Autocrine Motility Factor-Derived Tetradecapeptides as Dual-Action Chemosensitizers: Bridging Cell Competition...

Backgrounds: Autocrine motility factor (AMF) represents a paradoxical protein with dual roles in cancer progression and therapy. This study investigated AMF-derived tetradecapeptides as novel chemosensitizing agents to overcome multidrug resistance (MDR) in hematological malignancies (HMs). Methods: Seven AMF variants were screened for anticancer activity across 14 human cancer cell lines using MTT and Cell Counting Kit-8 (CCK-8) assays. Four tetradecapeptides (AMF-derived peptides AAP, HGP, HTP, and SKP) corresponding to the AMF206-219 region were designed and evaluated in three HM cell lines (HL-60, CCRF-CEM, IM-9) for growth inhibition, cellular internalization, reactive oxygen species (ROS) production, mitochondrial membrane potential, and gene/protein expression. Synergism with doxorubicin (DOX) and other chemotherapeutic agents was quantified by combination index (CI) analysis using the Chou-Talalay method and spheroid culture assays. Results: AMF variants demonstrated broad-spectrum anticancer activity, with HL-60 acute promyelocytic leukemia cells showing exceptional sensitivity. AMF variants functioned as cell competition-mediated killing signals, converting cancer cells into metabolically compromised "loser" cells through glucose metabolism disruption and oxidative stress induction. Among the four tetradecapeptides, AAP exhibited the most consistent growth inhibition across HM cell lines (HL-60, CCRF-CEM, IM-9) while maintaining gp78/AMFR-mediated cellular internalization. AAP treatment induced dose-dependent ROS production, mitochondrial membrane potential alterations, and p53 upregulation, though glucose-6-phosphate dehydrogenase (G6PD) suppression showed cell-type specificity. Remarkably, AAP demonstrated potent synergistic effects with DOX, reducing IC50 values by 28-53% across tested HM cells through multiple complementary mechanisms: enhanced intracellular DOX retention, elevated oxidative stress, and downregulation of multidrug resistance protein 1 (MDR1; P-glycoprotein) and multidrug resistance-associated protein 1 (MRP1) in specific cell types. AAP similarly synergized with daunorubicin but not with etoposide or cytarabine, suggesting selectivity for ROS-inducing anthracyclines. Conclusions: These findings establish AMF-derived peptides as promising human-origin chemosensitizers capable of overcoming multidrug resistance in HMs through multi-targeted mechanisms. The ability to reduce anthracycline dosing while maintaining efficacy offers potential for minimizing cardiotoxicity and other dose-limiting adverse effects, warranting further preclinical development toward clinical translation.