apigenin, inhibited IFN-γ-induced PD-L1 expression by MDA-MB-468 cells. Apigenin-mediated inhibition of IFN-γ-induced PD-L1 expression by MDA-MB-468 and 4T1 cells was associated with reduced phosphorylation of STAT1, which was early and transient at Tyr701 and sustained at Ser727. Apigenin-mediated inhibition of IFN-γ-induced PD-L1 expression by MDA-MB-468 cells also increased proliferation and interleukin-2 synthesis by PD-1-expressing Jurkat T cells that were co-cultured with MDA-MB-468 cells. Apigenin therefore has the potential to increase the vulnerability of BC cells to T cell-mediated anti-tumor immune responses.

Flavonoids are a group of polyphenols ubiquitously present in vegetables, fruits and herbal products, despite various known pharmacological activities, few researches have been done about the interaction of flavonoids with breast cancer resistance protein (BCRP). The present study was designed to investigate the inhibitory effects of 99 flavonoids on BCRP in vitro and in vivo and to clarify structure-activity relationships of flavonoids with BCRP. Eleven flavonoids, including amentoflavone, apigenin, biochanin A, chrysin, diosimin, genkwanin, hypericin, kaempferol, kaempferide, licochalcone A and naringenin, exhibited significant inhibition (>50%) on BCRP in BCRP-MDCKII cells, which reduced the BCRP-mediated efflux of doxorubicin and temozolomide, accordingly increased their cytotoxicity. In addition, co-administration of mitoxantrone with the 11 flavonoids increased the AUC_0-t of mitoxantrone in different extents in rats. Among them, chrysin increased the AUC_0-t most significantly, by 81.97%. Molecular docking analysis elucidated the inhibition of flavonoids on BCRP might be associated with Pi-Pi stacked interactions and/or potential Pi-Alkyl interactions, but not conventional hydrogen bonds. The pharmacophore model indicated the aromatic ring B, hydrophobic groups and hydrogen bond acceptors may play critical role in the potency of flavonoids inhibition on BCRP. Thus, our findings would provide helpful information for predicting the potential risks of flavonoid-containing food/herb-drug interactions in humans.

Drug resistance in chemotherapy is a serious obstacle for the successful treatment of cancer. Drug resistance is caused by various factors, including the overexpression of P‑glycoprotein (P‑gp, MDR1). The development of new, useful compounds that overcome drug resistance is urgent. Apigenin, a dietary flavonoid, has been reported as an anticancer drug in vivo and in vitro. In the present study, we investigated whether apigenin is able to reverse drug resistance using adriamycin‑resistant breast cancer cells (MCF‑7/ADR). In our experiments, apigenin significantly decreased cell growth and colony formation in MCF‑7/ADR cells and parental MCF‑7 cells. This growth inhibition was related to the accumulation of cells in the sub‑G0/G1 apoptotic population and an increase in the number of apoptotic cells. Apigenin reduced the mRNA expression of multidrug resistance 1 (MDR1) and multidrug resistance‑associated proteins (MRPs) in MCF‑7/ADR cells. Apigenin also downregulated the expression of P‑gp. Apigenin reversed drug efflux from MCF‑7/ADR cells, resulting in rhodamine 123 (Rho123) accumulation. Inhibition of drug resistance by apigenin is related to the suppression of the signal transducer and activator of transcription 3 (STAT3) signaling pathway. Apigenin decreased STAT3 activation (p‑STAT3) and its nuclear translocation and inhibited the secretion of VEGF and MMP‑9, which are STAT3 target genes. A STAT3 inhibitor, JAK inhibitor I and an HIF‑1α inhibitor decreased cell growth in MCF‑7 and MCF‑7/ADR cells. Taken together, these results demonstrate that apigenin can overcome drug resistance.

