The effect of novel hydrazone derivatives on human lung, triple-negative breast and skin cancer cell 2D and 3D cultures

Cancer remains one of the leading causes of death worldwide, with lung, breast, and skin cancers being among the types with the highest incidence rates [1]. Lung cancer accounts for 18.7% of all new cancer mortality cases and is the most prevalent type worldwide [1]. Triple-negative breast cancer accounts for 12-17% of all new breast cancer cases and is associated with an adverse prognosis [2]. Although melanoma represents only 1% of skin cancer incidence, it accounts for over 80% of all skin cancer deaths [3,4]. Targeted therapies have demonstrated significant therapeutic potential in both standalone and combination cancer treatments [5]. In this study, we evaluate the anticancer activity of newly synthesized potential kinase inhibitors — hydrazone derivatives — in lung adenocarcinoma cell line A549, triple-negative breast cancer cell line MDA-MB-231, and melanoma cell line IGR39. The impact on cell viability of 14 novel hydrazone derivatives, bearing substitutes pyridine and 1,2,4-triazole moieties, was screened using the MTT assay at 50 µM compound concentration. Selected compounds with significant cytotoxic activity were further studied to establish their EC50 values. Based on determined values, compounds IT160, IT164, IT165, IT166, IT167, and IT168 were identified as promising candidates for further evaluation. The most potent compound, IT168, demonstrated an EC50 profile closest to that of the reference compound sunitinib, a clinically approved kinase inhibitor, with compound IT168 EC50 value being approximately 4 µM (sunitinib 2 µM) to the cancerous cell lines and approximately 5 µM (sunitinib 8 µM) to non-cancerous human fibroblasts (HF). Following EC50 determination, we assessed selected compounds for their effect on cell migration using the wound healing assay at 50% and 10% of their corresponding EC50 values. Only sunitinib exhibited statistically significant inhibition of migration across the used cell lines. Additionally, we evaluated the impact on three-dimensional (3D) cell culture growth and cell viability, revealing that compound 168 significantly reduced cell viability in A549 cell line spheroids at a fixed 5 µM concentration, albeit with lower efficacy than sunitinib. Despite the lack of substantial effects on migration or spheroid growth, the results from cell viability assays suggest potential applications in combination therapy and they aid in further compound structural optimization. In conclusion, compounds IT160, IT164, IT165, IT166, IT167, and IT168 exhibited notable anticancer activity in 2D cultures, with compound IT168 showing the most promising results in the A549 cell line 3D models. Further studies are necessary to explore their mechanisms of action and potential therapeutic applications.