Metabolites 2012.2.303-336:doi:10.3390/metabo2020303 OPEN ACCESS metabolites ISSN2218-1989 www.mdpi.com/journal/metabolites/ Review A Historical Overview of Natural Products in Drug Discovery Daniel A.DiasSylvia Urbanand Ute Roessner Metabolomics Australia,School of Botany,The University of Melbourne,Parkville,Victoria 3010, Australia Sehool of Applied Senees (Discipline of Applied Chemistry)Health Inovation Research Institute(HIRi)RMIT University,G.P.O.Box 2476V,Melbourne,Victoria 3001,Australia Australian Centre for Plant Functional Genomics,School of Botany,The University of Melboure, Parkville,3010,Victoria,Australia *Author to whom correspondence should be addressed;E-Mail:ddias@unimelb.edu.au; Tel.:+61-3-8344-3318. Received:I March 2012:in revised form:31 March 2012/Accepted:31 March 2012/ Published:16 April 2012 Abstract:Historically,natural products have been used since ancient times and in folklore for the treatment of many diseases and illnesses.Classical natural product chemistry methodologies enabled a vast array of bioactive secondary metabolites from terrestrial and marine sources to be discovered.Many of these natural products have gone on to become current drug candidates.This brief review aims to highlight historically significant bioactive marine and terrestrial natural products,their use in folklore and dereplication techniques to rapidly facilitate their discovery.Furthermore a discussion of how natural product chemistry has resulted in the identification of many drug candidates:the application of natural product chemistry and finally adopting metabolomic profiling and dereplication approaches for the comprehensive study of natural product extracts will be discussed. Keywords:natural products;secondary metabolites;drug discovery;bioactivity: metabolomics;dereplication,plants;sponges;algae;fungi
Metabolites 2012, 2, 303-336; doi:10.3390/metabo2020303 metabolites ISSN 2218-1989 www.mdpi.com/journal/metabolites/ Review A Historical Overview of Natural Products in Drug Discovery Daniel A. Dias 1,*, Sylvia Urban 2 and Ute Roessner 1,3 1 Metabolomics Australia, School of Botany, The University of Melbourne, Parkville, Victoria 3010, Australia 2 School of Applied Sciences (Discipline of Applied Chemistry), Health Innovations Research Institute (HIRi) RMIT University, G.P.O. Box 2476V, Melbourne, Victoria 3001, Australia 3 Australian Centre for Plant Functional Genomics, School of Botany, The University of Melbourne, Parkville, 3010, Victoria, Australia * Author to whom correspondence should be addressed; E-Mail: ddias@unimelb.edu.au; Tel.: +61-3-8344-3318. Received: 1 March 2012; in revised form: 31 March 2012 / Accepted: 31 March 2012 / Published: 16 April 2012 Abstract: Historically, natural products have been used since ancient times and in folklore for the treatment of many diseases and illnesses. Classical natural product chemistry methodologies enabled a vast array of bioactive secondary metabolites from terrestrial and marine sources to be discovered. Many of these natural products have gone on to become current drug candidates. This brief review aims to highlight historically significant bioactive marine and terrestrial natural products, their use in folklore and dereplication techniques to rapidly facilitate their discovery. Furthermore a discussion of how natural product chemistry has resulted in the identification of many drug candidates; the application of advanced hyphenated spectroscopic techniques to aid in their discovery, the future of natural product chemistry and finally adopting metabolomic profiling and dereplication approaches for the comprehensive study of natural product extracts will be discussed. Keywords: natural products; secondary metabolites; drug discovery; bioactivity; metabolomics; dereplication, plants; sponges; algae; fungi OPEN ACCESS
Metabolites 2012.2 304 1.Introduction 1.1.Natural Products in History Natural products (secondary metabolites)have been the most successful source of potential drug leads [1-5].However,their recent implementation in drug discovery and development efforts have somewhat demonstrated a decline in interest [1].Nevertheless,natural products continue to provide unique structural diversity in comparison to standard combinatorial chemistry.which presents opportunities for discovering mainly novel low molecular weight lead compounds.Since less than 10%of the world's biodiversity has been evaluated for potential biological activity,many more useful natural lead compounds await discovery with the challenge being how to access this natural chemical diversity [3]. The earliest records of natural products were depicted on clay tablets in cuneiform from Mesopotamia (2600 B.C.)which documented oils from Cupressus sempervirens(Cypress)and Commiphora species(myrrh)which are still used today to treat coughs,colds and inflammation [3]. The Ebers Papyrus (2900 B.C.)is an Egyptian pharmaceutical record,which documents over 700 plant-based drugs ranging from gargles,pills,infusions,to ointments.The Chinese Materia Medica(1100 B.C.)