Production of Functional Hepatocyte Growth Factor (HGF) in Insect Cells Infected with an HGF-Recombinant Baculovirus in a Serum-Free Medium

Abstract

Three insect cell lines, SL-7B cells derived from Spodoptera litura, Sf9, and High Five (Hi-5) cells, were used for the production of pro-hepatocyte growth factor (pro-HGF). Cells were cultured and then infected with a recombinant HGF-containing baculovirus in a serum-free medium. In SL-7B cells, pro-HGF is synthesized and excreted from the cells and late in infection is converted to a heterodimeric form of HGF even when the cells are grown in serum free medium. Conversion of a single-chain form of HGF (pro-HGF) into an HGF heterodimer was unexpected, as pro-HGF is normally cleaved by a serum protease called HGF activator. The proliferation activity of heparin-affinity-purified HGF from serum-free culture supernatant of SL-7B cells is comparable to that obtained from HGF converted by serum proteases, suggesting that SL-7B cells produce a functionally analogous protease to correctly process pro-HGF. This work reports, for the first time, on the feasibility of properly processing pro-HGF to form functional HGF by proteases from invertebrate cells in serum-free media. Avoiding the supplementation of sera provides the advantages of a low production cost, zero contamination of infectious agents from sera, and simple downstream product purification. Experimental results further demonstrate that the conversion of pro-HGF by insect cells is cell-line-dependent, because proteases in Hi-5 or Sf9 cells could not process pro-HGF as efficiently and properly as those in SL-7B cells.

In vitro and in vivo bioactivity of recombinant canine hepatocyte growth factor

“…However, the rcHGF product obtained should be further purified before the studies start. The generation of rcHGF was higher than the previously reported production of rhHGF using insect (Wang et al, 2000;Yee et al, 1993) or mammalian cells (Nakamura et al, 1989). The use of other production systems, for instance the yeast Pichia pastoris (Liu et al, 2005) may yield even higher quantities of >500 mg/L recombinant HGF.…”

Secretory expression and characterization of a recombinant-deleted variant of human hepatocyte growth factor inPichia pastoris

“…Since the concentration of native HGF in plasma is very low, purification of HGF from plasma is very difficult. It was reported that HGF is expressed in foreign gene expression systems such as mammalian cells, CHO cells [22,23] , insect cells [24] , and gene therapy [25] . Dang et al [26] and Li et al [27] .…”

PET of c-Met in Cancer with 64Cu-Labeled Hepatocyte Growth Factor

“…There have been several earlier attempts to express biologically active recombinant human HGF (rh-HGF) in both Escherichia coli and insect cells (13, 14). Although biological activity of rh-HGF produced from E.coli has been reported to be equivalent to its native form (15), the absence of the disulfide bond formation and lack of molecular glycosylation can potentially compromise its applicability in vivo.…”