Comparative Analysis of Polyphenolic Compounds in Different Amaranthus Species: Influence of Genotypes and Harvesting Year.

Amaranth is a nutritionally valuable crop, as it contains phenolic acids and flavonoids, yielding diverse plant secondary metabolites (PSMs) like phytosterol, tocopherols, and carotenoids. This study explored the variations in the contents of seventeen polyphenolic compounds within the leaves of one hundred twenty Amaranthus accessions representing nine Amaranthus species. The investigation entailed the analysis of phenolic content across nine Amaranthus species, specifically A. hypochondriacus, A. cruentus, A. caudatus, A. tricolor, A. dubius, A. blitum, A. crispus, A. hybridus, and A. viridis, utilizing ultra performance liquid chromatography with photodiode array detection (UPLC-PDA). The results revealed significant differences in polyphenolic compounds among accessions in which rutin content was predominant in all Amaranthus species in both 2018 and 2019. Among the nine Amaranthus species, the rutin content ranged from 95.72 ± 199.17 µg g-1 (A. dubius) to 1485.09 ± 679.51 µg g-1 (A. viridis) in 2018 and from 821.59 ± 709.95 µg g-1 (A. tricolor) to 3166.52 ± 1317.38 µg g-1 (A. hypochondriacus) in 2019. Correlation analysis revealed, significant positive correlations between rutin and kaempferol-3-O-ß-rutinoside (r = 0.93), benzoic acid and ferulic acid (r = 0.76), and benzoic acid and kaempferol-3-O-ß-rutinoside (r = 0.76), whereas gallic acid showed consistently negative correlations with each of the 16 phenolic compounds. Wide variations were identified among accessions and between plants grown in the two years. The nine species and one hundred twenty Amaranthus accessions were clustered into six groups based on their seventeen phenolic compounds in each year. These findings contribute to expanding our understanding of the phytochemical traits of accessions within nine Amaranthus species, which serve as valuable resources for Amaranthus component breeding and functional material development.

Apoptosis of Kinetin Riboside in Colorectal Cancer Cells Occurs by Promoting ß-Catenin Degradation.

Kinetin riboside is a naturally produced cytokinin that displays strong antiproliferative activity in various human cancer cells. However, the mechanism of chemoprevention in colorectal cancer cells has not been elucidated. We used a cell-based reporter system to identify kinetin riboside as an antagonist of the Wnt/ß-catenin pathway, which is aberrantly upregulated in colorectal cancer. Kinetin riboside suppressed ß-catenin response transcription (CRT) by accelerating the degradation of intracellular ß-catenin via a proteasomal degradation pathway. Pharmacological inhibition of glycogen synthase kinase-3ß did not affect CRT downregulation. Kinetin riboside decreased the intracellular ß-catenin levels in colorectal cancer cells with mutations in adenomatous polyposis coli (APC) and ß-catenin. Consistently, kinetin riboside repressed expression of c-Myc and cyclin D1, ß-catenin/T-cell factor (TCF)-dependent genes, and inhibited the proliferation of colorectal cancer cells. In addition, kinetin riboside stimulated apoptosis, as measured by an increase in annexin V-FITC-stained cells. These findings suggest that kinetin riboside exerts its anti-cancer activity by promoting ß-catenin degradation and has significant potential as a chemopreventive agent for colorectal cancer cells.

Metabolome-Wide Reprogramming Modulated by Wnt/ß-Catenin Signaling Pathway.

