30 Bioprospecting

Dr. Felix Bast

1. Learning outcomes

1.1 To learn the definition and concept of bioprospecting

1.2 To know how sustainable bioprospecting can be an effective strategy to fund biological conservation initiatives

1.3 To learn about various legislations and acts for ethical bioprospecting.

1.4 To learn about current challenges future perspectives in bioprospecting

2. Concept Map

3. Description

3.1 Introduction

Biodiversity prospecting, or bioprospecting, is the systematic and sustainable exploration of biochemical and genetic resources from the nature, and its further development into valuable products. Planet Earth harbours diversity which is currently estimated to be around 10 million species of plants, microbes and animals; these organisms and its genetic and biochemical constitution had been shaped by the process of evolution by natural selection for the last 3.5 billion years. Chemical tapestry of living beings are immensely complex and a vast majority of biologically active natural products are impossible to be chemically synthesized. Organisms harbours a rich panoply of biochemical and genetic resources that could potentially of immense use to the humanity. It is estimated that more than 90 percentage of drugs currently in world’s market came directly from organisms, especially plants. Recent advances in recombinant DNA technology enables us to explore wild genes from the organisms and cloning them to other organisms for various benefits. However, bioprospecting is an immensely time consuming endeavour. As per the current estimates, only one in 250000 natural samples would yield a commercially viable drug, and the development would take decades to complete.

Under the umbrella term Bioprospecting all efforts of utilization of biodiversity for the benefit of human beings is covered (see the concept map). This include development of drugs either directly from the natural products, or the development from traditional medicinal candidates through in-vitro screening of biological activities or through animal testing, development of natural products into industrial products such as dyes, anti fouling compounds, paints, glues, cosmetics and so on, development of natural products to supplement agri-food industry through its direct use as food, food additives, dietary supplements, fodder and fertilizer, development of natural products for biofuel as well as for carbon capture and sequestration (CCS), exploration and utilization of biodiversity for aesthetics, tourism and horticulture, and exploration of biodiversity for wild genes, its cloning and further development of commercial products or Genetically Modified Organisms.

3.2 Ethics of bioprospecting

Ethics of the exploration of natural resources is indeed very complex. Many stakeholders would claim the resources as their indigenous, cultural, national and other privileges and the benefits arising if any would need to be shared in a fair manner. Historically, the explorers and investors for bioprospecting hailed from developed countries and much of the bioprospecting had been happening in the developing countries. Reason for this trend is very clear; much of the world’s biodiversity is concentrated in tropics due to a number of factors, the most obvious among them is ‘species-energy relationship’- tropics get the maximum solar irradiation and subsequently this region supports maximum biodiversity, including human population. Owing to higher human population and higher human population density, the countries in the tropics have remained poorer than scarcely populated non-tropics. A number of biodiversity hotspots have been described across the world-regions with extraordinary species richness and endemism (when some species are found at these locations that are not found elsewhere in the world), and almost the entire of these hotspots are situated in developing countries in the tropics. Some of the noteworthy countries that have historically been target of bioprospecting include Peru, Costa Rica, Brazil, Jamaica, Kenya, Madagascar and India. World’s ten megadiverse hotspots are presented in Table 1.

Table 1. Ten of the world’s megadiverse hotspots. Adapted from Mittermeier et al, 1999.

The explorers and the downstream bioprospecting industries from Europe had, until very recently, been in fact exploiting the tropical countries with virtually no benefit sharing. One example is the development of anticancer drugs vincristine and vinblastine. These drugs were discovered by scientists from American pharmaceutical company Eli Lilly and University of Western Ontario, Canada respectively, in 1960s from the plant Rosy Periwinkle, native to Madagascar. Until the discovery of these alkaloids, the plant had been part of the traditional folk medicine of Madagascar for the treatment of diabetes. The bio prospectors extracted these vinca alkaloids that later turned out to be commercially successful chemotherapeutic drugs. However, neither the American company nor the Canadian University shared anny of the generated revenue with Madagascar.

Another example is when American corporation RiceTec attempted to patent certain varieties of long-grain Indian rice varieties including Basmati rice. Since then, thanks to the intervention of Indian Government, most of the claims of patent were invalidated and international patents of this firm got revoked. Yet another example is when American pharmaceutical firm WR Grace and Company applied for potent anti-fungal drugs extracted from neem tree. In fact, neem tree extracts had traditionally been used for a wide variety of diseases as part of Ayurveda in India, and a number of groups opposed that patent application. Finally, in 2005, WR Grace and Company lost the final appeal for the rejection of the patent and the issue was resolved.

