Efficient Constitutive Production of Human Fibroblast Interferon by Hamster Cells Transformed with the IFN-β 1 Gene Fused to An SV40 Early Promoter

Abstract

The coding sequence of the human interferon (IFN)-β₁ gene, fused 60 bp downstream from the RNA start site of the SV40 early gene, was transfected into dihydrofolate reductase (DHFR)-deficient Chinese hamster ovary (CHO) cells together with a selectable DHFR gene. Most transformants continuously secreted IFN-β₁ into the medium. Induction did not stimulate expression of the fused SV40-IFN-β₁ gene. The role of the SV40 promoter was verified by transforming cells with the unmodified human IFN-β₁ gene, or by the IFN-β₁ coding region fused to another poly(rI):(rC)-inducible gene. In these cases, the transformants showed strictly inducible (not constitutive) IFN secretion. By selection for methotrexate resistance, CHO clones with a 10–20-fold amplification of the SV40-IFN-β₁ DNA were obtained. Such clones constitutively produce up to 350,000 units IFN/ml per 10⁶ cells/24 hr, i.e., over 10 times more than fully induced human fibroblasts. In continuous culture with daily changes of medium, accumulation of IFN-β₁ is constant at a rate of 300,000 molecules per cell/hr. Batches of up to 16 mg of IFN-β₁ produced by the transformed CHO cells were purified to homogeneity by affinity chromatography on monoclonal antibodies. This IFN appears identical in size, activity, and immunospecificity to the native human IFN-β₁ glycoprotein.