A family of signal transduction pathways known as wingless type (Wnt) signaling pathways is essential to developmental processes like cell division and proliferation. Mutation in Wnt signaling results in a variety of diseases, including cancers of the breast, colon, and skin, metabolic disease, and neurodegenerative disease; thus, the Wnt signaling pathways have been attractive targets for disease treatment. However, the complicatedness and large involveness of the pathway often hampers pinpointing the specific targets of the metabolic process. In our current study, we investigated the differential metabolic regulation by the overexpression of the Wnt signaling pathway in a timely-resolved manner by applying high-throughput and un-targeted metabolite profiling. We have detected and annotated 321 metabolite peaks from a total of 36 human embryonic kidney (HEK) 293 cells using GC-TOF MS and LC-Orbitrap MS. The un-targeted metabolomic analysis identified the radical reprogramming of a range of central carbon/nitrogen metabolism pathways, including glycolysis, TCA cycle, and glutaminolysis, and fatty acid pathways. The investigation, combined with targeted mRNA profiles, elucidated an explicit understanding of activated fatty acid metabolism (ß-oxidation and biosynthesis). The findings proposed detailed mechanistic biochemical dynamics in response to Wnt-driven metabolic changes, which may help design precise therapeutic targets for Wnt-related diseases.

Lucidin 3-methyl ether from Rubia philippinensis suppresses the proliferation of multiple myeloma cells through the promotion of ß-catenin degradation.

BACKGROUND: Constitutive accumulation of ß-catenin has been frequently observed in multiple myeloma. Extracts from genus Rubia plants exhibit cytotoxic activity against several types of cancer cells; however, little is known about their chemopreventive mechanisms and bioactive metabolites. PURPOSE: Purpose: The study aimed to identify the underlying antiproliferative mechanisms of Rubia philippinensis extract in multiple myeloma cells and the major active metabolites responsible for cytotoxic activity of R. philippinensis. METHODS: The effects of R. philippinensis extracts and lucidin 3-methyl ether on the Wnt/ß-catenin pathway were determined by cell-based reporter assay, Western blot analysis, and RT-PCR. The antiproliferative activity was evaluated by cell viability assay and apoptosis analysis in RPMI8226 and MM.1S multiple myeloma cells. RESULTS: R. philippinensis extracts inhibited Wnt/ß-catenin signaling and lucidin 3-methyl ether, an anthraquinone derivative, was identified as the major active metabolite responsible for the inhibition of Wnt/ß-catenin signaling. Lucidin 3-methyl ether induced ß-catenin phosphorylation at Ser33/Ser37/Thr41 residues and promoted proteasomal degradation of ß-catenin via a GSK-3ß-independent mechanism, thereby downregulating Wnt3a-induced ß-catenin response transcription (CRT). Moreover, lucidin 3-methyl ether repressed the expression of ß-catenin/T-cell factor (TCF)-dependent genes, such as cyclin D1, c-myc, and axin-2, thus inhibiting MM cell proliferation. Apoptosis was also elicited by lucidin 3-methyl ether, as indicated by the increase in the population of annexin V-FITC positive cells and caspase-3/7 activity in MM cells. CONCLUSION: These findings indicate that R. philippinensis and its active metabolite lucidin 3-methyl ether prevent cell proliferation through the suppression of the Wnt/ß-catenin pathway and exhibit potential as chemopreventive agents for the treatment of MM.

Root Bark of Morus alba L. and Its Bioactive Ingredient, Ursolic Acid, Suppress the Proliferation of Multiple Myeloma Cells by Inhibiting Wnt/ß-Catenin Pathway.

The root bark of Morus alba L. has cytotoxic activity against several types of cancer cells. However, little is known about its chemopreventive mechanisms and bioactive metabolites. In this study, we showed that M. alba L. root bark extracts (MRBE) suppressed ß-catenin response transcription (CRT), which is aberrantly activated in various cancers, by promoting the degradation of ß-catenin. In addition, MRBE repressed the expression of the ß-catenin/T-cell factor (TCF)-dependent genes, cmyc and cyclin D1, thus inhibiting the proliferation of RPMI-8226 multiple myeloma (MM) cells. MRBE induced apoptosis in MM cells, as evidenced by the increase in the population of annexin VFITC- positive cells and caspase-3/7 activity. We identified ursolic acid in MRBE through LC/mass spectrum (MS) and observed that it also decreased intracellular ß-catenin, c-myc, and cyclin D1 levels. Furthermore, it suppressed the proliferation of RPMI-8226 cells by stimulating cell cycle arrest and apoptosis. These findings suggest that MRBE and its active ingredient, ursolic acid, exert antiproliferative activity by promoting the degradation of ß-catenin and may have significant chemopreventive potential against MM.

