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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 Polyfunctional Spacers Based onBiodegradable PeptidesLyubov A. Yarinich 1,2 , Lyudmila S. Koroleva 1,2 ,Tatyana S. Godovikova 1,2 , and Vladimir N. Silnikov 11 Institute of Chemical Biology and Fundamental Medicine, Novosibirsk, 630090, RussianFederation; 2 Novosibirsk State University, Novosibirsk, 630090, Russian Federation.IntroductionResearch of drug delivery into cells is actively carried out at present. Release of drug fromcarrier is necessary after uptake of therapeutic construction in target cells. Peptides –substrates for cathepsins taking part in cancer progression has been described in theliterature [1]. Application of such peptides as biodegradable linkers let to release thetherapeutic in transformed cells. Human serum albumin can be used as a carrier for drugdelivery in tumor cells. Modification of Cys-34 sulfhydryl group in albumin permits toobtain bioconjugate of protein with the therapeutic [2]. Antisense oligonucleotides have awide application in molecular biology, as they give an opportunity to have selective effecton gene expression. We propose such approach to delivery of antisense oligonucleotides tocells.Results and DiscussionIt is well known that cathepsin B is carboxypeptidase and cleaves Ala-Leu dipeptidefragments from C-terminal of substrate [3]. Therefore the peptides (AlaLeu) 2 Gly,(AlaLeu) 2 , (AlaLeu) 3 have been synthesized as the first step. Synthesis of peptides wascarried out in solution using Boc-strategy. To obtain bifunctional compounds based onsynthesized peptides, different constructs have been proposed. The first variant wascoupling of diamine to the C-terminal of peptide and maleimide fragment to the N-terminal(Figure 1). 4,9-Dioxa-1,12-diaminododecane was coupled to peptide 1 as linker to increasehydrophilicity of the target conjugate. Further free amino group in 2 has been protected byethyl trifluoroacetate with subsequent deblocking of -amino group of alanine. Compound3 was reacted with pentafluorophenyl ester of N-maleimidohexanoic acid. Finallybifunctional derivative of pentapeptide with trifluoroacetic protection (4) has beensynthesized. After removing of TFA-group, opening of maleimide cycle with loosingreactivity related to sulfhydryl group of cysteine has been noticed.BocAlaLeuAlaLeuGlyOEt11. EtOH, NaOH2.OHO N , N C NO3. H 2 N OOEDiPANH 2OBocAlaLeuAlaLeuGlyNH2OO NH 21. F 3 C O , Et 3 NOH 2 N2. TFA, CH 2 Cl 21. NOOOAlaLeuAlaLeuGlyOFONHFFFOF , EDiPAO32. TFA, CH 2 Cl 2HNOCF 3OONONHAlaLeuAlaLeuGlyFig. 1. Synthetic scheme of maleimide derivative of the AlaLeuAlaLeuGly.OThe second variant was to couple flexible amino linker with maleimide fragment toC-terminus without modification of peptide N-terminal amino group (Figure 2). In this casemaleimide cycle has remained and number of synthetic steps has been reduced. At firstNHO4ONHOCF 3526