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PHARMACROPS: Vaccines - A new health food




                                  PART 1


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TITLE:  VACCINES - A NEW HEALTH FOOD

SOURCE: The Economist, UK

AUTHOR: 

URL:    http://www.economist.com/science/displaystory.cfm?story_id=9333426

DATE:   14.06.2007

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VACCINES - A NEW HEALTH FOOD

Growing vaccines in rice may make them more stable and more effective

GETTING two for the price of one is always a good bargain. And according to a paper in this week’s Proceedings of the National Academy of Sciences, that is what Tomonori Nochi of the University of Tokyo and his colleagues have done. Using genetic engineering, they have overcome two of the limitations of vaccines. One is that they are heat-sensitive and thus have to be transported along a ”cold chain” of refrigerators to the clinics where they are used. The other is that, although they stimulate immune responses inside the body, they often fail to extend that protection to the outside, where it might prevent bacteria and viruses getting inside in the first place.

In this context, the outside is not the skin: that is dry and hostile to germs. It is the damp and welcoming surface of places such as the lung and the gut that are at risk. Although these are casually called internal, technically they are not. Any nasties in the gut or lungs have to cross the walls of those organs before they can multiply inside the body.

Dr Nochi’s genetic engineering involved growing the vaccine in rice. To prove the principle, he chose cholera, but it should work with other vaccines as well. With cholera, the immune response is induced by what is known as the cholera toxin B-subunit. This is a protein, and Dr Nochi took the gene that encodes it and inserted that gene into the genome of rice. Next to the B-subunit gene itself, he inserted a second piece of DNA called a promoter. This, as its name suggests, promotes activity in an adjacent gene. Promoters themselves are activated by other molecules, and whether they are switched on or not depends on whether the cell they are in provides the necessary stimulation. In this case Dr Nochi picked a promoter that is active in the tissue of rice grains.

It was then just a question of growing the rice and feeding the resulting grains to some experimental mice to find out what would happen. The first thing that happened was that the grains protected the B-subunit from being broken down in the stomach, thus overcoming one of the regular bugbears of protein-based drugs: that they cannot be given by mouth, because they will be digested. This is a problem with today’s cholera vaccine which is indeed taken by mouth and therefore affords poor protection. When the B-subunits got to the intestines they did exactly what Dr Nochi hoped and induced the production of antibodies and the secretion of those antibodies into the mucous coating of the intestinal wall. Dr Nochi’s mice really were protected. When he fed them cholera toxin, they did not get sick.

On top of all this, he got as good a response with rice that had been stored at room temperature for 18 months as he did when he used fresh grains. For a vaccine against a disease that is found predominantly in poor countries—places that tend to lack refrigerators and have only intermittent power to run those that do exist—that is an enormous advance. If Dr Nochi’s finding can be translated into a product that is safe and effective for people, it will be a big boost to the health of the world’s poor.



                                  PART 2

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TITLE:  VACCINES ARE FOR DINNER

SOURCE: Proceedings of the National Academy of Sciences of the United States of America, USA

