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[SANET-MG] gm food animals part 3



gm food animals part 3
C. Outstanding Safety Issues
Food derived from genetically modified animals pose several kinds of health risks, whether heritable or not, and we do not recommend using them as food unless and until these risks have been assessed, and comprehensive studies show that they are safe beyond reasonable doubt. The health risks of food derived from genetically modified animals come from the specific proteins encoded by the transgenes, from the transgenic nucleic acids and vectors used for genetic modification, and from unintended effects of transgenesis and the cloning procedures used to produce a herd of transgenic animals, as the transgenic animals are often sterile or else do not breed true [79]. Non-heritable traits, in particular, include potent synthetic antigens for vaccination and powerful immune regulators with well-described side effects, while both heritable and non-heritable traits include growth hormones. The ingestion of foods with growth factors, vaccine antigens or immune regulators is likely to have untoward impacts on the immune system and development of human beings, especially the young. Many of the genes used to create transgenic food animals are synthetic approximations of the original gene, but deemed, mistakenly, to be “substantially equivalent” to the natural genes. The synthetic genes contain DNA sequences that have never existed in evolution, and by no stretch of the imagination can they be presumed safe. Synthetic genes are used, first of all, because bacterial genes are not readily translated in animals and plants. Bacteria use different codons for the same amino acids (codon bias), and so the gene sequence has to be modified to allow for that. Transgenes are often composites of different genes. For example, a synthetic transgene was made up of an antibacterial gene from Staphylococcus (lyphostatin) joined to a gene from a Streptococcus bacteriophage (virus of bacteria) encoding endolysin, which dissolves bacteria. The synthetic composite gene was used to modify cows, so they would produce milk that kills bacteria [80]. One main problem discussed was allergenic potential of the protein in milk. Proponents assured us that the cows modified with the synthetic gene were unlikely to be allergic to the toxin because it is a part of their genome, and thus recognized as self. But they failed to mention that children drinking the milk would not recognize the protein as ‘self’, and might well mount immune reactions against the protein, including allergy. Efforts were made to ‘humanize’ transgenic proteins by altering the genes specifying a protein’s glycosylation pattern to avoid immune reactions including allergy (allergy sites on proteins often have specific glycosylation), but that approach was only partly effective [81-83]. In view of the recent finding that a normally harmless bean protein turned into a potent immunogen when transferred to pea [82, 83] (Transgenic Pea that Made Mice Ill), there is a case for banning all GM food products until and unless they can be proven safe by adequate tests. This applies all the more so to transgenic animal food products, especially milk, which is consumed predominantly by infants and children. The profligate use of nucleic acids (RNAs and DNAs) in livestock is a source of deep concern, as it is already well known that they are to varying degrees capable of horizontal gene transfer and recombination with attendant risks of creating new viruses and bacteria that cause diseases, and of triggering cancer by integrating into genome sites that activate oncogenes as gene therapy clinical trials have made all too clear [84] (Gene Therapy Woes). Similarly, RNAi overload proved lethal to mice [56]; and it is not safe to presume that the RNAi used to modify animals will not affect those consuming the treated animals. The dangers of genetic engineering, especially the use of recombinant viral vectors and bacteria have been recognized by genetic engineers themselves before the lure of commercial exploitation swept aside these concerns [85] (Gene Technology and Gene Ecology of Infectious Diseases). We have continued to warn of the dangers of environmental releases of genetically modified nucleic acids in subsequent years, and constructs with recombination hotspots such as viral promoters [86-89] (Slipping through the regulatory net; Cauliflower Mosaic Viral Promoter - A Recipe for Disaster?; Hazards of Transgenic Plants Containing the Cauliflower Mosaic ...; CaMV 35S promoter fragmentation hotspot confirmed, and it is ...) There have been no studies addressing the unintended changes of genetic modification in transgenic animals, which may well create unexpected toxins or immunogens [79] (Fatal Flaws in Food Safety Assessment: Critique of the Joint FAO ...). Similarly, the cloning process is already known to result in unintended gross morphological as well as genetic defects [90] (What's Wrong with Assisted Reproductive Technologies?) that may compromise the safety of transgenic meat.

