Binding and biologic characterization of recombinant human serum albumin-eTGFBR2 fusion protein expressed in CHO cells.

Transforming growth factor-1 (TGF-1) signaling is involved in cell metabolism, growth, differentiation, carcinoma invasion and fibrosis development, which suggests TGF-1 can be treated as a therapeutic target extensively. Because TGF-1 receptor type (TGFBR2) is the directed and essential mediator for TGF-1 signals, the extracellular domain of TGFBR2 (eTGFBR2), binding partner for TGF-1, has been produced in a series of expression systems to inhibit TGF-1 signaling. However, eTGFBR2 is unstable with a short half-life predominantly because of enzymatic degradation and kidney clearance. In this study, a fusion protein consisting of human eTGFBR2 fused at the C-terminal of human serum albumin (HSA) was stably and highly expressed in Chinese Hamster Ovary (CHO) cells. The high and stable expression sub-clones with Ig kappa signal peptide were selected by Western blot analysis and used for suspension culture. After fed-batch culture over 8d, the expression level of HSA-eTGFBR2 reached 180mg/L. The fusion protein was then purified from culture medium using a 2-step chromatographic procedure that resulted in 39% recovery rate. The TGF-1 binding assay revealed that HSA-eTGFBR2 could bind to TGF-1 with the affinity constant (K-D of 1.42 x 10(-8) M) as determined by the ForteBio Octet System. In addition, our data suggested that HSA-eTGFBR2 exhibited a TGF-1 neutralizing activity and maintained a long-term activity more than eTGFBR2. It concluded that the overexpressing CHO cell line supplied sufficient recombinant human HSA-eTGFBR2 for further research and other applications.