ABSTRACT
The role of fatty acid synthesis in endothelial cells (ECs) remains incompletely characterized. We report that fatty acid synthase knockdown (FASNKD) in ECs impedes vessel sprouting by reducing proliferation. Endothelial loss of FASN impaired angiogenesis in vivo, while FASN blockade reduced pathological ocular neovascularization, at >10-fold lower doses than used for anti-cancer treatment. Impaired angiogenesis was not due to energy stress, redox imbalance, or palmitate depletion. Rather, FASNKD elevated malonyl-CoA levels, causing malonylation (a post-translational modification) of mTOR at lysine 1218 (K1218). mTOR K-1218 malonylation impaired mTOR complex 1 (mTORC1) kinase activity, thereby reducing phosphorylation of downstream targets (p70S6K/4EBP1). Silencing acetyl-CoA carboxylase 1 (an enzyme producing malonyl-CoA) normalized malonyl-CoA levels and reactivated mTOR in FASNKD ECs. Mutagenesis unveiled the importance of mTOR K1218 malonylation for angiogenesis. This study unveils a novel role of FASN in metabolite signaling that contributes to explaining the anti-angiogenic effect of FASN blockade.
Subject(s)
Fatty Acid Synthase, Type I/physiology , Human Umbilical Vein Endothelial Cells/metabolism , Malonyl Coenzyme A/metabolism , Retinal Neovascularization/pathology , TOR Serine-Threonine Kinases/metabolism , Acetyl-CoA Carboxylase/antagonists & inhibitors , Animals , Cell Line, Tumor , Cell Proliferation , Fatty Acid Synthase, Type I/antagonists & inhibitors , Fatty Acid Synthase, Type I/genetics , Human Umbilical Vein Endothelial Cells/cytology , Humans , Mechanistic Target of Rapamycin Complex 1/metabolism , Mice , Mice, Inbred C57BL , Mice, Knockout , Orlistat/therapeutic use , Protein Processing, Post-Translational , Retinal Neovascularization/drug therapyABSTRACT
Distinct structural changes of the α2,3/α2,6-sialic acid glycosidic linkages on glycoproteins are of importance in cancer biology, inflammatory diseases, and virus tropism. Current glycoproteomic methodologies are, however, not amenable toward high-throughput characterization of sialic acid isomers. To enable such assignments, a mass spectrometry method utilizing synthetic model glycopeptides for the analysis of oxonium ion intensity ratios was developed. This method was successfully applied in large-scale glycoproteomics, thus allowing the site-specific structural characterization of sialic acid isomers.
Subject(s)
Proteomics , Sialic Acids/chemistry , Carbohydrate Conformation , Chromatography, Liquid , Stereoisomerism , Tandem Mass SpectrometryABSTRACT
Herein we describe the synthesis of a focused library of compounds based on the structure of goniothalamin (1) and the evaluation of the potential antitumor activity of the compounds. N-Acylation of aza-goniothalamin (2) restored the in vitro antiproliferative activity of this family of compounds. 1-(E)-But-2-enoyl-6-styryl-5,6-dihydropyridin-2(1H)-one (18) displayed enhanced antiproliferative activity. Both goniothalamin (1) and derivative 18 led to reactive oxygen species generation in PC-3 cells, which was probably a signal for caspase-dependent apoptosis. Treatment with derivative 18 promoted Annexin V/7-aminoactinomycin D double staining, which indicated apoptosis, and also led to G2 /M cell-cycle arrest. In vivo studies in Ehrlich ascitic and solid tumor models confirmed the antitumor activity of goniothalamin (1), without signs of toxicity. However, derivative 18 exhibited an unexpectedly lower in vivo antitumor activity, despite the treatments being administered at the same site of inoculation. Contrary to its in vitro profile, aza-goniothalamin (2) inhibited Ehrlich tumor growth, both on the ascitic and solid forms. Our findings highlight the importance of in vivo studies in the search for new candidates for cancer treatment.
Subject(s)
Antineoplastic Agents/chemical synthesis , Aza Compounds/chemistry , Pyrones/chemistry , Acylation , Animals , Antineoplastic Agents/therapeutic use , Antineoplastic Agents/toxicity , Apoptosis/drug effects , Carcinoma, Ehrlich Tumor/drug therapy , Cell Line, Tumor , Cell Proliferation/drug effects , Drug Screening Assays, Antitumor , Female , G2 Phase Cell Cycle Checkpoints/drug effects , Humans , M Phase Cell Cycle Checkpoints/drug effects , Mice , Mice, Inbred BALB C , Reactive Oxygen Species/metabolism , Structure-Activity Relationship , Transplantation, HeterologousABSTRACT
The present work describes the preparation of three novel series of compounds based on the structure of goniothalamin, a natural styryl lactone which has been found to display cytotoxic and antiproliferative activities against a variety of cancer cell lines. A focused library of 29 novel goniothalamin analogues was prepared and evaluated against seven human cancer cell lines. While the γ-pyrones and the aza-goniothalamin analogues were less potent than the lead compound, 2,4-dimethoxy analogue 88 has shown to be more potent in vitro than goniothalamin against all cancer cell lines evaluated. Furthermore, it was more potent than doxorubicin against NCI-ADR/RES, OVCAR-03 and HT-29 while being less toxic to human keratinocytes (HaCat). The 3,5-dimethoxy analogue 90 and 2,4,5-trimethoxy analogue 92 also displayed promising antiproliferative activity when compared to goniothalamin (1). These results provide new elements for the design and synthesis of novel representatives of this family of natural compounds.