Synthesis and Application of Silica-Coated Quantum Dots in Biomedicine.

Quantum dots (QDs) are semiconductor nanoparticles with outstanding optoelectronic properties. More specifically, QDs are highly bright and exhibit wide absorption spectra, narrow light bands, and excellent photovoltaic stability, which make them useful in bioscience and medicine, particularly for sensing, optical imaging, cell separation, and diagnosis. In general, QDs are stabilized using a hydrophobic ligand during synthesis, and thus their hydrophobic surfaces must undergo hydrophilic modification if the QDs are to be used in bioapplications. Silica-coating is one of the most effective methods for overcoming the disadvantages of QDs, owing to silica's physicochemical stability, nontoxicity, and excellent bioavailability. This review highlights recent progress in the design, preparation, and application of silica-coated QDs and presents an overview of the major challenges and prospects of their application.

Absolute oral and subcutaneous bioavailability of ortho-topolin riboside in mice.

Among essential phytohormones playing a pivotal role in regulating growth and development, ortho-topolin riboside (oTR) exerts the most substantial anti-tumor potency in various cancer cell lines. This study was designed to establish a quantitative determination method for oTR in mouse plasma using high-performance liquid chromatography coupled with tandem mass spectrometry (HPLC-MS/MS), to validate the analytical method including stability, and to characterise its pharmacokinetic behaviour in mice. After simple protein precipitation with acetonitrile including kinetin riboside (internal standard), oTR was eluted on a reversed-phase column using a mobile phase of water and acetonitrile (3:7 v/v, including 0.1% formic acid). The protonated precursor ion [M+H]+ and major fragment ion were confirmed at m/z 374.06 and 241.99 for oTR, and 348.23 and 216.06 for the IS, respectively. oTR was stable under bench and storage conditions. The analytical method met the criteria of FDA-validated bioanalytical methods and was successfully applied to a pharmacokinetic study for the first time following oral, subcutaneous, and intravenous administrations. While oTR was merely absorbed by an oral route, 90% of the absolute subcutaneous bioavailability was observed.

Ilimaquinone inhibits neovascular age-related macular degeneration through modulation of Wnt/ß-catenin and p53 pathways.

Neovascular age-related macular degeneration (nAMD) is a common cause of irreversible vision loss in the elderly. Anti-vascular endothelial growth factor has been effective in treating pathological ocular neovascularization, but it has limitations including the need for repeated intraocular injections for the maintenance of therapeutic effects in most patients and poor or non-response to this agent in some patients. in vitro cellular studies were conducted using retinal pigment epithelial cell lines (ARPE-19 and hTERT-RPE1), human umbilical vein endothelial cells (HUVECs), and human umbilical vein smooth muscle cells (HUVSMCs). in vivo efficacy of ilimaquinone (IQ) was tested in laser-induced choroidal neovascularization mouse and rabbit models. Tissue distribution study was performed in male C57BL6/J mice. IQ, 4,9-friedodrimane-type sesquiterpenoid isolated from the marine sponge, repressed the expression of angiogenic/inflammatory factors and restored the expression of E-cadherin in retinal pigment epithelial cells by inhibiting the Wnt/ß-catenin pathway. In addition, it selectively inhibited proliferation and tube formation of HUVECs by activating the p53 pathway. Topical and intraperitoneal administration of IQ significantly reduced choroidal neovascularization in rabbits and mice with laser-induced choroidal neovascularization. Notably, IQ by the oral route of exposure was highly permeable to the eyes and suppressed abnormal vascular leakage by downregulation of ß-catenin and stabilization of p53 in vivo. Our findings demonstrate that IQ functions through regulation of p53 and Wnt/ß-catenin pathways with conceivable advantages over existing cytokine-targeted anti-angiogenic therapies.

