Synthesis of Flavonols and Assessment of Their Biological Activity as Anticancer Agents.

A series of flavanols were synthesized to assess their biological activity against human non-small cell lung cancer cells (A549). Among the sixteen synthesized compounds, it was observed that compounds 6k (3.14 ± 0.29 µM) and 6l (0.46 ± 0.02 µM) exhibited higher potency compared to 5-fluorouracil (5-Fu, 4.98 ± 0.41 µM), a clinical anticancer drug which was used as a positive control. Moreover, compound 6l (4'-bromoflavonol) markedly induced apoptosis of A549 cells through the mitochondrial- and caspase-3-dependent pathways. Consequently, compound 6l might be developed as a candidate for treating or preventing lung cancer.

Synthesis of selenophene-containing flavonols and 2-styrylchromones: Evaluation of their activities compared with selenophene-containing chalcones as potential anticancer agents.

Previously, we documented the synthesis and assessed the biological effects of chalcones containing selenium against HT-29 human colorectal adenocarcinoma cells, demonstrating their significant potential. As research on selenium-containing flavonoids remains limited, this article outlines our design and synthesis of three selenium-based flavonols and three 2-styrylchromones. We conducted evaluations of these compounds to determine their impact on human lung cancer cells (A549, H1975, CL1-0, and CL1-5) and their influence on normal lung fibroblast MRC5 cells. Additionally, we included selenium-based chalcones in our testing for comparative purposes. Our findings highlight that the simplest compound, designated as compound 1, exhibited the most promising performance among the tested molecules.

Design, synthesis and bioactivity evaluation of fisetin derivatives as potential anti-inflammatory agents against LPS-induced acute lung injury.

Acute lung injury (ALI) refers to a common and life-threatening disease attributed to inflammation. However, effective drug treatments have been rare for ALI. Natural products have been considered as a vital source of drug discovery which indicates that it's a workable method to find new anti-inflammatory drugs in natural products. Inspired by the various biological activities of fisetin, we reported the design and synthesis of a series of fisetin derivatives which were also evaluated for their anti-inflammatory activities in J774A.1 macrophages. Most of the obtain derivatives could effectively inhibit the release of IL-6 and TNF-α in vitro experiments without cytotoxicity. The most promising compound 5b exhibited significant in vivo anti-inflammatory activity in the model of LPS-induced ALI in mice. On the whole, this study could provide novel candidates for the treatment of ALI.

Synthesis, inverse docking-assisted identification and in vitro biological characterization of Flavonol-based analogs of fisetin as c-Kit, CDK2 and mTOR inhibitors against melanoma and non-melanoma skin cancers.

Due to hurdles, including resistance, adverse effects, and poor bioavailability, among others linked with existing therapies, there is an urgent unmet need to devise new, safe, and more effective treatment modalities for skin cancers. Herein, a series of flavonol-based derivatives of fisetin, a plant-based flavonoid identified as an anti-tumorigenic agent targeting the mammalian targets of rapamycin (mTOR)-regulated pathways, were synthesized and fully characterized. New potential inhibitors of receptor tyrosine kinases (c-KITs), cyclin-dependent kinase-2 (CDK2), and mTOR, representing attractive therapeutic targets for melanoma and non-melanoma skin cancers (NMSCs) treatment, were identified using inverse-docking, in vitro kinase activity and various cell-based anticancer screening assays. Eleven compounds exhibited significant inhibitory activities greater than the parent molecule against four human skin cancer cell lines, including melanoma (A375 and SK-Mel-28) and NMSCs (A431 and UWBCC1), with IC50 values ranging from 0.12 to < 15 µM. Seven compounds were identified as potentially potent single, dual or multi-kinase c-KITs, CDK2, and mTOR kinase inhibitors after inverse-docking and screening against twelve known cancer targets, followed by kinase activity profiling. Moreover, the potent compound F20, and the multi-kinase F9 and F17 targeted compounds, markedly decreased scratch wound closure, colony formation, and heightened expression levels of key cancer-promoting pathway molecular targets c-Kit, CDK2, and mTOR. In addition, these compounds downregulated Bcl-2 levels and upregulated Bax and cleaved caspase-3/7/8 and PARP levels, thus inducing apoptosis of A375 and A431 cells in a dose-dependent manner. Overall, compounds F20, F9 and F17, were identified as promising c-Kit, CDK2 and mTOR inhibitors, worthy of further investigation as therapeutics, or as adjuvants to standard therapies for the control of melanoma and NMSCs.

