Substitution Kinetics, Albumin and Transferrin Affinities, and Hypoxia All Affect the Biological Activities of Anticancer Vanadium(V) Complexes.

Limited stability of most transition-metal complexes in biological media has hampered their medicinal applications but also created a potential for novel cancer treatments, such as intratumoral injections of cytotoxic but short-lived anticancer drugs. Two related V(V) complexes, [VO(Hshed)(dtb)] (1) and [VO(Hshed)(cat)] (2), where H2shed = N-(salicylideneaminato)-N'-(2-hydroxyethyl)-1,2-ethanediamine, H2dtb = 3,5-di-tert-butylcatechol, and H2cat = 1,2-catechol, decomposed within minutes in cell culture medium at 310 K (t1/2 = 43 and 9 s for 1 and 2, respectively). Despite this, both complexes showed high antiproliferative activities in triple-negative human breast cancer (MDA-MB-231) cells, but the mechanisms of their activities were radically different. Complex 1 formed noncovalent adducts with human serum albumin, rapidly entered cells via passive diffusion, and was nearly as active in a short-term treatment (IC50 = 1.9 ± 0.2 µM at 30 min) compared with a long-term treatment (IC50 = 1.3 ± 0.2 µM at 72 h). The activity of 1 decreased about 20-fold after its decomposition in cell culture medium for 30 min at 310 K. Complex 2 showed similar activities (IC50 ≈ 12 µM at 72 h) in both fresh and decomposed solutions and was inactive in a short-term treatment. The activity of 2 was mainly due to the reactions among V(V) decomposition products, free catechol, and O2 in cell culture medium. As a result, the activity of 1 was less sensitive than that of 2 to the effects of hypoxic conditions that are characteristic of solid tumors and to the presence of apo-transferrin that acts as a scavenger of V(V/IV) decomposition products in blood serum. In summary, complex 1, but not 2, is a suitable candidate for further development as an anticancer drug delivered via intratumoral injections. These results demonstrate the importance of fine-tuning the ligand properties for the optimization of biological activities of metal complexes.

Vanadium(V) Pyridine-Containing Schiff Base Catecholate Complexes are Lipophilic, Redox-Active and Selectively Cytotoxic in Glioblastoma (T98G) Cells.

Two new series of complexes with pyridine-containing Schiff bases, [VV O(SALIEP)L] and [VV O(Cl-SALIEP)L] (SALIEP=N-(salicylideneaminato)-2-(2-aminoethylpyridine; Cl-SALIEP=N-(5-chlorosalicylideneaminato)-2-(2-aminoethyl)pyridine, L=catecholato(2-) ligand) have been synthesized. Characterization by 1 H and 51 V NMR and UV-Vis spectroscopies confirmed that: 1) most complexes form two major geometric isomers in solution, and [VV O(SALIEP)(DTB)] (DTB=3,5-di-tert-butylcatecholato(2-)) forms two isomers that equilibrate in solution; and 2) tert-butyl substituents were necessary to stabilize the reduced VIV species (EPR spectroscopy and cyclic voltammetry). The pyridine moiety within the Schiff base ligands significantly changed their chemical properties with unsubstituted catecholate ligands compared with the parent HSHED (N-(salicylideneaminato)-N'-(2-hydroxyethyl)-1,2-ethanediamine) Schiff base complexes. Immediate reduction to VIV occurred for the unsubstituted-catecholato VV complexes on dissolution in DMSO. By contrast, the pyridine moiety within the Schiff base significantly improved the hydrolytic stability of [VV O(SALIEP)(DTB)] compared with [VV O(HSHED)(DTB)]. [VV O(SALIEP)(DTB)] had moderate stability in cell culture media. There was significant cellular uptake of the intact complex by T98G (human glioblastoma) cells and very good anti-proliferative activity (IC50 6.7±0.9 µM, 72 h), which was approximately five times higher than for the non-cancerous human cell line, HFF-1 (IC50 34±10 µM). This made [VV O(SALIEP)(DTB)] a potential drug candidate for the treatment of advanced gliomas by intracranial injection.

