Synthesis, characterization, and biological investigation of thiosemicarbazone-ruthenium(II) complexes

Four novel ruthenium complexes, Ru1-Ru4, were synthesized and characterized, with the general formula [Ru(L)(bipy)(dppb)]PF₆, where bipy = 2,2'-bipyridine, dppb = 1,4-bis(diphenylphosphine)butane, and L corresponds to morpholine-vanilline thiosemicarbazone ligands (VMTSC){(L₁ = 2-(3-methoxy-4-(2-morpholinoethoxy)benzylidene)-hydrazine-1-carbothioamide; L₂ = 2-(3-methoxy-4-(2-morpholinoethoxy)benzylidene)-N-methylhydrazine-1-carbothioamide; L₃ = 2-(3-methoxy-4-(2-morpholinoethoxy)benzylidene)-N-ethylhydrazine-1-carbothioamide; L₄ = 2-(3-methoxy-4-(2-morpholinoethoxy)benzylidene)-N-phenylhydrazine-1-carbothioamide)}. The complexes were characterized by molar conductance, IR, and UV-Vis spectroscopies, mass spectrometry, NMR (¹H, HSQC, COSY, ¹³C-DEPTQ, and ³¹P-{¹H}), and cyclic voltammetry. The purity of the compounds was confirmed by elemental analysis (CHN). In vitro cytotoxicity studies revealed that all compounds exhibited better cytotoxicity toward lung (A549) and breast (MDA-MB-231) tumor cells than cisplatin, showing lower effects on non-cancerous lung (MRC-5) and breast (MCF-10 A) cells. Among the compounds, Ru4 demonstrated the highest cytotoxicity and selectivity against triple-negative MDA-MB-231 cells (IC₅₀ = 1.30 ± 0.03 μM) with the highest selectivity index (SI = 8.3). Clonogenic experiment revealed a cytostatic effect for Ru4. Morphology assays revealed significant changes in the shape of MDA-MB-231 cells, indicating the cell death process. This result was confirmed by live/dead fluorescence assay, showing an increased dead cell population. The wound-healing experiment confirmed that Ru4 was able to inhibit cell migration, showing its potential as a metastasis inhibitor. DNA interaction studies show that Ru4 has a weak interaction with ct-DNA. Human serum albumin assay indicated a moderately spontaneous interaction between Ru4 and the protein, stabilized by hydrophobic interactions. Overall, findings suggest that Ru4 is a promising candidate for future studies in the development as an anticancer compound against breast cancer.