Preparation of Pluronic F127 hydrogel loaded with BIF1-iRGD recombinant protein for its targeted anti-cancer effects

Pluronic F127-based hydrogel is a fair formulation for increasing the protein stability and half-life without decreasing its biological activity. The present study aimed to prepare FP127 hydrogel loaded with the recombinant BIF1-iRGD protein as a new immunotoxin with targeting potential of cancer cells.

BIF1-iRGD in 19% w/v of FP127 was prepared by the cold method, and its in vitro release was determined. MTT and flow-cytometry assays were performed to evaluate the cytotoxic and apoptotic effects of BIF1-iRGD and BIFl-iRGD-hydrogel against 4T1 cells. The tumor size of 4T1 Balb-C mice was evaluated, and H&E staining was used for histopathology evaluation.

BIFl-iRGD release followed the first-order model. The toxicity of BIFl-iRGD was less than that in the formulation as a hydrogel after 48 and 72 h of treatment. The null hydrogel showed no toxicity compared to the untreated cells. After 24 h, cells treated with BIFl-iRGD-hydrogel and BIFl-iRGD around their 48-h IC₅₀ value developed apoptosis at about 55% and 35%, respectively. The tumor size decreased over time in the treated mice during the 20 days after the last injection. Also, no significant difference was observed between the effectiveness of the two groups. Tumor sections of mice treated with BIF1-iRGD and BIF1-iRGD-hydrogel had necrotic parts of about 65% and 70%, respectively.

BIF-iRGD-hydrogel showed in vivo anticancer effects with increased toxicity in comparison to the native protein. However, the investigation of protein distribution and probable cytotoxic effect on vital organs must be noted.