Tuning CpG motif position in nanostructured DNA for efficient immune stimulation

Area of Science:

• Immunology
• Nanotechnology
• Molecular Biology

Background:

• Polypod-like nanostructured DNA (polypodna) are effective for delivering cytosine-phosphate-guanine (CpG) oligodeoxynucleotides (ODNs) to immune cells.
• The immunostimulatory activity of single-stranded CpG ODNs depends on motif sequence and position, but this is less understood for nanostructured DNA.

Purpose of the Study:

• To investigate how the position of a CpG motif within polypodna affects its immunostimulatory activity.
• To compare the immune response generated by polypodna with CpG motifs located in different regions.

Main Methods:

• Designed four series of polypodna with a single potent CpG motif at varying positions and 2-5 CpG-free ODNs.
• Assessed immunostimulatory activity by measuring tumor necrosis factor-α release in Toll-like receptor-9 (TLR9)-positive murine macrophage-like RAW264.7 cells.
• Compared cellular uptake of different polypodna structures.

Main Results:

• Polypodna with the CpG motif in the 5 eal-overhang induced significantly higher tumor necrosis factor-α release compared to those with the motif in the double-stranded region.
• Cellular uptake was similar across different polypodna designs, indicating position-dependent activity.
• The relative immunostimulatory potency of CpG ODNs changed upon incorporation into polypodna structures.

Conclusions:

• CpG motif location within nanostructured DNA is a critical factor for immune stimulation, alongside sequence.
• Designing effective CpG motif-containing nanostructured DNA requires careful consideration of both motif placement and ODN sequence.
• These findings provide insights for developing novel immunomodulatory nanomedicines.