Synthesis and Characterization of Pt(IV) Fluorescein Conjugates to Investigate Pt(IV) Intracellular Transformations

Abstract

Pt(IV) anticancer compounds typically operate as prodrugs that are reduced in the hypoxic environment of cancer cells, losing two axial ligands in the process to generate active Pt(II) species. Here we report the synthesis of two fluorescent Pt(IV) prodrugs of cisplatin in order to image and evaluate the Pt(IV) reduction process in simulated and real biological environments. Treatment of the complexes dissolved in PBS buffer with reducing agents typically encountered in cells, glutathione or ascorbate, afforded a 3- to 5-fold fluorescence turn-on owing to reduction and loss of their fluorescein-based axial ligands, which are quenched when bound to platinum. Both Pt(IV) conjugates displayed moderate cytotoxicity against human cancer cell lines, with IC50 values higher than that of cisplatin. Immunoblotting and DNA flow cytometry analyses of one of the complexes, Pt(IV)FL2, revealed that it damages DNA, causing cell cycle arrest in S or G2/M depending on exposure time, with ultimately triggering of apoptotic cell death. Fluorescence microscopic studies prove that Pt(IV)FL2 enters cells intact and undergoes reduction intracellularly. The results are best interpreted in terms of a model in which the axial fluorescein ligands are expelled through lysosomes, with the platinum(II) moiety generated in the process binding to genomic DNA, which results in cell death.

Conjugation of Cisplatin Analogues and Cyclooxygenase Inhibitors to Overcome Cisplatin Resistance

“…The conjugates are among the most active compounds of the class of platinum(IV) carboxylato complexes, most of which exhibited potencies similar to that of cisplatin. [10, 15c, d, 26a, 31] Only few of the previously reported derivatives have shown highly increased cytotoxicities, with activities in the sub-micromolar range. [15d, 32] Especially potent, however, are the benzoate derivatives, which resemble the structure of the ibuprofen conjugates and showed activities similar to 4 and 6 .…”

Cellular trafficking, accumulation and DNA platination of a series of cisplatin-based dicarboxylato Pt(IV) prodrugs

“…They are kinetically quite inert and should reach the tumor cells intact. Here they undergo reduction by biomolecules, whose low molecular weight (MW) prototypes are ascorbic acid (AA) and reduced glutathione (GSH) [3][4][5]. However, the reduction of Pt(IV) complexes by a still unidentified high MW cellular component plays an important role [6].…”

Conjugates of Cisplatin and Cyclooxygenase Inhibitors as Potent Antitumor Agents Overcoming Cisplatin Resistance

“…The conjugates are some of the most active in the class of platinum(IV) carboxylato complexes, most of which exhibit potencies similar to that of cisplatin. [17a,22] 1 and 2 were even able to completely overcome cisplatin resistance in MDA-MB-231 breast cancer cells, with an up to 400-fold lower IC 50 value than that of cisplatin.…”