Instead of randomly screening for potent biological activity of natural products, many of the bioprospecting companies and research groups target traditional and folk medicines for the samples, as chances are very high that these ethnobotanical remedies might harbour biologically active compounds, selected by hundreds of years of trial-and-error scenario of folk medicine. The bioprospecting firms usually negotiate the rates with the indegenous communities for per sample rates for the sample acquisition of these traditional medicines and herbal remedies, with no guaranteed downstream royalties or any further benefit sharing. Over exploitation of a number of medicinal plants, unfortunately, lead to the extinctions of a number of valuable species. However, under privileged countries of the tropics could not able to fund conservation drives, as they had a number of other imminent issues that need to be prioritized. Environmental conservationists proposed bioprospecting, if fair benefit sharing is ensured, as a potential revenue generator for the implementation of conservation measures in tropics.

3.3 Governmental legislations

There had been no inter-governmental legislations or legal frameworks until 1992 Earth Summit in Rio de Jeneiro, Brazil when United Nations adopted Convention of Biological Diversity, CBD that ensures fair benefit sharing between all the parties involved with the bioprospecting. There were in total 196 countries and European Union (notable exception is USA) ratified the CBD. The CBD made it compulsory to obtain Prior Informed Consents (PICs) from the party providing biological resources, to protect indigenous and traditional knowledge of the communities involved. The convention also made agreements for technology transfer, impact assessment, technical and scientific corporation, education and public awareness and provision for financial resources. The treaty was latter modified to include transparent legal frameworks for equitable benefit sharing in the meeting held at Nagoya, Japan in 2010, a supplementary agreement commonly known as Nagoya Protocol. The protocol entered into force in 2014 and had been ratified by 96 parties including European Union till date (exceptions include US, Canada and Russia). These protocols made it compulsory to have Prior Informed Consents (PIC) from the supplier of bioresource (for example, plant samples) before collecting the biological materials. Implementation of this legislation by the ratified countries dramatically reduced illegal accession of biological materials, a term known as biopiracy. Post CBD, such cases of suspected biopiracy could be trialled at International Court of Justice.

The consequences of these two legislations were in fact mixed; while it constituted the first international legal framework for benefit sharing, the framework resulted in huge bureaucratic huddles to an extent that the pharmaceutical companies have rather started turning to combinatorial synthetic chemistry rather than bioprospecting for the search of new drugs.

3.4 The scope of bioprospecting

Bioprospecting had been widely considered as a value creator for the global biodiversity resources, and the revenue generated can be effectively routed to fund conservation programmes of the poor third world countries of the world. One key issue in this regard is the value of biological resources and the way to quantify it. Of course, no unambiguous way exists to quantify the value of the bioresources and estimates have largely been subjective judgements. The estimate ranges between USD 21 per hectare of land to USD 9177 per hectare. A reason for this disparity is that the latter estimate was based upon the data from biodiversity hotspots- epicentres of biodiversity with unusual species richness and endemism. More commonly, companies negotiate for per sample price with the supplying countries. In the early 1990s Merck entered a deal with Costa Rican government for USD 1 million for a certain number of samples. In 2007, GlaxoSmith Kline signed a pact with Brazilian firm for the collection of 30,000 samples (that would translate to approximately USD 100 per sample). Novartis had an agreement with Brazilian government for USD 4 million for 10,000 samples. However, both of these agreements had to be cancelled as post Nagoya protocol these agreements attracted a huge political controversy in Brazil.

Classically, bioprospecting pharmaceutical firms target traditional/folk alternative medicines, as these herbal formulations might contain active biochemical. Reason for this is obvious; after tens of thousands of years of human evolution, we have identified and selected useful plants and employed them in traditional medicine. Use of traditional biological materials for modern medicine is called ethnomedicine or ethnopharmacology. Study of a particular region’s plants for its human use through traditional knowledge of indigenous people is called ethnobotany. There are numerous examples; the drug quinine is developed from cinchona tree used in folk medicine of Peru, Artemisinin and its derivatives were isolated from traditional Chinese medicines, and alkaloids morphine and codine from opium poppy used in various indigenous medicines worldwide. The compound Podophyllotoxin isolated from Podophyllum peltatum– a herb used in indigenous tribes of North America for treating warts- had been developed into anticancer drug etoposide. To overcome a number of obstacles during rigorous drug development and formal approval processes, herbal formulations are often developed into polymolecular botanical supplements, functional foods and neutraceuticals; an example is “BGR-34”, a purportedly antidiabetic formulation developed by CSIR, India.

