Synergistic antitumor effects of recombinant Interleukin-21 and 5-fluorouracil: A novel therapeutic approach against hepatocellular carcinoma.

Interleukin-21 (IL-21) is a pleiotropic cytokine that enhances both humoral and cellular immunity by increasing B-cell proliferation, T-cell effector function, and natural killer (NK) cell cytotoxicity. Studies have demonstrated its potential as an anti-cancer agent in the treatment of melanoma, neuroblastoma, fibrosarcoma, hepatocellular carcinoma (HCC) and non-small-cell lung carcinoma. The aim of this study was to evaluate the cytotoxic effect of recombinant human IL-21 (rhIL-21) protein, both alone and in combination with 5-fluorouracil, a potent anticancer agent, on the HepG2 cell line. To achieve this, the protein was synthesized in the periplasmic space of Escherichia coli and subsequently purified. The MTT viability assay results demonstrated that treating HepG2 cells with rhIL-21 alone resulted in 54.81 % and 59.75 % cytotoxicity after 24 and 48 h respectively, with corresponding IC50 values of 108 ng/mL and 68.90 ng/mL. Furthermore, the level of cytotoxicity remained constant at concentrations above 150 ng/mL of rhIL-21. Treatment with 25 µg/mL of 5-fluorouracil alone resulted in 21.09 % and 27.71 % cytotoxicity after 24 and 48 h, respectively. However, combining 5-fluorouracil with 150 ng/mL of rhIL-21 significantly increased toxicity, reaching 81.15 % and 89.34 % after 24 and 48 h, respectively - an improvement of approximately 60 %. This underscores the synergistic potential of rhIL-21 in combination therapy, with IC50 values of 25 µg/mL of 5-fluorouracil and 30.96 or 15.79 ng/mL of rhIL-21 after 24 or 48 h, respectively. This regimen demonstrated strong cytotoxic effects against HepG2 cells while showing low toxicity towards normal L929 fibroblasts, suggesting its potential safety and therapeutic applicability.