Title:
On designing stable magnetic vectors as carriers for malaria DNA vaccine.

Abstract:
Superparamagnetic iron oxide nanoparticles (SPIONs) can be used as therapeutic and diagnostic agents due to their unique magnetic characteristics, provided that they are stable in physiological conditions. Here, the assembly of different magnetic vector configurations comprising SPIONs, polyethylenimine (PEI), and hyaluronic acid (HA), acting as carriers for malaria DNA vaccine encoding Plasmodium yoelii merozoite surface protein MSP1-19 (VR1020-PyMSP1-19), and their stability in different cell media were investigated. The order of assembly affected vector size, surface charge, stability, and ability to bind and release DNA. Generally, all vectors showed relatively small size of less than 200 nm in water, whereas higher degree of aggregation was observed immediately after transferring to high-ionic strength media such as 150 mM NaCl buffer and RPMI 1640 culture media (Roswell Park Memorial Institute medium). However, the pre-addition of HA to DNA effectively reduced the extent of aggregation in serum-free RPMI 1640 with sizes of almost all complexes remaining below 90 nm, particularly at HA:PEI charge ratio of 100%. The presence of fetal bovine serum (FBS) in RPMI 1640 culture media further converted the surface charge of vectors from positive to negative, decreasing the size to smaller than 50 nm. Partial disassembly of some vectors was observed in water, in RPMI, and in RPMI supplemented with 10% FBS after incubation for 1h, but not in NaCl buffer, indicating that incubation of complexes in NaCl buffer prior to transfection may limit the intracellular release of plasmid DNA. DNase sensitivity assay showed that plasmid DNA vaccine encoding the PyMSP1-19 in all configurations preserved their structural integrity without damage, even after DNase I treatment for 30 min. This study demonstrated that structurally well-defined magnetic gene carriers could be designed to improve malaria DNA vaccine delivery systems, particularly for in vivo applications.