AUTHOR: David W. Pascual

URL:    http://www.pnas.org/cgi/content/full/104/26/10757

DATE:   26.06.2007

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VACCINES ARE FOR DINNER

Transgenic plants have been sought not only as bioreactors but also as potential scaffolds for oral vaccines. Tobacco was initially exploited for the successful expression of Streptococcus mutans surface protein A as a potential dental caries vaccine (1) and for hepatitis B surface antigen (2) as a vaccine bioreactor alternative for viral hepatitis B. Today, a number of edible plants (Table 1), including potatoes, tomatoes, maize, and soybeans, have been genetically modified to express a variety of vaccine targets, including hepatitis B surface antigen (2), Norwalk virus particles (3, 4), heat-labile enterotoxin B subunit (5, 6), and others (7–11) that can benefit both humans (6–8) and livestock (9–11). The advantage of edible vaccines is that often the plant products, whether leaves, fruit, or seed, can be readily consumed with limited or no processing. Viable oral platforms such as edible products suitable for human use are in demand to deliver vaccines (12). Obviously, the fact that food products are consumed obviates many of the health concerns that arise in the oral vaccination of humans (as reviewed in ref. 13). Edible vaccines can also have the advantage of circumventing cold-storage issues (the ”cold chain”), because plant tissues can be dried or, as when the seeds are targeted, have low moisture content. Likewise, water- or oil-based plant extracts can provide additional storage convenience. In this issue of PNAS, Nochi et al. (14) describe a rice-based oral vaccine that potentially addresses many of these topics. At hand is the need for vaccine development and strategies to aid underserved nations with the ability to produce vaccines locally in a cost-effective manner. Because rice is produced in many such areas, this current work shows the feasibility of propagating rice-based vaccines that are truly edible vaccines, unlike in the earlier work with tobacco that ultimately provided the mechanisms for edible vaccine development (1, 2). In addition, this approach breaks the cold-chain barrier that for many conventional vaccines drives up cost and creates storage problems. In fact, it is estimated that removal of this barrier could give an added benefit of as much as $300 million per year, which could provide vaccines for an additional 10 million children (15). In this regard, the work by Nochi et al. shows that their transgenic rice was stable for >18 months at room temperature or at 4°C. Because these vaccines are needle-free (16), they have the added advantage of eliminating the associated waste and potential for dissemination of bloodborne infections. The propagation of transgenic rice also has a lesser impact on the environment because of rice’s limited pollen scattering, unlike with maize or wheat (13). Thus, Nochi et al. have addressed many of the issues that prevent vaccines from coming to market, including cost, cold-chain, needle-associated, and regulatory concerns.

The authors’ examination of the type of vaccine produced is also a significant achievement. A major obstacle to furthering the field of edible vaccines is the need to produce effective immunity. Typically, vaccines applied to mucosal surfaces in the absence of adjuvant fail to stimulate an immunogenic response and resort to the default pathway, or tolerance (as reviewed in ref. 12). Obviously, we need to be tolerant of food; the genetic modification of foods may be potentially problematic and could result in food allergies, thus jeopardizing future consumption of the unmodified food in the diet. For rice, a major worldwide staple, this would be especially problematic. Nochi et al. (14) nicely show that oral immunization with the transgenic rice encoding the cholera toxin B subunit (CTB) does not stimulate a serum IgG response against rice storage protein or, presumably, a secretory IgA response. This will be a key element in the success of transgenic rice. Moreover, they show that the encoded CTB is resistant to the gut environment because of its expression in endosperm that normally is resistant to gastrointestinal digestion. In overcoming these obstacles, Nochi et al. show modest fecal IgA and serum anti-CTB antibody responses after the mice were orally immunized with the equivalent of 75–150 µg of rice-generated CTB per dose, given six times over a 10-week vaccination schedule. When compared with CTB rice-immunized mice, the mice orally dosed with recombinant CTB showed equivalent serum IgG antibody responses but weak fecal IgA responses. This latter finding is surprising, but the authors did show protection against native cholera toxin challenge, as evidenced by reduced intestinal water content (diarrhea). This level of protection was equivalent to that in mice given the rice CTB, whereas mice given native rice or PBS were not protected.

The selection of CTB as a candidate vaccine for testing in rice is appropriate because of its adjuvant properties (17). Nochi et al. (14) were able to show that the rice CTB could enter via the gut sampling cells or M cells (18), which are responsible for continually sampling the gut contents for aberrant antigens. A number of pathogens can also enter via these cells to cause infection, thus subverting this defensive mechanism. A major obstacle for successful oral transgenic plant vaccines is the potential outcome of tolerization rather than immunity, but this is true with any oral vaccine given in the absence of mucosal adjuvant. Tolerance, or lack of responsiveness, occurs when a particular antigen or vaccine is fed without costimulation of the innate immune system. Consequently, instead of being immunized, the host becomes actively unresponsive, and subsequent challenge with the antigen leaves the host unresponsive. Taking advantage of this known behavior, Takagi et al. (19) produced transgenic rice that encoded a fusion protein between soybean seed-storage protein glycinin AlaB1b and known allergen peptides from Japanese cedar pollen for targeted expression in the rice seed endosperm. When mice were fed with this transgenic rice expressing the pollen peptides known to be reactive for T cells, they were resistant to Japanese cedar pollen challenge, showing reduced serum histamine release, reduced allergic IgE antibodies, and reduced Th2 cells that support the allergic response. The authors also showed that cooking the transgenic rice did not affect its ability to tolerize the host (19). Although this outcome would be expected because T cell peptides are required for tolerization, the investigators show that oral feeding with transgenic rice can potentially treat autoimmune diseases (19), which is an advantage of unadjuvanted oral vaccines (12). CTB also has been used to induce oral tolerance (reviewed in ref. 17). Although immunity or tolerance can be driven by CTB, presumably by interaction with host M cells on Peyer’s patches, a major limitation of any edible vaccine will be the required coadministration of a mucosal adjuvant, unless mucosal adjuvants can be successfully coexpressed with the desired edible vaccine. Being able to demonstrate immunity using their transgenic rice represents a significant accomplishment for Nochi et al.

