CHARACTERIZATION OF A MONOCLONAL ANTIBODY-PURIFIED RECOMBINANT FACTOR-IX PRODUCED IN HUMAN HEPATOMA (HEPG2) CELL-CULTURES AFTER RETROVIRAL VECTOR-MEDIATED TRANSFER

Hemophilia B is a leading target for gene therapy, and hepatocytes are natural cells for transduction with specific factor IX vectors. Howev er, transduced hepatocytes do not produce sufficient factor IX for det ailed characterization. Genetic mutations or changes in protein struct ure can occur during vector construction, packaging or transduction th at may adversely affect function or metabolism. In this study, we eval uated the molecular structure, immunologic and biologic properties, an d the metabolic fate of a monoclonal antibody (MoAb)-purified recombin ant factor TX produced by HepG2 cells transduced with a CMV promoter-c ontrolled factor IX vector. We confirmed that monoclonal antibody (MoA b)-purified recombinant human (r) factor IX shared antigenic sites wit h MoAb-purified native human (n) factor IX, as determined by Western b lotting and inhibition assays. Protein chemical studies revealed a hig h degree of similarity between r and n factor IX. When infused into ra t,the biologic half-life of r factor IX was at least as long, if not l onger than that of n factor IX. Finally, enhanced r factor IX producti on was noted in EGF- and factor XII-treated cultures. These data indic ate that r factor IX produced by transduced HepG2 cells had no structu ral changes that adversely affected its function, immunoreactivity, an d metabolic fate.