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Tylophorine reduces protein biosynthesis and rapidly decreases cyclin D1, inhibiting vascular smooth muscle cell proliferation in vitro and in organ culture
Abstract
Background:
Tylophorine (TYL) is an alkaloid with antiproliferative action in cancer cells. Vascular smooth muscle cell (VSMC) proliferation and neointima formation contribute to restenosis after percutaneous coronary interventions.
Hypothesis/purpose:
Our goal was to examine the potential of TYL to inhibit VSMC proliferation and migration, and to dissect underlying signaling pathways.
Study design and methods:
TYL was administered to platelet-derived growth factor (PDGF-BB)-stimulated, serum-stimulated, quiescent and unsynchronized VSMC of rat and human origin. BrdU incorporation and resazurin conversion were used to assess cell proliferation. Cell cycle progression was analyzed by flow cytometry of propidium iodide-stained nuclei. Expression profiles of proteins and mRNAs were determined using western blot analysis and RT-qPCR. The Click-iT OPP Alexa Fluor 488 assay was used to monitor protein biosynthesis.
Results:
TYL inhibited PDGF-BB-induced proliferation of rat aortic VSMCs by arresting cells in G1 phase of the cell cycle with an IC50 of 0.13 µmol/l. The lack of retinoblastoma protein phosphorylation and cyclin D1 downregulation corroborated a G1 arrest. Inhibition of proliferation and cyclin D1 downregulation were species- and stimulus-independent. TYL also decreased levels of p21 and p27 proteins, although at later time points than observed for cyclin D1. Co-treatment of VSMC with TYL and MG132 or cycloheximide (CHX) excluded proteasome activation by TYL as the mechanism of action. Comparable time-dependent downregulation of cyclin D1, p21 and p27 in TYL- or CHX-treated cells, together with decreased protein synthesis observed in the Click-iT assay, suggests that TYL is a protein synthesis inhibitor. Besides proliferation, TYL also suppressed migration of PDGF-activated VSMC. In a human saphenous vein organ culture model for graft disease, TYL potently inhibited intimal hyperplasia.
Conclusion:
This unique activity profile renders TYL an interesting lead for the treatment of vasculo-proliferative disorders, such as restenosis.
... Anti-cancer compounds of the class of phenanthroindolizidine alkaloids (PAs) originate from the plant family Apocynaceae, including members of the genus Tylophora, which are endogenous to (sub-)tropical Africa, Asia, and Australia [50,51]. Naturally occurring PAs and their synthetic analogues show a broad range of action including anti-asthmatic [52], antiparasitic [53], antibacterial [54], antifungal [55], antiviral [56], anti-inflammatory [57], antiangiogenic [58,59], and anti-proliferative properties in vitro and in vivo, e.g., in a hepatocellular carcinoma xenograft model [60]. The anti-tumor mechanism has been described to depend on the blockade of cell signaling, e.g., NFκB in HepG2 [61] or HIF in T47D [62], and also the suppression of DNA replication and protein synthesis [63,64]. ...
Triple-negative breast cancer (TNBC), representing the most aggressive form of breast cancer with currently no targeted therapy available, is characterized by an inflammatory and hypoxic tumor microenvironment. To date, a broad spectrum of anti-tumor activities has been reported for phenanthroindolizidine alkaloids (PAs), however, their mode of action in TNBC remains elusive. Thus, we investigated six naturally occurring PAs extracted from the plant Tylophora ovata: O-methyltylophorinidine (1) and its five derivatives tylophorinidine (2), tylophoridicine E (3), 2-demethoxytylophorine (4), tylophoridicine D (5), and anhydrodehydrotylophorinidine (6). In comparison to natural (1) and for more-in depth studies, we also utilized a sample of synthetic O-methyltylophorinidine (1s). Our results indicate a remarkably effective blockade of nuclear factor kappa B (NFκB) within 2 h for compounds (1) and (1s) (IC50 = 17.1 ± 2.0 nM and 3.3 ± 0.2 nM) that is different from its effect on cell viability within 24 h (IC50 = 13.6 ± 0.4 nM and 4.2 ± 1 nM). Furthermore, NFκB inhibition data for the additional five analogues indicate a structure-activity relationship (SAR). Mechanistically, NFκB is significantly blocked through the stabilization of its inhibitor protein kappa B alpha (IκBα) under normoxic as well as hypoxic conditions. To better mimic the TNBC microenvironment in vitro, we established a 3D co-culture by combining the human TNBC cell line MDA-MB-231 with primary murine cancer-associated fibroblasts (CAF) and type I collagen. Compound (1) demonstrates superiority against the therapeutic gold standard paclitaxel by diminishing spheroid growth by 40% at 100 nM. The anti-proliferative effect of (1s) is distinct from paclitaxel in that it arrests the cell cycle at the G0/G1 state, thereby mediating a time-dependent delay in cell cycle progression. Furthermore, (1s) inhibited invasion of TNBC monoculture spheroids into a matrigel®-based environment at 10 nM. In conclusion, PAs serve as promising agents with presumably multiple target sites to combat inflammatory and hypoxia-driven cancer, such as TNBC, with a different mode of action than the currently applied chemotherapeutic drugs. Keywords: triple-negative breast cancer (TNBC); phenanthroindolizidine alkaloids; O-methyltylophorinidine; nuclear factor kappa B (NFκB); hypoxia-inducible factor (HIF); tumor microenvironment (TME); 3D spheroid; cancer-associated fibroblast (CAF) Citation: Reimche, I.; Yu, H.; Ariantari, N.P.; Liu, Z.; Merkens, K.; Rotfuß, S.; Peter, K.; Jungwirth, U.; Bauer, N.; Kiefer, F.; et al. Phenanthroindolizidine Alkaloids Isolated from Tylophora ovata as Potent Inhibitors of Inflammation, Spheroid Growth and Invasion of Triple-Negative Breast Cancer. Int. J. Mol. Sci. 2022, 23, 10319. https://
... This class of alkaloids have been reported for downregulation of cyclin-A2 protein, the mechanism by which these alkaloids induce cell cycle arrest at G 1 phase of the cell cycle [26]. The alkaloids have also been shown to successfully inhibit the proliferation of vascular smooth muscles through the cyclin-D1 downregulation [27]. Moreover, other involved bio-mechanisms, i.e., c-JUN-mediation, downregulation of NF-κB signaling, and inactivation of the Akt pathways are also reported as the potential biomechanism involved in elicitation of the anticancer activity of tylophorine alkaloids [24,26,28]. ...
