Proceedings book download - 5Z.com

Proceedings of the 31 st European Peptide SymposiumMichal Lebl, Morten Meldal, Knud J. Jensen, Thomas Hoeg-Jensen (Editors)European Peptide Society, 2010Design and Synthesis of Guanidinium-Rich MolecularTransporters for Drug DeliveryTatyana Dzimbova 1 , Kaloyan Georgiev 2 , and Tamara Pajpanova 11 Institute of Molecular Biology, Bulgarian Academy of Sciences, Sofia, Bulgaria; 2 MedicalUniversity of Varna, Faculty of Pharmacy, Varna, BulgariaIntroductionFor many therapeutic applications, it has become more important to find syntheticcompounds that have ability to transport a variety of drugs and cargo molecules into cellsand tissues.Microscopic studies revealed that oligoarginines and other arginine-rich peptides areefficiently taken up by cells. The molecule with the highest efficiency turned out to beoctaarginine, whereas peptides of < 5 and >12 arginine residues showed only negligibletranslocation. A direct comparison of nonamers composed of arginine, histidine, lysine orornithine revealed that arginine residues were most effective in penetrating the plasmamembrane because of their guanidinium group [1]. It was also shown that some branchedoligoarginines exhibited potent in vivo antiangiogenic activity and powerful in vitroangiogenesis inhibition of basic fibroblast growth factor (bFGF)-stimulated proliferation.These results suggest that arginine oligomers may be a useful tool for angiogenic diseases.To evaluate this hypothesis, a series of octamers were prepared that incorporated argininemimetics with either, oxy- or sulfoguanidino side chain (Figure 1). In addition lysineoligomers were also synthesized.Further antiproliferative and cytotoxic potential of the forming amino acid analogueswere examined in a panel of human tumor cell lines, as well as against VEGF-stimulatedhuman umbilical vein endothelial cells (HUVECs).Results and DiscussionYH NNHHNN H 2SynthesisPeptide oligomers (Cav) 8 , (NCav) 8 , (sArg) 8 and(sLys) 8 were obtained by conventional Fmoc solidphase chemistry on a Rink-amide resin withDIC/HOBt activation. Amino acids, Fmoc-Cav(Boc)-OH, Fmoc-Cav(NO 2 )-OH, Fmoc-NCav-OH, Fmoc-sArg(Boc)-OH and FmocsLys(Boc)-OHwere synthesized as previouslydescribed with minor modification [2-4]. TheFmoc moiety was deprotected by 20%piperidine/DMF and the activation was catalyzedwith NMP/DMF (0.4 M) for each cycle. The crudepeptides were purified by C18 reverse-phaseMPLC and their purity was checked by analyticalHPLC. Electro-spray MS were in agreement withthe expected results.Antiproliferative and cytotoxic determinationThe cytotoxic activity of compounds Cav, NNC, NNA and NsArg was studied in a panelof human tumor cell lines, as follows: HL-60 (acute myeloid leukemia), K-562 and LAMA-Enrichment factor (arbitrary units)Arginine2.0 *1.51.00.5COOHN H 2YH NNHNHXnCOOHAgrinine analogsX = O, SO 2 ; n = 1-3Y = H, NO 2NH 2H 2 NH 2 NCOOHLysine0.0Control 1 2 3 CAV ART-OHNH 2NHXnCOOHLysine analogsFig. 1. lysine analoguesFig. 1. Arginine and lysine analogs(Cav: X=O, n=2; NCav: X=O,n=1; sArg: X=SO 2 , n=2; NsArg:X=SO 2 , n=1; NNC: X=O, Y=NO 2 ,n=2; NNA: Y=NO 2 ).Fig. 2. Enrichment of the cytosole of HUVECs withmono and oligonucleosomal DNA-fragments aftertreatment with compounds NNC (1), NNA (2), NsArg(3), and Cav (at 500 µM) or ART-OH (at 50 µM), asassessed by a “Cell Death Detection” ELISA kit. Theasterisk indicates p