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Biotechnology and Society---Part
XVIII
Biopharmaceuticals
If you are planning for one
year, grow rice; for 20 years grow trees; for centuries, grow men - Chinese
Proverb
 Introduction:
Biopharmaceuticals have been produced for over 20 years using a variety of
recombinant techniques including bacterial, yeast/fungi and mammalian cell
culture systems. With the unravelling of the human genome and the concomitant
explosion of proteomics there will be an ever-increasing need for production of
several therapeutic proteins for various indications. The current production
methods and facilities will not be adequate to meet such needs. It is thus
prudent to consider alternative transgenic technologies, which are efficient as
well as economical.
Transgenic animals, chicken
(eggs) and plants are three such vehicles ripe for consideration. Production of
human proteins in transgenic plants has economic and qualitative benefits,
including reduced health risks from pathogen contamination such as prions and
mammalian viruses. The plant viruses are not known to cross into mammalian/human
species. The yields can be high and if expressed in seeds, lead to processing
convenience. The cultivation, harvesting and storage would follow established
good agricultural practices while the processing and purification would follow
the same methods that are currently used in mammalian cell culture systems.
The
upstream portion of the production of such proteins involves relatively little
capital investment as compared to a bioreactor. There is thus a considerable
saving in upfront costs of production. It is estimated that in a mammalian cell
culture system, the upstream and downstream costs are about the same. Thus,
using transgenic plants could be a considerable cost advantage.
Although there are differences
in codon usage and post-translational processing between plants and mammals,
these are minimal compared with mammals and microorganisms. The glycosylation
pattern is different too. Some carbohydrate moieties are unique to plants and
may present an antigenic challenge to the immune system when administered on a
regular basis. This is significant in the case of glycoproteins, although there
are ways to overcome such impediments. Nonglycosylated proteins when expressed
in plants are identical in all respects to those of mammalian origin. Even when
antibodies are produced in plants without the glycosylation a majority of
antibodies perform their function with utmost fidelity.
 Technology:
There are two transformation approaches used to produce recombinant
pharmaceuticals in plants. The first approach uses either a particle bombardment
technology or the bacterium Agrobacterium tumifaciens to incorporate the foreign
gene permanently in the plant tissue thereby guaranteeing perpetual propagation.
Several plants such as tobacco, rice, wheat, maize, oil seed rape and others
have been subjected to this method of transformation. The second strategy is to
infect the nontransgenic plants with modified recombinant viruses, which would
express the foreign gene(s) during their replication in the host. One could also
direct the foreign gene to be expressed in a tissue-specific manner such as in
the seeds. Examples of the latter type would be corn or oil seed rape.
Full-scale production will
likely involve grain and oilseed crops such as corn, rice, wheat, soybeans and
rapeseed. Proteins stored in seeds remain desiccated and have been shown to
retain their potency and quantity intact.
Companies and products in
development: As of this writing no protein produced in a transgenic animal
or plant has yet been approved for therapeutic use by the FDA. However, Monsanto
Protein Technologies (MPT), a unit of Monsanto Corporation, did take
a transgenic plant-produced antibody to the clinic in phase-II trials. The
product failed in the clinic but not because of its plant origin. This company
is also in research contract arrangements with several other biotech companies
to produce different antibodies in transgenic corn for different indications.
 Several
plant biotechnology companies have been in operation for more than 10 years and
developed several therapeutic proteins and enzymes for therapeutic use, which
are in various stages of clinical trials. Some products such as avidin, and ?-glucuronidase
are used in diagnostics. An enzyme, trypsin, produced in transgenic corn,
is used in a bioprocessing step in the production of insulin. The
products in development range from antibodies to enzymes, and cytokines to
growth factors.
CropTech Corporation*
(Blacksburg, VA) had 3 products in clinical production and 6 in the pre-clinical
stage. Glucocerebrosidase (for Gaucher’s disease) production in
transgenic tobacco using a technology whereby the production of the enzyme is
induced upon harvesting and crushing the tobacco leaves (wound healing mechanism
caused by an injury) has been patented by the company. This post-harvest
expression of a product has been termed MeGA-PharM® technology. Using tobacco
as the host, other products such as urokinase, human serum albumin
and iduronidase have also been expressed.