The phenanthroindolizidine alkaloid (-)-tylophorine has been reported for its significant anticancer activity working through different biomechanistic pathways. The current study aimed to evaluate the anticancer activity of phenanthroindolizidine alkaloids isolated from Tylophora indica. Six phenanthroindolizidine alkaloid (compounds 1–6) in addition to septicine (7), chlorogenic acid (8), and chlorogenic acid methyl ester (9) were isolated from Tylophora indica using different chromatographic techniques including vacuum liquid chromatography (VLC) and preparative high performance liquid chromatography (HPLC). The isolated compounds structures’ were determined using various spectro-analytical techniques, i.e., 1H-NMR, 13C-NMR, and mass spectrometry. The isolates’ structural stereochemistry and structural geometries were determined with the help of chiroptical techniques together with comparisons with the available standard samples. The in vitro anti-proliferative activity on three different cell lines, MCF-7, HepG2, and HCT-116 were evaluated. Among all the isolated compounds, tylophorinidine (5) was the most active cytotoxic agent with the lowest IC50 values at 6.45, 4.77, and 20.08 μM against MCF-7, HepG2, and HCT-116 cell lines, respectively. The bioactivities were also validated by the in vitro kinase receptors inhibition assay. Compound (5) also exhibited the highest activity with lowest IC50 values (0.6 and 1.3 μM against the Aurora-A and Aurora-B enzymes, respectively), as compared with all the isolated alkaloidal products. The structure activity relationship on the molecular properties, molecular attributes, and bioactivity levels were analyzed, interrelated, and the molecular docking studies on two different receptors, Aurora-A and Aurora-B, were determined, which provided the confirmations of the bioactivity with receptor-ligand geometric disposition, energy requirements, lipophilicity, and detailed the binding pharmacophore involvements responsible for bioactivity elicitations.
... Tylophorine (1) potently inhibited the PDGF-BB-induced proliferation of rat aortic vascular smooth muscle cells (VSMCs) in a concentration-dependent manner with an IC 50 of 0.13 lM, by arresting cells in G1 phase of the cell cycle as well as migration in activated VSMCs. It could decrease de novo protein synthesis, result in downregulation of cyclin D1 and prevent neointimal thickening in human saphenous vein organ culture (Joa et al. 2019). Anti-angiogenic results indicated tylophorine (1) and antofine (2) could be used as promising lead compounds for further studies towards the treatment of vasculo-proliferative disorders, such as restenosis. ...
Naturally occurring phenanthroindolizidine and phenanthroquinolizidine alkaloids (PIAs and PQAs) are two small groups of herbal metabolites sharing a similar pentacyclic structure with a highly oxygenated phenanthrene moiety fused with a saturated or an unsaturated N-heterocycle (indolizidine/quinolizidine moieties). Natural PIAs and PQAs only could be obtained from finite plant families (such as Asclepiadaceae, Lauraceae and Urticaceae families, etc.). Up to date, more than one hundred natural PIAs, while only nine natural PQAs had been described. PIA and PQA analogues have been applied to the development of potent anticancer agents all along because of their excellent cytotoxic activity. However, in the last two decades, other great biological properties, such as anti-inflammatory and antiviral activities were revealed successively by different pharmacological assays. Especially because of their potent antiviral activity against coronavirus (TGEV, SARS CoV and MHV) and tobacco mosaic virus, PIA and PQA analogues have attracted much pharmaceutical attention again, some of them have been used to present interesting targets for total or semi synthesis, and structure-activity relationship (SAR) study for the development of antiviral agents. In this review, natural PIA and PQA analogues obtained in the last two decades with their herbal origins, key spectroscopic characteristics for structural identification, biological activity with possible SARs and application prospects were systematically summarized. We hope this paper can stimulate further investigations on PIA and PQA analogues as an important source for potential drug discovery.
Bioactive compounds isolated from plant sources are the secondary metabolites that have toxicological or pharmacological properties depending upon their structure. Recently, these compounds have received more attention due to their diverse role in providing protection against various types of pathogenic and non-pathogenic diseases. These miraculous bioactive compounds also pursue pharmacological activities against several microbial pathogens and are also known to exhibit antidiabetic, antipyretic, anticancerous, antidiuretic, and antioxidant properties. Some bioactive compounds extracted from plants such as phenolic compounds, anthocyanidins, flavonoids, tannins, and flavones are found in significantly high concentration in the biosphere, and certain others are present in less amount but their pharmacological as well as commercial importance cannot be denied. The activities of bioactive compounds like flavonoids, polyphenols, not only functions to inhibit microbial growth but also they act synergistically with other drugs which make them ideal for alternative cancer remedies. This chapter provides a general insight into secondary metabolites having antimicrobial and anticancerous properties, their plant source, and their mode of actions.KeywordsBioactive compoundsAntimicrobialAnticancerousHuman health
Angioplasty often fails due to the abnormal proliferation of vascular smooth muscle cells (VSMCs). Success rates of angioplasty may increase following the administration of an agent that effectively ameliorates aberrant vascular remodeling. Icariside II(ICS-II) is a natural flavonol glycoside extract from the Chinese herbal medicine Epimedii that possesses several medicinal qualities that are beneficial in humans. Nevertheless, the role of ICS-II in addressing aberrant vascular remodeling have yet to be clarified. The current investigation studies the molecular effects of ICS-II on balloon-inflicted neointimal hyperplasia in rats in vivo and on platelet-derived growth factor (PDGF)-induced vascular proliferation in primary rat aortic smooth muscle cells (VSMCs) in vitro. ICS-II was found to be as effective as rapamycin, the positive control used in this study. ICS-II inhibited neointimal formation in injured rat carotid arteries and notably reduced the expression of Wnt7b. ICS-II significantly counteracted PDGF-induced VSMCs proliferation. Cell cycle analysis showed that ICS-II triggered cell cycle arrest during the G1/S transition. Western blot analysis further indicated that this cell cycle arrest was likely through Wnt7b suppression that led to CCND1 inhibition. In conclusion, our findings demonstrate that ICS-II possesses significant anti-proliferative qualities that counteracts aberrant vascular neointimal hyperplasia. This phenomenon most likely occurs due to suppression of the Wnt7b/CCND1 axis.
Cancer is a frightful disease that still poses a 'nightmare' worldwide, causing millions of casualties annually due to one of the human race's most significant healthcare challenges that requires a pragmatic treatment strategy. However, plants and plant-derived products revolutionize the field as they are quick, cleaner, eco-friendly, low-cost, effective, and less toxic than conventional treatment methods. Plants are repositories for new chemical entities and have a promising cancer research path, supplying 60% of the anticancer agents currently used. Alkaloids are important chemical compounds that serve as a rich reservoir for drug discovery and development. However, some alkaloids derived from natural herbs display anti-proliferation and antimetastatic activity on different forms of cancer, both in vitro and in vivo. Alkaloids have also been widely formulated as anticancer medications, such as camptothecin and vinblastine. Still, more research and clinical trials are required before final recommendations can be made on specific alkaloids. This review focuses on the naturally-derived bioactive alkaloids with prospective anticancer properties based on the information in the literature.