The greater protein content of rice is an advantage over some of the starch-based edible vaccines described previously (Table 1) and for heat-labile enterotoxin B subunit (LTB; ref. 5). LTB is very similar to CTB in exhibiting adjuvant activity. When maize-derived LTB was fed in three 1.0-mg doses to human volunteers, seven of nine volunteers produced a serum IgG response, whereas only four individuals showed a fecal IgA response (6). Protein-based seeds such as soybeans have the unique advantage of a high protein content (35–40%), as opposed to rice and maize that have 8–10% protein. When LTB was expressed in soybeans, as much as 2.4% of the total seed protein was LTB (20). Mice fed or parenterally immunized with soybean-derived LTB showed IgG and IgA anti-LTB antibody responses that could protect against diarrhea. Thus, these collective studies demonstrate, regardless of the plant-derived vaccine used, the importance of developing coexpressed mucosal adjuvants with the edible vaccine. Although Nochi et al. (14) show no anti-rice storage protein response, neither the expressed CTB nor the recombinant CTB have the adjuvant potency of native cholera toxin. However, when mice were fed wild-type rice in conjunction with native cholera toxin, an anti-rice storage protein antibody response was elicited (14), suggesting that the expression of highly potent mucosal adjuvants may be problematic if these were to stimulate immune responses to the food product, be it rice, maize, wheat, or soybeans. Perhaps plant-derived alternative adjuvants need to be sought, or alternative adjuvants need to be added exogenously to the prepared edible vaccine.

The future of edible vaccines will depend on the feasibility of producing sufficient quantities of immunogenic vaccines. The edible vaccines that innately possess immunogenicity and do not require additional adjuvant will probably be the first successful vaccines for human or livestock use. The selection of the transgenic plant platform will largely depend on the vaccine and the region where it will be propagated. The grain-based vaccines, as described by Nochi et al. (14), will be the most likely candidates because pollen dispersion or potential contamination of normal food supplies with transgenic pollen can be controlled. The future use of these vaccines also will depend on the development of stable transgenic lines that effectively maintain the vaccine expression for subsequent plant generations. In addition, edible vaccines may have to withstand food processing and possibly cooking. Nevertheless, a demand for the development of edible vaccines persists because they can eliminate many of the problems associated with conventional vaccines, including storage issues, injection risks and associated waste, high production costs, and ease of distribution in underserved areas. Who knows? It may nice to have a little vaccine with supper tonight!

 

Acknowledgements

This work was supported by Public Health Service Grants AI-41123, AI-55563, and AI-56286 and in part by Montana Agricultural Station and U.S. Department of Agriculture Formula funds.

 

References

1 Curtiss, RI & Cardineau, CA. (1990) World Patent App WO 90/02484.

2 Mason, HS, Lam, DM & Arntzen, CJ. (1992) Proc Natl Acad Sci USA 89, 11745–11749.[Abstract/Free Full Text]

http://www.pnas.org/cgi/ijlink?linkType=ABST&journalCode=pnas&resid=89/24/11745

3 Mason, HS, Ball, JM, Shi, JJ, Jiang, X, Estes, MK & Arntzen, CJ. (1996) Proc Natl Acad Sci USA 93, 5335–5340.[Abstract/Free Full Text]

http://www.pnas.org/cgi/ijlink?linkType=ABST&journalCode=pnas&resid=93/11/5335

4 Zhang, X, Buehner, NA, Hutson, AM, Estes, MK & Mason, HS. (2006) Plant Biotechnol J 4, 419–432.[CrossRef][Medline]

5 Mason, HS, Haq, TA, Clements, JD & Arntzen, CJ. (1998) Vaccine 16, 1336–1343.[CrossRef][ISI][Medline]

6 Tacket, CO, Pasetti, MF, Edelman, R, Howard, JA & Streatfield, S. (2004) Vaccine 22, 4385–4389.[CrossRef][ISI][Medline]