The green beet (Beta vulgaris var. cicla L.) and red beetroot (B. vulgaris var. rubra L.) contain phytochemicals that have beneficial effects on human health. Specifically, the green beet contains apigenin, vitexin, vitexin-2-O-xyloside and vitexin-2-O-rhamnoside, while the red beetroot is a source of betaxanthins and betacyanins. These phytochemicals show considerable antioxidant activity, as well as antiinflammatory and antiproliferative activities. Vitexin-2-O-xyloside, in combination with betaxanthins and betacyanins, exerts antiproliferative activity in breast, liver, colon and bladder cancer cell lines, through the induction of both intrinsic and extrinsic apoptotic pathways. A significant body of evidence also points to the role of these phytochemicals in the downregulation of the pro-survival genes, baculoviral inhibitor of apoptosis repeat-containing 5 and catenin beta-1, as well as the genes controlling angiogenesis, hypoxia inducible factor 1A and vascular endothelial growth factor A. The multi-target action of these phytochemicals enhances their anticancer activity. Vitexin-2-O-xyloside, betaxanthins and betacyanins can be used in combination with conventional anticancer drugs to reduce their toxicity and overcome the multidrug resistance of cancer cells. In this review, we describe the molecular mechanisms that enable these dietary phytochemicals to block the proliferation of tumor cells and inhibit their pro-survival pathways. Copyright © 2017 John Wiley & Sons, Ltd.

In the course of our screening for natural estrogenic compounds from Occidental medicinal herbs, the extracts of several herbs were found to show proliferative activity in MCF-7 (an estrogen-sensitive breast cancer cell line). Among these active herbs, the methanolic extract from the aerial parts of Petroselinum crispum (parsley) showed potent estrogenic activity, which was equal to that of isoflavone glycosides from soybean. Through bioassay-guided separation, we isolated several flavone glycosides and a new flavone glycoside, 6"-acetylapiin, with estrogenic activity together with a new monoterpene glucoside, petroside. The structures of 6"-acetylapiin and petroside were characterized by the chemical and physicochemical evidence. Estrogenic activities of these flavone glycosides were found to be enhanced by removal of their glycoside moieties. The EC50 values (concentration needed to enhance the MCF-7 proliferation 50% compared to non-estrogen treated cell) of their aglycones are as follows, apigenin (1.0 microM), diosmetin (2.9 microM), and kaempferol (0.56 microM). The estrogenic activities of these flavones are nearly equal to those of the isoflavones, daidzein (0.61 microM) and genistein (0.60 microM). The methanolic extract of parsley, apiin, and apigenin restored the uterus weight in ovariectomized mice when orally administered for consecutive 7 days.

AKR1B10 is an NADPH-dependent reductase that plays an important function in several physiological reactions such as the conversion of retinal to retinol, reduction of isoprenyl aldehydes, and biotransformation of procarcinogens and drugs. A growing body of evidence points to the important role of the enzyme in the development of several types of cancer (e.g., breast, hepatocellular), in which it is highly overexpressed. AKR1B10 is regarded as a therapeutic target for the treatment of these diseases, and potent and specific inhibitors may be promising therapeutic agents. Several inhibitors of AKR1B10 have been described, but the area of natural plant products has been investigated sparingly. In the present study almost 40 diverse phenolic compounds and alkaloids were examined for their ability to inhibit the recombinant AKR1B10 enzyme. The most potent inhibitors-apigenin, luteolin, and 7-hydroxyflavone-were further characterized in terms of IC50, selectivity, and mode of action. Molecular docking studies were also conducted, which identified putative binding residues important for the interaction. In addition, cellular studies demonstrated a significant inhibition of the AKR1B10-mediated reduction of daunorubicin in intact cells by these inhibitors without a considerable cytotoxic effect. Although these compounds are moderately potent and selective inhibitors of AKR1B10, they constitute a new structural type of AKR1B10 inhibitor and may serve as a template for the development of better inhibitors.

Apigenin is a flavonoid, which has been proved to possess effective anti-cancer bioactivities against variety of cell lines. However, little is known about its effect on the cell-surface and the interaction between cell-surface and the reacting drug. In this study, human breast cancer line (MCF-7) was selected to be as a cell model to investigate the effects of apigenin on cell growth, proliferation, apoptosis, cellular morphology, etc. MTT assay showed that the growth inhibition induced by apigenin was in a dose-dependent manner when treated with different concentrations of apigenin while had little cytotoxic effects on human normal cells (MCF-10A). Fluorescence-based flow cytometry was used to detect cellular apoptosis and ROS production. The results showed that 80 µM apigenin could effectively induce apoptosis and overproduction of ROS in MCF-7 cells. Here, atomic force microscopy (AFM) was utilized to detect the shapes and membrane structures of MCF-7 cells at cellular or subcellular level. The results showed that the control MCF-7 cells presented typical elongated-spindle shapes with abundant pseudopodia, while after treated with apigenin, the cells shrunk and became round, the pseudopodia diminished. Moreover, the images of ultrastructure indicated that the cell membrane was composed of nanoparticles of 49 nm, but with the treated concentrations of apigenin increasing, the sizes of membrane particles significantly increased to 400 nm. These results can improve our understanding of apigenin, which can be potentially developed as a new agent for treatment of cancers.

