Cancer significantly threatens global health, with projections estimating a rise to 28.4 million cases worldwide by 2040 [1]. Characterized by unregulated growth, relentless proliferation, and invasion and metastasis into normal cellular structures [[2], [3], [4]], cancer cells pose a unique challenge to medical science. Chemotherapy remains a critical component of therapeutic strategies. Since the advent of cisplatin, platinum-based drugs have assumed a pivotal role in treatment modalities. However, their clinical utility is often overshadowed by severe side effects and the emergence of drug resistance [[5], [6], [7]]. Consequently, there is an urgent imperative to develop and identify alternatives to platinum-based chemotherapeutics.

Over recent decades, organometallic Ru(II/III) compounds have emerged as potential alternatives to Pt-based drugs, extensively explored in oncological research [[8], [9]]. Notably, three ruthenium complexes, TLD-1433, NAMI-A, and KP1019, have progressed to clinical trials [[10], [11]]. Half-sandwich Ru(II) complexes, characterized by the general formula [LRu(II)(η6-arene/p-cymene)(X)]+ (where L = organic ligands, X = halogen), have demonstrated significant anticancer properties [[12], [13], [14], [15], [16]]. In particular, RM175, RAPTA-C, and RAPTA-T have reached pre-clinical evaluation stages [12,[17], [18]]. However, the clinical success of ruthenium-based therapeutics in tumor management remains elusive. Consequently, the conception and optimization of Ru(II) complexes, emphasizing enhanced anticancer efficacy and target specificity, are paramount for the advancement of Ru-based drugs. Simultaneously, by modulating various ligands (L), these Ru(II) complexes can achieve structural diversity, augmented antiproliferative potency, and targeted anti-tumor specificity [8,12]. Mitochondrion, a vital cellular organelle, is intricately involved in numerous eukaryotic cellular processes, including ATP and ROS production, calcium ion regulation, immune response mediation, and apoptosis [[19], [20]]. Therefore, targeting the mitochondria has gained prominence in cancer therapy strategies, with certain half-sandwich organometallic Ru(II) compounds containing N,N and N,O-donor ligands notably triggering apoptosis through induced mitochondrial dysfunction [18,[21], [22], [23]].

Pyrazolone derivatives frequently feature in medicinal chemistry, contributing to diverse drug designs [[24], [25], [26]]. Numerous studies have documented the utility of these derivatives in formulating antitumor [27], antioxidant [28], antidiabetic [29], and antimicrobial agents [30]. Specifically, 5-phenyl-2-(pyridin-2-yl)-2,4-dihydro-3H-pyrazol-3-ones (PDPO), a prevalent subclass, typically exist as 3-phenyl-1-(pyridin-2-yl)-1H-pyrazol-5-ols, exhibiting weak cytotoxicity against cancerous cells [[31], [32], [33]]. Recent investigations by R. L. Biannic et al. identified PDPO derivatives as potential immune checkpoint inhibitors capable of disrupting PD-1/PD-L1 interactions [33], while K. Tsuganezawa et al. provided evidence supporting the efficacy of PDPO derivatives as inhibitors of human COQ7 [34]. B. Kupcewicz et al. synthesized Cu(II) complexes with PDPO ligands and evaluated their anticancer properties; unfortunately, they did not exhibit significant cytotoxicity [31]. Considering the biological properties of Ru(II) complexes and PDPO characteristics, we postulated that integrating PDPO with a dichloro(η6-p-cymene) ruthenium(II) dimer could lead to the development of more potent Ru(II) anticancer agents, potentially inducing mitochondrial dysfunction.

Six pyrazalone derivatives (PDPO1-PDPO6) and their corresponding Ru(II) complexes (Ru1-Ru6) with incorporated η6-p-cymene moieties were engineered, synthesized, and assessed for in vitro anti-cancer efficacies. These PDPO ligands exhibit a notable characteristic of prototropic tautomerism, attributed to the unsubstituted position on pyrazole C-4 [32], a feature that complicates the structures of the associated Ru(II) complexes and restricts their use in coordination chemistry. Motivated by the nuanced prototropic tautomerism of PDPO, the complex structures were rigorously analyzed using 1H NMR, 13C NMR, FT-IR, HRMS, and X-ray single crystal diffraction methods. In light of the distinct hydrolysis traits of half-sandwich Ru(II) complexes, the hydrolysis behavior of bioactive complexes (Ru1-Ru3, Ru5-Ru6) was examined via UV–vis spectroscopy to uncover their activation mechanisms [22,35]. Ru2 emerged as the most potent anti-cancer agent and was evaluated for its role in inducing apoptosis and cell cycle arrest in the HepG2 cell line. Further investigation employed Western blot analysis to study the expression of cell cycle-associated proteins, including P21, cyclin D, and CDK4, offering insights into the mechanism of HepG2 cell cycle arrest triggered by Ru2. Post Ru2 administration, the intracellular reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) of HepG2 cells were measured, enhancing the comprehension of the anti-cancer mechanisms exhibited by the active complexes.