7 Mercenier, A, Wiedermann, U & Breiteneder, H. (2001) Curr Opin Biotechnol 12, 510–515.[CrossRef][ISI][Medline]

8 Tacket, CO. (2004) Expert Rev Vaccines 3, 529–531.[CrossRef][Medline]

9 Streatfield, SJ. (2005) Rev Sci Tech 24, 189–199.[ISI][Medline]

10 Rice, J, Ainley, WM & Shewen, P. (2005) Anim Health Res Rev 6, 199–209.[CrossRef][Medline]

11 Piller, KJ, Clemente, TE, Jun, SM, Petty, CC, Sato, S, Pascual, DW & Bost, KL. (2005) Planta 222, 6–18.[CrossRef][ISI][Medline]

12 Lavelle, EC & O’Hagan, DT. (2006) Expert Opin Drug Delivery 3, 747–762.[CrossRef]

13 Streatfield, SJ. (2005) Expert Rev Vaccines 4, 591–601.[CrossRef][ISI][Medline]

14 Nochi, T, Takagi, H, Yuki, Y, Yang, L, Masumura, T, Mejima, M, Nakanishi, U, Matsumura, A, Uozumi, A & Hiroi, T, et al. (2007) Proc Natl Acad Sci USA 104, 10986–10991.[Abstract/Free Full Text]

http://www.pnas.org/cgi/ijlink?linkType=ABST&journalCode=pnas&resid=104/26/10986

15 Das, P. (2004) Lancet Infect Dis 4, 719.[Medline]

16 Giudice, EL & Campbell, JD. (2006) Adv Drug Delivery Rev 58, 68–89.[CrossRef][ISI][Medline]

17 Lycke, N. (2005) Curr Mol Med 5, 591–597.[CrossRef][ISI][Medline]

18 Corthesy, B. (2007) J Immunol 178, 27–32.[Abstract/Free Full Text]

http://www.pnas.org/cgi/ijlink?linkType=ABST&journalCode=jimmunol&resid=178/1/27

19 Takagi, H, Hiroi, T, Yang, L, Tada, Y, Yuki, Y, Takamura, K, Ishimitsu, R, Kawauchi, H, Kiyono, H & Takaiwa, F. (2005) Proc Natl Acad Sci USA 102, 17525–17530.[Abstract/Free Full Text]

http://www.pnas.org/cgi/ijlink?linkType=ABST&journalCode=pnas&resid=102/48/17525

20 Moravec, T, Schmidt, MA, Herman, EM & Woodford-Thomas, T. (2007) Vaccine 25, 1647–1657.[CrossRef][ISI][Medline]

21 Choi, NW, Estes, MK & Langridge, WH. (2005) Mol Biotechnol 31, 193–202.[CrossRef][ISI][Medline]

Companion article to this Commentary:

>From the Cover: Rice-based mucosal vaccine as a global strategy for cold-chain- and needle-free vaccination

Tomonori Nochi, Hidenori Takagi, Yoshikazu Yuki, Lijun Yang, Takehiro Masumura, Mio Mejima, Ushio Nakanishi, Akiko Matsumura, Akihiro Uozumi, Takachika Hiroi, Shigeto Morita, Kunisuke Tanaka, Fumio Takaiwa, and Hiroshi Kiyono

PNAS 2007 104: 10986-10991.

[Abstract]

http://www.pnas.org/cgi/content/abstract/104/26/10986

[Full Text]

http://www.pnas.org/cgi/content/full/104/26/10986



                                  PART 3

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TITLE:  GM SAFFLOWER WITH HUMAN PRO-INSULIN

SOURCE: Institute of Science in Society, UK

AUTHOR: Joe Cummins

URL:    http://www.i-sis.org.uk/gmSaffloweHumanPro-Insulin.php

DATE:   18.07.2007

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GM SAFFLOWER WITH HUMAN PRO-INSULIN

Regulators show cavalier disregard for the safety of threatened species as well as human beings in proposed release of the GM pharm crop. Prof. Joe Cummins

This report has been submitted to the USDA on behalf of ISIS. Please circulate widely

Proposed release of transgenic safflower shrouded in secrecy

USDA-APHIS conducted an Environmental Assessment (EA) [1] in response to an application (06-363-103r), received from SemBioSys, Inc to field test a transgenic safflower (Carthamus tinctorius) line 4438-5A that produces human pro-insulin. The transgenic safflower was engineered to express an oleosin-human pro-insulin protein exclusively in its seed. The field site (<1 acre) is located on private property in Lincoln County, WA, and will be surrounded on all sides by a 50 ft fallow strip. The exact location of the site is withheld from the public; but the application and risk assessment are open for public comment at http://www.regulations.gov/fdmspublic/component/main until 23 July 2007.

