If everything goes to plan with the researchers at the São Paulo company Nanocore Biotecnologia, it should place on the market, within a three-year period, the first immunotherapeutic medicine made from recombinant DNA entirely developed in Brazil.. To achieve this goal, the company, with its head office in the city of Campinas, needs to build an ultra-clean certified area for the manufacture of the product and to root for the clinical tests with the drug, already underway, achieving the longed for results. “We’re highly optimistic” says the pharmacist from Minas Gerais, José Maciel Rodrigues Júnior, the executive director at Nanocore. “We’ve already been technologically very successful in the stages of production, purification and quality control. The DNA immunotherapy is being used in a clinical study with patients suffering from cancer of the head and neck and we’ve already managed a single dose formula.”
Medicines made from recombinant DNA, also known as genetic vaccines or plasmid DNA, are a new hope for various illnesses that have no treatment or as more efficient prophylactic methods. Contrary to the majority of traditional vaccines, which are developed from microorganisms – a virus or bacterium – that cause the illness, used in an attenuated or inactive form, therefore without pathogenic activity, the DNA immunotherapies base themselves only on a fragment – normally a protein – of the microorganism that unchains in the human organism the protective immunological response against such an ailment. Another difference is that, in the case of genetic vaccines, the individual does not receive a readymade vaccine by only the production messenger. Thus the DNA itself goes on to produce the vaccine in the organism. These medicines demonstrate the potential for improving the organism´s response against determined infections and against some types of tumors.
In order to have success with this therapy, the production of genetic material and recombinant protein with a degree of purity that attends to the demands of the regulating organs, such as the National Health Surveillance Agency (ANVISA) in Brazil and the Food and Drug Administration (FDA), of the United States, responsible for the analysis and liberation of medicines in that country, are essential. Throughout the world, various research groups are attempting to make these types of treatment viable. “There are at least 300 ongoing clinical studies” informs Maciel. But, according to the researcher, except for a DNA vaccine of animal use to protect horses from the West Nile fever virus, no other vaccine has been commercially launched.
Besides the risks that involve treatments that use drugs formulated from genetic manipulation – in the past, some patients submitted to experimental treatment with different modalities of genetic therapy died or developed cancer ” the difficulty in transforming this treatment into a reality resides in the complexity of its development. The first step consists of the identification of the microorganism´s protein that causes the illness or a tumor, which can serve as a target against which the vaccination will induce immunological protection. This protein, known as an antigen, is a molecule that brings into action the responses of the immunological system. Next, the researchers need to find, by way of genetic sequencing process, the gene that codifies this protein.
“Once the gene has been identified, we need to clone it in order to multiply the number of its copies. For this to happen, we insert the gene into a plasmodium, a DNA fragment in the shape of a ring. This plasmodium is placed in a non-pathogenic bacterium that, for its part, is inserted into a culture medium. Through a fermentation process, these bacteria multiply and the same thing occurs with the plasmids, thus generating a large quantity of copies of the gene that will be used in the vaccine” explains the biomedical doctor Sandra Aparecida dos Santos, responsible for the Biotechnological Division at the company Nanocore. The final stage of the process is purification, which is done with the breaking away of the bacteria and the isolation of the plasmid. Then it is placed in micro-spheres, with diameter higher than 1 micrometer (1 millimeter divided by a thousand), made from a bio-polymer that slowly degrades itself within the human organism. These micro-spheres, also manufactured by Nanocore, are made up into a matrix system, in which the DNA is found internally dispersed, and they are demonstrating a wide application in the field of immunization and of the controlled liberation of medicines into the human organism.
Clinical business
Located at the Techno Park Campinas, an entrepreneurial condominium alongside the Anhangüera highway, 100 kilometers from the city of São Paulo, the company Nanocore, founded some three years ago, is receiving financial assistance through the Small Business Innovation Research (PIPE) program and has already registered a patent for the formulation of the medicine. The company now hopes to close a business deal with investors in order to give continuance to the clinical studies on humans and to initiate the production of the vaccine on a large scale. It will need to enlarge its installations and to construct a certified area with the so called GMP (good manufacturing practices) conditions. The location needs to have an extremely aseptic environment, with fewer particles in the air than a surgery room, to have rigid control of the production flow and to be endowed with an adequate effluent outlet system. In the opinion of Maciel, advanced negotiations are under way with the National Economic and Social Development Bank (BNDES) in order to obtain financing for the project, budgeted at a few million reais.
