The Performance of Minicircle DNA Versus Parental Plasmid in p53 Gene Delivery Into HPV-18-Infected Cervical Cancer Cells

Abstract

Minicircle DNA (mcDNA) has been suggested as a vanguard technology for gene therapy, consisting of a nonviral DNA vector devoid of prokaryotic sequences. Unlike conventional plasmid DNA (pDNA), this small vector is able to sustain high expression rates throughout time. Thus, this work describes the construction, production, and purification of mcDNA-p53 and its precursor parental plasmid (PP)-p53 for a comparative study of both DNA vectors in the growth suppression of human papillomavirus (HPV)-18-infected cervical cancer cells. First, live cell imaging and fluorescence microscopy studies allowed to understand that mcDNA-p53 vector was able to enter cell nuclei more rapidly than PP-p53 vector, leading to a transfection efficiency of 68% against 34%, respectively. Then, p53 transcripts and protein expression assessment revealed that both vectors were able to induce transcription and the target protein expression. However, the mcDNA-p53 vector performance stood out, by demonstrating higher p53 expression levels (91.65 ± 2.82 U/mL vs. 74.75 ± 4.44 U/mL). After assuring the safety of both vectors by viability studies, such potential was confirmed by proliferation and apoptosis assays. These studies confirmed the mcDNA-p53 vector function toward cell cycle arrest and apoptosis in HPV-18-infected cervical cancer cells. Altogether, these results suggest that the mcDNA vector has a more promising and efficient role as a DNA vector than conventional pDNA, opening new investigation lines for cervical cancer treatment in the future.