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    Dr. Sachin Rustgi
    Dr. Sachin Rustgi

    Discussion with Assistant Research Professor Sachin Rustgi on the Genetic Modification of Wheat to Make it Safe for Celiacs

    Reviewed and edited by a celiac disease expert.

    Journal of Gluten Sensitivity Spring 2014 Issue

    Discussion with Assistant Research Professor Sachin Rustgi on the Genetic Modification of Wheat to Make it Safe for Celiacs - Image: CC--Clare Black
    Caption: Image: CC--Clare Black

    Celiac.com 11/21/2016 - This article is the result of an email exchange between Scott Adams and Dr. Sachin Rustgi, which took place between January and March, 2014.

    Scott Adams:
    For many years researchers have known that a non-genetically modified, celiac safe wheat does, in fact, exist. Please see:

    Celiac.com Sponsor (A12):
    I believe that what you are actually doing, which is supported by an approximately $900K corporate grant (if I recall correctly), is to create a GMO version that you can patent in order to make money selling the seeds. This may not be necessary, as what you seek already exists naturally, and I did explain this to your cooperator years ago.

    Sachin Rustgi:
    We are aware of these publications mentioned in your post. It is unfortunate that some of these research papers make broad claims not fully supported by the data presented in these reports. This practice is damaging to society in these two ways: i) These publications mislead the public, which gives rise to misconceptions or myths, making it difficult for the general public to accept other innovative ideas. ii) It could even negatively impact public health if the results were blindly accepted and changes were made in routine eating practices without having careful scientific scrutiny of the findings. The popular press and media is partly responsible, because without assessing the credibility of results, they pick broad claims from these publications and serve them to the public in language laden with emotional impact, which the public receives and bases their opinion on. This is also true for the general claims made in the publications cited earlier.

    Different celiac patients are sensitive to different ‘gluten' proteins (prolamins). If one feeds peripheral blood cells sampled from a patient or a small group of patients (from a specific geographical location) with gluten proteins derived from a wheat genotype, it is expected either to see a reaction (monitored by the production of interferon gamma) or no apparent effect. But in the latter case it does not mean that the wheat genotype is non-toxic to all celiac patients. Because the sample is not a good representative of the genetic variability for disease susceptibility available in the global population, and is likely representing the prevalent disease predisposition allele present in a population inhabiting a particular geographical area or a common disease predisposition allele existing in a larger population (like the one that interacts with the immunogenic 33-mer peptide derived from alpha 2 gliadin). Thus, these T-cell based assays using cell-lines restricted to specific gliadin epitopes are not sufficient to claim general low-toxicity of wheat lines for all celiac patients. I also recommend that readers consult the following publications: Kasarda DD (2007) Letter to the editor: Triticum moncoccum and celiac disease. Scandinavian Journal of Gastroenterology 42(9):1141-1142; Vaccino P, et al. (2009) A catalogue of Triticum monococcum genes encoding toxic and immunogenic peptides for celiac disease patients. Mol Genet Genomics 281(3):289–300.

    The results of screening hexaploid wheat material under the Celiac Disease Consortium (CDC) funded projects in the Netherlands resulted in a number of publications (Molberg et al. 2005; van Herpen et al. 2006; van den Broeck et al. 2009; van den Broeck et al. 2010). But the authors of these publications never claimed that the material can be used generally. Rather, they suggested these lines to have ‘low-toxicity', as they are devoid of specific epitopes or gluten proteins. Thus, they are good for consumption by a group of celiac patients who share a specific susceptibility allele. We have summarized this material and associated limitations in our publication under two headings, "Wheat Genotypes Naturally Deficient in Immunogenic Gluten Peptides" and "Discussion" [consult Wen et al. 2012 Proc Natl Acad Sci U S A 109(50):20543-20548 for details].

    In addition, there is a misconception that with breeding for improved yield, protein content and quality has enhanced the toxicity of the wheat lines, which has resulted in higher incidence of disease over the last couple of decades. Although careful analysis of the facts suggests that nothing has changed over time other than eating habits, procedures of disease diagnosis (became more sophisticated) and public awareness grew, which might have resulted in this increase in the estimated number of celiac patients [also consult Kasarda (2013) J Agric Food Chem 61:1155-1159; Brouns et al. (2013) J Cereal Sci 58:209-215].

    However, it is possible to identify low toxicity wheat lines showing reduced accumulation of certain prolamins or immunogenic epitopes, but these lines are not good for general use by celiac patients making labeling of these lines a nightmare, because with the present technology it almost impossible to make recommendations to the patients that they are sensitive only to a specific gluten protein and thus can consume a particular wheat variety.