Large Scale Biology
Corporation (Vacaville, CA) has developed a technology known as GeneWare®,
which utilises modified viruses carrying a human gene to infect tobacco mosaic
virus (TMV)-susceptible regular tobacco plants. The human protein is expressed
in the leaf tissues in the next few days, which can then be harvested and
processed. High expression is claimed (up to 10 per cent of total soluble
proteins). This company is also working on producing peptide antibiotics in
tobacco. Such antibiotics may be too toxic for the bacterial hosts. Efforts are
going on to produce single chain antibodies in transgenic tobacco for eventual
treatment of non-Hodgkin’s lymphoma.
Medicago, Inc. (Quebec,
Canada) has expressed insulin, IL-2, and glutenin in alfalfa plant
tissues. ProdiGene (College Station, TX) has produced avidin, aprotinin,
and ?-glucuronidase in transgenic corn. In addition, this company is working
with Eli Lilly & Co to produce different proteases, which are used in
manufacturing other therapeutic proteins. Epicyte Pharmaceuticals (San
Diego, CA) is developing antibodies (“plantibodies’) against herpes simplex
virus 1 and 2 infection and for contraception (anti-sperm antibody as a
prophylactic) using transgenic corn. Planet Biotechnology, Inc. (Mountain
View, CA) is conducting clinical trials for CaroRx, a monoclonal antibody
produced in transgenic tobacco that is directed against Streptococcus mutans, an
oral bacterium that causes dental caries contributing to tooth decay.
Meristem Therapeutics
(France) is producing clinical quantities of gastric lipase for exocrine
pancreatic insufficiency in phase-II a trials for the Belgian pharmaceutical
concern, Solvay. Meristem is also undertaking efforts to produce
hemoglobin in tobacco and in corn. A novel technology, known as Oleosin-fusion
gene technology, is used by SemBioSys Genetics (Calgary, Canada) to
produce hirudin, an anticoagulant originally isolated from the leech,
Hirudo medicinalis. Canola, an oilseed rape, is used as the host.
Production economics: It
is estimated that the cost of production of a human therapeutic protein via
mammalian cell culture is 50 per cent of the total at the Bioreactor level and
50 per cent for downstream purification. A typical therapeutic protein costs $
200/g of pure protein at a production level of 200 kg/year. If the same protein
is produced in transgenic corn, the cost is expected to be about $ 50/g at a
production level of 100 kg of a monoclonal antibody. At a 100,000 kg/year level
the cost will be $ 4.5/g. The major cost difference is in the upstream part
while the purification costs are comparable, since the purification steps are
the same in both cases.
Bioequivalence: The corn
seed-derived antibody (produced by MPT) was found to be equivalent to the
mammalian-derived antibody in biochemical properties, serum and urine clearance,
and effectiveness on target-tissue binding. The corn seed-derived antibody met
purity, safety and potency requirements necessary for use in human clinical
evaluation. It was also demonstrated that seed stored under controlled
conditions provide viable seed for at least seven years. In addition, the
quality and yield of the antibody produced by milling the corn right after
harvest and after storage for 20 months remained the same.
Conclusion:
Transgenic plant technology presents a viable alternative for
economical production of human therapeutic proteins. It is considered
imminent that plant-derived human therapeutic proteins will be in the
market in a few years. The “points to consider” and “guidance”,
published by the FDA, in place for transgenic animal-produced
therapeutics are being adapted for plant-derived therapeutics. If the
purity of the plant-produced material can be maintained on a par with
that of mammalian cell-culture derived product, then the plant-derived
product will have an advantage in terms of cost benefit.
Croptech Corporation went
out of business in 2003 for lack of continued financing.
Definitions:
Codon: A sequence of
three nucleotide bases that specifies an amino acid or a signal to start or stop
a function.
Glycosylation: The process of attaching a carbohydrate molecule to
another molecule.
Glycoprotein: A protein which has a pendant carbohydrate at one or more
points.
Prion: A Proteinacious infectious particle which causes “mad cow
disease”
Transgenic: A species in which a foreign gene is incorporated.
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