Pro-insulin is the precursor to insulin, normally made in the beta cell of the islets of Langerhans of the human pancreas. The protein is synthesized in the endoplasmic reticulum (membrane stacks within the cell), where it is folded and two sulphydryl (-SH) groups are oxidized into a disulphide bond (-S-S-). It is then transported to the Golgi apparatus (a special organelle) where it is packaged into secretory vesicles, and processed by a series of proteases into mature insulin. Mature insulin has 39 less amino acids; 4 are removed altogether, and the remaining 35 amino acids - the C-peptide - are cut out from the middle of the pro-insulin molecule; the two ends segments - the B chain and A chain - remain connected by the disulphide bond formed earlier [2, 3].

A patent application [4] describes the genetic modifications for high expression of human insulin in plants, including shortening the C-peptide by four amino acids. The APHIS report [1] notes further that the human pro-insulin has two amino acids removed for stability in plants plus 11 C terminal amino acids added to ensure retention of the protein in the endoplasmic reticulum of the plant seed cell. The pro-insulin sequence was fused to the Arabidopsis oleosin gene, to be exclusively expressed in seeds. Expression of the fused gene was controlled by the phaseolin promoter and terminator sequences from common bean. The bean promoter drives seed-specific transcription of the synthetic pro-insulin. A selectable marker is regulated by the parsley ubquitin promoter and terminator, and was deemed confidential business information even though it is said to be the most commonly used selectable marker in plants, and had been used in many previous field trials [1]. Animal feeding tests evaluating the toxicity of the neither the synthetic pro-insulin nor the marker gene and its proteins were included with the EA.

 

Site of release in area with threatened species

The area selected for the transgenic safflower field test releases - ”sagebrush steppe” - is dry and dominated by sagebrush. Resident animals include the sage grouse, sage sparrows, loggerhead shrikes, and even the once ubiquitous black-tailed hare or ”jackrabbit”. According to USAD/APHIS [1], the threatened species in the test area also include bald eagle, pygmy rabbits, Columbian white tailed deer and grey wolf, and the plant species Spalding’s catchfly and Ladies’ tresses. Pygmy rabbits are the most threatened species, the Columbia pygmy rabbit feeds mainly on sagebrush and its number may be as low as 30 or less. There has been limited success in breeding the rabbits in captivity [5, 6]. The pygmy rabbit is likely to feed on the transgenic safflower seeds with potentially detrimental (even fatal) consequences. The USDA/APHIS report claims there will be no toxicity from ingesting seeds from the transgenic safflower, from contact or from inhaling dust and debris [1]. Even if that were true - and there is evidence ignored by APHIS suggesting that the ingested pro-insulin from transgenic safflower is active (see below) - the disruption of the habitat of the pygmy rabbit by human activities and transportation is likely to drive the threatened animals to extinction. APHIS displays a cavalier disregard for the threatened species, ignoring studies that do not support their conclusions.

 

Evidence of potential harm to threatened species ignored

There is at least one report showing that transgenic pro-insulin can effectively reduce blood glucose in rats. Feeding a bracken fungus, Ganoderma lucium, modified with a gene for human pro-insulin to diabetic rats reduced their blood glucose [7]; presumably the modified fungus cell wall and endoplasmic reticulum prevent rapid degradation of pro-insulin, allowing the transgenic organism to deliver insulin to the diabetic animal. Cholera toxin pro-insulin fusion proteins were produced in lettuce and tobacco plants; and when powdered transgenic plant preparations were fed to diabetic mice, oral tolerance to insulin was produced, preventing the autoimmune degradation of insulin-producing beta cells in the pancreas [8]. Human insulin produced in Arabidopsis seeds was activated by exposure to the common digestive enzyme trypsin [9]. The APHIS report presumes that human pro-insulin will be degraded too rapidly for it to become activated when ingested by animals, but the studies cited show that may not the case. Furthermore, functional argentine peptides were found to enhance intestinal absorption of insulin such peptides may be encountered commonly in anti-microbial peptides [10]. Seed debris may produce dust that contains human pro-insulin, and it is worth noting that inhaled insulin is an available option for human therapy [11]. The APHIS report dismisses the possibility that inhaled debris and dust from the transgenic safflower could be active, but provides no experimental evidence to support that conclusion.