(1) Background: Extensive research has focused on flavan-3-ols, but information about the bioactivities of green tea flavonols is limited. (2) Methods: In this study, we investigated the antioxidative, anti-inflammatory, and anticancer effects of flavonol glycosides and aglycones from green tea using in vitro cell models. The fractions rich in flavonol glycoside (FLG) and flavonol aglycone (FLA) were obtained from green tea extract after treatment with tannase and cellulase, respectively. (3) Results: FLG and FLA contained 16 and 13 derivatives, respectively, including apigenin, kaempferol, myricetin, and quercetin, determined by mass spectrometry. FLA exhibited higher radical-scavenging activity than that of FLG. FLG and FLA attenuated the levels of intracellular oxidative stress in neuron-like PC-12 cells. The treatment of RAW 264.7 murine macrophages with FLG and FLA significantly reduced the mRNA expression of inflammation-related genes in a dose-dependent manner. Furthermore, FLG and FLA treatments decreased the viability of the colon adenoma cell line DLD-1 and breast cancer cell line E0771. Moreover, the treatment with FLG or FLA combined with paclitaxel had synergistic anticancer effects on the DLD-1 cell line. (4) Conclusions: Flavonols from green tea exerted beneficial effects on health and may be superior to flavan-3-ols.

Ten compounds, neopulchellin (1), 6α- hydroxyneopulchellin (2), β-sitosterol-3-O-β-D-glucoside (3), apigenin (4), quercitin (5), eupafolin (6), kaempferol-3-methoxy-7-O-α-L-rhamnoside (7), apigenin-7-O-β-D-glucopyranoside (8), α-amyrin (9) and β-sitosterol (10), were isolated from the leaves of Gaillardia aristata by applying bioassay guided fractionation. The cytotoxicity was traced against two human cancer cell lines (breast (MCF7) and colon (HCT116)). The highest cytotoxicity was revealed by compounds 1 and 2 (isolated from chloroform extract); with IC(50) values of 0.43, 0.32 µg mL(-1) against MCF7 and 0.46, 0.34 µg mL(-1) against HCT116, respectively. Compounds 9 and 10 (isolated from the n-hexane extract) exhibited lower IC(50) values of 3.05, 2.35 µg mL(-1) against MCF7 and 3.05, 2.35 µg mL(-1) against HCT116, respectively, while compounds 4-7 obtained from the ethyl acetate extract revealed the lowest cytotoxicity. Identification of the aforementioned compounds was carried out on the basis of their physico-chemical properties and spectral analysis (UV, EI/MS, 1D and 2D).

There is a well-established association between alcohol consumption and breast cancer risk. About 4% of the breast cancers in developed countries are estimated to be attributable to drinking alcohol. The mechanism of tumor promotion by alcohol remains unknown. Recent studies from our laboratory and others showed the ability of mammary tissue to bioactivate ethanol to mutagenic/carcinogenic acetaldehyde and free radicals. Xanthine oxidoreductase (XOR) is an enzyme involved in those biotransformation processes. In the present study, we provide evidence of the ability of different natural polyphenols and of folic acid derivatives to inhibit the biotransformation of alcohol to acetaldehyde by rat breast cytosolic XOR. Folic acid and dihydrofolic acid, at concentrations of 10 microM, inhibited 100% and 84%, respectively, of the cytosolic acetaldehyde formation. Thirty-five polyphenols were tested in these initial experiments: ellagic acid, myricetin, quercetin, luteolin, and apigenin inhibited 79-95% at 10 microM concentrations. The remaining polyphenols were either less potent or noninhibitory of acetaldehyde formation at similar concentrations in these screening tests. Results are relevant to the known preventive effects of folic acid against alcohol-induced breast cancer and to their potential preventive actions if added to foods or alcoholic beverages.