Vanadium Chloro-Substituted Schiff Base Catecholate Complexes are Reducible, Lipophilic, Water Stable, and Have Anticancer Activities.

A hydrophobic Schiff base catecholate vanadium complex was recently discovered to have anticancer properties superior to cisplatin and suited for intratumoral administration. This [VO(HSHED)(DTB)] complex, where HSHED is N-(salicylideneaminato)-N'-(2-hydroxyethyl)-1,2-ethanediamine and the non-innocent catecholato ligand is di-t-butylcatecholato (DTB), has higher stability compared to simpler catecholato complexes. Three new chloro-substituted Schiff base complexes of vanadium(V) with substituted catecholates as co-ligands were synthesized for comparison with their non-chlorinated Schiff base vanadium complexes, and their properties were characterized. Up to four geometric isomers for each complex were identified in organic solvents using 51V and 1H NMR spectroscopies. Spectroscopy was used to characterize the structure of the major isomer in solution and to demonstrate that the observed isomers are exchanged in solution. All three chloro-substituted Schiff base vanadium(V) complexes with substituted catecholates were also characterized by UV-vis spectroscopy, mass spectrometry, and electrochemistry. Upon testing in human glioblastoma multiforme (T98g) cells as an in vitro model of brain gliomas, the most sterically hindered, hydrophobic, and stable compound [t1/2 (298 K) = 15 min in cell medium] was better than the two other complexes (IC50 = 4.1 ± 0.5 µM DTB, 34 ± 7 µM 3-MeCat, and 19 ± 2 µM Cat). Furthermore, upon aging, the complexes formed less toxic decomposition products (IC50 = 9 ± 1 µM DTB, 18 ± 3 µM 3-MeCat, and 8.1 ± 0.6 µM Cat). The vanadium complexes with the chloro-substituted Schiff base were more hydrophobic, more hydrolytically stable, more easily reduced compared to their corresponding parent counterparts, and the most sterically hindered complex of this series is only the second non-innocent vanadium Schiff base complex with a potent in vitro anticancer activity that is an order of magnitude more potent than cisplatin under the same conditions.

Solution- and gas-phase behavior of decavanadate: implications for mass spectrometric analysis of redox-active polyoxidometalates.

Decavanadate (V10O28 6- or V10) is a paradigmatic member of the polyoxidometalate (POM) family, which has been attracting much attention within both materials/inorganic and biomedical communities due to its unique structural and electrochemical properties. In this work we explored the utility of high-resolution electrospray ionization (ESI) mass spectrometry (MS) and ion exclusion chromatography LC/MS for structural analysis of V10 species in aqueous solutions. While ESI generates abundant molecular ions representing the intact V10 species, their isotopic distributions show significant deviations from the theoretical ones. A combination of high-resolution MS measurements and hydrogen/deuterium exchange allows these deviations to be investigated and interpreted as a result of partial reduction of V10. While the redox processes are known to occur in the ESI interface and influence the oxidation state of redox-active analytes, the LC/MS measurements using ion exclusion chromatography provide unequivocal evidence that the mixed-valence V10 species exist in solution, as extracted ion chromatograms representing V10 molecular ions at different oxidation states exhibit distinct elution profiles. The spontaneous reduction of V10 in solution is seen even in the presence of hydrogen peroxide and has not been previously observed. The susceptibility to reduction of V10 is likely to be shared by other redox active POMs. In addition to the molecular V10 ions, a high-abundance ionic signal for a V10O26 2- anion was displayed in the negative-ion ESI mass spectra. None of the V10O26 cations were detected in ESI MS, and only a low-abundance signal was observed for V10O26 anions with a single negative charge, indicating that the presence of abundant V10O26 2- anions in ESI MS reflects gas-phase instability of V10O28 anions carrying two charges. The gas-phase origin of the V10O26 2- anion was confirmed in tandem MS measurements, where mild collisional activation was applied to V10 molecular ions with an even number of hydrogen atoms (H4V10O28 2-), resulting in a facile loss of H2O molecules and giving rise to V10O26 2- as the lowest-mass fragment ion. Water loss was also observed for V10O28 anions carrying an odd number of hydrogen atoms (e.g., H5V10O28 -), followed by a less efficient and incomplete removal of an OH• radical, giving rise to both HV10O26 - and V10O25 - fragment ions. Importantly, at least one hydrogen atom was required for ion fragmentation in the gas phase, as no further dissociation was observed for any hydrogen-free V10 ionic species. The presented workflow allows a distinction to be readily made between the spectral features revealing the presence of non-canonical POM species in the bulk solution from those that arise due to physical and chemical processes occurring in the ESI interface and/or the gas phase.