(Wu Shi Er Bing Fang.contains 52 prescriptions),Shennong Herbal (-100 B.C.. 365 drugs)and the Tang Herbal (659 A.D.,850 drugs)are documented records of the uses of natural products [3].The Greek physician,Dioscorides,(100 A.D.),recorded the collection,storage and the uses of medicinal herbs,whilst the Greek philosopher and natural scientist,Theophrastus(300 B.C.) dealt with medicinal herbs.During the Dark and Middle Ages the monasteries in England,Ireland, France and Germany preserved this Western knowledge whilst the Arabs preserved the Greco-Roman knowledge and expanded the uses of their own resources,together with Chinese and Indian herbs unfamiliar to the Greco-Roman world [3].It was the Arabs who were the first to privately own pharmacies (8th century)with Avicenna,a Persian pharmacist,physician,philosopher and poet. contributing much to the sciences of pharmacy and medicine through works such as the Canon Medicinae [3]. 1.2.Medicinal Plants in Folklore The use of natural products as medicines has been described throughout history in the form of traditional medicines,remedies,potions and oils with many of these bioactive natural products still being unidentified.The dominant source of knowledge of natural product uses from medicinal plants is a result of man experimenting by trial and error for hundreds of centuries through palatability trials or untimely deaths,searching for available foods for the treatment of diseases [6,7].One example involves the plant genus Salvia which grows throughout the southwestern region of the United States as well as northwester Mexico and which was used by Indian tribes of souther California as an aid in childbirth [61.Male newborn babies werecooked"in the hot Salvia ashes as it was believed that these babies consistently grew to be the strongest and healthiest members of their respective tribes and are claimed to have been immune from all respiratory ailments for life [6]. The plant,Alhagi maurorum Medik (Camels thorn)secretes a sweet,gummy material from the stems and leaves during hot days []This gummy sap is called"consists mostly of
Metabolites 2012, 2 304 1. Introduction 1.1. Natural Products in History Natural products (secondary metabolites) have been the most successful source of potential drug leads [1–5]. However, their recent implementation in drug discovery and development efforts have somewhat demonstrated a decline in interest [1]. Nevertheless, natural products continue to provide unique structural diversity in comparison to standard combinatorial chemistry, which presents opportunities for discovering mainly novel low molecular weight lead compounds. Since less than 10% of the world’s biodiversity has been evaluated for potential biological activity, many more useful natural lead compounds await discovery with the challenge being how to access this natural chemical diversity [3]. The earliest records of natural products were depicted on clay tablets in cuneiform from Mesopotamia (2600 B.C.) which documented oils from Cupressus sempervirens (Cypress) and Commiphora species (myrrh) which are still used today to treat coughs, colds and inflammation [3]. The Ebers Papyrus (2900 B.C.) is an Egyptian pharmaceutical record, which documents over 700 plant-based drugs ranging from gargles, pills, infusions, to ointments. The Chinese Materia Medica (1100 B.C.) (Wu Shi Er Bing Fang, contains 52 prescriptions), Shennong Herbal (~100 B.C., 365 drugs) and the Tang Herbal (659 A.D., 850 drugs) are documented records of the uses of natural products [3]. The Greek physician, Dioscorides, (100 A.D.), recorded the collection, storage and the uses of medicinal herbs, whilst the Greek philosopher and natural scientist, Theophrastus (~300 B.C.) dealt with medicinal herbs. During the Dark and Middle Ages the monasteries in England, Ireland, France and Germany preserved this Western knowledge whilst the Arabs preserved the Greco-Roman knowledge and expanded the uses of their own resources, together with Chinese and Indian herbs unfamiliar to the Greco-Roman world [3]. It was the Arabs who were the first to privately own pharmacies (8th century) with Avicenna, a Persian pharmacist, physician, philosopher and poet, contributing much to the sciences of pharmacy and medicine through works such as the Canon Medicinae [3]. 1.2. Medicinal Plants in Folklore The use of natural products as medicines has been described throughout history in the form of traditional medicines, remedies, potions and oils with many of these bioactive natural products still being unidentified. The dominant source of knowledge of natural product uses from medicinal plants is a result of man experimenting by trial and error for hundreds of centuries through palatability trials or untimely deaths, searching for available foods for the treatment of diseases [6,7]. One example involves the plant genus Salvia which grows throughout the southwestern region of the United States as well as northwestern Mexico and which was used by Indian tribes of southern California as an aid in childbirth [6]. Male newborn babies were “cooked” in the hot Salvia ashes as it was believed that these babies consistently grew to be the strongest and healthiest members of their respective tribes and are claimed to have been immune from all respiratory ailments for life [6]. The plant, Alhagi maurorum Medik (Camels thorn) secretes a sweet, gummy material from the stems and leaves during hot days [8]. This gummy sap is called “manna” and consists mostly of