We have used a tissue culture system based on breast carcinoma cell lines to investigate a large number of naturally occurring compounds and beverages for steroid hormone agonist and antagonist activity. The cell lines used, T-47D and BT-474, produce prostate specific antigen (PSA) upon stimulation with androgens, progestins, glucocorticoids and mineralocorticoids. This biomarker is secreted and can be measured in the tissue culture supernatant with very high sensitivity by an immunofluorometric procedure. Steroid hormone antagonist activity can be assessed with the same system by adding the candidate antagonist first and then stimulating the cells with a known agonist. By using this system we have identified three natural compounds, apigenin, naringenin and syringic acid which exhibited weak progestational activity and eleven other compounds which exhibited weak antiandrogenic/antiprogestational activity. Our study indicates that a significant number of natural compounds have the ability to bind to steroid hormone receptors and act as weak blockers. A fewer number of compounds not only bind to the receptors but they also mediate transcriptional activity, acting as agonists. The agonists and antagonists were active at levels around 10(-5) M, in accordance with previous reports for other phytochemicals. In comparison to synthetic and natural steroid hormones, the biological activity of these compounds is weaker by a factor of approximately 10(4)-fold.

The flavone apigenin and the mycotoxin zearalenone are two major compounds found in the human diet which bind estrogen receptors (ERs), and therefore influence ER activity. However, the underlying mechanisms are not well known. To unravel the molecular mechanisms that could explain the differential effect of zearalenone and apigenin on ER-positive breast cancer cell proliferation, gene-reporter assays, chromatin immunoprecipitation (ChIP) experiments, proliferation assays and transcriptomic analysis were performed. We found that zearalenone and apigenin transactivated ERs and promoted the expression of estradiol (E2)-responsive genes. However, zearalenone clearly enhanced cellular proliferation, while apigenin appeared to be antiestrogenic in the presence of E2 in both ER-positive breast cancer cell lines, MCF-7 and T47D. The transcriptomic analysis showed that both compounds regulate gene expression in the same way, but with differences in intensity. Two major sets of genes were identified; one set was linked to the cell cycle and the other set was linked to stress response and growth arrest. Our results show that the transcription dynamics in gene regulation induced by apigenin were somehow different with zearalenone and E2 and may explain the differential effect of these compounds on the phenotype of the breast cancer cell. Together, our results confirmed the potential health benefit effect of apigenin, while zearalenone appeared to be a true endocrine-disrupting compound.

Natural flavonoids with methoxy substitutions are metabolized by CYP1 enzymes to yield the corresponding demethylated products. The present study aimed to characterize the metabolism and further antiproliferative activity of the hydroxylated flavonoids apigenin, luteolin, scutellarein, kaempferol and quercetin in CYP1 recombinant enzymes and in the CYP1 expressing cell lines MCF7 and MDA-MB-468, respectively. Apigenin was converted to luteolin and scutellarein, whereas kaempferol was metabolized only to quercetin by recombinant CYP1 enzymes. Luteolin metabolism yielded 6 hydroxyluteolin only by recombinant CYP1B1, whereas CYP1A1 and CYP1A2 were not capable of metabolizing this compound. Molecular modeling demonstrated that CYP1B1 favored the A ring orientation of apigenin and luteolin to the heme group compared with CYP1A1. The IC50 of the compounds luteolin, scutellarein and 6 hydroxyluteolin was significantly lower in MDA-MB-468, MCF7 and MCF10A cells compared with that of apigenin. Similarly, the IC50 of quercetin in MDA-MB-468 cells was significantly lower compared with that of kaempferol. The most potent compound was luteolin in MDA-MB-468 cells (IC50 = 2 ± 0.3 μM). In the presence of the CYP1-inhibitors α-napthoflavone and/or acacetin, luteolin activation was lessened. Taken collectively, the data demonstrate that the metabolism of hydroxylated flavonoids by cytochrome P450 CYP1 enzymes, notably CYP1A1 and CYP1B1, can enhance their antiproliferative activity in breast cancer cells. In addition, this antiproliferative activity is attributed to the combined action of the parent compound and the corresponding CYP1 metabolites.