Cytotoxic Activity of Aplykurodin A Isolated From Aplysia kurodai against AXIN1-Mutated Hepatocellular Carcinoma Cells by Promoting Oncogenic ß-Catenin Degradation.

Dysregulation of the Wnt/ß-catenin signaling pathway is involved in the development of human hepatocellular carcinoma and has thus emerged as a therapeutic target for this malignant tumor. In this study, we employed sensitive cell-based assays to identify aplykurodin A isolated from Aplysia kurodai as an antagonist of Wnt/ß-catenin signaling. Aplykurodin A inhibited ß-catenin responsive transcription, which was stimulated by a Wnt3a-conditioned medium or a glycogen synthase kinase 3ß inhibitor by accelerating intracellular ß-catenin degradation. Aplykurodin A downregulated the level of oncogenic ß-catenin and decreased the expression of ß-catenin-dependent gene, leading to inhibition of human hepatoma Hep3B and SNU475 cell proliferation. Moreover, apoptosis and autophagy were elicited by aplykurodin A, as indicated by an increase the number of Annexin V-FITC-stained cells and the formation of microtubule-associated protein 1 light chain 3 puncta, respectively, in Hep3B and SNU475 cells. Our findings suggest that aplykurodin A provides a novel therapeutic strategy for human hepatocellular carcinoma via stimulation of oncogenic ß-catenin degradation.

Cytotoxic activity of broussochalcone a against colon and liver cancer cells by promoting destruction complex-independent ß-catenin degradation.

Aberrant activation of ß-catenin-response transcription (CRT) is a well-recognized characteristic of colorectal and liver cancers and thus a potential therapeutic target for these malignancies. Broussonetia papyrifera (paper mulberry) has been used as a herbal medicine to treat various diseases. Using a sensitive cell-based screening system, we identified broussochalcone A (BCA), a prenylated chalcone isolated from Broussonetia papyrifera, as an antagonist of CRT. BCA accelerated the turnover of intracellular ß-catenin that was accompanied by its N-terminal phosphorylation at Ser33/37/Thr41 residues, marking it for ubiquitin-dependent proteasomal degradation. Pharmacological inhibition of glycogen synthase kinase-3ß could not abrogate BCA-mediated degradation of ß-catenin. BCA decreased the intracellular ß-catenin levels in colon and liver cancer cells with mutations in ß-catenin, adenomatous polyposis coli, and Axin. BCA repressed the expressions of cyclin D1, c-Myc, and Axin2, which are ß-catenin/T-cell factor-dependent genes, and thus decreased the viability of colon and liver cancer cell. Moreover, apoptosis was elicited by BCA, as indicated by the increase in the population of Annexin V-FITC positive cells and caspase-3/7 activities in colon and liver cancer cells. These findings indicate that BCA exerts its cytotoxic effects by promoting phosphorylation/ubiquitin-dependent degradation of ß-catenin and may potentially serve as a chemopreventive agent for colonrectal and liver cancers.

CGK062, a small chemical molecule, inhibits cancer upregulated gene 2­induced oncogenesis through NEK2 and ß­catenin.