Metabolites 2012.2 305 melezitose,sucrose and invert sugar and it has been documented and claimed by the Ayurvedic people that the plant aids in the treatment of anorexia,constipation,dermatosis,epistaxis,fever,leprosy,and obesity []It was also used by the Israelis who boiled the roots and drank the extract as it stopped bloody diarrhea.The Konkani people smoked the plant for the treatment of asthma,whilst the Romans used the plant for nasal polyps []The plant Ligusticum Linnaeus found in Northern Europe and eastern north america was eaten raw first thing in the morning and was believed to protect a person from daily infection [the root was a cure for flatulence [10-12],an aphrodisiac [12]and was used as a sedative in the Faeroe Islands 10,13.Atropa belladonna Linnaeus (deadly nightshade)is found in central and Southern Europe,Western Asia,North Africa,North America and New Zealand. Its notoriously poisonous nature(three berries are sufficient to kill a child)firmly excluded it from the folk medicine compilation and seemed to have been accepted as dangerous to handle or to experiment with「141 1.3.Medicinal Natural Products from Other Sources Used in Folklore The fungus Piptoporus,which grows on birches was steamed to produce charcoal,valued as an antiseptic and disinfectant [15].Strips of P.betulinus were cut and used for staunching bleeding and were also found to make very comfortable corn pads [16].Another example is the fungus Agaricus campestris Linnaeux ex Fries(field mushroom)found in the northern and southern temperate zones and the Caribbean.A.campestris,had reportedly been stewed in milk to soothe cancer of the throat [17]. As early as the 17th-18th century,lichens had been used as dyes and were far more valued thar in folklore has been well documented [18].Lichens have been used as the raw materials for perfumes and cosmetics,medicine from the time of the early Chinese and Egyptian civilizations ]Wel known examples include Usnea dillenius ex Adanson which was traditionally used for curing diseases of the scalp and is still sold in pharmacies as an ingredient in anti-dandruff shampoos and in Ireland to treat sore eyes 9].The lichenU.Stirton was mixed with tobacco and butter,boiled and then cooled and applied as a lotion [14].Parmelia omphalodes(Linnaeus)Acharius,which is abundant in the British Isles,was used in brown dyes.In the highlands it was traditionally sprinkled on stockings at the start of a journey to prevent inflammation of the feet [20,21]and in Ireland it was used as a cure for bad sores under the chin as well as for burns and cuts [14]. By comparison,the marine environment has very few reported applications in traditional medicine The red algae Chondrus crispus and Mastocarpus stellatus were sources of a beverage,which was popular as a folk cure for colds,sore throats,chest infections including tuberculosis.The alga was also boiled in milk or water and used for kidney trouble and burns [22,23].Furthermore,three spoonfuls of the juice of the red alga Porphyra umbilicalis (Linnaeus)Kutzing,taken every morning followed by fasting for three weeks was found to be effective against cancers,in particular breast cancer 24]. P.umbilicalis has also been described in the Aran Islands for easing indigestion.and was also boiled and given to cows to relieve their springtime constipation [10,25]
Metabolites 2012, 2 305 melezitose, sucrose and invert sugar and it has been documented and claimed by the Ayurvedic people that the plant aids in the treatment of anorexia, constipation, dermatosis, epistaxis, fever, leprosy, and obesity [8]. It was also used by the Israelis who boiled the roots and drank the extract as it stopped bloody diarrhea. The Konkani people smoked the plant for the treatment of asthma, whilst the Romans used the plant for nasal polyps [8]. The plant Ligusticum scoticum Linnaeus found in Northern Europe and Eastern North America was eaten raw first thing in the morning and was believed to protect a person from daily infection [9]; the root was a cure for flatulence [10–12], an aphrodisiac [12] and was used as a sedative in the Faeroe Islands [10,13]. Atropa belladonna Linnaeus (deadly nightshade) is found in central and Southern Europe, Western Asia, North Africa, North America and New Zealand. Its notoriously poisonous nature (three berries are sufficient to kill a child) firmly excluded it from the folk medicine compilation and seemed to have been accepted as dangerous to handle or to experiment with [14]. 