Successful Expression of DNA-Based Vaccine Construct Encoding Human Papillomavirus Type ۱۶ E۷ Fused to Heat Shock Protein B۱ in Eukaryotic Cells

Introduction: Developing potent therapeutic vaccines against human papillomaviruses (HPVs) is crucial for the effective management of various HPV-associated cancers. DNA-based vaccines are attractive due to their safety, stability, and capacity to elicit a targeted immune response against specific antigens. Heat shock proteins (HSPs) can enhance the efficacy of DNA vaccines when used as adjuvants. In this study, we created a recombinant DNA molecule by fusing the HPV۱۶ e۷ gene with either the hspB۱ or hsp۲۷ gene and assessed its expression in a eukaryotic cell line. Methods: Initially, we constructed a recombinant eukaryotic expression vector by inserting the hsp۲۷-e۷ fusion gene into the pcDNA۳.۱ (-) vector. The concentration and purity of the sample were evaluated using NanoDrop spectrophotometry. We cultured human embryonic kidney ۲۹۳T (HEK-۲۹۳T) cells in RPMI ۱۶۴۰ medium and transfected them with the pcDNA۳.۱-hsp۲۷-e۷ construct using Lipofectamine ۲۰۰۰ transfection reagent. After ۴۸ hours, we assessed the expression of the Hsp۲۷-E۷ fusion protein by western blotting using an anti-E۷ monoclonal antibody. Results: We successfully subcloned the hsp۲۷-e۷ fusion gene into the pcDNA۳.۱ (-) vector, and enzymatic digestion confirmed a distinct ~۹۷۵ bp band on an agarose gel. The concentration and purity of the recombinant DNA vector in a ۱۰ mL culture were measured to be ۲۱۰ ng/µL and ۱.۸۶, respectively. Furthermore, the expression of the Hsp۲۷-E۷ fusion protein in HEK-۲۹۳T cells was confirmed by Western blot analysis, which detected a distinct band of approximately ۳۸ kDa. Conclusion: Our in vitro findings demonstrate successful expression of the DNA construct encoding the hsp۲۷-e۷ gene, which can be utilized as a DNA vaccine for future in vivo investigations.

Keywords:

Human papillomavirus (HPV) , E۷ , Small heat shock protein , DNA-based vaccine , Recombinant DNA construct , Eukaryotic cells