Optimal production and in vitro activity of recombinant endostatin from stably transformed Drosophila melanogaster S2 cells

Recombinant plasmids containing a complementary deoxyribonucleic acid codin g mouse endostatin were transfected and stably expressed in Drosophila mela nogaster Schneider 2 (S2) cells. Stably transformed polyclonal cell populat ions expressing recombinant endostatin were isolated after 4 wk of selectio n with hygromycin B. Recombinant endostatin expressed in the stably transfo rmed S2 cells under the influence of the Drosophila BiP protein signal sequ ence was secreted into the medium. Recombinant endostatin was also purified to homogeneity using a simple one-step Ni2+ affinity fractionation method. Purified recombinant endostatin inhibited endothelial cell proliferation i n a dose-dependent manner. The concentration at maximum inhibition for reco mbinant endostatin was approximately 1.8 mug/ml. The stably transformed S2 cells produced 18 mg recombinant endostatin/L 7 d after induction with 5 mu M CdCl2. Sodium butyrate supplementation (2.5 mM) increased recombinant end ostatin production by 17%. These findings demonstrate optimal production an d in vitro activity of recombinant endostatin from stably transformed D. me lanogaster S2 cells.