1.3. Medicinal Natural Products from Other Sources Used in Folklore The fungus Piptoporus betulinus, which grows on birches was steamed to produce charcoal, valued as an antiseptic and disinfectant [15]. Strips of P. betulinus were cut and used for staunching bleeding and were also found to make very comfortable corn pads [16]. Another example is the fungus Agaricus campestris Linnaeux ex Fries (field mushroom) found in the northern and southern temperate zones and the Caribbean. A. campestris, had reportedly been stewed in milk to soothe cancer of the throat [17]. As early as the 17th–18th century, lichens had been used as dyes and were far more valued than oriental spices. To date there are no lichen derived drugs approved on the market but their applications in folklore has been well documented [18]. Lichens have been used as the raw materials for perfumes and cosmetics, medicine from the time of the early Chinese and Egyptian civilizations [19]. Well known examples include Usnea dillenius ex Adanson which was traditionally used for curing diseases of the scalp and is still sold in pharmacies as an ingredient in anti-dandruff shampoos and in Ireland to treat sore eyes [19]. The lichen U. subfloridana Stirton was mixed with tobacco and butter, boiled and then cooled and applied as a lotion [14]. Parmelia omphalodes (Linnaeus) Acharius, which is abundant in the British Isles, was used in brown dyes. In the highlands it was traditionally sprinkled on stockings at the start of a journey to prevent inflammation of the feet [20,21] and in Ireland it was used as a cure for bad sores under the chin as well as for burns and cuts [14]. By comparison, the marine environment has very few reported applications in traditional medicine. The red algae Chondrus crispus and Mastocarpus stellatus were sources of a beverage, which was popular as a folk cure for colds, sore throats, chest infections including tuberculosis. The alga was also boiled in milk or water and used for kidney trouble and burns [22,23]. Furthermore, three spoonfuls of the juice of the red alga Porphyra umbilicalis (Linnaeus) Kützing, taken every morning followed by fasting for three weeks was found to be effective against cancers, in particular breast cancer [24]. P. umbilicalis has also been described in the Aran Islands for easing indigestion. and was also boiled and given to cows to relieve their springtime constipation [10,25]
Metabolites 2012.2 306 1.4.Primary and Secondary Metabolites (Natural Products) The biosynthesis and breakdown of proteins,fats,nucleic acids and carbohydrates,which are essential to all living organisms,is known as primary metabolism with the compounds involved in the pathways known as"primary metabolites"[26].The mechanism by which an organism biosynthesizes compounds called 'secondary metabolites'(natural products)is often found to be unique to an organism or is an expression of the individuality of a species and is referred to as "secondary metabolism"26.27].Secondary metabolites are generally not essential for the growth,development or reproduction of an organism and are produced either as a result of the organism adapting to its surrounding environment or are produced to act as a possible defense mechanism against predators to assist in the survival of the organism [26.28].The biosynthesis of secondary metabolites is derived from the fundamental processes of photosynthesis,glycolysis and the Krebs cycle to afford biosynthetic intermediates which,ultimately,results in the formation of secondary metabolites also known as natural products [26].It can be seen that although the number of building blocks are limited the formation of novel secondary metabolites is infinite.The most important building blocks employed in the biosynthesis of secondary metabolites are those derived from the intermediates:Acetyl coenzyme A(acetyl-CoA),shikimic acid,mevalonic acid and 1-deoxyxylulose-5-phosphate.They are involved in countless biosynthetic pathways,involving numerous different mechanisms and reactions (e.g.,alkylation,decarboxylation,aldol,Claisen and Schiff base formation [26]. It is hypothesized that secondary metabolism utilizes amino acids and the acetate and shikimate pathways to produce"shunt metabolites"(intermediates)that have adopted an alternate biosynthetic route.leading to the biosynthesis of secondary metabolites Modifications in the biosynthetic pathways may be due to natural causes (e.g..viruses or environmental changes)or unnatural causes (e.g.,chemical or radiation)in an effort to adapt or provide longevity for the organism [29].It is the unique biosynthesis of these natural products,produced by the countless number of terrestrial and marine organisms,which provides the characteristic chemical structures that possess an array of biological activities. 2.Historically Important Natural Products Traditional medicinal practices have formed the basis of most of the early medicines followed by subsequent clinical.pharmacological and chemical studies [5].Probably the most famous and well known example to date would be the synthesis of the anti-inflammatory agent,acetylsalicyclic acid(1) (aspirin)derived from the natural product,salicin (2)isolated from the bark of the willow tree Salix alba L.[30].Investigation of Papaver somniferum L.(opium poppy)resulted in the isolation of several alkaloids including morphine(3),a commercially important drug,first reported in 1803(Figure 1). It was in the 1870s that crude morphine derived from the plant P.somniferum,was boiled in acetic anhydride to yield diacetylmorphine (heroin)and found to be readily converted to codeine(painkiller). Historically,it is documented that the Sumerians and Ancient Greeks used poppy extracts medicinally whilst the Arabs described opium to be addictive [30].Digitalis purpurea L.(foxglove)had been traced back to Europe in the 10th century but it was not until the 1700s that the active constituent digitoxin()a cardiotonic glycoside was found to enhance cardiac conduction,thereby improving the
