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N-[2-Methyl-5-(triazol-1-yl)phenyl]pyrimidin-2-amine derivatives were synthesized and evaluated in vitro for their potential use as inhibitors of Bcr-Abl. The design is based on the bioisosterism between the 1,2,3-triazole ring and the amide group. The synthesis involves a copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC) as the key step, with the exclusive production of anti-(1,4)-triazole derivatives. One of the compounds obtained shows general activity similar to that of imatinib; in particular, it was observed to be more effective in decreasing the fundamental function of cdc25A phosphatases in the K-562 cell line.

Modern combinatorial chemistry is used to discover compounds with desired function by an alternative strategy, in which the biological target is directly involved in the choice of ligands assembled from a pool of smaller fragments. Herein, we present the first experimental result where the use of in situ click chemistry has been successfully applied to probe the ligand-binding site of Abl and the ability of this enzyme to form its inhibitor. Docking studies show that Abl is able to allow the in situ click chemistry between specific azide and alkyne fragments by binding to Abl-active sites. This report allows medicinal chemists to use protein-directed in situ click chemistry for exploring the conformational space of a ligand-binding pocket and the ability of the protein to guide its inhibitor. This approach can be a novel, valuable tool to guide drug design synthesis in the field of tyrosine kinases. KEYWORDS: ligand-binding site, Abl tyrosine kinase, click chemistry, drug design synthesis T he findings that Bcr-Abl (cytoplasmic tyrosine kinase) is the cause of the leukemic phenotype and that the tyrosine kinase activity of Abl is fundamental for Bcr-Abl-mediated transformation have made this kinase an important target for the development of specific therapies. In the recent past, advances in the selective inhibition of Bcr-Abl kinase activity led to the development of several active compounds, and in particular, imatinib mesylate (Gleevec) is the one that currently represents the front-line therapy of CML. Considering our interest in the discovery of new inhibitors of tyrosine kinases, 1−5 we have recently reported the design and preparation of a small collection of quality N-[2-methyl-5-(triazol-1-yl)phenyl]pyrimidin-2-amine derivatives. 6 The antiproliferative activity in the micromolar range pointed out the efficacy of the compound named FA030 (Figure 1, IC 50 = 0.89 μM on the K-562 cell line). This one showed a potent antienzymatic activity against recombinant Abl kinase (IC 50 = 0.9 ± 0.1 μM). The ADME prediction suggested no significant difference between the behavior exhibited by FA030 and that of imatinib that differs for an amide group in the place of the triazole ring. 6 Moreover, the binding mode was very similar to that of imatinib. FA030 seems to have six hydrogen bonds with the protein, and the majority of contacts are mediated by van der Waals interactions. The triazole ring is involved in two hydrogen bonds with the carbonyl groups of Asp 381 and His 361.Over the last years, target-guided in situ synthesis 7 has attracted our attention because it proved to be a captivating and efficient approach to drug discovery. We applied this strategy to the formation of bivalent compounds that are able to target the tubulin/microtubules dynamic system. 8 Thus, we successfully used tubulin as a target to influence the composition of the mixture of a dynamic combinatorial library by exploiting the disulfide bond exchange reaction. 9 In this scenario, we turned our attention to in situ click chemistry...

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The preparation of fluorescent nanoparticles containing squalenoyl-paclitaxel and a squalene-based fluorescein derivative is presented. The formation of self-assembled heteronanoparticles was confirmed by using the quasielastic light scattering (QELS) technique. The internalization in A549 human lung carcinoma cells was verified by microscopy analysis. Finally, paclitaxel in nanoassemblies maintains its ability to target microtubules.The recent advances in nanotechnology and nanomaterials have been integrated into analytical chemistry for the design of large numbers of fluorescent chemical and biological probes. In recent years instead of using single organic dye probes, these organic fluorophores were encapsulated in a particle matrix. These particles are much brighter than the dyes alone because one particle contains several dyes molecules. In addition, these dyes are more photostable because the entrapment enhances the stability and biocompatibility of the fluorophores. The small size of the nanoparticles allows a high signal-to-noise ratio response and signal amplification, therefore improving the analytical sensitivity and the response time. More importantly, their molecular size minimizes physical perturbation of living cells. Generally, fluorescent nanoparticles show superior optical properties, which facilitate their application for single molecule imaging and long-term tracking of biological molecules in vivo.[1] Our continued interest in the field of chemical approaches to target cancer cells [2][3][4][5][6][7][8][9] has led us to study the preparation of a novel class of squalene conjugated with paclitaxel, podophyllotoxin, camptothecin, and epothilone A. These conjugates were all functionalized with a squalene tail that enabled them to self-assemble in water, and to secure the release of the drug inside the cells through a disulfide-containing linker.[10] The need to trace the delivery of the nanoassemblies and to demonstrate the internalization of the drugs prompted us to prepare fluorescent heteronanoassemblies by mixing a paclitaxel-squalene conjugate and a fluorescein-squalene conjugate (Figure 1).We planned to use the recently described squalenoyl-paclitaxel 5 that was recognized as being able to act as a prodrug and to self-assemble.[11] The reaction of 1,1',2-tris-norsqualene alcohol 4 with fluorescein isothiocyanate (FITC) in the presence of sodium hydride gave the stable compound 6. To verify the ability of compound 6 (fluorescein conjugate) to form nanoassemblies and of compounds 5 and 6 to form heteronanoparticles, [12] five different nanosuspensions were prepared by the solvent displacement method. This consists in dissolving both the compounds in ethanol and removing organic solvent in vacuo after the addition of MilliQ water (Figure 2).Measurement of the mean diameter and polydispersity index by using quasielastic light scattering (QELS) confirmed the presence of self-assemblies (Table 1). It was evident that by decreasing the amount of fluorescein-squalene conjugate 6, the mean diameter...