Metal-based Theranostic and Therapeutic Agents for The Targeted Treatment and Imaging of Tumors

Abstract

The present PhD thesis resulted from the undertaking of two separate research projects, both anyway being strictly related to the rational design of metal complexes with potential therapeutic and/or diagnostic applications. The first project (discussed in Chapter 2) dealt with the design of vitamin B12-functionalized platinum(II)-based metallotheranostic agents for the targeted treatment and imaging of tumors. Vitamin B12 (cyanocobalamin) is an essential nutrient with very low bioavailability. Upon cell internalization, it undergoes conversion into its cofactors methylcobalamin (used to produce methionine) and adenosylcobalamin (a coenzyme involved in Krebs cycle). Compared with normal ones, tumor cells show higher accumulation of vitamin B12 to support their abnormal proliferation, and such increased demand for cyanocobalamin can be exploited for the tumor-specific delivery of therapeutic/diagnostic agents by functionalizing vitamin B12 with suitable metallodrugs and/or luminescent probes. In this context, we here report on the development of fluorescent vitamin B12-metallodrug conjugates of the type [FLUO-B12-{M}] in which cyanocobalamin is functionalized at the 5'-site of the ribose unit with a fluorophore (FLUO: Rhodamine 6G), whereas the CoIII-cyano moiety is coordinated to a metal-based anticancer scaffold ({M}: Pt(II) substrate). The rationale behind the proposed designing approach is based on the evidence that vitamin B12 is converted into its cofactors (methylcobalamin or adenosylcobalamin) inside the cell through the reduction of CoIII to CoII and the subsequent release of the cyano group. Therefore, by binding platinum(II)-containing bioactive substrates[4] to the cyanocobalamin CoIII-CN moiety, should the overall bioconjugate accumulate preferentially in the tumor cells, the cytotoxic species {CN-metallodrug} would be released directly into the diseased site where it can exert its anticancer activity without affecting healthy tissues. Additionally, the fluorophore attached at the 5'-ribose moiety would allow the transport and biodistribution to be followed and assessed by fluorescence spectroscopy. Four novel cyanocobalamin-platinum(II) derivatives were successfully generated and fully characterized, including the evaluation of their lipophilicity and luminescent properties. Although exhibiting low antiproliferative activity (IC50 = 40-70 M), both fluorescent vitamin B12-platinum(II) conjugates showed enhanced capability to inhibit cells viability compared with the inactive metal precursors and non-fluorescent vitamin B12-platinum(II) analogues, confirming the beneficial effect of the functionalization with the rhodamine scaffold not only for imaging purposes but also in a view to improving their biological activity. The second project (discussed in Chapter 3) involved the design of dual-action platinum(II)-and gold(III)-based selective G-quadruplex (G4) DNA binders for the targeted anticancer chemotherapy. In recent years, G4 nucleic acids have gained growing importance because of their potential involvement in preventing cancer cell proliferation and immortalization, and capability to inhibit oncogene transcription and expression. Therefore, the rational design of small molecules capable of selectively stabilizing G4 structures is a promising strategy to the development of potent anticancer drugs preferably targeting cancer cells over normal ones. Accordingly, the aim of the proposed project was to design metallodrugs in which a metal-based anticancer agent is conjugated to selected G4-targeting fused heterocyclic ligands (e.g. 1,8-naphthalimide derivatives). The goal is to take advantage of the G4 binding properties of the heterocyclic scaffold to generate targeted chemotherapeutics capable of stabilizing G4 and, thus, blocking tumor cell growth/proliferation. Remarkably, should preferential accumulation of the overall metal conjugate into tumor cells be achieved, the cytotoxic metal-containing moiety would also exert its antiproliferative activity only against diseased tissues without affecting healthy ones, thus acting as a “dual-action” anticancer agent. Six novel 1,8-naphthalimide-containing metal complexes based on platinum(II)- or gold(III)-dithiocarbamato scaffolds were successfully generated and fully characterized, including the evaluation of their lipophilicity, stability in solution and luminescent properties. A number of biological studies were carried out and the best performer, the gold(III) complex Au2' was shown to: (i) IG50 values the low micromolar or sub-micromolar range, (ii) trigger the generation of large amounts of ROS without inducing mitochondria depolarization, and (iii) to be quickly taken up by cells and homogeneously localized within the entire cellular space.