Malaria is one of the most dangerous communicable diseases, which can affect infants, children and adults of any sex and any age group. According to a report by the World Health Organization (WHO), malaria is believed to kill a child every 30 seconds; and has affected more than 1 billion people worldwide. More than 90 per cent deaths are in Sub-Sahara Africa and Sub- Asian regions.
Female Anopheles mosquito is the carrier of malaria parasite (virus) that bites human to enter victim’s liver through blood stream, where they grow and divide. Malaria parasite that causes chronic or acute infestation of red blood cells (RBCs) belongs to the genus Plasmodium falciparum or P. vivax.
Unlike other enteric or rheumatic fever it does not cause extreme abdominal pain, pain in joints or headache but malaria fever is characterized clinically by skin rashes, high grade fever, weakness, loss of RBCs (anemia) and enlargement of spleen. One of the main effects of this parasite is to cause destruction of RBCs and organs that produce RBCs. Malaria antibodies can appear after a couple of days but may remain undetectable depending upon the extent of infection.
Several clinical assays are currently known for detection of malaria auto-antibodies but allergy remains asymptomatic until exposure to total load of Merozoites. Malaria sporozoite invades liver and escapes into blood stream to produce tons of daughter cells called merozoites. This cycle of growth continues to invade our blood stream and enters the saliva of biting mosquito to release a whole new cycle of infection. Some parasites become pre-matured in farm animals before entering human body thus causing more destruction of erythrocytes.
Department of Infections and Tropical Disease at London School of Hygiene and Tropical Medicine; and Global Clinical R&D of Glaxosmithkline Biologicals have successfully cleared clinical trials after two decades of research for a novel Virus like Particle (VLPs) vaccine to fight malaria. Currently, malaria treatment includes cheap synthetic drugs like Chloroquine.
A common practice in malaria-endemic regions is to build-up an immune response for P. falciparum by continuous exposure in small doses from infants to adults. Passive immunity in adults builds a strong antibody response against the surface proteins of infected erythrocytes. Parasite Plasmodium falciparum responsible for the most common form of malaria is observed to mutate rapidly and has developed resistance to traditional small molecule medicines like chloroquine.
This drawback of Chloroquine has become a major issue in treatment of malaria. In addition, parasite's favored mode of transport, via the Anopheles gambiae mosquito, has also developed its own resistance to insecticides. Gradual acquisition of protective immune responses from this parasite in infants and growing adults formed a conceptual framework for development of malaria vaccine by using immune responses produced by P. falciparum antigens.
GSK, one of the world’s leading pharmaceutical company, has been dedicated to development of malaria vaccine over two decades in association with Merck research laboratories, School of Bioengineering, University of Queensland, Austria, Medical Biology Centre, Belfast, UK, National institute of health, Rockville, Maryland and many other public-private sectors. VLPs has proved to become the most promising technology which uses genetic engineering and protein engineering techniques to produce recombinant fusion proteins containing antigenic signals for host cells and tissues.
These structural viral proteins are inactivated from being replicated in the host cells but capable of producing a protective immune response when a person is infected with the parasite. These structural mimics of VLP proteins make them efficient means for delivering antigenic signals to produce antibodies for generation of an immune response and fight infection causing parasite.
Bacterial artificial chromosome (BACs) and Yeast artificial chromosomes (YACs) can be selected for creation of recombinant proteins known as vectors. These vectors containing recombinant proteins are expressed in insects, plant or mammalian cells and tissues for expression of genes encoding antibodies to generate appropriate immune response for malaria.
Such physical characteristics of VLPs have been exploited for purification in bio-laboratories to produce monoclonal (Mab) and polyclonal (Pab) antibodies. US National Institute for Health and Walter Reed Army Institute of Research (WRAIR) identified a circumsporozoite protein (CSP) with likely target for immune responses with gene encoded Plasmodium falciparum for Anopheles mosquito.