Synthesis of Isorhamnetin-3-O-Rhamnoside by a Three-Enzyme (Rhamnosyltransferase, Glycine Max Sucrose Synthase, UDP-Rhamnose Synthase) Cascade Using a UDP-Rhamnose Regeneration System.

Isorhamnetin-3-O-rhamnoside was synthesized by a highly efficient three-enzyme (rhamnosyltransferase, glycine max sucrose synthase and uridine diphosphate (UDP)-rhamnose synthase) cascade using a UDP-rhamnose regeneration system. The rhamnosyltransferase gene (78D1) from Arabidopsis thaliana was cloned, expressed, and characterized in Escherichia coli. The optimal activity was at pH 7.0 and 45 °C. The enzyme was stable over the pH range of 6.5 to 8.5 and had a 1.5-h half-life at 45 °C. The Vmax and Km for isorhamnetin were 0.646 U/mg and 181 µM, respectively. The optimal pH and temperature for synergistic catalysis were 7.5 and 25 °C, and the optimal concentration of substrates were assayed, respectively. The highest titer of isorhamnetin-3-O-rhamnoside production reached 231 mg/L with a corresponding molar conversion of 100%. Isorhamnetin-3-O-rhamnoside was purified and the cytotoxicity against HepG2, MCF-7, and A549 cells were evaluated. Therefore, an efficient method for isorhamnetin-3-O-rhamnoside production described herein could be widely used for the rhamnosylation of flavonoids.

Synthetic flavonols and flavones: A future perspective as anticancer agents.

A series of flavonoid derivatives, flavones (F1-F3) and flavonols (OF1-OF3) were synthesized. Their structures were confirmed through various spectroscopic techniques and elemental analysis. These were then tested for cytotoxic activity against mouse fibroblast (NIH 3T3), human endothelial cervical (HeLa) and breast (MCF7) cell lines in vitro by MTT assay. The flavonol series showed prominent potentials than the flavones. The compound OF2 in flavonols exhibited greater potentials MCF7 cell with IC50 value of 0.96µM and OF3 has 1.04µM. In contrast, the OF3 exhibited higher activity against HeLa cell with IC50 value of 0.51µM and OF2 has 1.06µM. The compounds OF2 and OF3 exhibited activity against mouse fibroblast (NIH 3T3) cell with IC50 values 2.48 and 1.24µM. The OF1 was found to be moderate to inactive against all cells. Cytotoxic screening of the tested flavones, F1 to F3 were also active against all cells but the activity was less in comparison to flavonol series of compounds suggestion the possible involvement of hydroxyl (OH) at position 3 in case of flavonols. These results indicated a cheering scaffold that may lead to innovation of potent anti-breast cancer activity.

Discovery of a Prenylated Flavonol Derivative as a Pin1 Inhibitor to Suppress Hepatocellular Carcinoma by Modulating MicroRNA Biogenesis.

Peptidyl-prolyl cis-trans isomerase Pin1 plays a crucial role in the development of human cancers. Recently, we have disclosed that Pin1 regulates the biogenesis of miRNA, which is aberrantly expressed in HCC and promotes HCC progression, indicating the therapeutic role of Pin1 in HCC therapy. Here, 7-(benzyloxy)-3,5-dihydroxy-2-(4-methoxyphenyl)-8-(3-methylbut-2-en-1-yl)-4H-chromen-4-one (AF-39) was identified as a novel Pin1 inhibitor. Biochemical tests indicate that AF-39 potently inhibits Pin1 activity with an IC50 values of 1.008 µm, and also displays high selectivity for Pin1 among peptidyl prolyl isomerases. Furthermore, AF-39 significantly suppresses cell proliferation of HCC cells in a dose- and time-dependent manner. Mechanistically, AF-39 regulates the subcellular distribution of XPO5 and increases miRNAs biogenesis in HCC cells. This work provides a promising lead compound for HCC treatment, highlighting the therapeutic potential of miRNA-based therapy against human cancer.

Structure-activity relationship and pharmacokinetic studies of 3-O-substitutedflavonols as anti-prostate cancer agents.

