Disposition and gene expression characteristics in solid tumors and skeletal muscle after direct injection of naked plasmid DNA in mice.

Previous studies have suggested that direct injection of naked plasmid DNA (pDNA) into solid tumors can be a useful method for in vivo gene transfer into tumor cells. To gain more insight into this approach, we studied the disposition and gene expression characteristics of naked pDNA after intratumoral injection by direct comparison with those after intramuscular injection in mice. pDNA encoding reporter genes were directly injected into subcutaneous solid tumor models and skeletal muscles. Biodistribution studies using radiolabeled pDNA showed that the elimination of pDNA from the injection site was relatively fast and a part of the pDNA was absorbed from the lymphatic system after both local injections. Confocal microscopic studies using fluorescein-labeled pDNA demonstrated that pDNA distributed efficiently throughout the muscle tissue whereas pDNA localization in the tumor tissue was restricted. Characterization of gene expression clarified the variation in expression level between tumor preparations and some factors affecting the expression level in the tumor. Reporter gene expression was significantly inhibited by simultaneous administration of some polyanions in both cases, suggesting that a specific mechanism may be involved in the naked pDNA uptake by muscle and tumor cells. These findings provide useful information for direct naked pDNA delivery into solid tumors.

Morphological study on Mycobacterium bovis BCG Tokyo 172 cell wall skeleton (SMP-105).

Mycobacterial cell wall consists of rigid cell wall skeleton (CWS), a mycoloyl arabinogalactan peptidiglycan complex, in which mycoloyl structure varies by the mycobacterial species diversely, whereas the arabinogalactan peptidoglycan structure is consistent comparatively. The CWS of Mycobacterium bovis BCG has long been expected as a potent adjuvant for immunotherapy of malignant tumor. Although the chemical structure of CWS has been established in the last few decades, the physicochemical properties of CWS having highly amphipathic micelle structure with very long mycoloyl and carbohydrate chains are not unveiled. In this study, the ultrastructure of CWS of M. bovis BCG Tokyo 172 (SMP-105), suspended in several solvents with different polarity, was investigated with a particle size analyzer, a transmission electron microscope (TEM) and other techniques. As a result, the particle size was about 4.7 to 67.8 microm in physiological saline, but it became smaller and more compact when suspended in hydrophobic solvents. TEM images showed two different morphological forms distinctively: double folded sheet structure in hydrophilic conditions and multilayered rolled sheet structure in hydrophobic conditions. These studies have revealed characteristic surface features of SMP-105, the hydrophobic moiety occupying dominant space and the hydrophilic moiety smaller space, respectively, which may lead to the acceleration of immunological studies on this product.