The mechanisms through which cancer­upregulated gene 2 (CUG2), a novel oncogene, affects Wnt/ß­catenin signaling, essential for tumorigenesis, are unclear. In this study, we aimed to elucidate some of these mechanisms in A549 lung cancer cells. Under the overexpression of CUG2, the protein levels and activity of ß­catenin were evaluated by western blot analysis and luciferase assay. To examine a biological consequence of ß­catenin under CUG2 overexpression, cell migration, invasion and sphere formation assay were performed. The upregulation of ß­catenin induced by CUG2 overexpression was also accessed by xenotransplantation in mice. We first found that CUG2 overexpression increased ß­catenin expression and activity. The suppression of ß­catenin decreased cancer stem cell (CSC)­like phenotypes, indicating that ß­catenin is involved in CUG2­mediated CSC­like phenotypes. Notably, CUG2 overexpression increased the phosphorylation of ß­catenin at Ser33/Ser37, which is known to recruit E3 ligase for ß­catenin degradation. Moreover, CUG2 interacted with and enhanced the expression and kinase activity of never in mitosis gene A­related kinase 2 (NEK2). Recombinant NEK2 phosphorylated ß­catenin at Ser33/Ser37, while NEK2 knockdown decreased the phosphorylation of ß­catenin, suggesting that NEK2 is involved in the phosphorylation of ß­catenin at Ser33/Ser37. Treatment with CGK062, a small chemical molecule, which promotes the phosphorylation of ß­catenin at Ser33/Ser37 through protein kinase C (PKC)α to induce its degradation, reduced ß­catenin levels and inhibited the CUG2­induced features of malignant tumors, including increased cell migration, invasion and sphere formation. Furthermore, CGK062 treatment suppressed CUG2­mediated tumor formation in nude mice. Taken together, the findings of this study suggest that CUG2 enhances the phosphorylation of ß­catenin at Ser33/Ser37 by activating NEK2, thus stabilizing ß­catenin. CGK062 may thus have potential for use as a therapeutic drug against CUG2­overexpressing lung cancer cells.

Longan fruit increase bone mineral density in zebrafish and ovariectomized rat by suppressing RANKL-induced osteoclast differentiation.

BACKGROUND: The receptor activator of nuclear factor-kappa B ligand (RANKL)-induced nuclear factor-kappa B (NF-κB) signaling pathway plays essential roles in osteoclast differentiation and may serve as an attractive target for the development of therapeutics for osteoporosis. PURPOSE: This study aimed to identify plant extracts that attenuated RANKL-induced NF-κB signaling pathway and examine their anti-osteoporotic effects in animal model systems. METHODS: Osteoclast differentiation was determined by western blot analysis, RT-PCR, and tartrate-resistant acid phosphatase (TRAP) assay. The effect of Longan (Dimocarpus longan Lour.) fruit extract (LFE) on bone mineral density was evaluated by calcein staining in zebrafish and micro-CT analysis in ovariectomized (OVX) rat. RESULTS: LFE nullified RANKL-induced down-regulation of inhibitor of NF-κB, which keeps NF-κB sequestered in the cytosol, thereby inhibiting translocation of NF-κB to the nucleus, in RAW264.7 cells. In addition, LFE decreased the nuclear levels of c-Fos and nuclear factor of activated T-cells c1, which play crucial roles in RANKL-induced osteoclast differentiation, in RAW264.7 cells. LFE repressed RANKL-activated cathepsin K and TRAP expression in RAW264.7 cells, resulting in a reduction of the number of TRAP-positive multinucleated cells, without cytotoxicity. Furthermore, LFE increased bone mineralization in zebrafish and prevented bone loss in OVX rat. CONCLUSION: Collectively, our findings suggest that LFE exerts its anti-osteoporotic activity through inhibition of osteoclast differentiation and may have potential as a herbal therapeutic or preventive agent for the treatment of osteoporosis.

Protopine isolated from Nandina domestica induces apoptosis and autophagy in colon cancer cells by stabilizing p53.

The tumor suppressor p53 plays essential roles in cellular protection mechanisms against a variety of stress stimuli and its activation induces apoptosis or autophagy in certain cancer cells. Here, we identified protopine, an isoquinoline alkaloid isolated from Nandina domestica, as an activator of the p53 pathway from cell-based natural compound screening based on p53-responsive transcription. Protopine increased the p53-mediated transcriptional activity and promoted p53 phosphorylation at the Ser15 residue, resulting in stabilization of p53 protein. Moreover, protopine up-regulated the expression of p21WAF1/CIP1 and BAX, downstream genes of p53, and inhibited the proliferation of HCT116 colon cancer cells. Apoptosis was elicited by protopine as indicated by caspase-3/7 activation, poly ADP ribose polymerase cleavage, and increased population of Annexin V-FITC-positive cells. Furthermore, protopine induced the formation of microtubule-associated protein 1 light chain 3 (LC3) puncta and LC3-II turnover, typical biochemical markers of autophagy, in HCT116 cells. Our findings suggest that protopine exerts its antiproliferative activity by stimulating the p53 pathway and may have potential as a chemopreventive agent for human colon cancer.

