HIGH-LEVEL EXPRESSION OF SECRETED GLYCOPROTEINS IN TRANSFORMED LEPIDOPTERAN INSECT CELLS USING A NOVEL EXPRESSION VECTOR

An expression cassette for continuous high-level expression of secrete d glycoproteins by transformed lepidopteran insect cells has been deve loped as an alternative to baculovirus and mammalian cell expression s ystems. The expression cassette utilizes the promoter of the silkmoth cytoplasmic actin gene to drive expression from foreign gene sequences , and also contains the ie-1 transactivator gene and the HR3 enhancer region of BmNPV to stimulate gene expression. Using an antibiotic-resi stance selection scheme, we have cloned a Bm5 (silkmoth) cell line ove rexpressing the secreted glycoprotein juvenile hormone esterase (JHE-K K) at levels of 190 mg/L in batch suspension cultures. A baculovirus ( AcNPV) expressing the same gene under the control of the p10 promoter of AcNPV produced only 4 mg/L active JHE in static cultures of infecte d Sf21 cells. A cloned Bm5 cell line overexpressing a soluble isoform of the alpha-subunit of the granulocyte-macrophage colony simulating f actor receptor (solGMR alpha) was also generated and produced five tim es more solGMR alpha in static cultures than a cloned BHK cell line ob tained by transformation with a recombinant expression cassette utiliz ing the human cytomegalovirus (CMV) enhancer-promoter system. Finally, we show that recombinant protein expression levels in transformed Bm5 cells remain high in serum-free media, that expression is stable even in the absence of antibiotic selection, and that lepidopteran cells o ther than Bm5 may be used equally efficiently with this new expression cassette for producing recombinant proteins. (C) 1998 John Wiley & So ns, Inc.