Increased proliferation of vascular smooth muscle cells (VSMCs) contributes to the pathogenesis of atherosclerosis (AS), and the insulin like growth factor 2 (IGF2) is involved in AS through effects on VSMCs growth and migration. The IGF2 mRNA-binding protein 1(IGF2BP1) is a secreted protein that can bind to IGF2 and regulate its localization, however, whether IGF2BP1 could regulate VSMCs proliferation remains to be elucidated. This study aimed to investigate the role of IGF2BP1 in VSMCs proliferation and uncover the potential mechanism. Primary human aortic VSMCs that transfected with or without shRNA-IGF2BP1 were stimulated by platelet-derived growth factor-BB (PDGF-BB), and then cell proliferation, intracellular Ca ²⁺ level, cell apoptosis and the expression of IGF2BP1, calmodulin (CaM) and cell cycle-related proteins were detected. RNA pull down assay was used to determine the interaction between IGF2BP1 and nuclear factor of activated T cells isoform-3 (NFATc3). We found that PDGF-BB promoted cell proliferation and enhanced IGF2BP1 protein expression in a concentration-dependent manner. The 10 μg/L PDGF-BB significantly increased intracellular Ca ²⁺ level, NFATc3, CaM and calcineurin A protein expression, TUNEL-positive cells, and the expression of cell cycle-related proteins cyclin D1/E1/B1. However, knockdown of IGF2BP1 significantly blunted all these effects induced by PDGF-BB. In addition, IGF2BP1 could bind to NFATc3 RNA. Collectively, knockdown of IGF2BP1 could inhibit PDGFBB- induced VSMCs proliferation via targeting NFATc3/Ca ²⁺ /calmodulin pathway and disturbing the effect of NFATc3/ on cell cycle.
Tylophora indica is a potential remedial climber of Asclepiadaceae. Roots and leaves of the plant contain several alkaloids including tylophorine, tylophorinine and tylophrinidine. Major alkaloid, tylophorine, found in T. indica possessed several properties such as immunosuppressive, antitumor, antifeedant, antibacterial, actifungal, antiamoebic, diuretic and hepatoprotective activities. In addition to this, tylophorine provides positive stimulation to adrenal cortex. Biotechnological production of tylophorine was fulfilled by inducing hairy roots mediated by Agrobacterium rhizogenes (A4 strain). It was followed by its growth in liquid suspension culture that could yield maximum biomass and tylophorine production. This type of liquid suspension culture yielded 9.8±0.21 mgl-1 tylophorine within 4 to 6 weeks of incubation. Maceration technique employed for the extraction of tylophorine was the most viable and efficient protocol. Although many reports are available regarding the biotechnological production of tylophorine, its competent and economic production still continues as a problematic issue.
Objective:
Although SET(I2PP2A) and miRNAs are reported to play a pivotal role in lung cancer, the underlying mechanisms have remained obscure. To address this issue, we investigated how miRNAs and SET participate in the progression of lung cancer.
Methods:
miRNAs that target SET were predicted from multiple miRNA databases. Three human NSCLC cell lines and two normal lung cell lines were used to evaluate aberrant miRNA and SET expressions. A dual luciferase reporter assay system was employed to verify the interaction between miRNA and SET. Stable miRNA knockdown and SET overexpression in A549 cells were achieved through lentivirus transfection; the corresponding influences on lung cancer progression were also examined.
Results:
In this study, A549 was the sole cell line to lack SET/TAF-Iα expression, which was inversely correlated with the up-regulation of miR-21-5p. SET was subsequently revealed as the direct target site of miR-21-5p in A549 cells. The stable miR-21-5p knockdown and SET/TAF-Iα overexpression were shown to markedly enhance the expression of SET/TAF-Iα and to inhibit the migration, invasion, proliferation as well as the in vivo tumorigenicity of A549 cells.
Conclusion:
We suggest that SET/TAF-Iα might be a tumor suppressing factor regulated by miR-21-5p in lung adenocarcinoma. This might provide a target for lung adenocarcinoma therapy.
The C-19 quassinoid eurycomalactone (1) has recently been shown to be a potent (IC50 = 0.5 μM) NF-κB inhibitor in a luciferase reporter model. In this study, we show that 1 with similar potency inhibited the expression of the NF-κB-dependent target genes ICAM-1, VCAM-1, and E-selectin in TNFα-activated human endothelial cells (HUVECtert) by flow cytometry experiments. Surprisingly, 1 (2 μM) did not inhibit TNFα-induced IKKα/β or IκBα phosphorylation significantly. Also, the TNFα-induced degradation of IκBα remained unchanged in response to 1 (2 μM). In addition, pretreatment of HUVECtert with 1 (2 μM) had no statistically significant effect on TNFα-mediated nuclear translocation of the NF-κB subunit p65 (RelA). Quantitative RT-PCR revealed that 1 (0.5–5 μM) exhibited diverse effects on the TNFα-induced transcription of ICAM-1, VCAM-1, and SELE genes since the mRNA level either remained unchanged (ICAM-1, E-selectin, and VCAM-1 at 0.5 μM 1), was reduced (VCAM-1 at 5 μM 1), or even increased (E-selectin at 5 μM 1). Finally, the time-dependent depletion of a short-lived protein (cyclin D1) as well as the measurement of de novo protein synthesis in the presence of 1 (2–5 μM) suggested that 1 might act as a protein synthesis inhibitor rather than an inhibitor of early NF-κB signaling.
Objective
To explore the feasibility and safety of using tissue-type plasminogen activator (t-PA) to prevent graft restenosis after coronary artery bypass grafting (CABG).
Methods
In this prospective observational study, 37 patients underwent CABG between June 2009 and May 2013. These patients were grouped according to the anti-coagulation strategy after surgery: t-PA (n = 12) and conventional treatments (n = 25). In the t-PA group, the patients received acetylsalicylic acid (ASA) and clopidogrel plus intravenous infusion of t-PA (0.25 mg/kg/day) starting at 24 h after surgery and that lasted for 3 days. In the conventional group, the patients received only ASA and clopidogrel. 64-row spiral computed tomographic coronary angiography was performed at 1 week, 1, and 3 months after surgery to evaluate the patency of the graft vessel.
