Communication www.rsc.org/chemcomm CHEMCOMM New nucleoside based solid supports. Synthesis of 5A,3A-derivatized thymidine analogues† M. de Champdoré, a L. De Napoli, a G. Di Fabio, a A. Messere, b D. Montesarchio a and G. Piccialli* c a Dipartimento di Chimica Organica e Biochimica, Università degli Studi di Napoli ‘Federico II’, via Cynthia 4, 80126 Napoli, Italy b Dipartimento di Scienze Ambientali, Seconda Università di Napoli, via Vivaldi 43, 81100 Caserta, Italy c Facoltà di Scienze, Università del Molise, via Mazzini 8, 86170 Isernia, Italy. E-mail: picciall@unina.it; Fax: 39-081-674393; Tel: 39-081-674127/6/5 Received (in Cambridge, UK) 8th August 2001, Accepted 19th October 2001 First published as an Advance Article on the web 26th November 2001 Two new thymidine based polymeric supports, in which the nucleosides have been anchored through the thymine moiety to a b-hydroxy thioether functionalized resin via a Mitsu- nobu reaction, have been prepared. A simple and efficient solid-phase synthesis of 5A,3A-bis-derivatized thymidine ana- logues has so been developed, following methodologies well established in peptide and oligonucleotide chemistry and is here proposed for the preparation of a variety of different nucleoside conjugate products. Combinatorial chemistry allows the study of the properties of many classes of compounds much faster than a classical serial analysis. The combinatorial approach to the simultaneous polymer-supported synthesis of chemical compounds has allowed the rapid screening of a plethora of different substrates, speeding up lead identification and optimisation, and resulting in fundamental developments in biomedicinal chemistry. 1 Combinatorial syntheses are often carried out as sequential, high fidelity condensation and addition reactions on solid-phase linked scaffolds which are typically polyfunctional molecules. 2 In this frame nucleosides can be regarded as very useful scaffolds to be incorporated into polymeric supports for combinatorial libraries 3 since they contain functional groups which can be differently manipulated and stereogenic centres useful for a defined spatial presentation of the various substituents. In addition, the biomedical potential of nucleoside derivatives is well known. Therefore nucleosides are partic- ularly attractive as platforms for the design of primary screening libraries. Aiming at the synthesis of new functionalized solid supports for combinatorial libraries of chimeric biomolecules incorporating nucleoside moieties (e.g. nucleoside analogues, conjugates or other hybrids), we turned our attention to ad hoc modified polymeric supports linking suitably protected nucleo- side derivatives in which the ribosidic functions are susceptible of further modifications. In order to attach to polymeric supports nucleosides through the base, we have investigated the possibility of immobilizing thymidine derivatives in the solid phase by reacting the imino function of the thymine moiety. For this purpose, the Mitsunobu reaction 4 of aliphatic alcohols with amidic or imidic species may be particularly useful for an efficient N-alkylation of pyrimidine nucleoside derivatives; this approach has been also successfully exploited by us in order to glycosylate the N-3 position of uridine derivatives. 5 Starting from our previous experiments, we first tested the solid phase Mitsunobu condensation between sugar protected thymidine 3 and a commercially available hydroxy Tentagel ® resin (0.24 meq g 21 , 1, Scheme 1), a polymeric support largely employed in peptide and oligonucleotide synthesis. Such reaction gave satisfactory incorporation yields of the thymidine residue (80% yields by DMT test) but did not result in a practically feasible route, since no mild reaction conditions were found for the final detachment of the intact nucleoside from the solid phase. Therefore we have devised an alternative synthetic scheme involving the Mitsunobu reaction of b-hydroxyethylthioether Tentagel resin 2‡ with thymidine derivatives 3 and 4 (Scheme 1) to yield nucleoside derivatized supports 5 and 6, respectively. Such supports have been designed to allow a simple and efficient release of the nucleosidic material via a b-elimination reaction by mild alkaline treatment on the alkylthioethyl function in 5 or 6, once oxidized to sulfone. We report herein a general synthetic strategy to obtain a variety of thymidine- containing hybrid molecules by exploiting supports 5 and 6 which can form phosphoester or glycosidic linkages using the OH functions, and amide or phosphoramidate bonds exploiting the N 3 group, previously reduced to amine. Thymidine derivatives 3 and 4 were loaded onto resin 2 in the presence of PPh 3 and DEAD in THF–DCM to produce supports 5 and 6. The DMT test showed in both cases a typical loading of 0.17 meq g 21 (90% yield). To transiently mask the 3A- hydroxy moiety in support 5 (Scheme 1), the tert-butyldime- thylsilyl (TBDMS) group was employed in view of its well established orthogonality to DMT and its stability under both acidic and mild basic conditions as well as during the steps involved in the glycosidic 6 and phosphotriester 7 linkage formation. The best results in the removal of TBDMS were obtained under treatment with Et 3 N·3HF (overnight, rt) as verified by coupling the 3A-OH with 2A-deoxycytidine phosphor- amidite building block by standard automated phosphoramidite protocol 7 (Scheme 2). The coupling yield was almost quantita- tive (98%) as checked by DMT cation quantitation on 7 and in no case loss of nucleosidic material was observed. Then the thioether function of the support was oxidized with a mClPBA solution 8 (0.5 M in DCM, 1 h, rt) and, after detritylation of the 5A-OH groups, complete detachment from the support was achieved by treatment with conc. aq. ammonia solution (18 h, 60 °C) giving the desired 3A–3A phosphodiester linked dinucleo- tide 8. The crude detached material was purified by reverse phase HPLC and the identity of isolated 8 was ascertained by 1 H, 31 P NMR and MS data. † Electronic supplementary information (ESI) available: experimental details. See http://www.rsc.org/suppdata/cc/b1/b107200p/ Scheme 1 Solid support functionalization by the Mitsunobu approach. This journal is © The Royal Society of Chemistry 2001 2598 Chem. Commun., 2001, 2598–2599 DOI: 10.1039/b107200p