Stereo-Selective Pharmacokinetics of Ilimaquinone Epimers Extracted from a Marine Sponge in Rats.

An ilimquinone (IQ) mixture isolated from Hippiospongia metachromia, consisting of IQ and epi-ilimaquinone (epi-IQ), exerts anti-HIV, anti-microbial, anti-inflammatory, and anti-cancer effects. An HPLC-MS/MS method was developed for simultaneous determination of the two epimers in rat plasma, separating them using a biphenyl column. Ascorbic acid is added during the sample preparation to ensure the stability of both isomers. The plasma concentrations of the isomers were monitored following intravenous and oral administration of the IQ mixture in rats as well as the individual epimers that were separately orally administered. Compare to IQ, epi-IQ was much more stable in rat plasma, likely due to its configurations of decalin. Both substances decayed in more than bi-exponential pattern, with an elimination rate constant of 1.2 h-1 for IQ and 1.7 h-1 for epi-IQ. The epi-IQ was distributed more widely than IQ by about two-fold. Consequently, the clearance of epi-IQ was greater than that of IQ by about three-fold. The oral absolute bioavailability for IQ was 38%, and, that for epi-IQ, was 13%. Although the systemic exposure of IQ was greater than that of epi-IQ by ~8.7-fold, the clearance of each isomer was similar when administered either orally or intravenously, when normalized for bioavailability. The stereo-specific behavior of the isomers appears to originate from differences in both their tissue distribution and gastrointestinal permeability.

HPLC-MS/MS analysis of ilimaquinone and its application in a pharmacokinetic study in rats.

Ilimaquinone, a metabolite isolated from the marine sponge Hippiospongia metachromia, has antimicrobial, cytotoxic, anti-HIV, anti-inflammatory, and anti-cancer activities. A new quantitative analytical method for determination of ilimaquinone in rat plasma using HPLC-MS/MS was developed and validated. Ascorbic acid was added to ensure the stability of ilimaquinone in plasma. After protein precipitation using acetonitrile plus diclofenac as an internal standard, the analytes were chromatographed on a biphenyl column with a mobile phase of methanol and water (8:2, v/v, including 0.1% formic acid). This method was successfully applied in a pharmacokinetic study of ilimaquinone after oral administration in rats.

CCAAT/enhancer-binding protein-ß functions as a negative regulator of Wnt/ß-catenin signaling through activation of AXIN1 gene expression.

Axin1, a concentration-limiting component of the ß-catenin destruction complex, negatively regulates the Wnt/ß-catenin pathway. Axin1 concentration is reported to be regulated by proteasomal degradation; however, its transcriptional regulation has not yet been reported. Here, we demonstrated that CCAAT/enhancer-binding protein-ß (C/EBP-ß) activates axis inhibition protein 1 (AXIN1) gene expression, thereby attenuating Wnt/ß-catenin signaling. C/EBP-ß interacted with cis-regulatory element for C/EBP-ß in the 5'-upstream sequences of the AXIN1 gene and increased AXIN1 promoter activity. Functional analysis using Drosophila and zebrafish models established that C/EBP-ß negatively regulates the Wnt/ß-catenin pathway. Small-molecule-based up-regulation of C/EBP-ß induces AXIN1 gene expression and down-regulates the intracellular ß-catenin level, thereby inhibiting hepatoma cell growth. Thus, our findings provide a unique mechanistic insight into the regulation of Axin homeostasis and present a novel strategy for the development of anticancer therapeutics targeting Wnt/ß-catenin signaling.

