Cancer remains one of the leading causes of death worldwide, with millions of new cases diagnosed annually. Cisplatin is a major advance in chemotherapy, but its severe side effects and the development of resistance limit its long-term effectiveness. In this context, palladium(II) complexes have gained attention as structural analogues of platinum compounds because they have the potential to exhibit antitumor activity while reducing toxicity. Six novel palladium(II) complexes containing thiosemicarbazide derivatives and diphosphine ligands [1,3-bis(diphenylphosphine)propane (dppp) or 1,4-bis(diphenylphosphine)butane (dppb)] were synthesized and thoroughly characterized by FTIR, 1H NMR and 31P NMR, high-resolution mass spectrometry, UV–Vis spectroscopy, and single-crystal X-ray diffraction. The structural analyses confirmed distorted square-planar Pd(II) geometries featuring N,S-bidentate thiosemicarbazide and chelating bisphosphine ligands. The cytotoxicity of the complexes was evaluated against breast (MCF-7 and MDA-MB-231), prostate (DU-145), lung (A549), ovarian (A2780 and A2780cis), and non-tumor (MRC-5) cell lines using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assays. One of the complexes exhibited the highest cytotoxicity, with IC₅₀ values approaching 1 μM in ovarian and breast cancer cells. B3 was about 25–30 times more active and selective than cisplatin (SI ≈ 15). Additional tests demonstrated that B3 blocked colony formation and migration, triggered dose-dependent apoptosis, and exhibited minimal toxicity to non-tumor cells. Notably, B3 demonstrated significant activity against cisplatin-resistant ovarian cells (A2780cis) in three-dimensional (3D) spheroid cultures, indicating its potential under physiologically relevant conditions. Overall, the structural features represent a promising lead compound for developing next-generation palladium-based metallodrugs with improved selectivity and effectiveness against resistant tumor types.