Growth factors play a major role in medical research as components of cell culture media used for proliferation and differentiation of various cells. Their use is expected to increase dramatically as progress is made in stem cell research, tissue engineering and regenerative medicine. Furthermore, their application as biopharmaceuticals for human therapeutic use is becoming increasingly important.
Recombinant growth factors are commercially produced in different host expression systems such as bacteria, yeast, insect and mammalian cells. While these systems can be efficient, there is still a need for new production systems that are serum- and animal-free, more economical, and void of endotoxins and other contaminating components, such as pyrogenic and pro-inflammatory agents that may affect their use in biological systems and assays. A number of different plant systems are in development for the efficient production of protein biopharmaceuticals such as growth factors (for review see [1
]). Barley seeds and other cereals are plant systems that have been explored with good results for molecular farming [2
]. Using barley seeds for heterologous protein production is an attractive option because they are a cost-saving alternative that are easy to grow and with well-established post-harvest handling and processing methods [4
]. The majority of human proteins that have been produced in plants show significant structural, biochemical and functional similarity to proteins from animal cell systems [6
]. Furthermore, barley as a host system features genetic self-containment, as barley is a self-pollinating crop plant. Barley is G
afe (G.R.A.S.) by the FDA with low secondary metabolite content, such as phenolics (unpublished data). Presently, most plant produced biopharmaceuticals and other recombinant proteins have been expressed in tobacco which is known for the presence of alkaloids and other compounds potentially toxic to humans [7
]. Accumulating heterologous proteins in seeds presents many advantages as seeds are natural long-term storage vehicles of proteins within an inert environment. They can be stored for years without affecting the quality of the protein [4
] and they have a relatively simple protein profile with the major part belonging to few and dominant polypeptide classes that can be separated with relative ease from the target protein, thus facilitating downstream procedures considerably [9
Flt3 ligand is an essential hematopoietic growth factor involved in proliferation and differentiation of stem cells and in the development of NK cells, B cells, T cells, monocytes/macrophages and dendritic cells. Flt3 ligand may be of clinical value as it is, together with other growth factors and cytokines, considered a stimulant of the immune system through its effect on both dendritic and natural killer cells production (reviewed by [10
]). Recombinant human Flt3 ligand is commercially available from bacterial source and CHO cells. In the present paper, we describe the production of recombinant human Flt3 ligand in barley grain using the Orfeus expression system developed by ORF Genetics for production of recombinant proteins termed ISOkines.
To our knowledge, this is the first report describing expression and purification of a recombinant human growth factor from cereal seeds with biological activity comparable to commercially available counterparts. This is also the first publication on expression of Flt3 ligand in a plant system. The results demonstrate that barley is a proficient system for the production of active recombinant growth factors for cell culture, media preparation and medical application.