Metabolites 2012, 2 306 1.4. Primary and Secondary Metabolites (Natural Products) The biosynthesis and breakdown of proteins, fats, nucleic acids and carbohydrates, which are essential to all living organisms, is known as primary metabolism with the compounds involved in the pathways known as “primary metabolites” [26]. The mechanism by which an organism biosynthesizes compounds called ‛secondary metabolites’ (natural products) is often found to be unique to an organism or is an expression of the individuality of a species and is referred to as “secondary metabolism” [26,27]. Secondary metabolites are generally not essential for the growth, development or reproduction of an organism and are produced either as a result of the organism adapting to its surrounding environment or are produced to act as a possible defense mechanism against predators to assist in the survival of the organism [26,28]. The biosynthesis of secondary metabolites is derived from the fundamental processes of photosynthesis, glycolysis and the Krebs cycle to afford biosynthetic intermediates which, ultimately, results in the formation of secondary metabolites also known as natural products [26]. It can be seen that although the number of building blocks are limited, the formation of novel secondary metabolites is infinite. The most important building blocks employed in the biosynthesis of secondary metabolites are those derived from the intermediates: Acetyl coenzyme A (acetyl-CoA), shikimic acid, mevalonic acid and 1-deoxyxylulose-5-phosphate. They are involved in countless biosynthetic pathways, involving numerous different mechanisms and reactions (e.g., alkylation, decarboxylation, aldol, Claisen and Schiff base formation [26]. It is hypothesized that secondary metabolism utilizes amino acids and the acetate and shikimate pathways to produce “shunt metabolites” (intermediates) that have adopted an alternate biosynthetic route, leading to the biosynthesis of secondary metabolites [29]. Modifications in the biosynthetic pathways may be due to natural causes (e.g., viruses or environmental changes) or unnatural causes (e.g., chemical or radiation) in an effort to adapt or provide longevity for the organism [29]. It is the unique biosynthesis of these natural products, produced by the countless number of terrestrial and marine organisms, which provides the characteristic chemical structures that possess an array of biological activities. 2. Historically Important Natural Products Traditional medicinal practices have formed the basis of most of the early medicines followed by subsequent clinical, pharmacological and chemical studies [5]. Probably the most famous and well known example to date would be the synthesis of the anti-inflammatory agent, acetylsalicyclic acid (1) (aspirin) derived from the natural product, salicin (2) isolated from the bark of the willow tree Salix alba L. [30]. Investigation of Papaver somniferum L. (opium poppy) resulted in the isolation of several alkaloids including morphine (3), a commercially important drug, first reported in 1803 (Figure 1). It was in the 1870s that crude morphine derived from the plant P. somniferum, was boiled in acetic anhydride to yield diacetylmorphine (heroin) and found to be readily converted to codeine (painkiller). Historically, it is documented that the Sumerians and Ancient Greeks used poppy extracts medicinally, whilst the Arabs described opium to be addictive [30]. Digitalis purpurea L. (foxglove) had been traced back to Europe in the 10th century but it was not until the 1700s that the active constituent digitoxin (4), a cardiotonic glycoside was found to enhance cardiac conduction, thereby improving the
