Abstract
Cancer is the second leading cause of death worldwide, surpassed only by cardiovascular diseases. This study investigated the anticancer effects of recombinant Clostridium α-toxin on breast cancer, both in vitro and in vivo. The entire coding sequence of a codon-optimized α-toxin was designed, cloned into the pET28a (+) vector, and expressed as recombinant α-toxin in Escherichia coli (E. coli) BL 21(DE3) cells transformed with the recombinant plasmid. The recombinant α-toxin was purified using Ni²⁺ affinity chromatography, and its accuracy and purity were confirmed through sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot analysis. The anticancer effects of purified α-toxin were then assessed in vitro and animal models against MCF-7 breast cancer cells. Protein analysis confirmed the presence of a 48 kDa band corresponding to the recombinant α-toxin. Additionally, the IC₅&sub0; values of α-toxin against MCF-7 cells at 24, 48, and 72 h were 407.3±2.392 μg/mL, 287.3±5.411 μg/mL, and 258.1±4.671 μg/mL, respectively. In vivo, results demonstrated a significant reduction in mean cancer nodule size following α-toxin treatment (p<0.001). These findings suggest that α-toxin may serve as a promising candidate for breast cancer therapy.