The tumor microenvironment enhances the expression of cssDNA by modulating cell cycle signaling pathways via SKP2

Area of Science:

• Molecular Biology
• Gene Therapy
• Oncology

Background:

• Circular single-stranded DNA (cssDNA) is an emerging nucleic acid vector with potential in genetic disorder treatment, gene editing, and oncotherapy.
• cssDNA possesses unique attributes like high stability, structural simplicity, conformational flexibility, and low molecular mass, making it a promising gene therapy tool.

Purpose of the Study:

• To evaluate the expression efficiency of cssDNA compared to conventional plasmids in tumor cell lines.
• To investigate the impact of tumor microenvironment (TME) conditions on cssDNA expression.
• To elucidate the mechanisms underlying TME-modulated cssDNA expression, including cell cycle regulation and the role of SKP2.

Main Methods:

• Comparative analysis of cssDNA and plasmid expression in various tumor cell lines.
• Assessment of cssDNA expression under simulated TME conditions (glucose/glutamine deficiency, hypoxia).
• Cell cycle analysis and identification of key regulatory proteins (SKP2) involved in cssDNA expression modulation.

Main Results:

• cssDNA demonstrated superior gene expression efficiency compared to conventional plasmids across diverse tumor cell lines.
• Specific TME conditions (glucose deficiency, glutamine deficiency, hypoxia) enhanced cssDNA expression.
• TME conditions induced G1 cell cycle arrest, and cssDNA expression peaked during the late G2/M phase.
• S-phase kinase-associated protein 2 (SKP2) was identified as a critical regulator, with SKP2 inhibition enhancing cssDNA expression.

Conclusions:

• cssDNA is a highly efficient gene expression vector with advantages over traditional plasmids.
• Tumor microenvironment conditions modulate cssDNA expression through cell cycle perturbations and SKP2-dependent pathways.
• These findings support the development of cssDNA-based cancer therapies and future clinical translation.