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Herein we report the first example of an isoDGR-drug conjugate ( 2), designed to release paclitaxel selectively within cancer cells expressing integrin αVβ3. Conjugate 2 was synthesized by connecting the isoDGR peptidomimetic 5 with paclitaxel via the lysosomally cleavable Val-Ala dipeptide linker. Conjugate 2 displayed a low nanomolar affinity for the purified integrin αVβ3 receptor (IC50=11.0 n m). The tumor targeting ability of conjugate 2 was assessed in vitro in anti-proliferative assays on two isogenic cancer cell lines characterized by different integrin αVβ3 expression: human glioblastoma U87 (αVβ3+) and U87 β3-KO (αVβ3−). The isoDGR-PTX conjugate 2 displayed a remarkable targeting index (TI=9.9), especially when compared to the strictly related RGD-PTX conjugate 4 (TI=2.4). [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]