Computational discovery and preclinical validation of a novel nanodrug eEF2 kinase inhibitor for suppression of tumor growth in triple-negative breast cancer by induction of ferroptosis and apoptosis.

Significant genetic heterogeneity has hindered the identification of molecular targets and development of effective targeted therapies for triple negative breast cancer. Currently available targeted therapies are not curative for TNBC patients. Eukaryotic Elongation Factor-2 kinase (eEF2K) is a clinically significant proto-oncogenic therapeutic target linking this atypical alpha kinase to poor patient survival and a key driver of tumor growth and progression in TNBC, positioning it as a critical and emerging molecular target. Development of eEF2K inhibitors for clinical translation has been challenging due to the unknown three-dimensional structure and lack of potent and selective eEF2K inhibitors. Here, we employed a homology modeling, in silico physics-based molecular simulations studies to rationally design, synthesize and in vitro and in vivo identification a novel potent eEF2K inhibitor. The lead compound-2I demonstrated a potential to engage in covalent interactions with eEF2K enzyme, as suggested by in silico covalent docking and static interaction analyses, and significant in vitro inhibitory activity and suppressed primary and multidrug resistant TNBC cell proliferation at submicromolar concentrations, induced ferroptosis and apoptosis, while having no impact on normal breast epithelial cells. In vivo systemic injection of the eEF2K inhibitor encapsulated in single-lipid nanoparticles demonstrated remarkable therapeutic efficacy and suppressing tumor growth in multiple orthotopic TNBC xenograft models in mice with no sign of toxicity. eEF2K inhibition synergistically enhanced the efficacy of standard chemotherapeutics such as paclitaxel. Our results indicate that the novel eEF2K-targeted nanotherapy is safe and has a significant potential for clinical translation as a monotherapy or in combination with chemotherapy for treatment of patients with TNBC or other eEF2K-dependent solid cancers.