Peng Chen

The role of microorganisms in cancer is not a new concept—bacteria, parasites, and viruses have all been implicated in cancer development. Fusobacterium nucleatum , a member of the oral microbiota, has emerged as an important cancer-associated bacterium. Unbiased global genomic analyses have demonstrated that F. nucleatum is enriched in human colorectal cancer (CRC) tissues compared with noncancerous colorectal tissues, underscoring the urgent need for specific antimicrobial agents targeting F. nucleatum for CRC prevention and treatment. In this study, structure-based virtual screening successfully identified ebastine and ceritinib as antimicrobial agents targeting Fusobacterium nucleatum FtsZ (FnFtsZ). Their potent anti -F . nucleatum activity was confirmed through in vitro assays, including minimum inhibitory concentration determination and microscopic observation of filamentous morphology. Both ebastine and ceritinib exhibited significant antibacterial effects. Enzyme activity and polymerization assays demonstrated that their mechanism of action involves inhibition of FnFtsZ GTPase activity and disruption of FnFtsZ polymerization. Scanning electron microscopy (SEM) and light microscopy further confirmed that these compounds effectively disrupt cell division, leading to abnormal or halted division and inhibiting F. nucleatum proliferation. Overall, our findings identified ebastine and ceritinib as potential drugs against F. nucleatum proliferation, although further structural optimization is required. Additionally, our results demonstrate the effectiveness of an integrated computational-experimental approach targeting bacterial cell division in microbiome-associated pathologies. IMPORTANCE: Colorectal cancer (CRC) is strongly driven by the enrichment of the oncogenic bacterium Fusobacterium nucleatum in gut lesions. However, the lack of targeted anti- F. nucleatum drugs significantly limits microbe-related interventions for CRC. In this study, we repurposed two clinically approved drugs, ebastine and ceritinib, as highly effective inhibitors of FnFtsZ, the key protein responsible for bacterial cell division. We demonstrated that both drugs inhibit bacterial proliferation by suppressing FnFtsZ function and disrupting the cell division process. Our work offers safe, readily available candidate agents for controlling CRC-associated pathogenic bacteria and establishes a reliable computational-experimental workflow to screen division-targeted antimicrobials for microbiome-related cancers.

Background: Gastric cancer (GC) remains a leading cause of cancer-related mortality worldwide, highlighting the urgent need for effective and accessible therapies. Drug repurposing offers a cost-effective strategy to identify novel candidates from approved drugs. Terfenadine, a classical antihistamine with an established safety profile, has demonstrated antitumor activity in various malignancies; however, its efficacy and mechanism in GC have not been systematically explored. This study investigates the therapeutic potential of terfenadine in GC and elucidates its underlying molecular mechanisms. Methods: Cytotoxicity was assessed in AGS, HGC27, and MKN45 GC cell lines. Effects on proliferation, colony formation, migration, apoptosis, cell cycle distribution, and mitochondrial membrane potential were examined. Mechanisms were explored through bioinformatics analysis, molecular docking, and Western blotting. Synergy with 5-fluorouracil (5-Fu) was evaluated using checkerboard assays and analyzed by SynergyFinder. In vivo efficacy was validated in a patient-derived xenograft (PDX) model. Results: Terfenadine exhibited potent cytotoxicity against GC cells, with IC ₅₀ values of 5.14 μM (AGS), 3.95 μM (HGC27), and 5.01 μM (MKN45) at 48 h, demonstrating superior potency compared to 5-Fu in AGS and HGC27 cells. It significantly suppressed colony formation and migration, induced G0/G1 phase arrest via downregulation of CDK4/6 and phosphorylated Rb, and promoted mitochondrial apoptosis as evidenced by nuclear condensation and loss of mitochondrial membrane potential. Molecular docking predicted strong binding affinity to AKT (score: 9.09). Western blot analysis revealed that terfenadine treatment reduced the expression of PI3K, total and phosphorylated AKT, and mTOR, indicating modulation of the PI3K/AKT/mTOR pathway. Combination with 5-Fu produced synergistic cytotoxicity (synergy scores >10 b y Loewe and HSA models). In the PDX model, terfenadine (10 mg/kg) significantly suppressed tumor growth, reducing final tumor weight by 41.2% ( p < 0.001). Conclusion: This study demonstrates that terfenadine exerts multifaceted antitumor effects in GC through modulation of the PI3K/AKT/mTOR pathway, exhibits synergistic activity with 5-Fu, and shows in vivo efficacy in a clinically relevant PDX model. These findings support the repurposing of terfenadine as a promising therapeutic agent for GC.