Insect cell culture for industrial production of recombinant proteins

Abstract

Insect cells used in conjunction with the baculovirus expression vector system (BEVS) are gaining ground rapidly as a platform for recombinant protein production. Insect cells present several comparative advantages to mammalian cells, such as ease of culture, higher tolerance to osmolality and by-product concentration and higher expression levels when infected with a recombinant baculovirus. Here we review some of the recent developments in protein expression by insect cells and their potential application in large-scale culture. Our current knowledge of insect cell metabolism is summarised and emphasis is placed on elements useful in the rational design of serum-free media. The culture of insect cells in the absence of serum is reaching maturity, and promising serum substitutes (hydrolysates, new growth and production-enhancing factors) are being evaluated. Proteolysis is a problem of the BEVS system due to its lytic nature, and can, therefore, be a critical issue in insect cell bioprocessing. Several cell- or baculovirus proteases are involved in degradation events during protein production by insect cells. Methods for proteolysis control, the optimal inhibitors and culture and storage conditions which affect proteolysis are discussed. Finally, engineering issues related to high-density culture (new bioreactor types, gas exchange, feeding strategies) are addressed in view of their relevance to large-scale culture.

Growth of recombinant Drosophila melanogaster Schneider 2 cells producing rabies virus glycoprotein in bioreactor employing serum-free medium

“…On the other hand, the presence of 30 mM of NH 4 Cl had a great impact on recombinant protein production by HighFive TM , despite affecting cell growth only discreetly. Studies also show that alanine is generally produced when glucose and amino acids are in excess in culture medium as well as ammonium is formed when there is glucose limitation (Ikonomou et al 2003). This was probably not the case in the present work, since ammonium was produced in the presence of high glucose concentrations.…”

“…Ammonium production attained values up to 80 mg L -1 in runs 1, 2, 4 and 5, and around 67 mg L -1 in run 3. According to Ikonomou et al (2003), insect cells are not as sensible as mammalian cells to ammonium presence, since the addition of 10 mM of ammonium salts to culture medium did not affect Sf-9 and BTI-EAA cell lines. On the other hand, the presence of 30 mM of NH 4 Cl had a great impact on recombinant protein production by HighFive TM , despite affecting cell growth only discreetly.…”

“…For baculovirus expression system based on Trichoplusia ni cells, the literature reports that high ammonium concentrations reduces or even inhibits protein production (Yang et al 1996;Ikonomou et al 2003). Since the RVGP undergoes structural changes in acidic pH, loosing its native shape (Gaudin et al 1993;Gaudin 1997;Maillard and Gaudin 2002), pH and lactate concentration were monitored.…”

Evaluation of concentrated milk whey as a supplement for SF9 Spodoptera frugiperda cells in culture

“…In this system, the protein posttranslational modifications are similar to those found in mammalian cells (Verma et al 1998). In addition, insect cells present several advantages when compared to mammalian cells, such as ease of culture, higher tolerance to osmolality (Olejnik et al 2003) and much higher expression levels when infected with recombinant baculovirus (Ikonomou et al 2003). The use of baculovirus as an alternative pesticide in agriculture has increased steadily in the last years.…”

Designing Two Individual AcMNPV Polyhedrin-Plus Bac-to-Bac Expression System in order to Express GFP and CPV-VP2 in Insect Cells

“…One of the best systems, considered in production of recombinant proteins, is the use of baculoviruses in insect cell expression system. The system allows proper post-translational modifi cations; producing biosimilars of protein of interest (19)(20)(21)(22). Here, Bacto-Bac baculovirus expression vector system (BEVS) was used for heterologous expression in in insect cells.…”