POLYOMA-VIRUS PSEUDOCAPSIDS AS EFFICIENT CARRIERS OF HETEROLOGOUS DNAINTO MAMMALIAN-CELLS

Polyoma virus VP1 pseudocapsids, generated from a recombinant baculovi rus, have been successfully used to transfer exogenous DNA stably into rodent (rat-2) cells. To evaluate the efficiency and biological usefu lness of this route for introducing heterologous DNA into cells, the g ene for a transforming deletion mutant of the middle T antigen of poly oma virus, dl8 MT, was used initially. Whereas the amount of DNA packa ged together with pseudocapsids was found to be variable (2-30%), even at low efficiency its transfer as biologically functional information was high. The dl8 MT gene was stably transferred and integrated in lo w copy numbers into the host chromosome. Transformed cell lines (deriv ed from single foci) were shown to produce high levels of the correspo nding mutant protein, which was active in an in vitro protein kinase a ssay. In comparisons with the calcium phosphate DNA coprecipitation pr ocedure (or lipofectin route), the VP1 pseudocapsid approach was shown to have many advantages in terms of maintenance of DNA fidelity and i ncreased efficiency of gene expression. This system was also assessed for its ability to transfer into and express the chloramphenicol acety l transferase (CAT) gene in a human liver cell line. Here again, the a ssay for functional CAT expression showed the pseudocapsid transfer pr ocedure to compare favorably with lipofectin transfer. In another tran sient assay, a low-level endogenously expressed gene, p43, was complex ed with pseudocapsids and transferred into human embryo lung fibroblas ts, thereby increasing the expression levels. The ease of production o f VP1 pseudocapsids, coupled with their efficient transfer of biologic ally useful information, should make this route of gene delivery an at tractive proposition for further exploration with regard to gene thera py.