The cost of pharmaceuticals is exorbitantly high and continues to rise. Some pharmaceuticals, in addition to their high cost, are also limited in supply, making it impossible for them to be available to every patient that needs them. This is particularly problematic in developing countries, where both cost and availability hinder the distribution of pharmaceuticals to needy populations.
Several factors contribute to the high costs of producing pharmaceuticals, and result in the high price of the pharmaceuticals for the consumer. A major contributing factor is the lack of economical means of producing the product. This is particularly true for protein and peptide-based medications. Another contributing factor for some medications is the inability to administer therapeutically effective amounts of the pharmaceutical agent orally. Many pharmaceuticals can only be administrated by injection into a particular site in the body. For example, many immunization medications for the treatment of allergies or infectious diseases require administration by injection. Protein and peptide pharmaceuticals, such as human growth hormone and insulin, can often only be administered by injection. Another factor that contributes to the cost of many pharmaceutical medications is their delivery to the hospital or distribution site. Particularly in hot climates, delivery and storage of pharmaceutical medications requires expensive refrigeration equipment. This is a major challenge in developing countries, where such equipment is often unavailable.
Pharmaceutical proteins and peptides have been produced in a wide variety of hosts. Many therapeutic proteins have been produced in heterologous expression systems including prokaryotes such as Escherichia coli and Bacillus subtilis, and eukaryotes such as yeast, fungi, insect cells, animal cells, and transgenic animals. Bacterial expression systems are relatively easy to manipulate and the yield of the product is high. However, mammalian proteins often require extensive posttranslational modification for functional activity, which can be a limiting factor in bacterial expression systems. Cell culture systems such as mammalian, human, and insect cell culture systems are more convenient for the production of complex proteins. However, long lead times, low recovery of the product, possible pathogen transfer, and high capital and production costs present serious concerns. Transgenic animals may provide an unlimited supply of complex proteins. Unfortunately, this system is limited by the long period of time it takes to generate new and improved products and the risk of pathogen transfer to human subjects.
The economic and biochemical limitations to producing pharmaceutical proteins and peptides in prokaryotic and eukaryotic cells, including high production costs, low yields, secretion problems, inappropriate modifications in protein processing, difficulties scaling up to larger volumes, and contamination have led researchers to examine plants as new hosts for the large-scale production of proteins and peptides with the expectation of reduced cost. Production of proteins in transgenic plants is described, for example, in U.S. Pat. Nos. 5,750,871; 5,565,347; 5,464,763; 5,750,871; and 5,565,347. Although plants are less expensive to grow and harvest in bulk than prokaryotic and eukaryotic cells, expression of the foreign gene in plant cells is typically low. In addition, harvesting the plant typically requires breaking the plants, for example, by removing the leaves, separating the stems from the roots, or removing the roots. Such breakage usually results, a process that initiates wilting of the plant part and apoptosis of the plant. A plant undergoing apoptosis generates proteases that contribute to the degradation of the transgenically expressed protein before purification of the protein is even begun. Even if the plant is to be directly consumed, the activity of the expressed pharmaceutical protein may be reduced by harvest-induced intercellular degradation machinery.
Another major concern associated with producing foreign proteins in transgenic plants that are grown in open fields is the possibility of cross-pollination with plants in the wild, making it possible for the foreign protein to enter the food chain. The complexity of governmental regulations surrounding agricultural practices for transgenic plants makes it difficult to get new transgenic plants approved for agricultural use. Furthermore, the outdoor environment is impossible to control, making proper growth, development, and regulation of foreign gene expression difficult to guarantee. For example, the induction of a heat inducible, light inducible, hormone inducible, or chemically inducible promoter would be practically impossible in an outdoor environment. Of course, the outdoor temperature and light levels cannot be controlled. Additionally, hormones or chemicals sprayed on a plant are likely to be dispersed not onto the plant, but into the environment by wind and rain. Spaying crop fields is also quite costly.
Rodriguez et al. (see U.S. Pat. Nos. 5,888,789; 5,889,189; and 5,994,628) disclose production of proteins, including pharmaceutical proteins, in grains such as barley or rice. Malting is a process by which grain is germinated under controlled conditions and in contained facilities to produce a product, e.g., a foreign protein product. The harvested product is the malted grain, which is typically kiln-dried at between 120° F. and 130° F. The developmentally regulated amylase promoter typically drives expression of foreign proteins in this system. Although the foreign proteins can be expressed in high quantities in this system, harvesting and administration of the protein usually requires processing of the malted grain, which can alter the quality of the expressed foreign protein.
There exists the need for a controlled regulatable system for producing pharmaceutical proteins in plants that decreases the amount of intercellular degradation of the expressed protein upon harvest.