Summary, general discussion and outlook

Transcription

1 7 Summary, general discussion and outlook

2 Chapter INTRDUCTIN Chemotherapy has seen much improvement over the last decades in terms of curability and improving the quality of life of patients undergoing treatment [1]. There are, however, many forms of cancer that remain difficult to treat [2]. Cisplatin has proven to be very successful in the treatment of a narrow range of tumours, in particular testicular cancer [3]. Furthermore, it has provided an important basis for the development of other platinum-based chemotherapy [4]. Further advances in cancer chemotherapy can be expected to be made via two distinct strategies. Firstly, existing therapy can be improved through better drug formulation, drug targeting and synthesis and testing of structurally similar compounds. Secondly, the problem can be tackled by defining new targets and designing new drugs specific to these targets. Such an approach will become more apparent as the genetic understanding of cancer increases leading to the discovery of new cancer specific targets [5]. Although this approach may lead to the end of cisplatin in chemotherapy as we know it, it does not imply the end of platinum in medicine altogether. In future drugs, the platinum pharmacophore may also find use in specific agents for targets other than nuclear DNA. The research described in this thesis explores some of the possibilities to improve on the use of cisplatin as a drug. Either through targeting ligands, or through variation of ligands in a combinatorial fashion, platinum complexes with more specific modes of action compared to cisplatin are pursued. These studies have lead to the development of efficient (solid-phase) synthetic methods, as well as to the discovery of several compounds with promising biological properties. In this final chapter the results and findings from this thesis are summarised and evaluated, leading to some suggestions for future research in this field of medicinal chemistry. 90

3 Summary, general discussion and outlook 7.2 SUMMARY AND GENERAL CNCLUSINS The introductory Chapter 1 outlines the field of platinum antitumour chemistry to provide a background for the remainder of the thesis. Starting with the discovery of cisplatin, a summary of the mechanism of action of the drug is described, leading to the development of alternative platinum complexes for anticancer treatment. The uptake of platinum complexes is discussed and related to targeting, a topic that is dealt with extensively in Chapter 2. Finally targets other than nuclear DNA that play a role in cisplatin treatment are described to give background to the experiments carried out in Chapter 6. Chapter 2 gives an extensive literature overview of the different approaches that have been used to increase the efficacy of platinum drugs and reduce systemic toxicity through targeting at cellular or molecular level. The chapter provides insight in the state-of-the-art developments in this field, the main topic of the thesis. In chapters 3, 4, and 5 solid-phase synthesis is employed for the construction of new platinum complexes. Chapter 3 describes a new method to synthesise dinuclear platinum species on the solid phase. In Chapter 4 this method is used to assemble platinum drugs tethered to targeting peptides. The peptides aim to target a conjugated platinum drug-moiety either to a specific tissue or to nuclear DNA. The biological efficacy of these complexes is evaluated and it is observed that, although some of the compounds possess promising cytotoxic activity, the desired targeting effect is achieved to a limited extend only. Clearly, when forming a conjugate of two biologically active entities the resulting complexes do not necessarily display the sum of both activities. This may be due to interaction between the two ends of the molecule, or even due to a changed interaction of one entity with its target resulting from the presence of the other. In Chapter 5 a new solid-phase synthetic method is presented, making asymmetric platinum complexes readily available. Rather than through rational design, random variation is used in this chapter to construct a small series of new platinum complexes. This library revealed several complexes with promising activities during a biological screen, illustrating how combinatorial variation as opposed to rational design, can be a fruitful means for finding new platinum drugs. Indeed when the two hit compounds are evaluated in more detail, the complexes show surprisingly high activities in cell lines sensitive to cisplatin and less reduction in activity in a cisplatin resistant cell line. This result is encouraging, since acquired cisplatin resistance is a major obstacle faced by cisplatin in the clinic. In Chapter 6 the possibility of using the platinum pharmacophore for uses other than DNA binding and the triggering of apoptosis is considered. Complexes targeted towards the active 91

4 Chapter 7 site of cathepsin enzymes are synthesised with varying degrees of reactivity. These complexes are subsequently evaluated for their ability to inhibit cysteine proteases in general, and cathepsin enzymes in particular. In model NMR studies the affinity of the platinum moiety for thiol containing nucleophiles, as present in the active site of the target enzymes, is confirmed. This reactivity, however, does not lead to significant activity in the biological models studied. This lack of activity is ascribed mainly to a lack of affinity of the complexes for the active site of the enzymes studied. 7.3 UTLK AND FUTURE PRSPECTS The new compounds presented in chapters 4 and 6 contain rationally designed targeting functionalities that are supposed to induce selectivity, or increased target affinity compared to the parent compounds. From their biological evaluation, however, it can be seen that such a rational approach will not necessarily lead to useful new drug candidates. Indeed this trend is also seen in the compounds reviewed in Chapter 2. A great deal of platinum compounds designed for receptor-mediated targeting do not show much improvement in activity with regard to their parent compounds. To improve upon the compounds described in Chapter 6 the first and most obvious approach would be to synthesise compounds with an extended or more intricate targeting ligand. This would make the platinum have less influence on the properties of the complex as a whole, presumably leading to an improved delivery of the reactive platinum atom to the active site of the target enzymes. Compound 1 (Figure 7.1) for example, has a targeting ligand resembling that of known inhibitors more closely than the compounds presented in chapter 6. Secondly the non-targeting ligands surroundings the platinum atom can be varied. In the work presented in Chapter 6 only chloride and nitrogen-donating ligands are considered. It would be of interest to investigate the influence oxygen donors or sulfur donors have on the reactivity and biological activity of this class of compounds (2, Figure 7.1). ne could also envisage two targeting elements introduced cis or trans to each other across a platinum atom (3, Fig 7.1). In this case the reactive platinum is hidden inside a larger molecule. Such a complex would mimic the natural substrates of the protease enzymes more closely and may therefore be targeted more efficiently to the active site, possibly leading to good inhibitory properties. 92