Metabolites 2012.2 307 strength of cardiac contractibility.Digitoxin (4)and its analogues have long been used in the management of congestive heart failure and have possible long term detrimental effects and are being replaced by other medicines in the treatment of"heart deficiency"[30].The anti-malarial drug quinine (5)approved by the United States FDA in 2004,isolated from the bark of Cinchona succirubra Pav.ex Klotsch,had been used for centuries for the treatment of malaria,fever,indigestion,mouth and throa diseases and cancer.Formal use of the bark to treat malaria was established in the mid 1800s when the British began the worldwide cultivation of the plant [30].Pilocarpine (6)found in Pilocarpus jaborandi(Rutaceae)is an L-histidine-derived alkaloid,which has been used as a clinical drug in the treatment of chronic open-angle glaucoma and acute angle-closure glaucoma for over 100 vears In 1994,an oral formulation of pilocarpine (6)was approved by the FDA to treat dry mouth (xerostomia)which is a side effect of radiation therapy for head and neck cancers and also used to stimulate sweat glands to measure the concentrations of sodium and chloride(Figure 1)[31].In 1998. the oral preparation was approved for the management of Sjogren's syndrome,an autoimmune disease that damages the salivary and lacrimal glands. Figure 1.Acetylsalicyclic acid (1),Salicin (2),Morphine (3),Digitoxin (4).Quinine (5) and Pilocarpine(6). HO HO OH HO HO.H.CN 5 6 2.1.Natural Products from Fungi Macro and micro fungi have been part of human life for thousands of years.They were used as food (mushrooms).in preparation of alcoholic beverages (yeasts),medication in traditional medicine and for cultural purposes.Currently with the advances in microbiology,their uses have extended to enzymes,biological control,antibiotics and other pharmacologically active products [32]
Metabolites 2012, 2 307 strength of cardiac contractibility. Digitoxin (4) and its analogues have long been used in the management of congestive heart failure and have possible long term detrimental effects and are being replaced by other medicines in the treatment of “heart deficiency” [30]. The anti-malarial drug quinine (5) approved by the United States FDA in 2004, isolated from the bark of Cinchona succirubra Pav. ex Klotsch, had been used for centuries for the treatment of malaria, fever, indigestion, mouth and throat diseases and cancer. Formal use of the bark to treat malaria was established in the mid 1800s when the British began the worldwide cultivation of the plant [30]. Pilocarpine (6) found in Pilocarpus jaborandi (Rutaceae) is an L-histidine-derived alkaloid, which has been used as a clinical drug in the treatment of chronic open-angle glaucoma and acute angle-closure glaucoma for over 100 years. In 1994, an oral formulation of pilocarpine (6) was approved by the FDA to treat dry mouth (xerostomia) which is a side effect of radiation therapy for head and neck cancers and also used to stimulate sweat glands to measure the concentrations of sodium and chloride (Figure 1) [31]. In 1998, the oral preparation was approved for the management of Sjogren's syndrome, an autoimmune disease that damages the salivary and lacrimal glands. Figure 1. Acetylsalicyclic acid (1), Salicin (2), Morphine (3), Digitoxin (4), Quinine (5) and Pilocarpine (6). 2.1. Natural Products from Fungi Macro and micro fungi have been part of human life for thousands of years. They were used as food (mushrooms), in preparation of alcoholic beverages (yeasts), medication in traditional medicine and for cultural purposes. Currently with the advances in microbiology, their uses have extended to enzymes, biological control, antibiotics and other pharmacologically active products [32]
Metabolites 2012.2 308 Undoubtedly one of the most famous natural product discoveries derived from a fungus (microorganism)is that of penicillin (7)from the fungus,Penicillim nom discovered by Fleming in 1929 [33].A countercurrent extractive separation technique which produced 7 in high yields was required for experimentation that ultimately saved countless lives and won Chain and Florey (together with Fleming)the 1945 Nobel prize in Physiology and Medicine (Figure 2)[34].This discovery led to the re-isolation and clinical studies by Chain,Florey and co-workers in the early 1940s and commercialization of synthetic penicillins,which ultimately revolutionized drug discovery research [35-38]. After publication of the first clinical data on penicillin G(7)between 1942-1944 there was a worldwide endeavor to discover new antibiotics from microorganisms and bioactive natural products [39,40].Up until 1968,old methods for detecting B-lactams were still being utilized and it was concluded that all natural B-lactams had been discovered [39].Nevertheless,this was not the case as with the introduction.in the 1970s.of new screening methods.the production of bacterial strains supersensitive to B-lactams,tests for the inhibition of B-lactamases and specificity for sulphur- containing metabolites resulted in the discovery of novel antibiotic structural classes (norcardicins carbapenems and monobactams)including the isolation of the antibiotics.norcardicin (8).imipenem (9)and aztreonam (10),respectively (Figure 2)[39,41].There are presently nine B-lactams (two cephalosporins,six carbapenems and one penem)in clinical trials or undergoing drug registration, along with the novel class of broad spectrum antibiotics called the glycylcyclines [41]. Figure 2.Penicillin(7),Norcardicin(8),Imipenem(9)and Aztreonam(10). NOHH OH -OH 01 7 H H CHa COOH N.SO3H HO Macro fungi such as polypores are a large group of wood-rotting fungi of the phylum Basidiomycota (basidomvcetes)and Ascomycota.which are a major source of pharmacologically active substances There are about 25,000 species of basidiomycetes,of which about 500 are members of the Aphyllophorales [42].Approximately 75%of tested polypore fungi have shown strong antimicrobial activities and may constitute a viable source for the development of novel antibiotics.Many compounds have displayed antiviral,cytotoxic,antineoplastic,cardiovascular,anti-inflammatory. immune-stimulating and anticancer activities [42,43].Fungi are more commonly microorganisms, some of which can spend at least part of their life cycle inside plant tissues without causing any visible
