Hydrogel nanocarrier encapsulated recombinant IκBα as a novel anticancer protein therapeutics

Abstract

The NFkB overexpression triggers drug resistance in many types of cancer by up-regulating anti-apoptotic genes. IkBa, an endogenous protein and natural inhibitor of NFkB, blocks translocation of NFkB from cytoplasm to nucleus and thereby, sensitizes cancer cells to external stimuli. Herein, we have cloned a PCR amplified cDNA of IkBa, and expressed the recombinant GST tagged IkBa in Escherichia coli BL21 (DE3) cells. The recombinant GST-IkBa was purified by glutathione-agarose affinity chromatography and characterized by Western blot, MALDI-TOF, UV-Vis spectroscopy and circular dichroism spectroscopy. The GST-IkBa 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-IkBa 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-IkBa 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-IkBa as a novel recombinant protein therapeutic approach for cancer, which may further be regimented in combination therapy.

Synergistic Anticancer Activity of Fluorescent Copper Nanoclusters and Cisplatin Delivered through a Hydrogel Nanocarrier

“…Also, the polymer NC composite could easily be synthesized in the form of hydrogel nanocarrier, by using PVA as the cross linker, which is favorable for cellular uptake. 27 The hydrogel could be then turned into a powder, which showed stable fluorescence, owing to the NCs, for more than a month. Further, the emission due to the NCs was useful for imaging mammalian cells by optical microscopy and more importantly to probe the cells by commercial flow cytometer, without having to use any other dye.…”

“…The fluorescence of the composite was found to be sensitive to the pH of the medium, and the emission could be tuned reversibly according to the pH. Also, the polymer NC composite could easily be synthesized in the form of hydrogel nanocarrier, by using poly­(vinyl alcohol), PVA, as the cross-linker, which is favorable for cellular uptake . The hydrogel could be then turned into a powder, which showed stable fluorescence, owing to the NCs, for more than a month.…”

Chemosensitization of IκBα-overexpressing glioblastoma towards anti-cancer agents

“…IkBa was cloned in the pGEM-T Easy vector (Promega) as described previously. 40 The IkBa gene was transferred from pGEM-T Easy to the pCINeo vector (Promega) by digestion with restriction enzymes followed by ligation. IkBa cloned in the pGEM-T Easy vector was digested with Eco RI (cutting site 5 0 GAATTC 3 0 ) and Spe I (cutting site 5 0 ACTAGT 3 0 ).…”

Targeting Wnt Canonical Signaling by Recombinant sFRP1 Bound Luminescent Au-Nanocluster Embedded Nanoparticles in Cancer Theranostics

“…Studies have elaborated on the phenomenon of chemosensitization of cancer cells toward a conventional chemotherapeutic agent by overexpression of a relevant gene or addition of a therapeutic protein. , This fact has been exploited in this work, whereby the binding of protein to Chi-Au NC-Alg was found to considerably sensitize HeLa cells toward cisplatin. The MTT assay exhibited a sharp reduction in cell viability in the case of combination therapy (cisplatin with protein-NPs), as compared to that of only protein-NPs or only cisplatin treated cells (Figure A).…”

“…Recent material innovations have propagated a paradigm shift toward the unique concept of the codelivery of drug and therapeutic molecules conjugated to nanocarriers . Studies have revealed that these therapeutic molecules by themselves may not be toxic enough to eradicate cancer completely but that they may be used in combination with chemotherapy to help reduce the dosage of drugs and diminish their side effects. , Ligand-targeted NPs loaded with chemotherapeutic drugs are increasingly finding application in the remedy of cancer, with the common targets being receptors overexpressed by cancer cells, such as folate and transferrin receptors. − However, even though aberrant upregulation of the Wnt pathway is one of the highlights in cancer, to date there has been no report documenting the targeting of the Wnt pathway with NPs for cancer therapy.…”