Comments to the Proposed Draft Guideline for the Conduct of Food Safety Assessment of Foods Derived from Recombinant - DNA Animals We comment first on matters raised by codex and then some issues not discussed in the Codex food safety assessment.

Codex QUESTIONS FOR AN EXPERT CONSULTATION

Marker and Reporter Genes

What developments have occurred in the development and use of reporter and selectable marker genes? Selectable markers are commonly employed with both heritable and non-heritable genetic modifications of animals. Common reporter genes are green fluorescent protein, beta-glucuronidase and beta-galatosidase. Selectable markers have included herbicide tolerance genes, although they are not widely used.

Are there non-antibiotic resistance marker or reporter genes that have been demonstrated to be safe to humans in food products, and if so, what are they? A far as we know, none of the non-antibiotic resistance marker or reporter genes has been demonstrated as safe to humans in food products, while at least one of them, beta-glucuronidase, was found to have amino acid similarities to known allergens [81] (Are Transgenic Proteins Allergenic?). In prokaryote vectors and in applications such as non-heritable (epigenetic) modifications or in cloning animals from modified tissue cells, antibiotic resistance markers are commonly employed. With prokaryote expression systems for producing pharmaceuticals, the most common way to achieve selection in the absence of antibiotics is via complementation of an essential gene expressed in a plasmid vector in a strain with a defect in the same essential gene. Several authors have used the dapD gene, which has a role in the lysine biosynthetic pathway and in cell wall assembly. Cobra Therapeutics proposed a very promising system, the ‘operator repressor titration for antibiotic-free plasmid maintenance’, in which plasmid loss induces the down regulation of the essential dapD gene, and thus the death of the bacteria. Other systems such as pCOR, based on the complementation of an amber mutation, have also been established. Nevertheless, the requirement for a minimal medium for culture means these systems are less likely to be used for production depending on over-expression. The various complementation-based expression systems have the common drawback of being strain dependent, as genetic knockout or modification of an essential gene is not easily transferable from one strain to another and has to be done independently [91]. A luminescence gene cassette from the Photohabdus luminescens bacterium (an insect pathogen) provides a light emitting tracer gene that can be used with both prokaryote and eukaryote organisms [92].

When removal of specific DNA sequences is desired, are reliable and safe techniques available to do this on a routine basis? There is one system, the Crelox, used almost exclusively with both prokaryotes and eukaryotes. Lox sites are signals for site-specific recombination by the Cre recombinase enzyme. A pair of lox site flanks a marker gene or any gene to be removed. The Cre recombinase is driven by a promoter designed to respond to a signal such as an antibiotic or drug. The main problem with its use in higher animals and plants is that the genomes of the higher organism contain cryptic lox sites that are recognized by Cre recombinase, causing chromosomal instability in the genome. The Cre recombinase is effectively genotoxic [93, 94] (Terminator Recombinase Does Scramble Genomes) and should not be used.

Non-heritable Applications

Are there relevant differences from a food safety perspective between animals with heritable and nonheritable traits, and if so, what are they? Our review of the non-heritable techniques includes the use of DNA plasmids and viral vectors in both vaccination and in gene therapy to improve meat production or quality. It may appear that the food safety issues of heritable transgenic traits and non-heritable traits are different. Non-heritable traits are mainly based on DNA plasmids, bacterial vectors or viral vectors that do not theoretically integrate into the germline genome, though there is always a small probability that any DNA introduced into an organism may integrate into the germline genome, as the germ cells are not separated from somatic cells by any real physiological barrier that prevents horizontal gene transfer. On account of the unjustified presumption that the foreign genetic material will not be incorporated into the germline, there is a tendency for relaxed regulation, which is equally unjustified. Many of the recombinant DNA plasmids, bacterial vectors or viral vectors have been subject to clinical trials or even approved with little fanfare and public notification. It has been presumed that the recombinant genes and their protein products are not present in the milk or meat of treated animals but there is little published information to support that assumption, and that is perhaps the main danger.

