Multi-omics profiling reveals that targeting NEK3 synergizes with cuproptosis to promote M1 macrophage polarization in cervical cancer
Cervical cancer (CC) remains a significant clinical challenge, necessitating novel therapeutic strategies that target tumor cells and remodel the immunosuppressive microenvironment. Cuproptosis, a copper-dependent cell death process, may influence antitumor immunity, but its mechanisms remain unclear. This study examines how cuproptosis modulates macrophage polarization in cervical cancer, focusing on the kinase NEK3. We employed a multi-omics approach, combining RNA sequencing and proteomic profiling of CC cells treated with elesclomol and CU (ES + Cu) to induce cuproptosis. Functional validation was conducted through in vitro assays measuring cell viability, migration, invasion, ROS production, and mitochondrial membrane potential. The role of NEK3 was further elucidated using overexpression and knockdown models. Macrophage polarization was assessed using co-cultures, ELISA, and flow cytometry. Cuproptosis induction suppressed CC cell proliferation, migration, and invasion, accompanied by ROS accumulation and mitochondrial dysfunction. Multi-omics analyses revealed extensive reprogramming of cellular stress and proteostasis pathways. NEK3 was identified as a key suppressor of cuproptosis. While NEK3 overexpression inhibited cuproptosis, its knockdown enhanced sensitivity to this cell death pathway. Furthermore, NEK3 depletion in tumor cells promoted M1-like macrophage polarization via MAPK signaling and increased macrophage recruitment. In vivo, NEK3 knockdown synergized with cuproptosis induction, leading to significant tumor growth suppression and an increased M1/M2 macrophage ratio. Our findings establish NEK3 as a critical molecular link between cuproptosis and Antitumor immunity in cervical cancer. By regulating both cell death sensitivity and macrophage polarization, NEK3 represents a promising therapeutic target for synergizing with cuproptosis-inducing agents to achieve dual cytotoxic and immunomodulatory effects.