Thirty-eight 3-O-substituted-3',4'-dimethoxyflavonols and twenty-five 3-O-substituted-3',4',7-trimethoxyflavonols have been synthesized for systematic investigation on the structure-activity relationships of 3-O-substituted-3',4'-dimethoxyflavonols in three human prostate cancer cell models. Our findings indicate that incorporation of an appropriate amino group to 3-OH of 3',4'-dimethoxyflavonol and 3',4',7-trimethoxyflavonol through a 3- to 5-carbon linker can substantially improve the in vitro antiproliferative potency in three human prostate cancer cell models, but not in two non-neoplastic human epithelial cell models (MCF 10A and PWR-1E). 1-Methylpiperazine, pyrrolidine, and dibutylamine are optimal terminal amine groups that, in combination with a 3- to 5-carbon linker, are notably beneficial to the anti-proliferative potency of 3-O-substituted-3',4'-dimethoxyflavonols. It is worth noting that 3-O-(4-methylpiperazin-1-yl)propyl-3',4',7-trimethoxyflavonol (76) induces PC-3 cell death in a completely different way from 3-O-pyrrolidinopentyl-3',4',7-trimethoxyflavonol (81) even though they belong to 3-O-substituted-3',4',7-trimethoxyflavonols and exhibit similar potency in inhibiting PC-3 cell proliferation, suggesting that the mechanism of action for each specific 3-O-substitutedflavonol varies with different amino moiety. 3-O-(N,N-Dibutylamino)propyl-3',4'-dimethoxyflavonol (42) emerged as the most promising derivative due to its substantially improved potency in cell models, superior bioavailability in rats, and good selectivity of inhibiting prostate cancer cell proliferation over non-neoplastic human epithelial cell proliferation.

3-O-Substituted-3',4',5'-trimethoxyflavonols: Synthesis and cell-based evaluation as anti-prostate cancer agents.

Twenty-two 3-O-substituted-3',4',5'-trimethoxyflavonols have been designed and synthesized for their anti-proliferative activity towards three human prostate cancer cell lines. Our results indicate that most of them are significantly more potent than the parent 3',4',5'-trimethoxyflavonol in inhibiting the cell proliferation in PC-3 and LNCaP prostate cancer cell models. 3-O-Substituted-3',4',5'-trimethoxyflavonols have generally higher potency towards PC-3 and LNCaP cell lines than the DU145 cell line. Incorporation of an ethyl group to 3-OH of 3',4',5'-trimethoxyflavonol leads to 3-O-ethyl-3',4',5'-trimethoxyflavonol as the optimal derivative with up to 36-fold enhanced potency as compared with the corresponding lead compound 3',4',5'-trimethoxyflavonol, but with reversed PC-3 cell apoptotic response. Introduction of a dipentylaminopropyl group to 3-OH increases not only the antiproliferative potency but also the ability in activating PC-3 cell apoptosis. Our findings imply that modification on 3-OH of trimethoxyflavonol can further enhance its in vitro anti-proliferative potency and PC-3 cell apoptosis induction.

Antiproliferative activity and apoptosis induction by 3',4'-dibenzyloxyflavonol on human leukemia cells.

In this study, we investigated the effects of synthetic 3',4'-dibenzyloxyflavonol on viabilities of eight human tumor cells. It was cytotoxic against leukemia cells (HL-60, U-937, MOLT-3, K-562, NALM-6, Raji), with significant effects against P-glycoprotein-overexpressing K-562/ADR and Bcl-2-overexpressing U-937/Bcl-2 cells, but had no significant cytotoxic effects against quiescent or proliferating human peripheral blood mononuclear cells. The IC50 value for the leukemia HL-60 cells was 0.8 ± 0.1 µM. This indicates a 60-fold greater toxicity than the naturally occurring flavonol quercetin. Synthetic 3',4'-dibenzyloxyflavonol induced S phase cell cycle arrest and was a potent apoptotic inducer in human leukemia cells. Cell death was (i) mediated by the activation and the cleavage of initiator and executioner caspases; (ii) prevented by the pan-caspase inhibitor z-VAD-fmk; (iii) associated with the release of cytochrome c and with the phosphorylation of members of the mitogen activated protein kinases including p38MAPK, JNK/SAPK and ERK, and (iv) independent of the generation of reactive oxygen species. The synthetic 3',4'-dibenzyloxyflavonol is a potent cytotoxic compound against several human leukemia cells and might be useful in the development of new strategies in the fight against cancer.

A new class of flavonol-based anti-prostate cancer agents: Design, synthesis, and evaluation in cell models.