Sesterterpenoid and Steroid Metabolites from a Deep-Water Alaska Sponge Inhibit Wnt/ß-Catenin Signaling in Colon Cancer Cells.

The Wnt/ß-catenin signaling pathway is known to play critical roles in a wide range of cellular processes: cell proliferation, differentiation, migration and embryonic development. Importantly, dysregulation of this pathway is tightly associated with pathogenesis in most human cancers. Therefore, the Wnt/ß-catenin pathway has emerged as a promising target in anticancer drug screening programs. In the present study, we have isolated three previously unreported metabolites from an undescribed sponge, a species of Monanchora (Order Poecilosclerida, Family Crambidae), closely related to the northeastern Pacific species Monanchora pulchra, collected from deep waters off the Aleutian Islands of Alaska. Through an assortment of NMR, MS, ECD, computational chemical shifts calculation, and DP4, chemical structures of these metabolites have been characterized as spirocyclic ring-containing sesterterpenoid (1) and cholestane-type steroidal analogues (2 and 3). These compounds exhibited the inhibition of ß-catenin response transcription (CRT) through the promotion of ß-catenin degradation, which was in part implicated in the antiproliferative activity against two CRT-positive colon cancer cell lines.

Anti-proliferative activity of CGK012 against multiple myeloma cells via Wnt/ß-catenin signaling attenuation.

The aberrant activation of Wnt/ß-catenin signaling is involved in the development of multiple myeloma; thus, this signaling pathway is a potential target for the development of therapeutics for this malignancy. Here, we performed cell-based chemical screening and found that CGK012, a pyranocoumarin compound, suppressed the Wnt3a-CM-mediated activation of ß-catenin response transcription. CGK012 induced ß-catenin phosphorylation at Ser33/Ser37/Thr41, leading to proteasomal degradation and reducing the level of intracellular ß-catenin. Furthermore, CGK012 consistently decreased the amount of ß-catenin and repressed the expression of cyclin D1, c-myc, and axin-2 (downstream target genes of ß-catenin) in RPMI-8226 multiple myeloma cells. In addition, CGK012 inhibited the proliferation of RPMI-8226 cells and promoted apoptosis, as indicated by the increase in the population of Annexin V-FITC-stained cells and caspase-3/7 activity. These findings suggest that CGK012 could exert antiproliferative activity against multiple myeloma cells by attenuating the Wnt/ß-catenin pathway; thus, it may have potential as a therapeutic agent for multiple myeloma treatment.

LKY-047: First Selective Inhibitor of Cytochrome P450 2J2.

Highly selective cytochrome P450 CYP2J2 (CYP2J2) inhibitors suitable for reaction phenotyping are currently not available. (7S)-(+)-(4-Nitro-phenyl)-acrylic acid, 8,8-dimethyl-2-oxo-6,7-dihydro-2H,8H-pyrano[3,2-g]chromen-7-yl-ester (LKY-047), a decursin derivative, was synthesized, and its inhibitor potencies toward CYP2J2 as well as other cytochrome P450 (P450) enzymes in human liver microsomes (HLM) were evaluated. LKY-047 was demonstrated to be a strong competitive inhibitor of CYP2J2-mediated astemizole O-demethylase and terfenadine hydroxylase activity, with Ki values of 0.96 and 2.61 µM, respectively. It also acted as an uncompetitive inhibitor of CYP2J2-mediated ebastine hydroxylation with a Ki value of 3.61 µM. Preincubation of LKY-047 with HLMs and NADPH did not alter inhibition potency, indicating that it is not a mechanism-based inhibitor. LKY-047 was found to be a selective CYP2J2 inhibitor with no inhibitory effect on other human P450s, such as CYPs 1A2, 2A6, 2B6, 2C8, 2C9, 2C19, 2D6, 2E1, and 3A (IC50 > 50 µM). These in vitro data support the use of LKY-047 as a selective CYP2J2 inhibitor with potential application in the identification of P450 isoforms responsible for drug metabolism in reaction phenotyping assays.