    Thus, this trait is an obvious candidate for genetic engineering. Two major achievements in this direction are:

    • i) Gil-Humanes J, Pistón F, Tollefsen S, Sollid LM, Barro F (2010) Effective shutdown in the expression of celiac disease-related wheat gliadin T-cell epitopes by RNA interference. Proc Natl Acad Sci USA 107(39):17023–17028.
    • ii) Wen et al. (2012) Structural genes of wheat and barley 5-methylcytosine DNA glycosylases and their potential applications for human health. Proc Natl Acad Sci U S A 109(50):20543-20548.

    Collectively, as different celiac patients have sensitivities for different gliadins and glutenins, it is almost impossible to breed wheat lines safe for all celiac patients using conventional breeding approaches. A second issue is the identification of a product's suitability for a group of celiac patients and its labeling, which would be a great logistic challenge. After careful scrutiny of the literature and that the Codex Alimentarius Commission declared that all wheat, barley and rye species including spelt (Triticum spelta L.), khorasan or kamut (T. polonicum L.), durum, einkorn (T. monococcum), triticale, tritordium and their hybrids are immunogenic, and should be avoided by celiac patients (also consult Open Original Shared Link) add to that challenge. Moreover, some individuals are now known to be sensitive to oat gluten proteins (however, all oat varieties are not toxic, e.g., PrOatina™), and in rare cases, some are even sensitive to maize gluten proteins.

    According to the latest (August 2, 2013) FDA recommendations any product having

    Another issue that I want to raise is genetic modification. Why can't we look at transgenics more objectively without having negative feelings toward the technology before we start? Of course, researchers should first look for a solution in nature, and that's what we did, but there is no perfect solution available in nature. Although, the approaches we undertook are inspired by nature, the only way to deliver them is through biotechnology. This approach is where we are silencing the transcriptional regulator of all immunogenic prolamins. It was inspired by a mutation in a regulatory gene in barley. But, this mutation is ‘leaky'. That means it is not completely devoid of immunogenic prolamins. Similarly, the approach to express gluten-detoxifying enzymes in wheat grains was inspired by a barley enzyme that expresses during grain germination and degrades gluten proteins, along with a similar enzyme (but with complementary function) from the black mold Aspergillus, which naturally grows on bread slices. Another example is enzymes secreted by Lactobacillus species, a cocktail of acidifying and proteolytic lactic acid bacteria traditionally used for long-time fermentation by sourdough. These natural enzymes are capable of detoxifying gluten but express at a wrong time or a wrong location or are industrially inapplicable.

    We have no intention to patent the technology or the product. We will license the varieties to the Washington Grain Commission, which is a general trend at Washington State University.

    Scott Adams:
    From my perspective, your endeavor faces two big problems:
    What you end up with will be a genetically modified form of wheat, which is not allowed in Europe and other places (the list seems to be growing here);
    What you end up with will still be called wheat, and according to current laws in the USA, cannot be labeled "gluten-free." This is a huge issue that would also be true for einkorn.

    It would likely be a much easier process to further test einkorn's safety in celiac patients than to create a new genetic variant (that would really be similar to einkorn...right?), then have to go through the same process of testing.

    Sachin Rustgi:
    I agree with you on the first point, but as I mentioned previously, this debate about the so-called ‘GMOs' will settle down with time, which has happened in the past with several other technologies. For instance, people initially learned to make genetic crossbreeds in the 18th century, but the general acceptance of this technology as a breeding tool had to wait until the rediscovery of the Mendel's laws in 1900. Embracing this technology resulted in the production of hybrid maize that significantly boosted its yield.
    The second example is the reluctance to use induced mutagenesis in plant breeding, which is now well accepted and used as a standard procedure to increase genetic variability. The major advantage of shifting to this technology was the production of semi-dwarf rice and wheat cultivars that resulted in the ‘green revolution'. Similarly, the general public will accept the transgenic approach, as there is no other way to meet the growing demand for quality food. I foresee the outcome as an ‘evergreen revolution'.

    Moreover, if you look in depth at the outcome of using any of the above mentioned procedures, it is always a genetically modified organism, but it is up to us where we would like to draw a line.

    We are not in favor of releasing ‘reduced or low-toxicity' wheat lines because, as I mentioned earlier, sensitive to, at present. there is no way of telling patients which gluten protein(s) they are sensitive to. In addition, the wheat varieties are not marketed on the basis of their protein composition (however, it is possible to determine the protein profile of a wheat variety). Thus, our ultimate goal is to develop ‘celiac-safe' wheat genotypes completely devoid of immunogenic prolamins or expressing large quantities of gluten detoxifying enzymes. In the former case, the gluten-level of the wheat line is expected to be lower than or equal to the FAO recommended limit of 20 ppm, allowing its labeling as a ‘gluten-free' commodity. In the latter case, the genotype will contain the dietary enzyme supplement within its grains, and hence, will be labeled differently, and will serve as a natural dietary therapy for celiac patients. (These grains or derived flours can be blended into normal flour to bake different products).

