Open Access
Human papillomavirus L1 protein expressed in tobacco chloroplasts self-assembles into virus-like particles that are highly immunogenic
(Wiley, 2008) Fernández San Millán, Alicia; Martín Ortigosa, Susana; Hervás Stubbs, Sandra; Corral-Martínez, Patricia; Seguí-Simarro, José M.; Gaétan, Julien; Coursaget, Pierre; Veramendi Charola, Jon; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
Cervical cancer is the second most prevalent cancer in women worldwide. It is linked to infection with human papillomavirus (HPV). As the virus cannot be propagated in culture, vaccines based on virus‐like particles have been developed and recently marketed. However, their high costs constitute an important drawback for widespread use in developing countries, where the incidence of cervical cancer is highest. In a search for alternative production systems, the major structural protein of the HPV‐16 capsid, L1, was expressed in tobacco chloroplasts. A very high yield of production was achieved in mature plants (~3 mg L1/g fresh weight; equivalent to 24% of total soluble protein). This is the highest expression level of HPV L1 protein reported in plants. A single mature plant synthesized ~240 mg of L1. The chloroplast‐derived L1 protein displayed conformation‐specific epitopes and assembled into virus‐like particles, visible by transmission electron microscopy. Furthermore, leaf protein extracts from L1 transgenic plants were highly immunogenic in mice after intraperitoneal injection, and neutralizing antibodies were detected. Taken together, these results predict a promising future for the development of a plant‐based vaccine against HPV.
Open Access
Producción de albúmina humana en cloroplastos transgénicos de tabaco
(2001) Fernández San Millán, Alicia; Mingo Castel, Ángel; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
Open Access
Antibiotic-free chloroplast genetic engineering: an environmentally friendly approach
(Cell Press, 2001) Daniell, Henry; Wiebe, Peter O.; Fernández San Millán, Alicia; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
Open Access
Expression of recombinant proteins lacking methionine as N-terminal amino acid in plastids: human serum albumin as a case study
(Elsevier, 2007) Fernández San Millán, Alicia; Farrán Blanch, Inmaculada; Molina Azcona, Andrea; Mingo Castel, Ángel; Veramendi Charola, Jon; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
Open Access
Chloroplast transgenic approach for the production of antibodies, biopharmaceuticals and eddible vaccines
(CSIRO Publishing, 2001) Daniell, Henry; Dhingra, A.; Fernández San Millán, Alicia; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
Open Access
Transformación nuclear vs cloroplástica para la producción de albúmina humana (HSA)
(SECIVTV, 2003) Farrán Blanch, Inmaculada; Fernández San Millán, Alicia; Mingo Castel, Ángel; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
Open Access
An in vitro tuberization bioassay to assess maturity class of new potato clones
(Taylor and Francis, 2000) Veramendi Charola, Jon; Sota, V.; Fernández San Millán, Alicia; Villafranca Rodríguez, María José; Martín-Closas, L.; Pelacho, A.M.; Mingo Castel, Ángel; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
Open Access
A chloroplast transgenic approach to hyper-express and purify human serum albumin, a protein highly susceptible to proteolytic degradation
(Wiley / Blackwell, 2003) Fernández San Millán, Alicia; Mingo Castel, Ángel; Miller, Michael; Daniell, Henry; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua; Gobierno de Navarra / Nafarroako Gobernua
Human Serum Albumin (HSA) accounts for 60% of the total protein in blood serum and it is the most widely used intravenous protein in a number of human therapies. HSA, however, is currently extracted only from blood because of a lack of commercially feasible recombinant expression systems. HSA is highly susceptible to proteolytic degradation in recombinant systems and is expensive to purify. Expression of HSA in transgenic chloroplasts using Shine-Dalgarno sequence (SD), which usually facilitates hyper-expression of transgenes, resulted only in 0.02% HSA in total protein (tp). Modification of HSA regulatory sequences using chloroplast untranslated regions (UTRs) resulted in hyper-expression of HSA (up to 11.1% tp), compensating for excessive proteolytic degradation. This is the highest expression of a pharmaceutical protein in transgenic plants and 500-fold greater than previous reports on HSA expression in transgenic leaves. Electron micrographs of immunogold labelled transgenic chloroplasts revealed HSA inclusion bodies, which provided a simple method for purification from other cellular proteins. HSA inclusion bodies could be readily solubilized to obtain a monomeric form using appropriate reagents. The regulatory elements used in this study should serve as a model system for enhancing expression of foreign proteins that are highly susceptible to proteolytic degradation and provide advantages in purification, when inclusion bodies are formed.
Open Access
Plant-based antibodies and virus-like particles: a leap towards new therapeutic development
(Nova Science Publishers, 2008) Obregón, Patricia; Fernández San Millán, Alicia; Veramendi Charola, Jon; Agronomía, Biotecnología y Alimentación; Agronomia, Bioteknologia eta Elikadura; Institute for Multidisciplinary Research in Applied Biology - IMAB
The generation of therapeutic antibodies and fusion proteins for medical application is one of the fastest growing areas of the pharmaceutical industry with more than 150 therapeutic antibodies and fusion proteins currently either in clinical trial or use. At the same time, the use of virus-like particles has become an interesting tool in the fight against viral infections. Thus, some devastating high-incidence diseases such as HIV or cancer are currently chosen as clear targets for this type of therapeutical strategy. However, the high production cost of the current manufacturing systems of these molecules is a latent hurdle to overcome. With the advent of biotechnology, transgenic plants have emerged as a more economical new strategy for recombinant protein production. Antibodies and virus-like particles have been demonstrated to be well expressed in plants. In addition, the achieved protein expression level of most of them in the plant system has been reported to be compatible with that established for commercial viability. These facts make the use of plants for the generation of these types of recombinant molecules a very promising strategy to the development of lower cost biopharmaceuticals. In consequence, it could lead to exert important economical and medical implications as being affordable for developing countries where the incidence of infectious diseases is the highest. The development and production of these therapeutic molecules in plants is reviewed in this chapter, and the medical implications, advantages and limitations of both the plant-system and plant-derived molecules for practical use are discussed.