Periodic Reporting for period 1 - TSPO METALLODRUGS (Novel, highly effective and selective anticancer metallodrugs)
Reporting period: 2018-02-01 to 2020-01-31
This project was aimed at the development of novel, highly effective anticancer metallodrugs that selectively target the mitochondria of cancer cells by combining the favourable cytotoxic properties of gold with the capability of TSPO binders to act as delivery vehicles for antimitochondrial agents.
Cancer cells becoming resistant in the course of treatment is a major challenge in cancer chemotherapy. Anticancer agents that target the mitochondria have the potential to bypass resistance mechanisms and to (re )activate cell death pathways that are mutated or lacking in cancer cells. They are also anticipated to act selectively on tumor cells, as these have a deregulated mitochondrial metabolism. TSPO is a relatively small protein located at the outer mitochondrial membrane and is overexpressed in many tumors. It plays a fundamental role in mitochondrial biochemistry and has a regulatory function in mitochondria-mediated cell death pathways. Thioredoxin reductase (TrxR) is a key enzyme for the maintenance of the cellular redox balance and has a mitochondrial isoform. Its inhibition leads to an increase in reactive oxygen species (ROS) and to a decrease in mitochondrial thiols, both events that trigger cell death. The antitumor activity of many gold complexes is due to the ability of gold(I) to inhibit TrxR.
Cancer cells becoming resistant in the course of treatment is a major challenge in cancer chemotherapy. Anticancer agents that target the mitochondria have the potential to bypass resistance mechanisms and to (re )activate cell death pathways that are mutated or lacking in cancer cells. They are also anticipated to act selectively on tumor cells, as these have a deregulated mitochondrial metabolism. TSPO is a relatively small protein located at the outer mitochondrial membrane and is overexpressed in many tumors. It plays a fundamental role in mitochondrial biochemistry and has a regulatory function in mitochondria-mediated cell death pathways. Thioredoxin reductase (TrxR) is a key enzyme for the maintenance of the cellular redox balance and has a mitochondrial isoform. Its inhibition leads to an increase in reactive oxygen species (ROS) and to a decrease in mitochondrial thiols, both events that trigger cell death. The antitumor activity of many gold complexes is due to the ability of gold(I) to inhibit TrxR.
Six new TSPO binding ligands with specific metal binding sites and 15 new anticancer metallodrug candidates were synthesized and characterized by multinuclear NMR spectroscopy, infra-red spectroscopy and mass spectrometry and their purity was confirmed by elemental analysis and high performance liquid chromatography (HPLC). UV/Vis spectroscopic and HPLC studies confirmed the stabilities of the drug candidates under physiological conditions and in cell culture medium. The interactions of the gold complexes with biological sulfur ligands were studied using the amino acid cysteine as a model for sulfur-containing biomolecules. The results showed that phosphorus and N-heterocyclic carbene co-ligands are easily replaced by cysteine, while the TSPO ligands remain coordinated to gold. Three Au complexes were shown to be highly active towards human bladder cancer cell lines (5637, T24, Caki) with IC50 values in the 0.1 to 5.0 μM range. A correlation between cellular uptake and cytotoxicity in bladder cancer cells could be established. Structure–activity relationship studies showed that compounds with a phosphorus co-ligand are the most active compounds. The cytotoxic Au complexes were shown to be effective inhibitors of TrxR. TSPO molecular docking studies confirmed that all ligands and complexes can interact with the TSPO binding pocket. Thus, the newly synthesized Au complexes represent the first example for combining TrxR inhibition and TSPO binding.
In addition to gold complexes, complexes with Pt(IV) and Cu(II), two other metal ions widely studied in anticancer metalldrug research, were developed and investigated. Two Pt(IV) complexes exhibited IC50 values in breast cancer cells which were 3 - 13 fold lower than that of the reference metallodrug cisplatin and showed selectivity for cancer cells over non-cancerous cells. It was shown for the first time that Pt(IV) pro-drugs for cisplatin with a TSPO binder in an axial position exert cytotoxicity in MCF-7 cells via a multimodal mechanism combining the effects of DNA damage, depolarisation of the mitochondrial membrane potential, swelling and breaking down of the mitochondria and induction of oxidative stress due to enhanced ROS production. Two Cu complexes were synthesized that are more active than the reference metallodrug toward cisplatin-sensitive and cisplatin-resistant ovarian cancer cell lines with IC50 values in the low micromolar range (1.4 to 6.0 μM).
In addition to gold complexes, complexes with Pt(IV) and Cu(II), two other metal ions widely studied in anticancer metalldrug research, were developed and investigated. Two Pt(IV) complexes exhibited IC50 values in breast cancer cells which were 3 - 13 fold lower than that of the reference metallodrug cisplatin and showed selectivity for cancer cells over non-cancerous cells. It was shown for the first time that Pt(IV) pro-drugs for cisplatin with a TSPO binder in an axial position exert cytotoxicity in MCF-7 cells via a multimodal mechanism combining the effects of DNA damage, depolarisation of the mitochondrial membrane potential, swelling and breaking down of the mitochondria and induction of oxidative stress due to enhanced ROS production. Two Cu complexes were synthesized that are more active than the reference metallodrug toward cisplatin-sensitive and cisplatin-resistant ovarian cancer cell lines with IC50 values in the low micromolar range (1.4 to 6.0 μM).
The synthesized Au complexes are the first examples of Au complexes designed to interact simultaneously with the overexpressed TSPO receptor and thioredoxin reductase in cancer cells and thus represent a new class of potential Au- and Cu-based metallodrugs. Furthermore, it was shown for the first time that Pt(IV) pro-drugs for cisplatin with a TSPO binder in an axial position exert cytotoxicity in MCF-7 cells that are notoriously not very sensitive to the benchmark Pt drug cisplatin via a multimodal mechanism combining the effects of DNA damage, loss of the mitochondrial membrane potential and ROS generation.
It is expected that the work carried out during the project will mainly achieve scientific impact through the generation of new knowledge, as the results of the structure-activity relationship and mechanistic studies can inform further cancer research.
It is expected that the work carried out during the project will mainly achieve scientific impact through the generation of new knowledge, as the results of the structure-activity relationship and mechanistic studies can inform further cancer research.