Metabolites 2012, 2 308 Undoubtedly one of the most famous natural product discoveries derived from a fungus (microorganism) is that of penicillin (7) from the fungus, Penicillium notatum discovered by Fleming in 1929 [33]. A countercurrent extractive separation technique which produced 7 in high yields was required for the in vivo experimentation that ultimately saved countless lives and won Chain and Florey (together with Fleming) the 1945 Nobel prize in Physiology and Medicine (Figure 2) [34]. This discovery led to the re-isolation and clinical studies by Chain, Florey and co-workers in the early 1940s and commercialization of synthetic penicillins, which ultimately revolutionized drug discovery research [35–38]. After publication of the first clinical data on penicillin G (7) between 1942–1944 there was a worldwide endeavor to discover new antibiotics from microorganisms and bioactive natural products [39,40]. Up until 1968, old methods for detecting β-lactams were still being utilized and it was concluded that all natural β-lactams had been discovered [39]. Nevertheless, this was not the case as with the introduction, in the 1970s, of new screening methods, the production of bacterial strains supersensitive to β-lactams, tests for the inhibition of β-lactamases and specificity for sulphurcontaining metabolites resulted in the discovery of novel antibiotic structural classes (norcardicins, carbapenems and monobactams) including the isolation of the antibiotics, norcardicin (8), imipenem (9) and aztreonam (10), respectively (Figure 2) [39,41]. There are presently nine β-lactams (two cephalosporins, six carbapenems and one penem) in clinical trials or undergoing drug registration, along with the novel class of broad spectrum antibiotics called the glycylcyclines [41]. Figure 2. Penicillin (7), Norcardicin (8), Imipenem (9) and Aztreonam (10). N N S O O H N N O O CO2H H H OH H NOH O CO2H H2N N O COOH HO S N H H H N H O N O SO3H 7 9 10 O 8 OH NH CH3 N N S H2N HO O O Macro fungi such as polypores are a large group of wood-rotting fungi of the phylum Basidiomycota (basidomycetes) and Ascomycota, which are a major source of pharmacologically active substances. There are about 25,000 species of basidiomycetes, of which about 500 are members of the Aphyllophorales [42]. Approximately 75% of tested polypore fungi have shown strong antimicrobial activities and may constitute a viable source for the development of novel antibiotics. Many compounds have displayed antiviral, cytotoxic, antineoplastic, cardiovascular, anti-inflammatory, immune-stimulating and anticancer activities [42,43]. Fungi are more commonly microorganisms, some of which can spend at least part of their life cycle inside plant tissues without causing any visible
Metabolites 2012.2 309 sign of infection or disease [44,45].They have been found to inhabit trees,grasses,algae and herbaceous plants and live in the intercellular spaces of plant stems,petioles,roots and leaves withou affecting the host organism [46].Collectively these fungi are known as endophytes.Novel bioactive secondary metabolites derived from fungal sources have yielded some of the most important natural products for the pharmaceutical industry [3]. In 1953.Edmund Kornfeld first isolated vancomycin(11)a glycopeptide antibiotic produced in cultures of Amycolatopsis orientalis which is active against a wide range of gram-positive organisms such as Staphvlococci and Streptococci and against gram-negative bacteria,mycobacteria and fungi and was approved by the FDA in 1958(Figure 3).It is used for the treatment of severe infection and against susceptible organisms in patients hypersensitive to penicillin (7)[5].The macrolide erythromycin (12)from Saccharopolyspora ervthraea is an antibacterial drug.which contains a 14-membered macrocycle composed entirely of propionate units(Figure 3).Erythromycin (12)has broad spectrum activities against gram-positive cocci and bacilli and is used for mild to moderate. upper and lower respiratory tract infections [5,26].Currently there are three semisynthetic ketolide derivatives of erythromycin (12).cethromycin (ABT-773.RestanzaTM).EP-420 (by Enanta Pharmaceuticals)and BAL-19403(by Basilea)which are in clinical development [1]. Figure 3.Vancomvcin (11)and Erythromvcin (12). 9 -C OH CH HOOC NH OH 11 Single cell viruses represent the smallest existing life forms causing cold,influenza,ebola and SARS.Presently,there seems to be a limited number of antiviral natural products or synthetically derived analogues from fungi [47].Betulinic acid (13),a triterpenoid obtained from the bark of Betula pubescens was originally identified as a weak inhibitor of HIV replication [48,49].Betulinic acid can inhibit topoisomerase I and is being evaluated in Phase I trials as a cancer chemo-preventive agent (Figure 4)[50].Bevirimat (PA-457)(14),extracted from a Chinese herb Syzygium claviflorum is in Phase IIb clinical trials and is believed to inhibit the final step of the HIV Gag protein processing [51] Ganoderic acid B(15),isolated from the fruiting bodies and spores of Ganoderma lucidum,displayed significant anti-HIV-1 protease activity with an ICso value of 20 uM (Figure 4)[52]