Two series of apigenin [5,7-dihydroxy-2-(4-hydroxyphenyl)-4H-chromen-4-one] derivatives, 3a-3j and 4a-4j, were synthesized. The apigenin and alkyl amines moieties of these compounds were separated by C₂ or C₃ spacers, respectively. The chemical structures of the apigenin derivatives were confirmed using ¹H-NMR, ¹³C-NMR, and electrospray ionization mass spectroscopy. The in vitro antibacterial and antiproliferative activities of all synthesized compounds were determined. Among the tested compounds, 4a-4j displayed significant antibacterial activity against the tested strains (Staphylococcus aureus, Bacillus subtilis, Escherichia coli, and Pseudomonas aeruginosa). Additionally, 4i showed the best inhibitory activity with minimum inhibitory concentrations of 1.95, 3.91, 3.91, and 3.91 μg/mL against S. aureus, B. subtilis, E. coli, and P. aeruginosa, respectively. The antiproliferative activity of the apigenin derivatives was evaluated by an MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] assay. We determined that 4a-4j displayed better growth inhibition activity against four human cancer cell lines, namely, human lung (A549), human cervical (HeLa), human hepatocellular liver (HepG2), and human breast (MCF-7) cancer cells, than the parent apigenin. Compound 4j was found to be the most active antiproliferative compound against the selected cancer cells. Structure-activity relationships were also discussed based on the obtained experimental data.

In the present study, aqueous extracts of Salvia africana, Salvia officinalis 'Icterina' and Savia mexicana origin were screened for their phenolic composition and for antibacterial, antioxidant, anti-inflammatory and cytotoxic properties. The three aqueous extracts contained distinct phenolic compounds, with S. africana presenting the highest total levels (231.6 ± 7.5 μg/mg). Rosmarinic acid was the dominant phenolic compound in all extracts, yet that of S. africana origin was characterized by the present of yunnaneic acid isomers, which overall accounted for about 40% of total phenolics. In turn, S. officinalis 'Icterina' extract presented glycosidic forms of apigenin, luteolin and scuttelarein, and the one obtained from S. mexicana contained several simple caffeic acid derivatives. S. africana aqueous extract exhibited high antioxidant potential in four methods, namely the DPPH^• (2,2-diphenyl-1-picrylhydrazyl) scavenging ability, iron-reducing power, inhibition of β-carotene bleaching and of thiobarbituric acid reactive substances (TBARS), for which EC₅₀ values were equal or only 1.3-3.1 higher than those of the standard compounds. Moreover, this extract was able to lower the levels of nitric oxide (NO) production in lipopolysaccharide (LPS)-activated RAW 264.7 macrophages (EC₅₀ = 47.8 ± 2.1 μg/mL). In addition, the three sage aqueous extracts showed promising cytotoxic effect towards hepatocellular HepG2, cervical HeLa, and breast carcinoma cells MCF-7. Overall this study highlights the potential of three little-exploited Salvia species, with commercial value for applications in food or pharmaceutical industries.

The expression of transmembrane transporter multidrug resistance-associated protein 1 (MRP1) confers the multidrug-resistant phenotype (MDR) on cancer cells. Since the activity of the other MDR transporter, P-glycoprotein, is sensitive to membrane perturbation, we aimed to check whether the changes in lipid bilayer properties induced by flavones (apigenin, acacetin) and flavonols (morin, myricetin) were related to their MRP1 inhibitory activity. All the flavonoids inhibited the efflux of MRP1 fluorescent substrate from human erythrocytes and breast cancer cells. Morin was also found to stimulate the ATPase activity of erythrocyte ghosts. All flavonoids intercalated into phosphatidylcholine bilayers as judged by differential scanning calorimetry and fluorescence spectroscopy with the use of two carbocyanine dyes. The model of an intramembrane localization for flavones and flavonols was proposed. No clear relationship was found between the membrane-perturbing activity of flavonoids and their potency to inhibit MRP1. We concluded that mechanisms other than perturbation of the lipid phase of membranes were responsible for inhibition of MRP1 by the flavonoids.