Are there specific food safety questions (e.g. with regard to types of vectors) that should be considered relative to the assessment of safety of food from animals containing heritable versus non-heritable traits? We have stressed in our review that the both heritable and non-heritable modifications pose the same kinds of risks, from the products of the transgenes, from the nucleic acids and vectors used in genetic modification, and from unexpected effects of transgenesis, and in the case of heritable modifications, from the cloning procedures. Non-heritable traits, in particular, include potent synthetic antigens for vaccination and powerful immune regulators, while both heritable and non-hertable traits include growth hormones. These contaminants in foods are likely to have adverse impacts on the immune system and development of human beings, especially the young. There does not seem to be much published information on the fate of vectors or the transgenes in the treated animals, or the potential for horizontal gene transfer and recombination to create new pathogens. RNAi overload proved lethal to mice; and it is not safe to presume that the RNAi used to modify animals will not affect those consuming the treated animals. Non-heritable genetic modifications are more threatening than heritable modifications because of its widespread use without the necessary risk assessments. It is also highly likely that meat or milk of recombinant animals will not even be labelled in the market, as they do not fall under the rubric of genetic modification. The profligate use of nucleic acids (RNAs and DNAs) in livestock is a source of deep concern, as it is already well known that they are to varying degrees capable of horizontal gene transfer and recombination with attendant risks of creating new viruses and bacteria that cause diseases, and of triggering cancer by integrating into genome sites that activate oncogenes. There have been no studies addressing the unintended changes of genetic modification in transgenic animals or of cloning, which may well create unexpected toxins or immunogens.

Substantial equivalence has no value and is misleading
The Codex Draft Guideline states:” The concept of substantial equivalence is a key step in the safety assessment process.” We take issue with that statement. “Substantial equivalence” is often used as a starting point to structure the safety assessment of a new food in the most undiscerning and reductionist way. For example, comparisons are made in the gross composition of proteins, carbohydrates and fats, or in amino acid compositions, which generally show little or no difference; and so it allows the proponent to focus on the transgene product(s) only [79]. Moreover, the comparators are completely arbitrary. Instead of comparing the transgenic variety with the variety from which it has been derived, companies have been allowed to compare the transgenic variety with the entire species, or indeed with whole category of foodstuffs from many different species, as in the case of edible oils for example. Although there have been attempts to improve on establishing substantial equivalence by incorporating profiles of total protein, metabolites and transcripts, the technical hurdles involved in comparing and interpreting patterns are insurmountable, and no official requirements are enforced. In this way, unintended, untoward effects of the modifications will not be revealed unless specific tests other than those used for establishing substantial equivalence are carried out. Examples are tests for toxicity, allergenicity and immunogenicity. Substantial equivalence therefore has nothing to say about the safety of the transgenic food product, and it would be highly misleading to assume it does.

Synthetic genes are not substantially equivalent to the natural
One important fact ignored by the Codex guidelines, which also disposes of the concept of substantial equivalence is that the recombinant animals are constructed using synthetic versions of natural genes that often involve composites of different genes, with different nucleic acid sequences as well as changes in amino acid sequence. The changes in nucleic acid sequence will lead to differences in the recognition of the gene by nucleosomes and histones. Changes in amino acids will give proteins with different conformations that would affect the proteins’ interactions with other proteins, and are likely to be regarded as foreign by the host’s immune system, as well as by humans eating the transgenic food. Furthermore, these proteins specify potent antigens, growth factors, cytokines or other signal proteins that have potent biological effects and can in no way be regarded as safe.

Transgenes exchanged between closely related species are not substantially equivalent Even when genes are transferred between closely related species, glycosylation patterns change during protein processing, and could have catastrophic consequences for the human consumer [82, 83] (as discussed in the literature review). Codex should abolish the discredited concept of substantial equivalence once and for all, in recognition that it is highly misleading when used as a key concept in safety assessment. We do not recommend using genetically modified animals and animal products as food, until and unless they can be proven to be safe by comprehensive safety evaluations, whether the genetic alterations are heritable or non-heritable.

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