Abstract: Oral squamous cell carcinoma (OSCC) poses a significant global health challenge due to its high recurrence rate and resistance to cisplatin. While DNA repair pathways contribute to chemoresistance, the role of scaffold proteins in stabilizing repair enzymes remains largely unexplored. This study aimed to elucidate the function of CCDC43 in cisplatin sensitivity and to identify a natural product targeting the CCDC43-PNKP axis. Bioinformatic analyses were conducted using bulk and single-cell transcriptomic and proteomic datasets. Molecular assays investigated the CCDC43-PNKP interaction and the impact of neferine on this axis. Xenograft models assessed the efficacy of combined neferine and cisplatin treatment. Machine learning identified CCDC43 as a key prognostic marker and driver of cisplatin resistance. CCDC43 was upregulated in OSCC epithelial cells and associated with poor survival. CCDC43 bound and stabilized PNKP by inhibiting ubiquitin-dependent degradation, thereby enhancing DNA repair and reducing cisplatin sensitivity. Through artificial intelligence-based virtual screening and experimental validation, neferine was found to disrupt the CCDC43-PNKP interaction, promoted PNKP degradation, increased cisplatin-induced DNA damage, and suppressed OSCC growth without acute toxicity. Collectively, the CCDC43-PNKP axis represents a critical therapeutic target in OSCC. Neferine is a promising ethnopharmacological agent capable of improving cisplatin sensitivity by targeting this pathway.