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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, 2010Functionalized PNA Backbone Building Blocks Eligible forDiels-Alder Click Chemistry in Molecular ImagingRüdiger Pipkorn, Manfred Wießler, Waldemar Waldeck,Mario Koch, and Klaus BraunGerman Cancer Research Center, INF 280, D-69120, Heidelberg, GermanyIntroductionTherapies of cancer via “old fashioned” chemotherapeutics normally lead to undesiredadverse reactions followed by abortion of the therapy. Modern chemotherapeutics aremarked by their cell specific interaction with DNA without harming the surroundinghealthy tissue. This progress in chemotherapeutic development led to molecular therapiesdemanding molecular diagnostics [1-3]. The open questions concerning the transport ofimaging components in blood circulation, the differentiation between tumor andsurrounding healthy tissue, and the local enrichment within target tissue and target cells canbe answered as follows: Development and design of the needed molecules led to modularlybuilt, functional molecules which require not only synthesis and quality insurance, but alsoattributes like fast, specific, quantitative and irreversible ligation of these singlemodules[4]. Numerous approaches for improving the ligation of components finally led toour special “click chemistry”. Furthermore, a combination of several selectable differentligation compounds, either diagnostic compounds or therapeutic agents in one multistepreaction is possible by the Diels-Alder reaction with inverse electron demand (DAR inv ).Results and DiscussionThe synthesis of the functionalized PNA backbone for the DAR inv was carried out in thesteps as described here. The commonly used synthesis of the desired PNA building blocksis shown in Figure 1. The chemical synthesis of the “Reppe anhydride”-PNA buildingblock tetracyclo-[5.4.2 1,7 .O 2,6 .O 8,11 ]3,5-dioxo-4-aza-9,12-tridecadiene, Fmoc protected, isshown in Figure 2.Using the solidH 2NNH Cl 2+OOH 2NHNOOFmoc-Succinimidphase synthesis (SPPS)we can prepare functionalmodular PNAbackbonepolymers forcoupling different activeagents or imaging moleculesin order to reachlocal concentrations atthe desired target siteuntil now unachievable.Using the SPPS weprepared a pentamer(shown in Figure 3)harboring a PNA’s amidebackbone with an additionalcysteine attachedto the amino terminus,which in turn will becoupled later to thecysteine of a cell membrane transport facilitating peptide (CPP) permitting an efficientcellular uptake imperative for biochemical studies.This pentamer represents the corresponding reacting partner (a dienophile compound)for the DAR inv mediated ligation to pharmacologically active substances or molecules forimaging in diagnostics harbouring the diene groups. After attachment of the pentamerloaded with the functional molecules to the CPP, which facilitates the passage acrossbiological membranes the modular conjugate forms a “BioShuttle” transporter variant asexemplified in Figure 4.HNFmocFig. 1. Synthesis of the Fmoc protected PNA backbonemodule.Fig. 2. Synthesis of PNA monomer functionalized with Reppeanhydride.HNOO206