5 Summary, general discussion and outlook N 3 Cl N N N N N N N 3 N N N 2 R 1 N 2 N R 2 3 Figure 7.1: New platinum-based cathepsin inhibitors containing a more elaborate targeting functionality, 1; oxygen donor leaving group, 2; or a second targeting element trans to the first targeting ligand, 3. Using the results and observations described in Chapter 4, including the generalised methodology for the construction of platinum peptide conjugates, compounds with a more elaborated targeting ligand may be designed. The lack of selectivity for the complexes targeted towards the α 9 β 1 -integrin-displaying cells is tentatively ascribed to the high lypophilicity of the compounds. It will be of interest to perform studies investigating the binding affinity of the platinum-peptide constructs for the target receptor. Furthermore, testing of compounds containing a hydrophilic element next to the integrin-binding peptide (4, Figure 7.2) will give insight as to whether changing the polarity will increase the target affinity of these constructs. The compounds targeted to the nucleus may benefit from a prodrug approach. By introducing a labile bond near the platinum-drug moiety in these complexes, similar to the complexes described by Kageyama [6], the platinum moiety can be released inside the nucleus (5, Figure 7.2). This would avoid the peptide to interfere with the mechanism of action of the platinum compound after delivery to the DNA. 2 N N N KKKGPLAEIDGIELGKKK 4 N 2 Cl Cl 2 N 2 N targeting peptide 5 N 2 2+ Figure 7.2: A hydrophilic integrin targeted platinum complex, 4 and a prodrug platinum-peptide conjugate, 5. 93

6 Chapter 7 ADEPT prodrugs are prodrugs of a very different nature, and present another promising class of compounds that may present clinically useful targeted platinum drugs. In ADEPT prodrugs an active drug moiety is linked to a hydrophilic group, preventing the prodrug from passive uptake across the cell membrane. The active drug can be released from the prodrug when a specific enzyme cleaves off the hydrophilic moiety. In ADEPT prodrug therapy this enzyme, fused to a cancer targeting antibody, is administered to the patient. Subsequently the prodrug is administered. As a result the active drug is primarily released at the target tissue. In a recent example [7] it is unambiguously illustrated that the ADEPT-prodrug principle is compatible with platinum antitumour drugs. The problem with this complex is, however, that the species released upon enzymatic activation is not a known platinum drug. Designing more intricate platinum-adept prodrugs in which no trace from the hydrophilic part is found after enzymatic activation is likely to lead to exciting new drugs (Figure 7.3). The polynuclear compounds described by Farrell [8] are suited for such an approach. For example, the N moiety in the aliphatic linker of complex 7 can serve as a handle to attach hydrophilic functionalities. In the case of compound 6 this functionality is cleaved as a whole upon enzymatic activation releasing the known active drug species 7. The synthesis of this class of compounds is currently ongoing in Leiden N 2 Cl N N 2 Cl β-glucuronidase N N 2 Cl N 2 Cl 7 + C 2 Figure 7.3: Platinum prodrug, 6, is converted to active dinuclear platinum drug, 7, upon activation by β- glucuronidase. Work resulting from the data presented in Chapter 5 may lead in two directions. Firstly the high success rate of the library described in the chapter may inspire the synthesis of a much larger library of asymmetric cisplatin analogues. By using an automated synthesiser it would be possible to prepare a large matrix of compounds, using a variety of N-Fmoc protected amino alcohols, or amino acids and a large number of commercial and non-commercial amines. By applying a cytotoxicity screen one can then rapidly identify the more potent 94

7 Summary, general discussion and outlook library members. A smaller-scale approach could also be envisaged. Inspired by the hit compound containing methylamine cis to ethanolamine (8, Figure 7.4) two more compounds containing a methylamine ligand cis to a functionalised amino-alcohol have been prepared on a large scale (9, 10, Figure 7.4) [9]. 2 N N 2 2 N N 2 2 N N Figure 7.4: Promising asymmetric platinum compounds Cytotoxicity tests performed with these compounds have revealed that these compounds are not only active in the cisplatin sensitive cell lines, but also in the cisplatin resistant cell line. Furthermore, a direct relationship between the uptake of the compounds and the cytotoxic behaviour was found. igher uptake leading to higher cytotoxicity can be an indication of lower side effects. Clearly these results are of great importance in the development of cisplatin analogues with improved properties compared to the parent compound. REFERENCES [1] S. Neidle, D. E. Thurston, Nat. Rev. Cancer 5 (2005) [2] M. A. Fuertes, C. Alonso, J. M. Perez, Chem. Rev. 103 (2003) [3] S. Schweyer, A. Soruri, A. eintze,. J. Radzun, A. Fayyazi, Int. J. ncol. 25 (2004) [4] J. Reedijk, Proc. Natl. Acad. Sci. U. S. A. 100 (2003) [5]. Yu, R. Jove, Nat. Rev. Cancer 4 (2004) [6] Y. Kageyama, Y. Yamazaki,. kuno, J. Inorg. Biochem. 70 (1998) [7] R. A. Tromp, S. van Boom, C. M. Timmers, S. van Zutphen, G. A. van der Marel,. S. verkleeft, J.. van Boom, J. Reedijk, Bioorg. Med. Chem. Lett. 14 (2004) [8]. Rauter, R. DiDomenico, E. Menta, A. liva, Y. Qu, N. Farrell, Inorg. Chem. 36 (1997) [9] S.. van Rijt, S. van Zutphen,. den Dulk, J. Brouwer, J. Reedijk manuscript in preparation. 95