    Two research groups, one in the US and the other in The Netherlands, are producing large quantities of these therapeutic enzymes in bacteria, and their utility as a dietary therapy for celiac patients is currently under advanced clinical trials. In this situation it will be the consumer's decision whether to get enzymes derived from a bacterium to be use as a food supplement or from wheat bread. These enzymes will not only be advantageous for celiac patients but will also prove beneficial to healthy individuals as these enzymes dramatically improve gluten digestibility and bioavailability. Healthy individuals, like celiac patients, cannot fully digest gluten proteins, but unlike celiacs, their intestines are impermeable to the undigested/partially digested gluten proteins, thus they are capable of flushing it out of their systems before it can induce an immune response. (Healthy individuals also do not carry disease predisposition alleles.) This indicates that even in healthy individuals the bioavailability of gluten proteins is low, which can be improved by feeding on these enzyme-fortified grains. It will also reduce how much must be eaten to get a similar amount of nutrition.

    As I mentioned previously, Triticum monococcum (popularly known as einkorn), is good for consumption by one group of patients, but the major difficulty is determining who can have it without causing damage to their intestines. Thus, we are continuously working toward obtaining wheat genotypes that will be safe for all celiac patients, not just for a sub-group of celiac patients. This will avoid problems with labeling and diagnosis.

    Scott Adams:
    If possible, I also just want to clarify my point #2, and get your reply to it. I believe that you said anything testing below 20ppm can be labeled "gluten-free" in the USA, but the new regulations are a bit more complicated (Open Original Shared Link):
    An ingredient that is any type of wheat, rye, barley, or crossbreeds of these grains;
    An ingredient derived from these grains and that has not been processed to remove gluten;
    An ingredient derived from these grains and that has been processed to remove gluten, if it results in the food containing 20 or more parts per million (ppm) gluten.

    I guess you could argue that the genetic modification process has removed the gluten, but this would be a legal argument that certainly isn't obvious in the new laws. For example, it is my understanding that beers which contain barley and have used an enzyme to render them gluten-free cannot, at present, be labeled "gluten-free" under the new law, even though they test below 20 ppm.

    Sachin Rustgi:
    I agree with you on the issue of labeling under the new regulations, but as you said it could be argued that these new wheat strains (devoid of immunogenic prolamins) should not be counted with the immunogenic wheat varieties, and should be classified as a new market class of wheat. The additional support for this argument comes from the preliminary feeding trials performed on transgenic gluten sensitive mice, and advanced trials performed under the NIH guidelines on existing gluten sensitive monkeys, and on interested celiac patients. Hopefully, the argument, supported by strong evidence, will foster reconsideration of the present labeling regulations.

    Scott Adams:
    What are the chances that cross-pollination of your celiac-safe variety of wheat by normal unsafe wheat will occur, and cause a percentage of the celiac-safe crop to become unsafe?

    Sachin Rustgi:
    Wheat is a strictly self-pollinated plant with a natural out-crossing rate of less than 4% in cultivated varieties (in exceptional cases up to 6.05% of out-crossing was reported). Out-crossing occurs mainly in the late emerging wheat spikes, which contribute very little to the total seed count of a plant. Moreover, wheat pollens are relatively heavier in comparison with the other grass pollens, thus could travel to a maximum of 1 m distance from the pollen source, and under optimal field conditions (20C and 60% relative humidity) can survive up to one half hour after release from anthers. Thus, if the APHIS (Animal and Plant Health Inspection Service) recommendations about isolation distances are followed a contamination of ‘celiac-safe' wheat with the ‘immunogenic' wheat can easily be avoided. In addition, a positive correlation between the rate of out-crossing in wheat and the length of flowering period was documented. Thus, the celiac-safe wheat genotypes can be selected for early and synchronously flowering phenotype to further reduce the rate of out-crossing.



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    Guest Susan Russell

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    Just to point out another problem that may be hard to control. After all the work to provide a gluten free wheat, cross contamination may occur in the harvesting and trucking of the product. Not many farmers and truckers will buy new equipment to make sure the wheat gets to market safely uncontaminated. (Storage elevators? Milling?)

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  • About Me

    Dr. Sachin Rustgi

    Dr. Sachin Rustgi, born in 1980, is currently affiliated as Assistant Research Professor with the Department of Crop & Soil Sciences at the Washington State University, Pullman. He holds a master's degree in Botany and a Ph.D. degree in Genetics and Plant Breeding. Immediately after graduating in 2006 he joined WSU, and since then, has been working there in various capacities. His research is a balance between applied and basic sciences with a focus on targeted improvement of cereals using tools of structural and functional genomics. His major research activities include development of natural dietary therapies for ‘gluten syndrome' by engineering wheat genotypes devoid of immunogenic-gluten proteins or expressing gluten-detoxifying enzymes, biomimetic engineering of wheat roots to produce an antifungal antibiotic pyrrolnitrin or an antimycotic enzyme endochitinase to armor wheat against the major root pathogens, and development of herbicide (imidazolinone) resistant barley cultivars.


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