Doxorubicin loaded on chitosan-protamine nanoparticles triggers apoptosis via downregulating Bcl-2 in breast cancer cells

Abstract Cancer-specific drug delivery is a reliable approach to evade undesirable side effects and increase the bioavailability of the drug in tumor cells. In the present study, we treated breast cancer cells MDA-MB-231 with doxorubicin (DOX) loaded on chitosan-protamine nanoparticles (CPNPs) to investigate the composite ability to induce apoptosis. CPNPs were prepared and characterized using FTIR spectroscopy, transmission electron microscope, and zeta potential determination. CPNPs showed a drug encapsulation efficiency (EE) of 21%, drug loading capacity (LC) of 3.65% and a particle size of 117 nm. In vitro release study indicated that DOX release from CPNPs-DOX was pH-dependent, where it released with rates 60.10%, 44.15% and 25.10% at pH 4.0, 6.8 and 7.4, respectively. Cells were treated with three concentrations (1, 2, and 3 μM) of either free DOX, doxorubicin loaded on CPNPs (CPNPs-DOX), or empty carrier for 48 h. Cell viability was assessed using MTT and trypan blue assays. Meanwhile, apoptosis rate using PI/Annexin V-FITC staining cell cycle analysis were performed using PI staining-based flow cytometry. MTT and trypan blue assays showed a significant decrease in the viability/count upon treating cells with DOX-CPNPs. Flow cytometry data revealed an arrest of the breast cancer cell at G2/M (47.18%) in the CPNPs-DOX treatment. Quantitative real time PCR analysis showed that CPNPs-DOX treatment has significantly downregulated Bcl-2 compared with free DOX treatment and control. These results indicate the efficiency of using CPNPs as a drug carrier for DOX in treating breast cancer cells, however, these conclusion needs further investigation.