Results
The mean stenosis severity of the saphenous vein grafts was lower in the t-PA group compared with the conventional group at 3 months after surgery (p < 0.05), but there was no significant difference at 1 week and 1 month (p > 0.05). The patency rate of the grafts was not significantly different between the two groups at 1 week, 1, and 3 months after surgery (p > 0.05).
Conclusion
Early application of t-PA after CABG was feasible and safe, and might help prevent early restenosis of SV grafts. Additional clinical randomized trials are necessary to address this issue.
Tylophorine analog DCB-3503 is a potential anticancer and immunosuppressive agent that suppresses the translation of cellular regulatory proteins, including cyclin D1, at the elongation step. However, the molecular mechanism underlying this phenomenon remains unknown. This study demonstrates that DCB-3503 preferentially binds to heat shock cognate protein 70 (HSC70), which is a determinant for cyclin D1 translation by binding to the 3′-untranslated region (3′ UTR) of its mRNA. DCB-3503 allosterically regulates the ATPase and chaperone activities of HSC70 by promoting ATP hydrolysis in the presence of specific RNA binding motifs (AUUUA) of cyclin D1 mRNA. The suppression of cyclin D1 translation by DCB-3503 is not solely caused by perturbation of the homeostasis of microRNAs, although the microRNA processing complex is dissociated with DCB-3503 treatment. This study highlights a novel regulatory mechanism of protein translation with AUUUA motifs in the 3′ UTR of mRNA by HSC70, and its activity can be allosterically modulated by DCB-3503. DCB-3503 may be used to treat malignancies, such as hepatocellular carcinoma or breast cancer with elevated expression of cyclin D1.
Medicinal plants have historically proven their value as a source of molecules with therapeutic potential, and nowadays still represent an important pool for the identification of novel drug leads. In the past decades, pharmaceutical industry focused mainly on libraries of synthetic compounds as drug discovery source. They are comparably easy to produce and resupply, and demonstrate good compatibility with established high throughput screening (HTS) platforms. However, at the same time there has been a declining trend in the number of new drugs reaching the market, raising renewed scientific interest in drug discovery from natural sources, despite of its known challenges. In this survey, a brief outline of historical development is provided together with a comprehensive overview of used approaches and recent developments relevant to plant-derived natural product drug discovery. Associated challenges and major strengths of natural product-based drug discovery are critically discussed. A snapshot of the advanced plant-derived natural products that are currently in actively recruiting clinical trials is also presented. Importantly, the transition of a natural compound from a "screening hit" through a "drug lead" to a "marketed drug" is associated with increasingly challenging demands for compound amount, which often cannot be met by re-isolation from the respective plant sources. In this regard, existing alternatives for resupply are also discussed, including different biotechnology approaches and total organic synthesis. While the intrinsic complexity of natural product-based drug discovery necessitates highly integrated interdisciplinary approaches, the reviewed scientific developments, recent technological advances, and research trends clearly indicate that natural products will be among the most important sources of new drugs also in the future.
Copyright © 2015. Published by Elsevier Inc.
Platelet-derived growth factor-BB (PDGF-BB) acts as a full mitogen for cultured aortic smooth muscle cells (SMC), promoting DNA synthesis and cell proliferation. In contrast, angiotensin II (Ang II) induces cellular hypertrophy as a result of increased protein synthesis, but is unable to drive cells into S phase. In an effort to understand the molecular basis for this differential growth response, we have examined the downstream effects of PDGF-BB and Ang II on regulators of the cell cycle machinery in rat aortic SMC. Both PDGF-BB and Ang II were found to stimulate the accumulation of G1 cyclins with similar kinetics. In addition, little difference was observed in the expression level of their catalytic partners, Cdk4 and Cdk2. However, while both factors increased the enzymatic activity of Cdk4, only PDGF-BB stimulated Cdk2 activity in late G1 phase. The lack of activation of Cdk2 in Ang II-treated cells was causally related to the failure of Ang II to stimulate phosphorylation of the enzyme on threonine and to downregulate p27Kip1 expression. By contrast, exposure to PDGF-BB resulted in a progressive and dramatic reduction in the level of p27Kip1 protein. The time course of p27Kip1 decline was correlated with a reduced rate of synthesis and an increased rate of degradation of the protein. Importantly, the repression of p27Kip1 synthesis by PDGF-BB was associated with a marked attenuation of Kip1 gene transcription and a corresponding decrease in Kip1 mRNA accumulation. We also show that the failure of Ang II to promote S phase entry is not related to the autocrine production of transforming growth factor-β1 by aortic SMC. These results identify p27Kip1 as an important regulator of the phenotypic response of vascular SMC to mitogenic and hypertrophic stimuli.
Tylophorine compounds have been the focus of drug development for decades. Tylophorine derivatives exhibit anti-cancer activities but their cellular targets remain unknown. We used a biotinylated tylophorine derivative to probe for the interacting cellular target(s) of tylophorine. Tylophorine directly binds to caprin-1 and consequently enhances the recruitment of G3BP1, c-Myc mRNA, and cyclin D2 mRNA to form a ribonucleoprotein complex. Subsequently, this tylophorine targeted ribonucleoprotein complex is sequestered to the polysomal fractions and the protein expressions of the associated mRNA-transcripts are repressed. Caprin-1 depleted carcinoma cells become more resistant to tylophorine, associated with decreased formation of the ribonucleoprotein complex targeted by tylophorine. Consequently, tylophorine downregulates c-Myc and cyclins D1/D2, causing hypophosphorylation of Rb and suppression of both processing-body formation and the Warburg effect. Gene expression profiling and gain-of-c-Myc-function experiments also revealed that the downregulated c-Myc contributes to the anti-oncogenic effects of tylophorine compounds. Furthermore, the potent tylophorine derivative dibenzoquinoline-33b elicited a similar effect, as c-Myc protein levels were also decreased in xenograft tumors treated with dibenzoquinoline-33b. Thus, tylophorine compounds exert anti-cancer activity predominantly by targeting and sequestering the caprin-1 protein and c-Myc mRNA associated ribonucleoprotein complex.
Anti-angiogenesis targeting VEGFR2 has been considered as an important strategy for cancer therapy. Tylophorine is known to possess anti-inflammatory and antitumor activity, but its roles in tumor angiogenesis, the key step involved in tumor growth and metastasis, and the involved molecular mechanism is still unknown. Therefore, we examined its anti-angiogenic effects and mechanisms in vitro and in vivo.
We used tylophorine and analyzed its inhibitory effects on human umbilical vein endothelial cells (HUVEC) in vitro and Ehrlich ascites carcinoma (EAC) tumor in vivo.