APHIS implies that wild animals would not be affected by human insulin [1], but rabbits were among the animals first used in the discovery of insulin, and continue to be used as experimental animals in current studies on insulin action [12]. Furthermore, birds [13] and snakes [14] also respond to human insulin; and it is probably safe to say that all of the threatened species, and human beings are potential victims of the release of food crops modified to produce human insulin. The APHIS report notes that grain crops surrounding the transgenic safflower plot will provide a more attractive ”free lunch” for birds and mammals than the transgenic safflower; that is a fallacious and dangerous assumption because the ’free lunch’ will attract both foragers and predators to the test site. Furthermore, the fallow strip around the test plot is unlikely to discourage browsers such as rabbits that feed at night to avoid predators.

 

Safe haven for pharm crops but deadly for humans and wild life

Eastern Washington State is rapidly being transformed into a haven for transgenic crops modified to produce pharmaceuticals. Along with previous safflower field test releases, large plantings of humanized barley are being tested. The exact locations of such tests are not disclosed and people living near the test sites are unaware of the potential hazards to their health. The impact of such developments on threatened species is also ignored and dismissed by APHIS. The APHIS report reads more like a public relations document for the company rather than an independent critical evaluation of the company proposal. This is potentially deadly for humans and wildlife, and the agency should be held to public account.

 

References

1 USDA-APHIS Environmental Assessment In response to permit application (06-363-103r), received from SemBioSys, Inc. for a field-test to produce human proinsulin (line 4438-5A) in genetically engineered safflower (Carthamus tinctorius) seeds U.S. Department of Agriculture Animal and Plant Health Inspection Service Biotechnology Regulatory Services 06_363103r 06/22/2007 http://www.regulations.gov/fdmspublic/component/main

2 Wikipedia Proinsulin 2007 http://en.wikipedia.org/wiki/Proinsulin

3 Davidson, H. Proinsulin processing. Cell Biochemistry and Biophysics 2004 Supplement, 143-57.

4 Molony M, Boothe J, Keone R, Nykiforuk C and Van Rooijen. Method for production of insulin in plants, 2005 US Patent 2005/0039235A1

5 Washington Department of Fish and Wildlife Pygmy Rabbit 1995

6 Hays D. Washington Department of Fish and Wildlife Washington Pygmy Rabbit 2003 Recovery Plan Update addendum to 1995 above

7 Ni T, Hu Y, Sun L, Chen X, Zhong J, Ma H and Lin Z. Oral route of mini-proinsulin-expressing Ganoderma lucidum decreases blood glucose level in streptozocin-induced diabetic rats. Int J Mol Med. 2007, 20(1), 45-51.

8 Ruhlman T, Ahangari R, Devine A, Samsam M and Daniell H.   Expression of cholera toxin B-proinsulin fusion protein in lettuce and tobacco chloroplasts--oral administration protects against development of insulitis in non-obese diabetic mice. Plant Biotechnol J. 2007, 5(4), 495-510.

9 Nykiforuk CL, Boothe JG, Murray EW, Keon RG, Goren HJ, Markley NA and Moloney MM. Transgenic expression and recovery of biologically active recombinant human insulin from Arabidopsis thaliana seeds. Plant Biotechnol J. 2006,:77-85.

10 Morishita M, Kamei N, Ehara J, Isowa K and Takayama K. A novel approach using functional peptides for efficient intestinal absorption of insulin. J Control Release 2007, 118(2), 177-84.

11 Guevara CA. Inhaled insulin for diabetes mellitus. N Engl J Med. 2007, 356(20):2106-7.

12 Barillas R, Friehs I, Cao-Danh H, Martinez JF, del Nido PJ. Inhibition of glycogen synthase kinase-3beta improves tolerance to ischemia in hypertrophied hearts. Ann Thorac Surg. 2007, 84(1), 126-33.

