藥學(xué)專(zhuān)業(yè)畢業(yè)論文外文翻譯--轉(zhuǎn)基因植物——生物醫(yī)藥的生產(chǎn)工廠(chǎng)（英文）

1、NATURE BIOTECHNOLOGY VOL 18 NOVEMBER 2000 http://biotech.nature.com 1151Biopharmaceuticals have traditionally been produced using a vari- ety of transgenic systems, including cultured mammalian cells, bacteria, and fun

2、gi1–3. In the future, demand for existing biophar- maceuticals (e.g., erythropoietin to treat anemia and insulin to treat diabetes), as well as new therapeutic proteins discovered through genomics efforts, is expected to

3、 rise considerably2. It is prudent, therefore, to evaluate alternative transgenic production systems and determine how the future availability of safe recombi- nant biopharmaceuticals can be ensured in a cost-effective m

4、an- ner. Producing therapeutic proteins in plants has many economic and qualitative benefits, including reduced health risks from pathogen contamination, comparatively high yields, and produc- tion in seeds or other stor

5、age organs2. The cultivation, harvesting, storage, and processing of transgenic crops would also use an exist- ing infrastructure and require relatively little capital investment2–4, making the commercial production of b

6、iopharmaceuticals an exciting prospect. Plants are potentially a cheap source of recombi- nant products4–6: Kusnadi et al.7 have estimated that the cost of producing recombinant proteins in plants could be 10- to 50-fold

7、 lower than producing the same protein by Escherichia coli fermen- tation, depending on the crop. Many recombinant therapeutic proteins are produced using mammalian expression systems. A big advantage of these, and of in

8、sect tissue culture systems, is that they correctly synthesize and process mammalian products. However, product yields are generally low, and the requirement for fetal bovine serum in the growth medium makes production e

9、xpensive2. In addition, cultured mammalian cells are sensitive to shear forces that occur during industrial-scale culture, and to variations in temperature, pH, dis- solved oxygen, and certain metabolites. This makes it

10、 necessary to control culture conditions carefully, because variation in cell growth can affect fermentation and product purity. Although bac- terial and fungal systems are more robust, they are not ideal for syn- thesiz

11、ing many mammalian proteins because of differences inmetabolic pathways, protein processing, codon usage, and the for- mation of inclusion bodies3. Although some differences exist in post-translation processing and in co

12、don usage between plants and mammals, these are few compared with differences between mam- mals and microorganisms2,8,9. Where differences in processing do represent a problem, it may be possible to engineer plants with

13、altered protein maturation pathways3,10. Biopharmaceuticals produced in cell culture systems have to be purified from the culture supernatant, an expensive process. Plants can be made to store proteins in seed endosperm,

14、 from where they can be easily extracted2,11. Nevertheless, purification is potentially an expensive step, and various methods are being developed to overcome this problem, including the expression of proteins as fusions

15、 with oleosin (see discussion below for hirudin4–6). An alternative approach is to cover the costs of purifi- cation with the income from the extraction of conventional prod- ucts, such as meal, oil, or starch. The costs

16、 of isolating human serum albumin from starch potatoes, for example, could be large- ly covered by concomitant starch production4,5,12. In addition, purification may not always be necessary, for example, in the case of e

17、dible vaccines. Plant-derived products, whether purified or not, are less likely to be contaminated with human pathogenic microorganisms than those derived from animal cells, because plants don’t act as hosts for human i

18、nfectious agents13. In this short review, we outline the main types of plant expres- sion systems employed for the agricultural production of biophar- maceuticals and provide an overview of the types of vaccine, anti- bo

19、dy, and therapeutic protein products currently under develop- ment (for a review on foreign protein production in plant tissue cul- ture, see ref. 14).Agricultural production systems Two transformation approaches are com

20、monly used to produce recombinant pharmaceuticals in plants8,10,15–17. In the first, stablyTransgenic plants as factories for biopharmaceuticalsGlynis Giddings*, Gordon Allison, Douglas Brooks, and Adrian CarterInstitute