Tylophorine significantly inhibited a series of VEGF-induced angiogenesis processes including proliferation, migration, and tube formation of endothelial cells. Besides, it directly inhibited VEGFR2 tyrosine kinase activity and its downstream signaling pathways including Akt, Erk and ROS in endothelial cells. Using HUVECs we demonstrated that tylophorine inhibited VEGF-stimulated inflammatory responses including IL-6, IL-8, TNF-alpha, IFN-gamma, MMP-2 and NO secretion. Tylophorine significantly inhibited neovascularization in sponge implant angiogenesis assay and also inhibited tumor angiogenesis and tumor growth in vivo. Molecular docking simulation indicated that tylophorine could form hydrogen bonds and aromatic interactions within the ATP-binding region of the VEGFR2 kinase unit.
Tylophorine exerts anti-angiogenesis effects via VEGFR2 signaling pathway thus, may be a viable drug candidate in anti-angiogenesis and anti-cancer therapies.
Tylophorine, a phenanthroindolizidine alkaloid, is the major medicinal constituent of herb Tylophora indica. Tylophorine treatment increased the accumulation of c-Jun protein, a component of activator protein 1 (AP1), in carcinoma
cells. An in vitro kinase assay revealed that the resultant c-Jun phosphorylation was primarily mediated via activated c-Jun N-terminal protein kinase (JNK). Moreover, flow cytometry indicated that ectopically overexpressed c-Jun
in conjunction with tylophorine significantly increased the number of carcinoma cells that were arrested at the G1 phase. The tylophorine-mediated downregulation of cyclin A2 protein levels is known to be involved in the primary G1 arrest. Chromatin immunoprecipitation and reporter assays revealed that tylophorine enhanced the c-Jun downregulation of
the cyclin A2 promoter activity upon increased binding of c-Jun to the deregulation AP1 site and decreased binding to the
upregulation activating transcription factor (ATF) site in the cyclin A2 promoter, thereby reducing cyclin A2 expression.
Further, biochemical studies using pharmacological inhibitors and RNA silencing approaches demonstrated that tylophorine-mediated
elevation of the c-Jun protein level occurs primarily via two discrete prolonged signaling pathways: (i) the NF-κB/PKCδ_(MKK4)_JNK cascade, which phosphorylates c-Jun and increases
its stability by slowing its ubiquitination, and (ii) the PI3K_PDK1_PP2A_eEF2 cascade, which sustains eukaryotic elongation
factor 2 (eEF2) activity and thus c-Jun protein translation. To the best of our knowledge, this report is the first to demonstrate
the involvement of c-Jun in the anticancer activity of tylophorine and the release of c-Jun translation from a global translational
blockade via the PI3K_PDK1_eEF2 signaling cascade.
Cells remove proteins by two processes: degradation and dilution due to cell growth. The balance between these basic processes is poorly understood. We addressed this by developing an accurate and noninvasive method for measuring protein half-lives, called "bleach-chase," that is applicable to fluorescently tagged proteins. Assaying 100 proteins in living human cancer cells showed half-lives that ranged between 45 minutes and 22.5 hours. A variety of stresses that stop cell division showed the same general effect: Long-lived proteins became longer-lived, whereas short-lived proteins remained largely unaffected. This effect is due to the relative strengths of degradation and dilution and suggests a mechanism for differential killing of rapidly growing cells by growth-arresting drugs. This approach opens a way to understand proteome half-life dynamics in living cells.
DCB-3503, a tylophorine analog, inhibits the growth of PANC-1 (human pancreatic ductal cancer cell line) and HepG2 (human hepatocellular cancer cell line) tumor xenografts in nude mice. The inhibition of growth leads to cancer cell differentiation instead of cell death. However, the mechanisms of action of tylophorine analogs is unknown.
In this study, we show that DCB-3503 suppresses the expression of pro-oncogenic or pro-survival proteins with short half-lives, including cyclin D1, survivin, beta-catenin, p53, and p21, without decreasing their mRNA levels. Proteasome inhibitor reversed the inhibitory effect of DCB-3503 on expression of these proteins. DCB-3503 inhibited the incorporation of radiolabeled amino acid and thymidine, and to a much lesser degree of uridine, in a panel of cell lines. The mechanism of inhibition of protein synthesis is different from that of cycloheximide (CHX) as assayed in cell culture and HeLa in vitro translation system. Furthermore, in contrast to rapamycin, DCB-3503 does not affect protein synthesis through the mTOR pathway. DCB-3503 treatment shifts the sedimentation profiles of ribosomes and mRNAs towards the polysomal fractions while diminishing monosome abundance, indicative of the inhibition of the elongation step of protein synthesis. Preferential down regulation of several studied proteins under these conditions is likely due to the relative short half-lives of these proteins.
The inhibitory effect of DCB-3503 on translation is apparently distinct from any of the current anticancer compounds targeting protein synthesis. Translation inhibitors with novel mechanism could complement current chemotherapeutic agents for the treatment of human cancers and suppress the occurrence of drug resistance.
The phenanthroindolizidine and phenanthroquinolizidine alkaloids, typified by tylophorine and cryptopleurine, are a family of plant-derived small molecules with significant therapeutic potential. The plant extracts have been used in herbal medicine and the isolated compounds have displayed a range of promising therapeutic activity such as anti-ameobicidal, anti-viral, anti-inflammatory and anti-cancer activity. Despite their therapeutic protential, no compounds in this class have fully passed clinical trials. Drawbacks include low in vivo anti-cancer activity, central nervous system toxicity and low natural availability. A number of biological effects of these compounds, such as protein and nucleic acid synthesis suppression, have been identified, but the specific biomolecular targets have not yet been identified. Significant effort has been expended in the synthesis and structure-activity-relationship (SAR) studies of these compounds with the hope that a new drug will emerge. This review will highlight important contributions to the isolation, synthesis, SAR and mechanism of action of the phenanthroindolizidine and pheanthroquinolizidine alkaloids.
