Synthesis and Biological Evaluation of RGD Peptidomimetic-Paclitaxel Conjugates Bearing Lysosomally Cleavable Linkers

• Published in: Chemistry : a European journal 2015-04, Vol.21 (18), p.6921-6929
• Main Authors: Dal Corso, Alberto, Caruso, Michele, Belvisi, Laura, Arosio, Daniela, Piarulli, Umberto, Albanese, Clara, Gasparri, Fabio, Marsiglio, Aurelio, Sola, Francesco, Troiani, Sonia, Valsasina, Barbara, Pignataro, Luca, Donati, Daniele, Gennari, Cesare
• Format: Article
• Language: English
• Subjects: Antigens; Antineoplastic Agents - chemical synthesis; Antineoplastic Agents - chemistry; Antineoplastic Agents - pharmacology; antitumor agents; Binding, Competitive; Biotechnology; Cell Line, Tumor; Cell Proliferation - drug effects; Cell Survival - drug effects; Chemistry; Conjugates; Devices; Diketopiperazines - chemistry; drug delivery; Drug Liberation; Drug Stability; Enzymes; Humans; Integrin alphaVbeta3 - metabolism; integrins; Leukemia; Ligands; Lysosomes - chemistry; Molecular Structure; Nanostructure; Oligopeptides - chemistry; Paclitaxel - analogs & derivatives; Paclitaxel - chemical synthesis; Paclitaxel - chemistry; Paclitaxel - pharmacology; Peptides; Peptides, Cyclic - chemical synthesis; Peptides, Cyclic - chemistry; Peptides, Cyclic - pharmacology; peptidomimetics; Peptidomimetics - chemical synthesis; Peptidomimetics - chemistry; Peptidomimetics - pharmacology; prodrugs; Receptors; Selectivity
• ISSN: 0947-6539; 1521-3765
• DOI: 10.1002/chem.201500158

Two small‐molecule–drug conjugates (SMDCs, 6 and 7) featuring lysosomally cleavable linkers (namely the Val–Ala and Phe–Lys peptide sequences) were synthesized by conjugation of the αvβ3‐integrin ligand cyclo[DKP–RGD]‐CH2NH2 (2) to the anticancer drug paclitaxel (PTX). A third cyclo[DKP–RGD]–PTX conjugate with a nonpeptide “uncleavable” linker (8) was also synthesized to be tested as a negative control. These three SMDCs were able to inhibit biotinylated vitronectin binding to the purified αVβ3‐integrin receptor at nanomolar concentrations and showed good stability at pH 7.4 and pH 5.5. Cleavage of the two peptide linkers was observed in the presence of lysosomal enzymes, whereas conjugate 8, which possesses a nonpeptide “uncleavable” linker, remained intact under these conditions. The antiproliferative activities of the conjugates were evaluated against two isogenic cell lines expressing the integrin receptor at different levels: the acute lymphoblastic leukemia cell line CCRF‐CEM (αVβ3−) and its subclone CCRF‐CEM αVβ3 (αVβ3+). Fairly effective integrin targeting was displayed by the cyclo[DKP–RGD]–Val–Ala–PTX conjugate (6), which was found to differentially inhibit proliferation in antigen‐positive CCRF‐CEM αVβ3 versus antigen‐negative isogenic CCRF‐CEM cells. The total lack of activity displayed by the “uncleavable” cyclo[DKP–RGD]–PTX conjugate (8) clearly demonstrates the importance of the peptide linker for achieving the selective release of the cytotoxic payload. It's all about selectivity: Two RGD peptidomimetic–paclitaxel (PTX) conjugates endowed with peptide linkers were prepared, and their activities as tumor‐homing devices were tested in vitro against two cancer cell lines expressing different levels of αvβ3 integrin. One of these conjugates (see scheme; AA: amino acid) was able to kill the αvβ3‐expressing tumor cells with remarkably increased selectivity relative to that of the free PTX.