Bioprospecting D 3.3 65 Drug discovery period Drug development period Drug marketing Line expansion period Decision to seek new agent for therapy Chemicals and natural products tested Lead identified Candidate chosen and confirmation tests run IND filed Clinical studies initiated NDA prepared and submitted NDA approved Drug launched Post-marketing surveillance initiated New clinical indications pursued Marketing support Chemical synthesis initiated, natural source products identified Analogs and derivatives tested IND plan established and initiated New dosage forms and formulations developed Clinical Phases I, II, III Phase IV Figure D 3.3-2 Pipeline concept of drug development. Explanation in text and in Table D 3.3-1. IND = Investigational New Drug Application, NDA = New Drug Application Source: modified on the basis of McChesney, 1996 Table D 3.3-1 Periods of time typically required for the development of new drugs. The figures relate to the development stages outlined in Fig. D 3.3-2. Source: McChesney, 1996 Stages of Time Average Direct clinical development frame period costs [years] [years] [1,000 US-$] 1. Project design up to IND development 0.5–2.5 1.5 400 2. Clinical Phase I 0.5–1.5 1.0 2,600 3. Clinical Phase II 1.0–5.0 2.5 11,400 4. Clinical Phase III 1.0–5.0 2.5 7,400 FDA testing ofthe NDA 1.0–5.0 2.5 2,200 Total development 4.0–19.0 10.0 24,000 basis, these amounts are 10–100 times higher. Collecting wild plants is often not justifiable ecologically, quite apart from the fact that there are often considerable fluctuations of substance content within a wild population (McChesney, 1996). One alternative is to produce medications by cultivating wild plants. Worldwide so far around 3,000 wild plant species have become established as cultivated plants and time periods of up to five years for herbaceous plants and over ten years for woody plants have to be calculated for transformation into a cultivated plant. A very effective example ofthe domestication of a wild plant on a large scale is Erythroxylum coca, that in 1990 produced at least 1,000 tonnes of cocaine (McChesney, 1996). Alternatively, in the context of a bioprospecting programme, existing cultivated plants can be found that can synthesize the desired substance or one of its pre-phases in sufficient quantity to cover the need in crop plantations. So far only 5–15 per cent ofthe higher plants have been investigated for possible bioactive substances (Balandrin et al, 1993). The broad scale screening of plants is juxtaposed with ethno-botany that makes targeted use of traditional natural medicinal knowledge (eg that ofthe shamans). Some firms in collaboration with indigenous experts are trying to extract new substances from the medicinal herbs ofthe Third World.Already around 10 per cent of all pharmaceuticals in Western medicine are based on medicinal plants from the Third World. Whereas 10,000–
66 D The use of genetic and species diversity 100,000 synthetic substances have to be tested to bring a new medicine onto the market, the same number of randomly collected plants is sufficient to develop on average five new medicines. The success rate of ethno-botany is probably even higher, for here the effect of traditional medicinal plants is already proven (SAG, 1997). In contrast to the long development story of Aspirin ® (Box D 3.3-2) the development of Taxol ® represents the modern bioprospecting approach (Box D 3.3-3). In the case of Taxol ® a horizontal genetic exchange, probably with a fungus in the bark ofthe yew tree, means that in future Taxol ® does not have to be produced from yew plantations but can be produced from cultures of this fungus. By identifying the gene for the substance of interest, eg through modern molecular diagnosis (eg differential display) and highly effective sequencing techniques, even more prospects open up. As will be explained with examples in Section D 3.3.4, the desired gene can be transferred (‘transfected’) into a cultivated plant, but also in an easily cultivatable microorganism. With this type of measure, economic viability can often be enhanced. D 126.96.36.199 Terrestrial microorganisms Terrestrial bacteria and fungi are predestined as producers of secondary metabolites as generally they live in symbiotic or parasitic relationships. As illustrated by the example of Taxol ® , increasingly microorganisms are being identified as the real source of substances which originally were wrongly ascribed to a parasitized plant (or animal) or one living in symbiosis.The poisonous ergot of rye alkaloids that are produced by sac fungi living on cereals (Ascomycetes ofthe Claviceps genus) are a classic example. Accordingly, the interest in bioprospecting microorganisms is growing, particularly since so far only a fraction ofthese have been identified and analysed (Table D 1.2-1). Consequently, some 50 per cent of individual projects supported under the BMBF’s support focus ‘Biotechnology 2000’ address the use of microorganisms andtheir products. A large number of highly effective antibiotic, cytostatic (cell-growth inhibitors) or immune-suppressant substances from various microorganisms have been used for years. Over 20 different antibiotics are produced from Streptomyces species alone. This family also produces a larger number of cancer blocking cytostatica and Box D 3.3-2 Aspirin ® – a one-hundred year history Back around 400 BC Hippocrates was prescribing an infusion from the bark ofthe white willow (Salix alba) against inflammation ofthe joints. The proponents ofthe signature theory in the Middle Ages used white willow tea to treat stiff joints and rheumatic pain because the willow had supple branches. The natural painkiller was forgotten for a while until in 1763, a Mr Stone from England described the anti-pyretic effects of willow bark. This finding was used in 1806 at the time ofthe Continental Blockade in Germany as an urgently needed substitute for China bark (Chinchona) that could no longer be imported.The following steps led to the manufacture of pure acetylsalicylic acid, the active substance in willow bark: • in 1828 J A Buchner isolated a yellowish mass from willow bark, that he named Salicin. • in 1829 the French apothecary Leroux isolated Salicin in crystallized form. • in 1838 R Piria manufactured salicylic acid from Salicin. At around the same time the Swiss apothecary Pagenstecher distilled salicylaldehyde from the flowers ofthe herbaceous meadow-sweet (Filipendula ulmaria; syn. Spiraea ulmaria); oxidation to salicylic acid followed. • in 1853 the chemist C F Gerhardt managed for the first time to synthesize an impure, but also non durable, acetylsalicylic acid. • in 1874, after H Kolbe had elucidated the chemical structure of salicylic acid, industrial production began. • in 1876 Ries and Stricker proved that the synthetic salicylic acid was suited to treating rheumatic fever and it cost ten times less than the natural substance. The chemical factory of Friedrich von Heyden (Radebeul- Dresden) andthe Elberfeld paint factory, previously Friedrich Bayer & Co., tried to develop a variety that would be tolerable to the stomach. In August 1897 the chemist Dr Felix Hoffmann (Friedrich Bayer & Compagnon) reported for the first time a chemically pure and durable acetylsalicylic acid (ASA) extracted by acetylisation. The preparation was registered under the name Aspirin ® and patented in 1899.The First World War, with its immense need for pain and fever medication, the renewed and now total import blockade on quinine and finally the great fever pandemic of 1918/19 all promoted the success of Aspirin ® . The British pharmacologist John R Vane received the Nobel prize for medicine in 1982 for elucidating the active mechanism of ASA. Currently, new forms of ASA with longer effect and reduced side effects are being developed, such as Celebrex ® and Vioxx ® ; they were approved for the US market in 1999. ASA is constantly being ascribed new indications, eg lowering the risk of heart attack or reducing the risk for certain types of cancer. Bayer AG took a long time to correctly place Aspirin ® in the market which for a long time now has not been under patent protection. Today in Bitterfeld 785 tonnes of acetylsalicylic acid is processed annually to produce Aspirin ® tablets (1.8 million per hour) – that is a third of total world Aspirin ® production with annual sales of US$425 million. Aspirin ® is the most frequently sold drug on all continents.