Bioconjugation of Cyclometalated Gold(III) Lipoic Acid Fragments to Linear and Cyclic Breast Cancer Targeting Peptides

Cell-targeting peptides (CTPs) are increasingly used in the field of cancer research due to their high affinity and specificity to cell or tissue targets. In the search for novel metal-based drug candidates, our research group is particularly focused on bioconjugates by utilizing peptides to increase the selectivity of cytotoxic organometallic compounds. Motivated by the relatively high cytotoxic activity of gold complexes, such as Auranofin (approved to treat rheumatoid arthritis), for the treatment of various diseases, we anticipated that gold peptide bioconjugates would present interesting candidates for novel breast cancer therapies. For this, we investigate the use of the natural compound lipoic acid (Lpa) as a bioconjugation handle to link Au complexes in the oxidation state +III to peptides using the dithiol moiety. Using this strategy, we have synthesized Au­(III) complex bioconjugates linked to the linear LTVSPWY peptide and two cyclic DfKRG and KTTHWGFTLG tumor-targeting peptides. Solid-phase peptide synthesis (SPPS) was used to prepare the peptides, with lipoic acid introduced N-terminally as a conjugation handle. After peptide cleavage, the metal complex was introduced in solution by first reducing the internal disulfide bond, followed by reaction with Au­(ppy)­Cl₂ (1, ppy: 2-phenyl-pyridine), to yield the Au­(III)–Lpa–peptide bioconjugates. The new bioconjugates were successfully synthesized, purified by semi-preparative HPLC, and characterized by ESI-MS. Au­(III)-peptide bioconjugates were tested as cytotoxic agents against two different human breast cancer cell lines (MCF-7 and MDA-MB-231) and normal human fibroblasts cells (GM5657T) and compared to cisplatin, the parent Au­(III) dichloride complex, and metal-free peptides. These in vitro data show that the Au­(III)-peptide bioconjugate 5, possessing the cyclic integrin-targeting RGD-derived peptide sequence in the structure, exhibits improved activity compared to the parent gold­(III) compound Au­(ppy)­Cl₂ (1) as well as to cisplatin or the metal-free peptide. Moreover, the excellent targeting properties of 5 are supported by the fact that a Au­(III)-peptide conjugate with the exact same peptide sequence, but a linear rather than the cyclic form of 5 exhibits 10 times lower cytotoxic activity.