While most of the bioprospecting is targeted towards traditional/folk medicine to improve chances of success, a few goes for random screening of biological resources, albeit the procedure is extremely time consuming. A famous example is US National Cancer Institute’s large-scale natural product screening for anticancer activities. The programme screened extracts of over 70000 plant species over the course of around 2 decades, and resulted in the development of just one drug, paclitaxel-a commercially successful chemotherapeutic agent and a blockbuster drug. On the other extreme, a dramatic example is when a Swiss microbiologist, on a family trip to Norway, collected a bit of soil and brought back to his lab in Switzerland. Further research lead to the development of commercially successful immunosuppressant cyclosporine-A. This blockbuster drug generates revenue of approximately USD 1 billion a year. As no agreements for fair trade existed between Switzerland and Norway, Norway lost its share in the profit of this blockbuster drug entirely to Switzerland. Another example of lucky finds include development of antibacterial drug rapamycin from soil samples from Easter Islands. Soil sample from Himalayas collected by a team led by Dr. Javed Agrewala of CSIR- IMTech revealed its potent immunosuppressive and anti-bacterial properties. Further research lead to the development of Caerulomycin A, a pyridine-derived broad-spectrum antiobiotic and immunosuppressant. Nostrum Pharmaceutical reportedly have brought this technology for USD 3 million, and Dr. Javed Aggrewala was honoured with prestigious Bhatnagar Award for his contribution by the Indian Government.

A major bias in play in global bioprospecting arena is that most of the explorations were targeted to that of the landmasses of the planet. However, more than 70% of earth’s surface is covered with the ocean and it harbours rich biodiversity. Only a very limited number of companies (for example, Diversa Corporation and Magellan BioScience, US and MarBank in Norway) are involved with bioprospecting of marine biological resources. Oceanic bioprospecting has its own dramatic success stories. One example is development of industrial enzyme Ultra-Thin from the genes isolated from archaebacterial living in deep-sea hydrothermal vent in Pacific Ocean. Gene coding for Alkaline Phosphatase enzyme collected from an oceanic Shrimp species, which later developed as a successful PCR Post reaction clean-up tool (Exo-SAP-IT). Heat labile variant of the same enzyme, Antarctic Phosphatase, had been originally isolated from Antarctic bio resource. Salinosporamide-A from marine bacterium (Salinospora) is a protease inhibitor and a potential anticancer therapeutical currently in early-stage clinical trials. Ecteinascidin 743 (trabectedin) is an approved anticancer drug developed from a marine tunicate sea squirt Ecteinascidia turbinata. Luminase pulp bleaching enzyme was originally collected from the genes of geothermal sediment samples from Kamchatka, Russia. At present, the world has only 5 manned submersible suitable for deep-sea diving for more than 3000 meters, a major huddle for the bioprospecting of world’s oceans. Bioprospecting of open sea is also extremely rare; an estimate suggest that 99% of all ocean exploration for bioprospecting happens within territorial waters (part of the ocean that lies within 12 nautical miles or 22.5 km from the coast).