Another challenge for the concrete realization of the first national recombinant DNA vaccine is exactly the success of the clinical trials. The vaccine was used in a clinical study for safety evaluation on 22 terminal patients suffering from cancer of the head and neck. The test, concluded in October of last year, was conducted by researchers from the Medical School of the University of São Paulo (USP). “Because of a confidentiality agreement, the results can’t, for now, be disclosed, but I can advance that they were promising” says Maciel, an ex-professor from the Federal University of Minas Gerais (UFMG) and the coordinator of Transfer Coordination and Technological Innovation of that university between 1998 and 2001.
Against the tick
As well as the genetic vaccine for immunotherapy treatment, the Nanocore company is working on other biotechnology research lines. ´since we already dominate the recombinant protein engineering technology, we decided to dedicate ourselves to the development of other bio-products, such as the Follicle Stimulating Hormone (FSH) for human and animal use, a diagnostic kit for hantavirus, the production of analogues of amylen – a substance secreted by the same glands that produce insulin in the pancreas ” used in the treatment of diabetes, and a vaccine to control ticks. This last mentioned is at an advanced stage and, if everything goes well, should be on the market next year” says Maciel. These projects are being developed in collaboration with professors Mari Sogayar and Luiz Tadeu Figueiredo, both from USP.
Similar vaccines already exist on the market for the control of the tick, Boophilus microplus, but, according to Maciel, they are problematic because they need to be administered in three to five doses in the first year of vaccination with a reinforcing dose every six months, bringing about a serious management problem. The Nanocore vaccine will be applied in a single dose “Another problem is that the existing vaccines don’t give effective protection to the national herds, since there exist different types of ticks in the world and the commercial vaccines are produced starting from a species of tick that is not predominant in Brazil” emphasizes the pharmacist Karla de Melo Lima, responsible for the Analytical Development Division at Nanocore. “We’re working on the production of the same protein used in foreign DNA vaccines, but cloned starting from Brazilian species of ticks” she says.
The commercially available vaccines against the tick are based on a recombinant protein, namely Bm86 – an antigen found in the tick´s intestinal membrane ” which remains inaccessible to the bovine immune system, not allowing for, in a natural manner, the development of immunity. The antibodies against this antigen in the vaccinated animals, together with other plasma components, are taken in by the tick with the blood, making it possible that the specifically formed antibodies unite to the intestinal protein, bringing about damage in the tick´s intestine. The final goal of the vaccination is not the direct death of this arachnoid, but the progressive control of the number of these small animals in successive generations, by way of the reduction of their reproductive capacity.”We don’t want to exterminate the tick because, indirectly, it´s responsible for conferring upon the cattle immunity towards other illnesses. The ideal is to maintain a minimal level of infestation that doesn’t cause any economic losses” says Karla.
The conclusion of the study on the efficiency of the vaccine, which is being developed in partnership with the Brazilian Agricultural Research Corporation (Embrapa) Beef Cattle Unit and with the UFMG, should occur by the end of this year. The field tests are being carried out on around 100 animals at different ranches. “This is the final stage with the objective of registering the vaccine with the Ministry of Agriculture. Our idea is to begin to produce it next year” says Maciel. The development of the product is going to fall into three phases. First the single dose vaccine will be made using foreign strains of tick. Starting from next year, the Nanocore company´s intention is to go on to use Brazilian strains. “In a third phase, we want to associate a second antigen to the vaccine, which will also be effective against the tick´s larval phase. The current antigen only acts during the adult phase” clarifies Karla. “We want the vaccine to impede the fixation of the larvae, not allowing the tick to complete its life cycle and to reproduce, generating other descendants.” Should the research at Nanocore prove to be successful, the Brazilian cattle breeders, will gain a strong ally in the fight to combat an infestation that, it is estimated, annually causes economic losses of US$ 1 billion to the country.
The Projects
1. The production of purified plasmid DNA and recombinant proteins on a large scale, for use in vaccines and diagnostic kits. (nº 01/08334-8); Modality Small Business Innovation Research (PIPE) Program; Coordinator José Maciel Rodrigues Júnior – Nanocore; Investment R$ 363,454.41 and US$ 21,000.00 (FAPESP)
2. Single vaccine dose against the bovine tick (nº 03/13390-0); Modality Small Business Innovation Research (PIPE) Program; Coordinator José Maciel Rodrigues Júnior – Nanocore; Investment R$ 469,080.00 and US$ 12,000.00 (FAPESP)
