Hydrogel nanocarrier encapsulated recombinant IκBα as a novel anticancer protein therapeutics

Abstract

The NFκB overexpression triggers drug resistance in many types of cancer by up-regulating anti-apoptotic genes. IκBα, an endogenous protein and natural inhibitor of NFκB, blocks translocation of NFκB from cytoplasm to nucleus and thereby, sensitizes cancer cells to external stimuli. Herein, we have cloned a PCR amplified cDNA of IκBα, and expressed the recombinant GST tagged IκBα in Escherichia coli BL21 (DE3) cells. The recombinant GST-IκBα was purified by glutathione-agarose affinity chromatography and characterized by Western blot, MALDI-TOF, UV-Vis spectroscopy and circular dichroism spectroscopy. The GST-IκBα recombinant protein was encapsulated within polyvinyl alcohol (PVA)/polyvinyl pyrrolidone (PVP) hydrogel nanocarrier (NC) and then characterized by transmission electron microscopy (TEM), field emission scanning electron microscopy (FESEM), Fourier transform infrared (FTIR) spectroscopy, dynamic light scattering (DLS) and zeta potential measurements. The pH-dependent protein release was performed in vitro to study the pH tunability of the hydrogel NCs. Furthermore, the therapeutic efficacy of GST-IκBα loaded hydrogel NCs was evaluated on HeLa (cervical carcinoma) cells by cytotoxicity assay and cell cycle analysis. TUNEL assay by flow cytometry confirmed apoptosis of HeLa cells. Administration of GST-IκBα by hydrogel NCs showed significant cell growth inhibition of drug resistant U87MG cells in combination with 5-FU. Our results essentially attributed the hydrogel NC-mediated administration of recombinant GST-IκBα as a novel recombinant protein therapeutic approach for cancer, which may further be regimented in combination therapy.