Four constructs from several were chosen based on sequences of P. falciparum and expressed for pre-Clinical Grade of Mab to central Repeat Region of CSP protein. One of the grades R32tet32 was selected for clinical development using E.coli for malaria vaccine with a dose range from 10µg to 800µg. Most vaccination program utilizes three doses on periodic intervals to offer immunity for 18 months.
Many medicinal product(s) brought to the market could not inhibit parasite P. falciparum from spreading the disease. The mutations caused in the parasite have been blamed on the widespread use of choloroquine for broadly indicative symptoms like flu, fever headache and even typhoid to an extent.
Chloroquine, a cheap synthetic drug; traditionally has been used for treatment of malaria but blocks parasite’s ability to digest host’s RBCs. Chloroquine is also shown to enter the digestive vacuole of malaria parasite that cause building up of toxic units which eventually kills the parasite P. falciparum. Several point mutations in the parasite have rendered the drug useless as it alters protein that transports chloroquine into the parasite.
Many hetrologouls (hybrid) systems developed for production of proteins using Virus Like Particles (VLPs) have not been possible to grow. Attenuated viruses in cell culture systems on large scale are still under research phase. Most VLPs generated a means to understanding the assembly or structure of viruses in spite of limited success of VLP production as a candidate vaccine. Virus like Particles (VLPs) is identified for delivery of antigens that tend to be recombinant fusion proteins consists antigenic signals fused to structural viral proteins (hybrid system).
VLPs spontaneously form into particles that are structurally similar to authentic viruses thereby making them efficient means in delivering antigenic signals to the immune system. VLPs are highly effective type of subunit vaccine, which mimic the structure of overall virus particle without the requirement of containing infectious genetic material as most of them completely lack DNA or RNA genome.
VLP preparations are based on the observation of the caspid protein that leads to spontaneous assembly of particles structurally similar to authentic viruses. These indicate the fact that lower doses of antigen relative to subunit vaccine might be capable of eliciting same protective response. For malaria treatment and vaccination programs, many and varied options are available to produce a substance using novel virus like particles (VLPs).
The active ingredient Artemisinin extracted and developed as Anti-malaria drug is associated with Chinese worm wood plant Artemisia annua. This product has a mode of action that pumps free radicals into the mosquito and by-passes resistances that are built up by other compounds for the parasite, thus called Artemisinin combitorial therapies (ACTs). Possible use of VLPs in delivering therapeutic agents is by systemic injection or oral administration.
Foreign proteins can be successfully expressed within target cells (Liver or intestinal mucosa) for in vivo applications to alter the life cycle of malaria. This approach is shown to have certain bioprocessing complexities, as modification of VLP greatly alter levels of production, size and product specific properties. For culturing large quantities, alternative source of nutrients becomes a necessity as development and formulation of culture medium is one of the most diverse steps in fermentation technology. Various sources of carbon, nitrogen and vitamins are necessary and no hard and fast rule applies in separation of properties; usually possible with complex materials such as yeast.Unicellular organism grown on defined media gives complete control over environmental parameters that can be tracked to classical genetic techniques and functions. In spite of being the most promising technologies currently in clinical trials for the delivery of these antigens, VLPs associate some clinical side effects and bioprocessing issues in manufacturing. There are currently a number of VLP-based malaria vaccines in clinical trials.
The current market demand for VLP is primarily as products for vaccination. In case for malaria vaccines the market according to Malaria Atlas Project (MAP) is 2.37 billion people that are at the risk of contracting malaria from Plasmodium falciparum and about one billion live under lower risk of infection. After success of malaria vaccine through VLP expression system, many scientists across the globe are using this technology to explore possibilities for other diseases like influenza, flu, alzheimer’s disease, chikungunya, cancer and tuberculosis.
(The author is MD of VMG Biotech Consultants, New Delhi, a premier biotechnology consultancy and Contract Research Organization.)