Enhancement of oncolytic virotherapy by vanadium(V) dipicolinates.

Oncolytic viruses rewire the immune system and can lead to long-lasting antitumor defenses against primary and metastatic tumors. However, results from clinical studies have shown heterogeneity in responses suggesting that multiplexed approaches may be necessary to consistently generate positive outcomes in patients. To this end, we explored the combination of oncolytic rhabdovirus VSV∆51 with vanadium(V) dipicolinate derivatives, which have already been explored for their antidiabetic properties in animal models. The combination of vanadium-based dipicolinate compounds with VSV∆51 significantly increased viral replication and cytotoxicity in the human renal cell carcinoma cell line 786-0. The effects of three vanadium(V)-dipicolinate coordination complexes ([VO2dipic]-, [VO2dipic-OH]- and [VO2dipic-Cl]- with -OH or -Cl in the para position) were compared to that of the simple salts using spectroscopy and speciation profiles. Like the vanadate salts and the vanadyl cation, all dioxovanadium(V) dipicolinate complexes tested were found to increase viral infection and cytotoxicity when used in combination with VSV∆51. Viral sensitization is dependent on the vanadium since free dipicolinate ligands exerted no effect on viral infection and viability. The ability of these complexes to interact with interfaces and the stability of the complexes were evaluated under physiological conditions. Results indicate that these complexes undergo hydrolysis in cell culture media thereby generating vanadate. The vanadium dipicolinate derivatives in the context of immunovirotherapy shares similarities with previous studies exploring the antidiabetic properties of the compounds. The synergy between vanadium compounds and the oncolytic virus suggests that these compounds may be valuable in the development of novel and effective pharmaco-viral therapies.

Linker Variation and Structure-Activity Relationship Analyses of Carboxylic Acid-based Small Molecule STAT3 Inhibitors.

The molecular determinants for the activities of the reported benzoic acid (SH4-54), salicylic acid (BP-1-102), and benzohydroxamic acid (SH5-07)-based STAT3 inhibitors were investigated to design optimized analogues. All three leads are based on an N-methylglycinamide scaffold, with its two amine groups condensed with three different functionalities. The three functionalities and the CH2 group of the glycinamide scaffold were separately modified. The replacement of the pentafluorobenzene or cyclohexylbenzene, or replacing the benzene ring of the aromatic carboxylic or hydroxamic acid motif with heterocyclic components (containing nitrogen and oxygen elements) all decreased potency. Notably, the Ala-linker analogues, 1a and 2v, and the Pro-based derivative 5d, all with (R)-configuration at the chiral center, had improved inhibitory activity and selectivity against STAT3 DNA-binding activity in vitro, with IC50 of 3.0 ± 0.9, 1.80 ± 0.94, and 2.4 ± 0.2 µM, respectively. Compounds 1a, 2v, 5d, and other analogues inhibited constitutive STAT3 phosphorylation and activation in human breast cancer and melanoma lines, and blocked tumor cell viability, growth, colony formation, and migration in vitro. Pro-based analogue, 5h, with a relatively polar tetrahydropyranyl (THP) ring, instead of the cyclohexyl, showed improved permeability. In general, the (R)-configuration Pro-based analogs showed the overall best profile, including physicochemical properties (e.g., microsomal metabolic stability, Caco-2 permeability), and in particular, 5d showed improved tumor-cell specificity.