A Pitstop-2 analog impairs viability of aggressive lung cancer cells by disrupting nuclear pore integrity.

We previously demonstrated that Pitstop-2, an inhibitor of clathrin-mediated endocytosis (CME), exhibits CME-independent inhibitory effects on nuclear pore complexes (NPCs). Pitstop-2 interferes with β-propeller folds in both clathrin coats and NPC scaffold proteins. NPCs are not only the mediators of all nucleocytoplasmic transport but are also involved in regulating fundamental cellular physiological processes, including gene expression and proliferation. Their upregulation is strongly associated with malignant transformation, as evidenced in our studies involving non-small cell lung cancer (NSCLC) cells. Therefore, herein, we set out to design and synthesize novel compounds using Pitstop-2 as a lead substance. Since the inhibition of NPC formation was recently shown to cause cancer cell death selectively, our efforts focused on designing compounds with enhanced inhibitory effects on NPCs. Among these, a Pitstop-2 analog, CSV-22, demonstrated the highest pharmacological potency and exhibited NPC-disruptive effects superior to those of Pitstop-2 at lower concentrations. Computational docking analysis revealed that CSV-22 interacts with β-folds in NPC scaffold proteins, which are essential for the structural and functional integrity of NPCs. Functional assays revealed that CSV-22 selectively impairs viability in highly metastatic NSCLC cells, with lower IC₅₀ values after 24-hour exposure. Transferrin uptake assays further suggest that CSV-22 does not significantly inhibit CME in NSCLC, distinguishing its mechanism from Pitstop-2. These findings position CSV-22 as a promising candidate for targeted cancer therapy.