[Description]:
Silvestrol is a eukaryotic translation initiation factor 4A (eIF4A) inhibitor isolated from the fruits and twigs of Aglaia foveolata.
[Related Catalog]:
[Target]
[In Vitro]
Silvestrol is a specific eIF4A-targeting translation inhibitor. Silvestrol exhibits significant cytotoxic activity against many human cancer cell lines, such as lung, prostate, and breast cancer with IC50 values ranging from 1 to 7 nM[1]. Silvestrol significantly reduces the number of LNCaP cell colonies. Silvestrol (30 nM, 120 nM) induces apoptosis in LNCaP cells, through the mitochondrial pathway. Apaf-1, Caspase-2, caspase-9, and caspase-10 are involved in Silvestrol-induced apoptosis but caspase-3 and 7 are not[2]. Silvestrol (50 nM) exerts an immediate inhibitory effect and causes near-static cell index compared with the control cells. Silvestrol (6.25 nM) enhances proliferation more than the vehicle control-treated cells, whereas a higher concentration of Silvestrol (50 nM) can inhibit cell proliferation. Silvestrol and episilvestrol display synergistic effects in combination with cisplatin[3]. Silvestrol induces caspase-3 activation and apoptotic cell death in a time- and dose-dependent manner. Silvestrol-mediated cell death is attenuated in ATG7-null mouse embryonic fibroblasts (MEFs) lacking a functional autophagy protein[4].
[In Vivo]
Silvestrol (1.5 mg/kg, i.p.) does not adversely affect production of human IgG by xenografted B-lymphocytes in mice. Silvestrol significantly prolongs survival compared to vehicle. There is no such lymphocyte infiltration detected in the spleens of any of the Silvestrol-treated mice, and nor do these animals exhibit any other obvious signs of lymphoma upon necropsy[5].
[Kinase Assay]
The Apo-ONETM homogeneous caspase-3/7 assay kit is used to measure the activities of caspase-3 and -7. Cells (7×104 cells/mL) are treated with Silvestrol (15 nM, 30 nM, 60 nM, 120 nM, and 240 nM) or etoposide for 24 h in a black 96-well plate. Etoposide, which is known to activate caspases-3/7 in LNCaP cells, is used as a positive control for this assay. At the end of the treatment, lysis buffer and the substrate (Z-DEVD-rhodamine 110) are mixed and added to the cells. Upon sequential cleavage and removal of the DEVD peptides by caspase-3 and -7 activity and excitation at 499 nm, the rhodamine 110-leaving group becomes intensely fluorescent. The emission maximum is 521 nm. The amount of fluorescent product generated is proportional to the amount of caspase-3 and -7 cleavage activity present in the sample. The samples are measured in triplicate. Caspase-3 and -7 activity is indicated by net fluorescence[2].
[Cell Assay]
The cells are seeded at a density of 7×104 cells/mL in 100-mm culture dishes and are treated with 30 nM or 120 nM concentrations of Silvestrol for 24 h. The cells are trypsinized, washed with PBS, and fixed with 1% (w/v) paraformaldehyde in PBS on ice for 30 min. After centrifugation, the cells are suspended in 70% (v/v) ethanol at −20°C until use. All ethanol is removed from the reaction tubes and cells are washed twice with PBS. The cells are processed for labeling with fluorescein-tagged deoxyuridine triphosphate nucleotide at 22°C-24°C overnight, washed, and incubated with propidium iodide/RNase A solution in the dark for 30 min at room temperature. The labeled cells are then analyzed by flow cytometry[2].
[Animal admin]
Mice[5] Peripheral blood mononuclear cells (PBMC) are injected intraperitoneally (IP) into SCID mice depleted of murine natural killer (NK) cells by pretreatment (plus weekly re-treatment) with anti-asialo (GM1). Engraftment is confirmed by hu-IgG ELISA. Treatments with vehicle (30% hydroxypropyl-β-cyclodextrin) or Silvestrol (1.5 mg/kg every 48 hr IP) begin 2 weeks post-engraftment[5].
[References]
[1]. Chambers JM, et al. Synthesis of biotinylated episilvestrol: highly selective targeting of the translation factors eIF4AI/II. Org Lett. 2013 Mar 15;15(6):1406-9.
[2]. Kim S, et al. Silvestrol, a potential anticancer rocaglate derivative from Aglaia foveolata, induces apoptosis in LNCaP cells through the mitochondrial/apoptosome pathway without activation of executioner caspase-3 or -7. Anticancer Res. 2007 Jul-Aug;27(4B):2175-83.
[3]. Daker M, et al. Inhibition of nasopharyngeal carcinoma cell proliferation and synergism of cisplatin with silvestrol and episilvestrol isolated from Aglaia stellatopilosa. Exp Ther Med. 2016 Jun;11(6):2117-2126.
[4]. Chen WL, et al. Silvestrol induces early autophagy and apoptosis in human melanoma cells. BMC Cancer. 2016 Jan 13;16:17.
[5]. Patton JT, et al. The translation inhibitor silvestrol exhibits direct anti-tumor activity while preserving innate and adaptive immunity against EBV-driven lymphoproliferative disease. Oncotarget. 2015 Feb 20;6(5):2693-708.
[6]. Wolfe AL, et al. RNA G-quadruplexes cause eIF4A-dependent oncogene translation in cancer. Nature. 2014 Sep 4;513(7516):65-70.
[7]. Wiegering A, et al. Targeting Translation Initiation Bypasses Signaling Crosstalk Mechanisms That Maintain High MYC Levels in Colorectal Cancer. Cancer Discov. 2015 Jul;5(7):768-781.
[8]. Todt D, et al. The natural compound silvestrol inhibits hepatitis E virus (HEV) replication in vitro and in vivo. Antiviral Res. 2018 Sep;157:151-158.
[Related Small Molecules]