Abnormal vascular smooth muscle cell (VSMC) proliferation contributes to the pathogenesis of restenosis. Thus, drugs interfering with cell cycle progression in VSMC are promising candidates for an antirestenotic therapy. In this study, we pharmacologically characterize N-5-(2-aminocyclohexyl)-N-7-benzyl-3-isopropyl-1(2)H-pyrazolo[4,3-d]pyrimidine-5,7-di-amine (LGR1406), a novel derivative of the cyclin-dependent kinase (CDK) inhibitor roscovitine (ROSC), in PDGF-BB-activated VSMC. Cell proliferation was quantified measuring DNA synthesis via 5-bromo-2'-deoxyuridine incorporation. Analysis of cell cycle distribution was done by flow cytometry using propidium iodide-stained nuclei. Key regulators of the cell cycle and relevant signaling pathways were dissected by Western blot analyses. In addition, in vitro kinase assays and in silico studies regarding the pharmacokinetic profile of both compounds were performed. LGR1406 shows a stronger (IC(50) = 3.0 muM) antiproliferative activity than ROSC (IC(50) = 16.9 muM), halting VSMCs in G(0)/G(1) phase of the cell cycle, whereas ROSC does not arrest but rather delays cell cycle progression. Neither of the compounds interferes with early PDGF-BB-induced signaling pathways (p38, extracellular signal-regulated kinase 1/2, c-Jun NH(2)-terminal kinase, Akt, signal transducer and activator of transcription 3), and both inhibit CDKs, with LGR1406 exerting a slightly higher potency against CDK1/2 and 4 than ROSC. Expression of cyclins A and E as well as hyperphosphorylation of the pocket proteins retinoblastoma protein and p107 are negatively affected by both compounds, although to a different extent. In silico calculations predicted a much higher metabolic stability for LGR1406 compared with ROSC. Altogether, ROSC derivatives, such as LGR1406 seem to be promising compounds for further development in antirestenotic therapy.
Tylophorine, a representative phenanthroindolizidine alkaloid from Tylophoraindica plants, exhibits anti-inflammatory and anti-cancerous growth activities. However, the underlying mechanisms of its anti-cancer activity have not been elucidated and its effects on cell cycle remain ambiguous. Here, we reveal by asynchronizing and synchronizing approaches that tylophorine not only retards the S-phase progression but also dominantly arrests the cells at G1 phase in HepG2, HONE-1, and NUGC-3 carcinoma cells. Moreover, tylophorine treatment results in down regulated cyclin A2 expression and overexpressed cyclin A2 rescues the G1 arrest by tylophorine. Thus, we are the first to report that the downregulated cyclin A2 plays a vital role in G1 arrest by tylophorine in carcinoma cells.
Despite the lower patency of venous compared with arterial coronary artery bypass grafts, approximately 50% of grafts used are saphenous vein conduits because of their easier accessibility. In a search for ways to increase venous graft patency, we applied the results of a previous pharmacological study screening for non-toxic compounds that inhibit intimal hyperplasia of saphenous vein conduits in organ cultures. Here we analyse the effects and mechanism of action of leoligin [(2S,3R,4R)-4-(3,4-dimethoxybenzyl)-2-(3,4-dimethoxyphenyl)tetrahydrofuran-3-yl]methyl (2Z)-2-methylbut-2-enoat, the major lignan from Edelweiss (Leontopodium alpinum Cass.).
We found that leoligin potently inhibits vascular smooth muscle cell (SMC) proliferation by inducing cell cycle arrest in the G1-phase. Leoligin induced cell death neither in SMCs nor, more importantly, in endothelial cells. In a human saphenous vein organ culture model for graft disease, leoligin potently inhibited intimal hyperplasia, and even reversed graft disease in pre-damaged vessels. Furthermore, in an in vivo mouse model for venous bypass graft disease, leoligin potently inhibited intimal hyperplasia.
Our data suggest that leoligin might represent a novel non-toxic, non-thrombogenic, endothelial integrity preserving candidate drug for the treatment of vein graft disease.
Platelet-derived growth factor-BB (PDGF-BB) acts as a full mitogen for cultured aortic smooth muscle cells (SMC), promoting DNA synthesis and cell proliferation. In contrast, angiotensin II (Ang II) induces cellular hypertrophy as a result of increased protein synthesis, but is unable to drive cells into S phase. In an effort to understand the molecular basis for this differential growth response, we have examined the downstream effects of PDGF-BB and Ang II on regulators of the cell cycle machinery in rat aortic SMC. Both PDGF-BB and Ang II were found to stimulate the accumulation of G(1) cyclins with similar kinetics. In addition, little difference was observed in the expression level of their catalytic partners, Cdk4 and Cdk2. However, while both factors increased the enzymatic activity of Cdk4, only PDGF-BB stimulated Cdk2 activity in late G(1) phase. The lack of activation of Cdk2 in Ang II-treated cells was causally related to the failure of Ang II to stimulate phosphorylation of the enzyme on threonine and to downregulate p27(Kip1) expression. By contrast, exposure to PDGF-BB resulted in a progressive and dramatic reduction in the level of p27(Kip1) protein. The time course of p27(Kip1) decline was correlated with a reduced rate of synthesis and an increased rate of degradation of the protein. Importantly, the repression of p27(Kip1) synthesis by PDGF-BB was associated with a marked attenuation of Kip1 gene transcription and a corresponding decrease in Kip1 mRNA accumulation. We also show that the failure of Ang II to promote S phase entry is not related to the autocrine production of transforming growth factor-beta1 by aortic SMC. These results identify p27(Kip1) as an important regulator of the phenotypic response of vascular SMC to mitogenic and hypertrophic stimuli.
Cyclin-dependent kinase inhibitor p27, a critical determinant for cell cycle progression, is an important regulation target of mitogenic signals during arterial injury. In this study, we show in rat aortic smooth muscle cells that PDGF-BB down-regulated p27 protein and mRNA in an ERK-dependent mechanism. Inhibition of ERK, but not other subtypes of the mitogen-activated protein kinase family, prevented the reduction of p27 protein and mRNA. Conversely, direct activation of ERK via adenovirus-mediated expression of a constitutively active form of MEK led to a reduction of p27 protein and mRNA, further supporting the central role of ERK in regulation of p27 expression. Rapamycin, which potently inhibited PDGF-induced activation of p70 S6 kinase as well as proliferation of smooth muscle cells, did not alter the expression of p27. To delineate the molecular mechanism underlying the p27 down-regulation, we examined the effect of PDGF-BB on p27 promoter activity as well as mRNA stability. Stimulation with PDGF-BB significantly shortened the half-life of p27 mRNA without affecting its promoter activity. To further understand the PDGF-stimulated p27 mRNA turnover, we inserted the 5'- and/or 3'-untranslated regions of p27 cDNA into a non-PDGF-responsive luciferase gene. Only those chimeric genes that contained the 3'-untranslated region responded to PDGF-BB with reduced expression. Moreover, inhibition of ERK completely prevented the effect of PDGF on the chimera expression. In summary, our data suggest that p27 is down-regulated by PDGF-BB in vascular smooth muscle cells through an ERK-dependent posttranscriptional mechanism.