Ephedrae herba suppresses hepatocyte growth factor-induced cancer cell motility by inhibiting tyrosine phosphorylation of the hepatocyte growth factor receptor, c-Met, and the PI3K/Akt pathway. Moreover, Ephedrae herba directly inhibits the tyrosine-kinase activity of c-Met. Ephedrine-type alkaloids, which are the active component of Ephedrae herba, do not affect hepatocyte growth factor-c-Met-Akt signalling, prompting us to study other active molecules in the herb. We recently discovered herbacetin glycosides and found that their aglycon, herbacetin, inhibits hepatocyte growth factor-c-Met-Akt signalling. This study revealed a novel biological activity of herbacetin. Herbacetin suppressed hepatocyte growth factor-induced motility in human breast cancer MDA-MB-231 cells by inhibiting c-Met and Akt phosphorylation and directly inhibiting c-Met tyrosine kinase activity. The effects of herbacetin were compared to those of kaempferol, apigenin, and isoscutellarein, all of which have similar structures. Herbacetin inhibition of hepatocyte growth factor-induced motility was the strongest of those for the tested flavonols, and only herbacetin inhibited the hepatocyte growth factor-induced phosphorylation of c-Met. These data suggest that herbacetin is a novel Met inhibitor with a potential utility in cancer therapeutics.

Elsholtzia splendens is a copper-tolerant plant species which grows on copper deposits in China. The generation of a valuable E. splendens biomass on specific contaminated sites has become one of the promising phytotechnologies. The simultaneous separations of apigenin, luteolin, and rosmarinic acid yielded in the ethyl acetate extracts of the flowering aerial parts was achieved by the use of a macroporous resin, polyamide, and silicagel columns during chromatography. Chemical identification confirmed the structures based on the spectra of FTIR, NMR, and HPLC/ESI-MS. The isolated compounds of purity above 98.3% were evaluated for their in vitro cytotoxic activities against human cancer cell lines including A549 (non-small lung), A431 (skin), and Bcap37 (breast). Among these compounds, luteolin and apigenin presented the best cytotoxic activities against A549, A431, and Bcap37 cells and, therefore, both could be the valuable products for the post-harvest processing of E. splendens biomass.

The Axl receptor tyrosine kinase (RTK), along with Tyro 3 and Mer, belongs to the TAM subfamily that promotes survival, stimulates proliferation and/or inhibits apoptosis. In various types of human cancer, including breast, lung and prostate cancer, Axl expression is increased and correlates with an advanced clinical stage. In this study, we examined whether apigenin has an effect on Axl expression, which in turn can affect cell proliferation. The treatment of the non‑small cell lung cancer (NSCLC) cells, A549 and H460, with apigenin decreased Axl mRNA and protein expression in a dose‑dependent manner. Axl promoter activity was also inhibited by apigenin, indicating that apigenin suppressed Axl expression at the transcriptional level. Upon treatment with apigenin, the viability of both the A549 and H460 cells was gradually decreased and the anti-proliferative effects were further confirmed by the dose‑dependent decrease in the clonogenic ability of the apigenin‑treated cells. Subsequently, we found that the viability and clonogenic ability of the cells treated with apigenin was less or more affected by transfection of the cells with a Axl-expressing plasmid or Axl targeting siRNA, compared to transfection with the empty vector or control siRNA, respectively. In addition, apigenin increased the expression of p21, a cyclin-dependent kinase inhibitor, but reduced the expression of X-linked inhibitor of apoptosis protein (XIAP). These cell cycle arrest and pro-apoptotic effects of apigenin were also attenuated or augmented by the up- or downregulation of Axl expression, respectively, which suggests that Axl is a novel target of apigenin through which it exerts its inhibitory effects on cell proliferation. Taken together, our data indicate that apigenin downregulates Axl expression, which subsequently results in the inhibition of NSCLC cell proliferation through the increase and decrease of p21 and XIAP expression, respectively.

Turnera diffusa Willd, commonly known as Damiana, is employed in traditional medicine as a stimulant, aphrodisiac, and diuretic. Its leaves and stems are used for flavoring and infusion. Damiana is considered to be safe for medicinal use by the FDA. Pharmacological studies have established the hypoglycemic, antiaromatase, prosexual, estrogenic, antibacterial, and antioxidant activity of T. diffusa. The aim of the present study was to evaluate the possible cytotoxic effect of extracts and organic fractions of this plant on five tumor cell lines (SiHa, C-33, Hep G2, MDA-MB-231, and T-47D) and normal human fibroblasts. The results show that the methanolic extract (TdM) displayed greater activity on MDA-MB-231 breast cancer cells (with an IC50 of 30.67 μg/mL) than on the other cancer cell lines. Four organic fractions of this extract exhibited activity on this cancer cell line. In the most active fraction (F4), two active compounds were isolated, arbutin (1) and apigenin (2). This is the first report of a cytotoxic effect by T. diffusa on cancer cells. The IC50 values suggest that the methanolic extract of T. diffusa has potential as an anticancer therapy.

