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To improve the biological profile of 20(S)-camptothecin, a novel class of 20-O-linked camptothecin glycoconjugates has been designed for preferential cellular uptake into tumor cells by an active transport mechanism. Such conjugates have been optimized for enhanced solubility, stabilization of the camptothecin lactone ring, sufficient hydrolytic and proteolytic stability, and for an overall improvement in tumor selectivity. The constitution of the peptide spacer has a major impact on stability and biological activity of the conjugates both in vitro and in vivo. Glycoconjugates 17-22 with valine residues at the linkage position to camptothecin are sufficiently stable and show good antitumor activity in vitro against HT29 and other tumor cell lines. Fluorescence microscopy and flow cytometry experiments indicate that glycoconjugates such as 19 are taken up into lysosomal compartments of the tumor cell line HT29 by an active transport mechanism. The steric configuration of the particular amino acid residues linked to the camptothecin moiety has a major impact on the in vivo activity of the corresponding glycoconjugates in the breast cancer xenograft MX-1 model. Inhibiting tumor growth by >96%, the glycoconjugates 19 and 21 show the best activity in this particular model and have been investigated more extensively. The glycoconjugate 19 compares favorably to topotecan 4 and glycoconjugate 21 with respect to toxicity against hematopoietic stem cells and hepatocytes. Based on its profile, 19 has been selected for clinical trials.

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753 20(S)-Camptothecin 1 is a potent cytotoxic agent acting by inhibition of topoisomerase I which has been isolated first by Wall et al. from the wood and bark of the Chinese tree Camptotheca acuminata [1]. Despite some early evidence for activity, clinical trials were suspended because of unfavourable properties of the compound such as toxicity, poor solubility and a species dependent opening of the lactone-E-ring to give the inactive carboxylate form [2]. To overcome these disadvantages a number of camptothecin analogues, prodrugs and delivery systems have been developed and have recently entered the clinics [3]. The tertiary 20-hydroxy group of camptothecin is an attractive functionality in the molecule for the attachment of carrier residues. However, the chemistry for modifying this particular tertiary hydroxy group is challenging because of the steric hindrance and the deactivation due to the neighboured lactone ring. 20-O-Alkyl esters of camptothecin have been prepared by acylation with anhydrides of organic acids to provide prodrugs with increased lactone ring stability [4]. However, the coupling yields decreased dramatically when α-branched organic acids were employed.To achieve increased circulatory retention as well as a continuous therapeutic release of camptothecin, GreenAbstract. To improve solubility and tumor selectivity of 20(S)-camptothecin the synthesis of 20-O-linked glycoconjugates 11A -G is described. Particular focus of the paper is the utilization of N-tert-butoxycarbonyl protected amino acid N-carboxy anhydrides (UNCAs) 2a -f for an efficient acylation of the sterically hindered and deactivated tertiary 20-hydroxy group of 20(S)-camptothecin 1. Depending on the solvent and on the side chain of the amino acid different extents of epimerization of the amino acids during the coupling reaction are observed; however, the epimers can easily be separated after removal of the tert-butoxycarbonyl protecting group and camptothecin amino acid conjugates 4B -E with L-and D-configured amino acids, respectively, are obtained. Particularly, bulky and β-branched amino acids can be atwald et al. utilized the tertiary 20-hydroxy group for esterification with non-immunogenic polyethylene glycol (PEG) 40 kDa dicarboxylic acid and derivatives e.g. in the presence of diisopropyl carbodiimide [5], a method previously described for the modification of paclitaxel [6].To increase water solubility several simple amino acid conjugates of camptothecin and its analogues have been synthesized: Vishnuvajjala et al. functionalized the 20-hydroxy group of camptothecin by chloroacetic acid in pyridine/4-dimethylamino pyridine followed by conversion into the corresponding iodoacetate and amidation with various secondary amines to obtain glycinates [7]. Furthermore, Wani et al. and Wadkins et al. synthesized 20-O-glycinates via coupling of N-tert-butoxycarbonylglycine and subsequent deprotection to obtain water soluble prodrugs [8].Due to the steric hindrance and the deactivation of the tertiary 20-hydroxy group the acylatio...

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