Eating transgenic tobacco prevents cervical cancer

Human Papillomaviruses (HPV) are the causative agents for cervical, skin, head and neck tumors. In developing countries, cervical cancer is one of the main causes of cancer-related deaths. However, Italian researchers have now developed an immunologically active, cost-efficient vaccine against HPV. Currently, more than 150 different types of Human Papillomaviruses are known. The most commonly found HPVs in cervical carcinomas are HPV 16 and 18. The cancer causing factors of these viruses are the proteins E6 and E7, which are known as onco-proteins. Onco-proteins represent a promising target for the development of a therapeutic vaccine against HPV-associated tumors. This has been affirmed by an experiment showing that mice eating crude plant extracts containing the onco-protein E7 are partially protected against HPV-induced cancers. Unfortunately, attempts to produce large amounts of E7 protein have practically failed until now, because E7 is a short-lived protein that is rapidly degraded. Nevertheless, some E7-based HPV vaccines are currently being explored and the first promising results have been disclosed, but they still need further improvement to ensure protection. One method to produce high amounts of E7 protein in a relatively short time is by genetically engineering plants. An advantage of transgenic plant systems is that they generally lack human pathogens, oncogenic DNA sequences and endotoxins, explained Dr. Rosella Franconi from ENEA (Ente per le nuove Tecnologie, Energia e Ambiente). “This minimizes health risks and lowers the production costs.” Plants can be genetically modified by either stable or transient transformation. With stable transformation, the introduced DNA integrates in the genome of the plant, whereas with transient transformation, the introduced gene sequences do not integrate into the genome of the plant. Ideal tools for transient transformation are viruses, whose genomes contain the new gene sequences of interest. When the plant is infected with the virus, the virus spreads and replicates and the genes introduced in its genome are proliferated in high amounts. The fact that viruses are usually not transmitted by pollen – which ensures genetic contamination of wild growing plants is prevented - is a further advantage of viral systems. In 2002, Dr. Franconi and her research teams from the IRE (the Cancer Institute “Regine Elena”) and the ISS (Istituto Superiore di Sanitŕ Laboratory of Virology), in Rome, produced the HPV16 E7 protein in the cytoplasm of tobacco (Nicotiana benthamiana) plants. To do so, they used the potato virus X (PVX). PVX is a safe tool for genetic engineering, because it does not infect animals, but does effectively transfer genetic material to a variety of plant species. Mice fed with crude extracts of transgenic tobacco plants showed strong immune responses and about 40 percent of the animals were protected from HPV-induced tumors. The researchers hypothesized that the low percentage of responders could be due to the low amount of E7 antigen in the plant extract vaccine dose. Only 3 to 4 microgram per gram fresh leaf of E7 protein were produced and the quantity administered to the mice was 20-fold lower than that known to prevent tumor growth. Dr. Franconi and her team therefore wondered if they could enhance the production levels of E7 by targeting it to the plant secretory pathway, which is used by cells to transport material to the outside. The researchers targeted the protein to the secretory pathway by adding a so-called signal sequence to the protein. Signal sequences guide proteins to specific areas for accumulation, for example to the endoplasmic reticulum, where the proteins are prepared and modified. This offered a natural way to pre-concentrate the E7 protein. After examining their results, Dr. Franconi’s team found that the E7 production level was five-fold higher (15 microgram protein per gram fresh leaf) compared to the production level in the cytoplasm, Dr. Franconi previously observed (The International Journal of Immunopathology and Pharmacology, in press). “This result may be related to a positive effect on protein stability,” Dr. Franconi told Checkbiotech. In the endoplasmic reticulum, there are few enzymes that degrade proteins, but many so called chaperons that help proteins fold correctly. An additional advantage of targeting E7 proteins to the secretory pathway is that the protein purification is further simplified, because the proteins might be naturally released from the roots or leaves of the plants. Since the E7 proteins accumulated in the spaces between the cells, referred to as the apoplast, their concentration was much higher and the efficacy of the plant vaccines was improved. Mice vaccinated with these plant extracts showed a stronger immune response that more effectively protected them from tumors than mice vaccinated with plant extracts producing the E7 protein in the cytoplasm. Dr. Franconi said, “We believe that a further enhancement of this anti-tumor immunity can be achieved by a combination of E7-containing extracts with immunostimulatory genes, or by application of two different antigens.” The fact that freeze-dried N. benthamiana leaf tissues containing high amounts of E7 antigen are stable for at least one year at room temperature, further improves production and storage of the vaccine. In addition, it offers the possibility to use such plant extracts as edible vaccines. “Edible plant based vaccines offer a palatable oral delivery system without the costly purification processes required for injectable vaccines,” Dr. Franconi explained. However, a great disadvantage of oral vaccine delivery is that the digestive system could degrade the antigens. This might limit the use of oral vaccines against pathogens that can survive in the harsh environment of the digestive system. By using a plant system for antigen production, however, the vaccine antigens inside the plant cells are naturally protected by the cell wall. Dr. Franconi affirmed, “Edible vaccines are now considered to be more stable and practical in the harsh environment of the gastrointestinal tract than purified vaccine antigens.” Oral immunization with tobacco plants, however, could raise the problem of toxicity, since many tobacco cultivars produce high levels of toxic alkaloids. Varieties with only low alkaloid amounts, however, have been shown to be safe production and delivery systems for therapeutic vaccine.s Mice fed with such tobacco leaves showed no symptoms of toxicity. Dr. Franconi’s work represents a significant step toward the development of a vaccine against cancers caused by the Human Papillomavirus type 16. Since HPV16 is an enormous problem - particularly in the developing world - a vaccine against this virus needs to be highly effective, temperature-stabile and easy to administer. Furthermore, it should be possible to produce the vaccine on a large scale and cost-efficiently. Plants, as both a production and delivery system for vaccines, meet these requirements. "Source":[ http://www.checkbiotech.org/root/index.cfm?fuseaction=news&doc_id=12544&start=1&control=211&page_start=1&page_nr=101&pg=1].

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