Flavonoids are a large class of polyphenolic compounds ubiquitously distributed in dietary plants with an array of biological activities. Flavonols are a major sub-class of flavonoids featuring a hydroxyl group at C-3. Certain natural flavonols, such as quercetin and fisetin, have been shown by in vitro cell-based and in vivo animal experiments to be potential anti-prostate cancer agents. However, the Achilles' heel of flavonols as drug candidates is their moderate potency and poor pharmacokinetic profiles. This study aims to explore the substitution effect of 3-OH in flavonols on the in vitro anti-proliferative potency against both androgen-sensitive and androgen-insensitive human prostate cancer cell lines. Our first lead flavonol (3',4'-dimethoxyflavonol), eight 3-O-alkyl-3',4'-dimethoxyflavonols, and six 3-O-aminoalkyl-3',4'-dimethoxyflavonols have been synthesized through aldol condensation and the Algar-Flynn-Oyamada (AFO) reaction. The WST-1 cell proliferation assay indicates (i) that all synthesized 3-O-alkyl-3',4'-dimethoxyflavonols and 3-O-aminoalkyl-3',4'-dimethoxyflavonols are more potent than the parent 3',4'-dimethoxyflavonol and the natural flavonol quercetin in suppressing prostate cancer cell proliferation; and (ii) that incorporation of a dibutylamino group to the 3-OH group through a three- to five-carbon linker leads to the optimal derivatives with up to 292-fold enhanced potency as compared with the parent flavonol. Flow cytometry analysis showed that the most potent derivative 22 can activate PC-3 cell cycle arrest at the G2/M phase and induce PC-3 cell apoptosis. No inhibitory ability of 22 up to 50µM concentration was observed against PWR-1E normal human epithelial prostate cells, suggesting its in vitro safety profile. The results indicate that chemical modulation at 3-OH is a vital strategy to optimize flavonols as anti-prostate cancer agents.

Profiling of Flavonol Derivatives for the Development of Antitrypanosomatidic Drugs.

Flavonoids represent a potential source of new antitrypanosomatidic leads. Starting from a library of natural products, we combined target-based screening on pteridine reductase 1 with phenotypic screening on Trypanosoma brucei for hit identification. Flavonols were identified as hits, and a library of 16 derivatives was synthesized. Twelve compounds showed EC50 values against T. brucei below 10 µM. Four X-ray crystal structures and docking studies explained the observed structure-activity relationships. Compound 2 (3,6-dihydroxy-2-(3-hydroxyphenyl)-4H-chromen-4-one) was selected for pharmacokinetic studies. Encapsulation of compound 2 in PLGA nanoparticles or cyclodextrins resulted in lower in vitro toxicity when compared to the free compound. Combination studies with methotrexate revealed that compound 13 (3-hydroxy-6-methoxy-2-(4-methoxyphenyl)-4H-chromen-4-one) has the highest synergistic effect at concentration of 1.3 µM, 11.7-fold dose reduction index and no toxicity toward host cells. Our results provide the basis for further chemical modifications aimed at identifying novel antitrypanosomatidic agents showing higher potency toward PTR1 and increased metabolic stability.

Synthesis and effects on cell viability of flavonols and 3-methyl ether derivatives on human leukemia cells.

Flavonoids are polyphenolic compounds which display an array of biological activities and are considered potential antitumor agents. Here we evaluated the antiproliferative activity of selected synthetic flavonoids against human leukemia cell lines. We found that 4'-bromoflavonol (flavonol 3) was the most potent. This compound inhibited proliferation in a concentration-dependent manner, induced apoptosis and blocked cell cycle progression at the S phase. Cell death was found to be associated with the cleavage and activation of multiple caspases, the activation of the mitogen-activated protein kinase pathway and the up-regulation of two death receptors (death receptor 4 and death receptor 5) for tumor necrosis factor-related apoptosis-inducing ligand. Moreover, combined treatments using 4'-bromoflavonol and TRAIL led to an increased cytotoxicity compared to single treatments. These results provide a basis for further exploring the potential applications of this combination for the treatment of cancer.

Structure-activity relationship studies of flavonol analogues on pollen germination.

Flavonoids are polyphenolic compounds required in the fertilization process in many, if not all, plants. However, the exact biological mechanism(s) and the interacting proteins are unknown. To determine the characteristics important in activating or inhibiting the pollination sequence, a structure-activity relationship analysis of natural and synthetic flavonols was conducted. Flavonol analogues were synthesized through a modified "one-pot" procedure that utilized a Baker-Venkataraman type rearrangement and a Suzuki-Miyaura cross-coupling of a halo-flavonol with an organotrifluoroborate. Of the flavonols tested, kaempferol was the only compound to act as a full agonist. The other smaller, less sterically hindered flavonols (galangin, kaempferide, and 4'-methyl flavonol) acted as partial agonists. Larger more hydrophobic flavonol analogues (3'- and 4'-benzoyl, 3'- and 4'-phenyl, and 3'- and 4'-iodo flavonols) had minimal or no agonist activity. Competition assays between kaempferol and these minimally activating flavonols showed that these analogues inhibited the action of kaempferol in a manner consistent with noncompetitive antagonism. The results suggest that steric hindrance is the most important factor in determining a good agonist. Hydrogen bonding also had a positive effect as long as the substituent did not cause any steric hindrance.

