Network Pharmacology-Guided Elucidation of Metformin and İrinotecan Mechanisms İn 2D and 3D Cancer Cell Models

Abstract

Cancer remains the second leading cause of death globally, following cardiovascular diseases, and continues to represent one of the most critical public health concerns of our time. Beyond its physiological impact, cancer also imposes significant psychological, social, and economic burdens on individuals and societies. Among women, breast and cervical cancers are particularly prevalent. While breast cancer development is associated with genetic predisposition, lifestyle, and socioeconomic factors, cervical cancer is primarily linked to high-risk human papillomavirus (HPV) infections, especially HPV-16 and HPV-18. Despite advances in chemotherapy, commonly used agents such as irinotecan often exhibit toxicity toward healthy cells, induce drug resistance, and face challenges due to the heterogeneous nature of cancer subtypes. These limitations underscore the need for safer, more effective, and targeted therapeutic alternatives. This study aims to explore the potential anticancer effects of metformin, a well-established antidiabetic agent, when used alone or in combination with irinotecan in breast and cervical cancer models. Additionally, it seeks to elucidate the underlying molecular mechanisms governing their cytotoxic and inhibitory effects through experimental and computational analyses. In this study, breast and cervical cancer cell lines were cultured under both two-dimensional (2D) and three-dimensional (3D) in vitro conditions. The cytotoxic effects of metformin and irinotecan, individually and in combination, were assessed through cell viability assays, and IC₅₀ values were determined. To evaluate their influence on tumor progression, migration assays were performed in both culture models. Furthermore, network pharmacology and molecular docking analyses were applied to identify shared molecular targets and key biological pathways potentially involved in the observed effects. The results revealed that both metformin and irinotecan exerted significant cytotoxic activity on the tested cancer cell lines. More importantly, their combined administration led to a stronger suppression of cancer cell proliferation, migration, and invasion compared with single-drug treatments. Network pharmacology analysis highlighted several common molecular targets, including SLC47A1, ACHE, HRH3, EGFR, F2, and NOS1, which are associated with critical biological processes such as apoptosis, angiogenesis, cell cycle control, and cellular stress regulation. Overall, this study demonstrates that the combined use of metformin and irinotecan enhances anticancer efficacy and may offer a promising therapeutic strategy for breast and cervical cancers. The integration of in vitro assays with computational approaches provides deeper mechanistic insights into how these drugs exert their effects, thereby supporting the potential repositioning of metformin as an adjuvant agent in cancer treatment.