This study examined the comparative anticancer effects of flavonoids and diazepam in the cultured cancer cells. In the SNU-C4 colorectal and MDA-MB-231 breast adenocarcinoma cells, apigenin and fisetin, flavonoids, and diazepam inhibited cancer cell survival concentration and incubation-time dependently. Diazepam consistently inhibited FAS activity, a known anticancer mechanism of flavonoids, in a concentration dependent manner. Unlike diazepam, in highly aggressive breast MDA-MB-231 cells known to have a nuclear/perinuclear located PBR, PK11195, a specific PBR ligand enhanced the proliferation of cells, and the proliferative effect of PK11195 was reversed by an addition of lovastatin, a HMG-CoA reductase inhibitor. Diazepam- and flavonoids-induced cytotoxic activity in both cancer cell lines was not reduced by the addition of 5-fluorouracil (5-FU), a chemotherapeutic agent. Like flavonoids, diazepam inhibited the release of vascular endothelial growth factor (VEGF) and granulocyte-macrophage-colony stimulating factor (GM-CSF) into supernatants of cultured in the SNU-C4 and MDA-MB-231 cells. In conclusion, this study provided in vitro information on the safe use of sedative in oncologic patients.

Evidence indicates sorghum may be protective against colon cancer; however, the mechanisms are unknown. Estrogen is believed to protect against colon cancer development by inducing apoptosis in damaged nonmalignant colonocytes. Three sorghum extracts (white, red, and black) were screened for estrogenic activity using cell models expressing estrogen receptor α (ER-α; MCF-7 breast cancer cells) and β [ER-β; nonmalignant young adult mouse colonocytes (YAMC)]. Black and white sorghum extracts had significant estrogenic activity mediated through both estrogen receptors at 1-5 and 5-10 μg/mL, respectively; but red sorghum did not. Activation of ER-β in YAMC reduced cell growth via induction of apoptosis. Only the black and red sorghums contained 3-deoxyanthocyanins; however, these compounds were non-estrogenic. Flavones with estrogenic properties, luteolin (0.41-2.12 mg/g) and apigenin (1.1-1.4 mg/g), and their O-methyl derivatives (0.70-0.95 mg/g) were detected in white and black sorghums, but not in the red sorghum. On the other hand, naringenin, a flavanone known to interfere with transcriptional activities of estrogen, was only detected in the red sorghum extract (as its 7-O-glycoside) at relatively high concentration (11.8 mg/g). Sorghum flavonoid composition has important implications on possible modes of chemoprotection by sorghum against colon carcinogenesis.

Acquired resistance to doxorubicin is a major hurdle in triple-negative breast cancer (TNBC) therapy, emphasizing the need to identify improved strategies. Apigenin and other structurally related dietary flavones are emerging as potential chemo-sensitizers, but their effect on three-dimensional TNBC spheroid models has not been investigated. We previously showed that apigenin associates with heterogeneous ribonuclear protein A2/B1 (hnRNPA2), an RNA-binding protein involved in mRNA and co-transcriptional regulation. However, the role of hnRNPA2 in apigenin chemo-sensitizing activity has not been investigated. Here, we show that apigenin induced apoptosis in TNBC spheroids more effectively than apigenin-glycoside, owing to higher cellular uptake. Moreover, apigenin inhibited the growth of TNBC patient-derived organoids at an in vivo achievable concentration. Apigenin sensitized spheroids to doxorubicin-induced DNA damage, triggering caspase-9-mediated intrinsic apoptotic pathway and caspase-3 activity. Silencing of hnRNPA2 decreased apigenin-induced sensitization to doxorubicin in spheroids by diminishing apoptosis and partly abrogated apigenin-mediated reduction of ABCC4 and ABCG2 efflux transporters. Together these findings provide novel insights into the critical role of hnRNPA2 in mediating apigenin-induced sensitization of TNBC spheroids to doxorubicin by increasing the expression of efflux transporters and apoptosis, underscoring the relevance of using dietary compounds as a chemotherapeutic adjuvant.

Keywords: Apoptosis; Breast cancer; Caspase-3; Caspases; Cell death; Dietary flavonoids; Drug bioavailability.