21、 of Biological Sciences, University of Wales, Aberystwyth, Cledwyn Building, Aberystwyth Ceredigion SY23 3DD, UK. *Corresponding author (gdg@aber.ac.uk).Received 12 November 1999; accepted 11 August 2000Plants have cons

22、iderable potential for the production of biopharmaceutical proteins and peptides because they are easily transformed and provide a cheap source of protein. Several biotechnology compa- nies are now actively developing, f

23、ield testing, and patenting plant expression systems, while clinical trials are proceeding on the first biopharmaceuticals derived from them. One transgenic plant-derived biophar- maceutical, hirudin, is now being commer

24、cially produced in Canada for the first time. Product purification is potentially an expensive process, and various methods are currently being developed to overcome this problem, including oleosin-fusion technology, whi

25、ch allows extraction with oil bodies. In some cases, deliv- ery of a biopharmaceutical product by direct ingestion of the modified plant potentially removes the need for purification. Such biopharmaceuticals and edible v

26、accines can be stored and distributed as seeds, tubers, or fruits, making immunization programs in developing countries cheaper and potentially easier to administer. Some of the most expensive biopharmaceuticals of restr

27、icted availability, such as glucocere- brosidase, could become much cheaper and more plentiful through production in transgenic plants.Keywords: biopharmaceuticals, GM crops, edible vaccine, antibodies, production system

28、s REVIEW© 2000 Nature America Inc. ? http://biotech.nature.com© 2000 Nature America Inc. ? http://biotech.nature.comNATURE BIOTECHNOLOGY VOL 18 NOVEMBER 2000 http://biotech.nature.com 1153One important target

29、 for current vaccine efforts is hepatitis B, a virus responsible for the majority of persistent viremia in humans that can cause chronic liver disease. The first commercially available recombinant vaccine was developed i

30、n yeast after concerns about the safety of serum-derived antigens. In many places, expense, together with a lack of facilities (e.g., refrigeration installations) has prohibited the use of recombinant vaccines. In 1992,

31、the World Health Organization (Geneva) and a consortium of other philan- thropic organizations began the Children’s Vaccine Initiative. As part of this effort, Thanavala et al.43 initially developed trans- genic tobacco,

32、 and then potatoes, that express hepatitis B surface antigen (HbsAg)43,45. By maximizing the immunogenicity of untreat- ed edible plant tissues, the group hopes to produce vaccines for developing countries. They have rec

33、ently shown that HbsAg trans- genic potatoes administered orally to mice can elicit humoral immune responses45. Ultimately, attention will focus on the modifi- cation of bananas, which are grown extensively throughout th

34、e devel- oping world and, in contrast to potatoes, can be eaten raw23. Banana vaccines, delivered as a purée, would cost just a small fraction of the price of traditional vaccines. In the future, banana vaccines cou

35、ld be produced against a range of diseases, including measles, polio, diph- theria, yellow fever, and certain types of viral diarrhea.Antibodies The alternative to inducing the immune system to produce antibod- ies is to

36、 deliver them directly. Although the therapeutic potential of antibodies has long been recognized (e.g., for individuals needing short-term protection against infectious agents), difficulties associ- ated with their prod

37、uction have limited clinical use8,40. In 1989, Hiatt et al.17 first demonstrated that functional antibod- ies could be produced in transgenic plants. Since then, a consider- able amount of effort has been invested in dev

38、eloping plants for antibody (or “ plantibody”) production8,9,11,17,31,33,40,42. Recombinant antibodies can be targeted to seeds and tubers9,11,31,33, and eventually it should be possible to store, transport, and administ

39、er antibodies in such plant tissues, which would be advantageous for immuniza- tion programs in developing countries9,33,47. Many of the antibodies produced in transgenic plants have appli- cations for human and animal h

40、ealth care8,33,40,48 (Table 2), and they may also prove useful for other applications such as bioremedia- tion9. Recombinant antibodies include fully assembled whole immunoglobulins17,48, antigen-binding fragments of imm