Since its introduction, the propidium iodide (PI) flow cytometric assay has been widely used for the evaluation of apoptosis in different experimental models. It is based on the principle that apoptotic cells, among other typical features, are characterized by DNA fragmentation and, consequently, loss of nuclear DNA content. Use of a fluorochrome, such as PI, that is capable of binding and labeling DNA makes it possible to obtain a rapid (the protocol can be completed in about 2 h) and precise evaluation of cellular DNA content by flow cytometric analysis, and subsequent identification of hypodiploid cells. The original protocol enhanced the capacity for a rapid, quantitative measure of cell apoptosis. For this reason, since its publication, the PI assay has been widely used, as demonstrated by the large number of citations of the original paper and/or the continuous use of the method in many laboratories.
Physiologically, following myocardial infarction (MI), retinoid levels elevate locally in the infarcted area. Whereas therapeutic systemic application of retinoids was shown to reduce the progression of ventricular dilatation and the onset of heart failure, the role of acute physiologically increased retinoids in the infarction zone is unknown to date. To reveal the role of local retinoids in the MI zone is the central aim of this study. Using human cell culture and co-culture models for hypoxia as well as various assays systems, lentivirus-based transgene expression, in silico molecular docking studies, and an MI model in rats, we analysed the impact of the retinoid all-trans retinoic acid (ATRA) on cell signalling, cell viability, tissue survival, heart function, and MI-induced death in rats. Based on our results, ATRA-mediated signalling does aggravate the MI phenotype (e.g. 2.5-fold increased mortality compared to control), whereas 5′-methoxyleoligin (5ML), a new agent which interferes with ATRA-signalling rescues the ATRA-dependent phenotype. On the molecular level, ATRA signalling causes induction of TXNIP, a potent inhibitor of the physiological antioxidant thioredoxin (TRX1) and sensitizes cells to necrotic cell death upon hypoxia. 5ML-mediated prevention of ATRA effects were shown to be based on the inhibition of cellular ATRA uptake by interference with the cholesterol (and retinol) binding motif of the transmembrane protein STRA6. 5ML-mediated inhibition of ATRA uptake led to a strong reduction of ATRA-dependent gene expression, reduced ROS formation, and protection from necrotic cell death. As 5ML exerted a cardioprotective effect, also independent of its inhibition of cellular ATRA uptake, the agent likely has another cardioprotective property, which may rely on the induction of TRX1 activity. In summary, this is the first study to show i) that local retinoids in the early MI zone may worsen disease outcome, ii) that inhibition of endothelial retinoid uptake using 5ML may constitute a novel treatment strategy, and iii) that targeting endothelial and myocardial retinoid uptake (e.g. via STRA6 inhibition) may constitute a novel treatment target in acute MI.
Since the first coronary angioplasty on Sept 16, 1977, the field of percutaneous coronary intervention has evolved rapidly. Now marking its 40th anniversary, percutaneous coronary intervention has become one of the most common medical procedures worldwide. Much of this progress has been due to the iteration and improvement of angioplasty technologies. Balloon angioplasty was limited by unpredictable procedural outcomes due to vessel dissection and recoil, and a high rate of restenosis. The introduction of stents resulted in more stable early results and lower rates of restenosis, although early stent thrombosis and neointimal hyperplasia causing vessel renarrowing were key limitations. Drug-eluting stents delivering antiproliferative agents significantly lowered the rates of restenosis, permitting widespread use of percutaneous coronary intervention in more advanced and complex disease. Although fully bioresorbable scaffolds have the potential to further improve long-term outcomes, they have not yet achieved results equivalent to those of conventional metallic drug-eluting stents in the early years after implantation. Progress in catheter technology did not occur in isolation, and the success of percutaneous coronary intervention is also due to important advances in intracoronary imaging, and adjunct pharmacotherapy—each of which is reviewed in other papers in this Series.
Background:
First-generation drug-eluting coronary stents (DES) were introduced in 2003 to 2004, and their use resulted in a considerable reduction in the development of in-stent restenosis at the cost of an increased risk of late stent thromboses.
Objectives:
This study followed clinical outcomes of patients included in a large randomized trial for 10 years to enable detection of late changes in annual event rates that could necessitate medical attention.
Methods:
A total of 2,098 unselected all-comer patients (50% with acute coronary syndrome) were randomly assigned to have a first-generation DES implanted. This study recorded the occurrence of a major adverse cardiac event (MACE) assessed as the composite of cardiac death, myocardial infarction, and target vessel revascularization. Stent thromboses were also assessed.
Results:
Of the 2,098 unselected patients, 73.1% were still alive after 10 years. During the follow-up period, MACE occurred in 346 (32.5%) in the group receiving a sirolimus-eluting stent and in 342 (33.1%) in the group receiving a paclitaxel-eluting stent (hazard ratio: 0.96; 95% confidence interval: 0.83 to 1.11; p = 0.60), with a steady annual rate of 2.6% after the first year. Definite, probable, and possible stent thrombosis appeared in 279 patients (13.3%), with no difference between stent types and with a steady annual rate of 1.3% after the first year.
Conclusions:
Among the surviving patients, the long-term annual MACE rate and the stent thrombosis rate appeared constant for both stent types, with no apparent late changes. Although there is no need for extraordinary medical attention for these patients, the absence of declines in annual event rates calls for continuous surveillance. (Danish Organization on Randomized Trials With Clinical Outcome II [SORT OUT II]; NCT00388934).
Cancer is responsible for millions of deaths throughout the world every year. Increased understanding as well as advancements in the therapeutic aspect seems suboptimal to restrict the huge deaths associated with cancer. The major cause responsible for this is high resistance as well as relapse rate associated with cancers. Several evidences indicated that cancer stem cells (CSC) are mainly responsible for the resistance and relapses associated with cancer.
Furthermore, agents targeting a single protein seem to have higher chances of resistance than
multitargeting drugs. According to the concept of network model, partial inhibition of multiple
targets is more productive than single hit agents. Thus, by fusing both the premises that CSC and
single hit anticancer drugs, both are responsible for cancer related resistances and screened
alkaloids for the search of leads having CSC targeting ability as well as the capability to
modulating multiple target proteins. The in silico experimental data indicated that emetine and
cortistatin have the ability to modulate hedgehog (Hh) pathway by binding to sonic hedgehog
(Hh), smoothened (Smo) and Gli protein, involved in maintenance CSCs. Furthermore,
solamargine, solasonine and tylophorine are also seems to be good lead molecules targeting
towards CSCs by modulating Hh pathway. Except solamargine and solasonine, other best lead
molecules also showed acceptable in silico ADME profile. The predicted lead molecules can be
suitably modified to get multitargeting CSC targeting agent to get rid of associate resistances.