Superior anticancer activity of halogenated chalcones and flavonols over the natural flavonol quercetin.

A series of chalcone and flavonol derivatives were synthesized in good yield by an eco-friendly approach. A pharmacological evaluation was performed with the human colorectal carcinoma cell line HCT116 and revealed that the anticancer activity of flavonols was higher when compared with that of the respective chalcone precursors. The antiproliferative activity of halogenated derivatives increases as the substituent in the 3- or 4-positon of the B-ring goes from F to Cl and to Br. In addition, halogens in position 3 enhance anticancer activity in chalcones whereas for flavonol derivatives the best performance was registered for the 4-substituted derivatives. Flow cytometry analysis showed that compounds 3p and 4o induced cell cycle arrest and apoptosis as demonstrated by increased S, G2/M and sub-G1 phases. These data were corroborated by western blot and fluorescence microscopy analysis. In summary, halogenated chalcones and flavonols were successfully prepared and presented high anticancer activity as shown by their cell growth and cell cycle inhibitory potential against HCT116 cells, superior to that of quercetin, used as a positive control.

Synthesis and biological evaluation of novel flavonols as potential anti-prostate cancer agents.

A library of flavonol analogues was synthesised and evaluated as potential anticancer agents against a human prostate cancer cell line, 22rν1. Compounds 3, 8 and 11 (IC(50) 2.6, 3.3 and 4.0 µM respectively) showed potent cancer cell growth inhibition, comparable to the lead compound 3',4',5'-trimethoxyflavonol (1) (IC(50) 3.1 µM) and superior to the naturally occurring flavonols quercetin (16) and fisetin (22) (both >15 µM). Results indicate that the 3',4',5'- arrangement of either hydroxy (OH) or methoxy (OMe) residues is important for growth arrest of these cells and that the OMe analogues may be superior to their OH counterparts. Compounds 1, 3, 8 and 11 warrant further investigation as potential agents for the management of prostate cancer.

Preparation of two sets of 5,6,7-trioxygenated dihydroflavonol derivatives as free radical scavengers and neuronal cell protectors to oxidative damage.

An unusual class of 5,6,7-trioxygenated dihydroflavonols (3a-e and 4a-j) were designed and prepared. Their antioxidative properties were assessed by examining their capacities in several in vitro models, including superoxide anion and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging, rat liver homogenate lipid peroxidation inhibition, PC12 cells protection from oxidative damage, and xanthine oxidase inhibition. These dihydroflavonols displayed positive quenching abilities towards O(2)(-) and DPPH free radicals, in which the majority exhibited superior antioxidant properties to Vitamin C. cis-Configurated compound (+/-)-3e demonstrated remarkable inhibition to LPO with an IC(50) value of 1.9+/-0.3 microM, which was apparently stronger than that of quercetin (IC(50)=6.0+/-0.4 microM). trans-Configurated dihydroflavonol (+/-)-4h exhibited significant protective effect on PC12 cells against oxidative damage with an EC(50) value of 41.5+/-5.3 microM, more effective compared to that of quercetin (EC(50)=81.8+/-8.7 microM). The 6-OH-5,7-dimethoxy analogue (+/-)-3d showed significant inhibition of xanthine oxidase with an IC(50) value of 16.0+/-0.8 microM, which is superior to that of allopurinol (IC(50)=23.5+/-2.0 microM). In addition to the hypothesized action mechanism of the bio-active compounds, 3D modeling was used to analyze the relationship between the minimized-energy structures and antioxidant activities.

Synthesis of Abyssinone II and related compounds as potential chemopreventive agents.

A facile and efficient approach to the synthesis of prenylated flavonoids as potential chemopreventive agents has been described. This features the synthesis of prenyl halide, prenylation of p-hydroxybenzaldehyde, formation of prenylated polyhydroxychalcone and cyclization of prenylated polyhydroxychalcone to flavanones (15) and (16), and flavonol (17) starting from isoprene (1). The structures of all three compounds have been characterized by NMR, IR and mass spectroscopy.