41、unoglobu- lins, and synthetic single-chain variable fragment gene fusions (scFv)9,31. Single-chain Fv antibodies are encoded by an artificial gene made by joining together light- and heavy-chain variable sequences9,11,31

42、,33.Plantibodies could be of particular benefit for topical immunotherapy8,16,40. Passive immunization of the mucosal sites could be effective against bacteria, fungi, and viruses8. Plantibodies against cell surface anti

43、gens of Streptococcus mutans have been shown to reduce tooth decay in animal models and humans44,45. Repeated large doses of antibody are required for topical passive immunotherapy, and transgenic plants could be useful

44、for produc- ing such large quantities40. Furthermore, it has been shown that transgenic plants can be generated that will efficiently assemble complex secretory antibod- ies, something previously thought difficult to ach

45、ieve in plants8,40. This was accomplished by constructing tobacco plants expressing four transgenes through crossing of plants transformed with single transgenes (gene stacking)17. The resulting quadruple transgenics eff

46、iciently assemble secretory immunoglobulin A (sIgA), the pre- dominant immunoglobulin that protects against microbial infection at mucosal sites8,40. Secretory IgA is both more efficient at binding antigens and more stab

47、le than the plantibodies initially produced, being more resistant to proteolysis. Because of this increased stabili- ty, treatment of all mucosal sites may be possible, including the gas- trointestinal tract8. The use of

48、 edible plants for this is obviously an attractive possibility40. The ability to assemble immunoglobulins is a major advantage that plants have over bacterial expression systems.Biopharmaceuticals Transgenic plants have

49、been constructed that express proteins such as enkephalins26, α-interferon2,24, human serum albumin12, and two of the most expensive drugs: glucocerebrosidase13,41 and granulo- cyte–macrophage colony-stimulating factor2,

50、13,41 (Table 3). Applied Phytologics (API; Sacramento, CA) has modified rice plants to pro- duce human α-1-antitrypsin, a protein of therapeutic potential in cystic fibrosis, liver disease, and hemorrhages. Trials of α-1

51、-antit- rypsin transgenic rice commenced in 1998, with protein extracted from malted grain. API hopes to have regulatory approval for trans- genic plant medical products by 2004. Gaucher’s disease is a recessively inheri

52、ted lysosomal storage dis- order resulting from deficiencies of lysosomal hydrolase glucocere- brosidase enzyme13. A drug developed from enzyme purified from human placentas is highly effective at reducing clinical sympt

53、oms. However, 10–12 tons/year of placentas are required to produce enough glucocerebrosidase for the average type I Gaucher’s patient, making it one of the world’s most expensive drugs41. A recent switch to production in

54、 mammalian cell culture systems has reduced this cost, but didn’t remove the drug from the “ most expensive” league. Glucocerebrosidase production in transgenic tobacco13,41 was patented by Cramer and colleagues at Virgi

55、nia Tech and StateREVIEWTable 2. The production of antibodies in transgenic plants. Goal Plant Protein Expression ReferencesystemsImmunoglobulins Synthesis of secretory immunoglobin for Tobacco Hybrid sIgA-G specific for

56、 AMT 9,40 treatment of dental caries S. mutans antigen II Synthesis of full-length IgG1 Tobacco IgG (Guy’s 13) specific for S. mutans AMT 17,40 surface protein (SA I/II) IgG assembly and secretion Tobacco IgG specific

57、for human creatine kinase AMT 11 Comparison of glycosylation in plant- and Tobacco IgG (Guy’s 13) specific for S. mutans AMT 10 animal-derived IgG1 surface protein (SA I/II)Single-chain Fv fragments Accumulation and sto

58、rage of protein in tubers Potato Phytochrome binding scFv AMT 11,21 Treatment of non Hodgkin’s lymphoma Tobacco scFv of IgG from mouse B-cell lymphoma AMT 19 Production of tumor-associated marker antigen Cereals scFvT84

59、.66 against carcinoembryogenic Particle 33 antigen bombardmentAMT, Agrobacterium mediated transformation.© 2000 Nature America Inc. ? http://biotech.nature.com© 2000 Nature America Inc. ? http://biotech.nature.