851. Development of Brain Tumor Organ Cultures for Characterizing Polymer/Plasmid DNA Mediated Gene Transfer

Abstract

Development of effective gene therapy requires an expeditious and reproducible test system. The syngenic RG2 rat brain tumor model satisfies the criteria for a clinically relevant animal model. Pluronic block copolymer L44 was formulated with a green fluorescent protein (GFP) expression plasmid, and sterotacticly injected into rat brain tumors. GFP positive cells were observed in the tumor. Yet, the numbers of transfected cells were insufficient for a therapeutic effect. Hence, the formulation requires other components to increase gene transfer efficiency. The turn around time for obtaining results from this animal model was not amendable to screening formulations for tumor gene transfer. Cell culture using rat glioma cell lines has proven to be advantageous in screening gene transfer vectors. Conversely, due to the two-dimensional single cell layer and volume of growth media necessary for tissue culture, it is not applicable for evaluating local administration of plasmid DNA. For this reason, an organ culture was developed to optimize screening. Rat brains tumors were sliced into 300-micron sections and cultured for 7days. Tumors were observed to grow over the 7-day period ultimately taking over the brain slice. Local injection of Poloxamer formulated GFP expression plasmid was shown to transfect tumor cells in the organ culture model. Presently, the brain organ cultures are being characterized for expression of normal and brain tumor cell markers. Comparison of staining patterns for biomarkers in the organ culture and in vivo brain tumors are being used to validate the organ culture. Concomitantly, formulation conditions are being tested to optimize gene transfer to the brain tumor.