Intimal hyperplasia is the leading cause of long-term failure in coronary artery bypass vein grafting, coronary artery stenting, angioplasty, arteriovenous fistula for dialysis, and allograft transplantation. Intimal hyperplasia is a product of vascular smooth muscle cell proliferation, migration through the internal elastic lamina, and deposition of extracellular matrix proteins driven by growth factors in the vasculature. This vascular pathology results in a progressive diminution of the vessel lumen and serves as a site for thrombosis and atherosclerotic lesions. A key cell type in the initiation of intimal hyperplasia is the vascular endothelial cell, which appears to have down-stream effects on the vascular smooth muscle proliferation and migration. Currently, the only means available for prevention of intimal hyperplasia is through inhibition of mammalian target of rapamycin (mTOR) with the immunosuppressant rapamycin. mTOR integrates up-stream signals from growth factors such as IL-2 and senses the cellular nutrient and energy levels and redox status. This presentation will discuss the potential means of preserving the vascular endothelial cell and, thereby, reducing the development of intimal hyperplasia in our open-heart surgical patients.
Tylophorine and related natural compounds exhibit potent antitumor activities. We previously showed that PBT-1, a synthetic C9-substituted phenanthrene-based tylophorine (PBT) derivative, significantly inhibits growth of various cancer cells. In this study, we further explored the mechanisms and potential of PBT-1 as an anticancer agent. PBT-1 dose-dependently suppressed colony formation and induced cell cycle G2/M arrest and apoptosis. DNA microarray and pathway analysis showed that PBT-1 activated the apoptosis pathway and mitogen-activated protein kinase signaling. In contrast, PBT-1 suppressed the nuclear factor kappaB (NF-kappaB) pathway and focal adhesion. We further confirmed that PBT-1 suppressed Akt activation accelerated RelA degradation via IkappaB kinase-alpha and down-regulated NF-kappaB target gene expression. The reciprocal recruitment of RelA and RelB on COX-2 promoter region led to down-regulation of transcriptional activity. We conclude that PBT-1 induces cell cycle G2/M arrest and apoptosis by inactivating Akt and by inhibiting the NF-kappaB signaling pathway. PBT-1 may be a good drug candidate for anticancer chemotherapy.
A simple resazurin-based cytotoxicity assay is presented for screening of cytotoxicity in hepatocytes and liver cell lines. Human hepatoma (HepG2) cells in 96-well culture plates were exposed to known toxic (cisplatin, 5-fluorouracil, ethionine, flufenamic acid, and diflunisal) and control (transplatin, 5-chlorouracil, methionine, and acetylsalicylic acid) compounds for 1-3 days, and resazurin (5 micromol/L) was added. A conventional short-term (1 h) assay was first performed, where cytotoxicity is indicated by decreased reduction of resazurin to its fluorescent product resorufin. Our improved assay consists of additionally measuring fluorescence 2-4 days later, when cytotoxicity is indicated by a striking increase in the concentration of resorufin, resulting from two distinct processes. First, viable liver-derived cells slowly convert resorufin to nonfluorescent metabolites. Fluorescence of control cell wells decreased to background during a 2- to 4-day exposure to resazurin. This metabolism of resorufin was largely blocked by dicumarol and to lesser extents by disulfiram and SKF525a. Second, dead or dying cells slowly convert resazurin to resorufin but do not further metabolize resorufin; thus this fluorescent metabolite accumulates to high levels in wells with dead cells by 2 to 4 days. A similar increase in fluorescence associated with cytotoxicity was observed in primary cultures of rat hepatocytes using the long-term resazurin-based assay. In addition to an improved signal relative to the short-term assay, the inversion of the fluorescent signal from high = alive short-term to high = dead long-term allows determination of two independent cytotoxicity endpoints after addition of one innocuous vital dye.
We previously demonstrated that platelet-derived growth factor (PDGF) induces the cyclin-dependent kinase inhibitor p21/WAF1 promoter in vascular smooth muscle cells (VSMC) via activation of a Sp1 site in VSMC. In this report, the role and relevance of the signaling pathway in the transcriptional regulation of p21WAF1 in VSMC was examined. PDGF stimulated the expression of p21WAF1 in VSMC, as evidenced by Immunoblot and Northern blot analyses. Treatment with PD98059, a specific MEK inhibitor, and the transient expression of VSMC with DN-MEK1 plasmid effectively down-regulated PDGF-induced p21WAF1 expression and promoter activity, respectively. Furthermore, the transactivation of PDGF-stimulated Sp1 was inhibited by treatment with PD98059 and the transient expression of VSMC with the DN-MEK1 plasmid. Finally, the transient transfection of VSMC with a dominant negative Ras (RasN17) suppressed PDGF-induced ERK activity, p21WAF1 expression, and promoter activity. The overexpression of RasN17 also abolished PDGF-stimulated Sp1 activity. In conclusion, the findings herein presented indicate that the activation of the Ras/ERK pathway contributes to the induction of p21WAF1 expression in VSMC. In addition, the transcription factor Sp1 that is involved in the Ras/ERK-mediated control of p21WAF1 regulation in VSMC in response to PDGF has now been identified.
C9-Substituted phenanthrene-based tylophorine derivatives (PBTs) (13-36) were synthesized and evaluated as in vitro anticancer agents against the human A549 lung cancer cell line. Twelve active compounds were further examined against DU-145 (prostate), ZR-751 (breast), KB (nasopharyngeal), and KB-Vin (multidrug resistant KB subline) human cancer cell lines. They showed potent cytotoxic activity against both wild type and matched multidrug resistant KB cell lines, and displayed notable selectivity toward DU-145 (prostate) and ZR-751 (breast) cancer cell lines. The mode of action of this class may be distinctly different from that of other cancer chemotherapeutic compounds. Three PBT analogs were also evaluated in a murine model. Compound 24b showed modest in vivo antitumor activity against human A549 xenograft in nude mice as well as potent in vitro cytotoxic activity, and thus, is a promising anticancer lead compound.
Long-term safety of drugeluting and bare-metal stents: evidence from a comprehensive network meta-analysis
- T Palmerini
- U Benedetto
- G Biondi-Zoccai
- D Della Riva
- L Bacchi-Reggiani
- P C Smits
- G J Vlachojannis
- L O Jensen
- E H Christiansen
- K Berencsi
- M Valgimigli
- C Orlandi
- M Petrou
- C Rapezzi
- G W Stone
Palmerini, T., Benedetto, U., Biondi-Zoccai, G., Della Riva, D., Bacchi-Reggiani, L., Smits,
P.C., Vlachojannis, G.J., Jensen, L.O., Christiansen, E.H., Berencsi, K., Valgimigli, M.,
Orlandi, C., Petrou, M., Rapezzi, C., Stone, G.W., 2015. Long-term safety of drugeluting and bare-metal stents: evidence from a comprehensive network meta-analysis. J. Am. Coll. Cardiol. 65, 2496-2507.