# Novel Drug Combinations in Lung Cancer: New Potential Synergies Between 5-FU and Repurposed Drugs
## 4 Discussion
Morphological analysis further supported these findings, revealing that higher concentrations of Resveratrol led to visible changes in cell phenotype, such as a reduction in cell number and alterations in cellular morphology. These effects were especially pronounced when Resveratrol was combined with the IC\({}_{50}\) concentration of 5-FU. The clonogenic assays provided additional insight, showing that Resveratrol significantly reduced the clonogenicity of A549 cells, particularly when combined with 5-FU. This indicates that Resveratrol not only enhances the immediate cytotoxic effects of 5-FU, but also impairs the long-term survival and proliferative capacity of lung cancer cells. Interestingly, the cell viability assays revealed that the combination of IC\({}_{50}\) 5-FU with Resveratrol did not lead to a significant further reduction in cell viability compared to IC\({}_{50}\) 5-FU alone. This suggests that while Resveratrol has strong anticancer properties, it may not enhance the immediate cytotoxic effects of 5-FU in this specific context. However, considering the severe side effects associated with 5-FU, the finding that combinations of Resveratrol with Levosimendan (50 uM Resveratrol + 1 uM Levosimendan and 100 uM Resveratrol + 0.1 uM Levosimendan) achieved similar reductions in cell viability is particularly promising. This suggests that the Resveratrol and Levosimendan combination could be a viable alternative treatment strategy, potentially offering similar anticancer efficacy with a reduced risk of side effects. The differential effects of Levosimendan and Resveratrol across various cancer cell lines have been studied. For instance, Levosimendan demonstrated significant anticancer activity in bladder cancer cells (UM-UC-5) at low concentrations (0.1 uM), where its combination with 5-FU resulted in a 59% reduction in cell viability [41]. However, in prostate cancer cells (PC-3), higher concentrations of Levosimendan (above 10 uM) were required to observe a substantial reduction in cell viability, indicating a dose-dependent effect that varies across cancer types. These findings highlight the importance of considering cancer subtype and drug concentration when evaluating therapeutic efficacy. Recent research on cervical cancer further elucidates Levosimendan's potential [60]. The study found that Levosimendan had a selective anticancer effect on HPV-negative C33A cells, with an IC\({}_{50}\) value of 58.42 uM. Although its potency was lower compared to cisplatin, a standard therapy for cervical cancer, Levosimendan selectivity and its ability to induce apoptosis in C33A cells make it a promising candidate for use as an adjuvant to standard treatments. The study also revealed that Levosimendan significantly reduced the migration and invasion of C33A cells, suggesting potential antim metastatic properties. The discrepancy in Levosimendan's effects across different cell lines may be attributed to variations in molecular pathways active in each type. Recent findings that Levosimendan activates nitric oxide (NO) production and induces cell-cycle disturbances, particularly the elevation of cells in the G1 phase and reduction in the S phase, provide insights into its mechanism of action [40; 60]. NO is a signaling molecule that plays a critical role in the regulation of blood flow, vascular permeability, and immune responses within the tumor microenvironment. By increasing NO production, Levosimendan may enhance apoptosis and promote cell cycle arrest, which reduces cancer cell proliferation. Additionally, Levosimendan inhibits phosphodiesterase 3 (PDE3), leading to an increase in intracellular cyclic AMP (cAMP) levels. Elevated cAMP levels are associated with antiproliferative effects and the inhibition of metastatic processes in several cancer models, which may explain the reduced clonogenic potential observed in A549 cells [40].