13 Remage-Healey L and Romero LM. Corticosterone and insulin interact to regulate glucose and triglyceride levels during stress in a bird. Am J Physiol Regul Integr Comp Physiol. 2001, 281(3), R994-1003.

14 Sidorkiewicz E and Skoczylas R. Effect of insulin on the blood sugar level in the grass snake (Natrix natrix L.). Comp Biochem Physiol A. 1974, 48(3), 457-64.



                                  PART 4

------------------------------- GENET-news -------------------------------

TITLE:  SAFFLOWER SEEDS SOWING A PROTEIN-PRODUCING FUTURE

SOURCE: in-Pharma Technologist, France

AUTHOR: Katrina Megget

URL:    http://in-pharmatechnologist.com/news/ng.asp?n=78254-sembiosys-pfizer-safflower-seed-lines-transgenic-technology-apo-a

DATE:   17.07.2007

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SAFFLOWER SEEDS SOWING A PROTEIN-PRODUCING FUTURE

17/07/2007 - The humble safflower seed is looking like it could be the next protein-producing factory after SemBioSys Genetics produced commercial levels of an atherosclerosis treatment.

The Canada-based biotechnology company has successfully modified safflower seed lines, and expressed and collected apolipoprotein A1 and its variant apolipoprotein A1 (Milano), collectively referred to Apo A1,which is a next-generation cardiovascular drug that targets the removal of atherosclerotic plaque from arteries.

The development represents a move into the $35bn cardiovascular drug market, but the seed lines have also proved successful in producing about 43 other therapeutic proteins at a fraction of the cost of conventional methods, SemBioSys chief scientific officer Dr Maurice Moloney told US-PharmaTechnologist.com.

The transgenic technology, which is looking as an increasingly popular tool for producing proteins, was the brainchild of Moloney, who developed the genetically modified ”Roundup Ready” crops.

In the safflower seed case, the Apo A1 gene is only expressed in the seed. The technology, which can be used in any oil seed, has been further developed so that the protein attaches itself to oil bodies in the seed making the recovery of the protein much easier as the oil floats out of the solution when the seeds are crushed to extract the protein.

”This dramatically reduces the cost of purification,” Moloney said.

The company decided to focus on safflower because it was a less common crop, compared to canola, so it was easier to biologically isolate the crop reducing the chances of cross pollination or the accidental mixing of seeds with non-transgenic ones, he said.

Meanwhile, developing the seeds as the protein-producing factories instead of another part of the plant, meant the company could utilize the long-term storage capabilities that seeds have as well as being able to adjust production of the protein depending on the market demand.

”This is difficult to do if you are dealing with conventional methods like fermentation,” Moloney said.

The cost advantages were also very promising over conventional methods, he said.

The cost to grow a ton of seed, about an acre of safflower which would then produce about 2kg of Apo A1, would be about $800. This is compared to the current cost which sits in the range of about $400 per gram of Apo A1.

Moloney estimated the technology could be developed to reduce the cost by 90 per cent.

As a result, the safflower seed technology would be particularly beneficial in the production of Apo A1, as large amounts of the protein were required for treatment, he said.

SemBioSys president and chief executive Andrew Baum said in a statement: ”We believe manufacturing capacity and cost are major commercialization issues for pharmaceutical companies developing Apo A1. Today’s announcement demonstrates that safflower seed is an enabling production vehicle for Apo A1 with the scale and economics necessary to allow for commercialization of this potentially transformative therapy.”

Apo A1 has been described as a promising new therapy for the treatment of cardiovascular disease. It is the major component of the good cholesterol, high density lipoprotein (HDL), which naturally removes atherosclerotic plaque from arteries.

Currently, Pfizer is developing Apo A1 via E.coli, with positive clinical trial results, while CSL and Borean Pharma have both recently confirmed the strong therapeutic potential of their respective Apo A1-based drug candidates, CSL-111 and Trimeric Apo A-I, in clinical and preclinical trials.

SemBioSys intends to scale up production of safflower-produced Apo A1 and perform the necessary preclinical work in 2008 in order to initiate clinical trials in 2009.

The company has also produced authentic human insulin in safflower, which is expected to enter clinical trials in early 2008.

SemBioSys is now planning to initiate development program partnerships.


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