3.5 Current status of bioprospecting

Owing to a number of issues, bioprospecting has come to a dramatic slowdown over the last two decades. The main reason for the current slowdown is increase in the bureaucratic huddles post the enforcement of various international agreements, especially CBD and Nagoya Protocol. Prior to these landmark agreements, bioprospectors from the developed countries would undertake explorations to the poor tropics and they would negotiate with the indigenous communities or local governments for the rates for per sample acquisition. The samples, thus acquired after relatively less efforts, would be brought back to their established labs and subsequently be used for various bioactivity screening, and ultimately, if successful, to the development of drugs. Post CBD and Nagoya protocols, however, sample acquisition became such a major huddle for the bioprospecting firms. It has increasingly became an impossible affair to acquire even a single sample from certain countries like Kenya that forbids all bioprospecting activities for the foreign companies. Another reason is the bioprospecting being extremely slow process that demand gargantuan efforts. Most of the world’s traditional medicines are now known to the scientists, and the bioprospecting efforts to develop drugs from these ethnobotanicals, although straightforward and heuristically advantageous option, have now became saturated. Random screening of natural products is indeed extremely labour intensive and time consuming. It is now becoming increasingly clear that bioprospecting is highly unlikely to become an economic driver to support biodiversity conservation measures by the poor country. Drug discovery companies these days increasingly rely on combinatorial chemistry opposed to natural products for the source of new drugs. This approach works first by identifying potential targets of specific diseases. For example, an enzyme involved in the development of cancer, that need to be inhibited. To inhibit an enzyme, its active site need to be blocked with an inhibitor molecule, that can perfectly bind with the site. As the three dimensional structure of this enzyme became available through analytical chemistry techniques such as NMR (Nuclear Magnetic Resonance) spectroscopy, a synthetic organic chemist can design an optimal molecule that can perfectly bind this target (its active site), like chiselling a perfect key to the lock. Synthesis of chemicals starting from scratch is called ‘total synthesis’. This combinatorial chemistry approach involves no natural products, and it is far straightforward and less laborious in comparison with traditional bioprospecting and screening approach. However, synthetic chemistry has its own limitations, and only small, structurally simple molecules can be synthesized as of today. As the natural products have been evolving through natural selection acting on them for billions of years, a richer and complex repertoire of structurally diverse compounds can be found in nature that are almost impossible to synthesize using synthetic chemistry approaches. However, with the advent of progress in structural chemistry, increasingly complex molecules are now being synthesized in world’s laboratories; for instance, taxol and cyclic depsipeptide.

One intuitive approach that combines the elements of bioprospecting and combinatorial chemistry approaches is that instead of screening the natural product extracts and purified compounds for various bioactivities, scientists construct a systematic, annotated and curated computational database of the structures of purified natural products. One such database is InterBioscreen (IBS, https://www.ibscreen.com/), perhaps the most extensive database of natural products with more than 50,000 accessions freely available to anyone. Other noteworthy databases of natural products include SuperNatural (http://bioinformatics.charite.de/supernatural/) and Drug Discovery Portal of University of Strathclyde, UK (www.ddp.strath.ac.uk). These database can be searched to find potential structural matches, (for example, an inhibitor molecule that perfectly bind with the three-dimensional structure of the enzyme that need to be targeted) for the biological pathways- a process known as virtual or in silico screening through molecular docking. For example, using the IBS, investors Singh and Bast, 2015) were able to identify natural products Myricetin and Epigallocatechol- two compounds from green tea- as potential inhibitors for a crucial pathway in the progression of many cancers, STAT-3 (Signal Transducer and Activator of Transcription-3).

4. Summary

4.1 Biodiversity prospecting or bioprospecting is an umbrella term that is used to refer the sustainable utilization of biodiversity for the betterment of human life. While the term is used most often to refer drug discovery using natural products, it also include all other utilizations including that for industrial applications, to support agri-food industry, for biofuel and carbon capture and sequestration, for aesthetical uses and also for the exploration of genes.

4.2 Two of the most important international legislations that govern sustainable biodiversity worldwide are CBD (UN Convention of Bio Diversity) and Nagoya Protocol. These agreements are intended as a legal framework for equitable benefit sharing and prior informed consent of resource provider. However, these legal frameworks have proven itself to be an impediment for the large-scale bioprospecting missions, through the resultant bureaucratic huddles.

4.3 A major bias in bioprospecting had been lesser exploration of world’s marine bioresurces, even through oceans cover more than 70 percent of earth’s surface. Notwithstanding minimal explorations towards deep oceans, a number of drugs and commercial products have been marketed, warranting further developments in this sector.

4.4 Bioprospecting had classically been projected as an economic driver for supporting conservation schemes of poor countries. However, post CBD and Nagoya Protocol implementation, bioprospecting had rather dramatically slowed down. At present vast majority of pharmacological industries resort to combinatorial chemistry assisted synthetic approach for the drug discovery. A novel two-step approach involving structural database of natural products and in-silico docking (screening) studies with potential targets of biological pathways are increasingly getting popular.

Cited References

Mittermeier, R.A., N. Myers, P. Robles-Gil, & C.G. Mittermeier (Eds.). (1999). Hotspots.
Earth’s Biologically Richest and Most Endangered Terrestrial Ecoregions.
CEMEX/Agrupación Sierra Madre, Mexico City.
Singh, P. & Bast, F. (2015). High-throughput virtual screening, identification and in vitro biological evaluation of novel inhibitors of Signal Transducer and Activator of Transcription 3. Medicinal Chemistry Research 24 (6) 2694-2708