Metabolites 2012, 2 309 sign of infection or disease [44,45]. They have been found to inhabit trees, grasses, algae and herbaceous plants and live in the intercellular spaces of plant stems, petioles, roots and leaves without affecting the host organism [46]. Collectively these fungi are known as endophytes. Novel bioactive secondary metabolites derived from fungal sources have yielded some of the most important natural products for the pharmaceutical industry [3]. In 1953, Edmund Kornfeld first isolated vancomycin (11) a glycopeptide antibiotic produced in cultures of Amycolatopsis orientalis which is active against a wide range of gram-positive organisms such as Staphylococci and Streptococci and against gram-negative bacteria, mycobacteria and fungi and was approved by the FDA in 1958 (Figure 3). It is used for the treatment of severe infection and against susceptible organisms in patients hypersensitive to penicillin (7) [5]. The macrolide erythromycin (12) from Saccharopolyspora erythraea is an antibacterial drug, which contains a 14-membered macrocycle composed entirely of propionate units (Figure 3). Erythromycin (12) has broad spectrum activities against gram-positive cocci and bacilli and is used for mild to moderate, upper and lower respiratory tract infections [5,26]. Currently there are three semisynthetic ketolide derivatives of erythromycin (12), cethromycin (ABT-773, Restanza™), EP-420 (by Enanta Pharmaceuticals) and BAL-19403 (by Basilea) which are in clinical development [1]. Figure 3. Vancomycin (11) and Erythromycin (12). Single cell viruses represent the smallest existing life forms causing cold, influenza, ebola and SARS. Presently, there seems to be a limited number of antiviral natural products or synthetically derived analogues from fungi [47]. Betulinic acid (13), a triterpenoid obtained from the bark of Betula pubescens was originally identified as a weak inhibitor of HIV replication [48,49]. Betulinic acid can inhibit topoisomerase I and is being evaluated in Phase I trials as a cancer chemo-preventive agent (Figure 4) [50]. Bevirimat (PA-457) (14), extracted from a Chinese herb Syzygium claviflorum is in Phase IIb clinical trials and is believed to inhibit the final step of the HIV Gag protein processing [51]. Ganoderic acid β (15), isolated from the fruiting bodies and spores of Ganoderma lucidum, displayed significant anti-HIV-1 protease activity with an IC50 value of 20 µM (Figure 4) [52]
Metabolites2012.2 310 Figure 4.Betulinic acid (13).Bevirimat (PA-457)(14)and Ganoderic acid B(15) HO 13 In 2002,amrubicin hydrochloride(16),related to the anthracycline,doxorubicin(17)(Adriamycin), was isolated from the fungus Streptomyces peucetius.Doxorubicin (17)is used to treat acute leukaemia,soft tissue and bone sarcomas,lung cancer,thyroid cancer and both Hodgkins and non-Hodgkins lymphomas (Figure 5)[5.26].Torrevanic acid (18)was isolated from an endophvte from the endangered tree,Torreya laxifolia [53]and was tested in several cancer cell lines and found to display 5-10 times greater potentency/cytotoxicity in cell lines that are sensitive to protein kinaseC causing cell death by apoptosis(Figure 5)[54]. Figure 5.Amrubicin hydrochloride(16),Doxorubicin(17)and Torreyanic acid(18). OH HO.C NH CH. H 00H6 H CH OH 。 17 2.2.Natural Products from Plants Plants have been well documented for their medicinal uses for thousands of years.They have evolved and adapted over millions of years to withstand bacteria,insects,fungi and weather to produce unique.structurally diverse secondary metabolites.Their ethnopharmacological properties have been used as a primary source of medicines for early drug discovery [55,56].According to the World Health Organization (WHO).80%of people still rely on plant-based traditional medicines for primary health care [57]and 80%of 122 plant derived drugs were related to their original ethnopharmacological purpose [58].The knowledge associated with traditional medicine (complementary or alternative herbal products)has promoted further investigations of medicinal plants as potential medicines and has led to the isolation of many natural products that have become well known pharmaceuticals. The most widely used breast cancer drug is paclitaxel(Taxol)(19),isolated from the bark of Taxus brevifolia (Pacific Yew).In 1962 the United States Department of Agriculture (USDA)first collected the bark as part of their exploratory plant screening program at the National Cancer Institute
Metabolites 2012, 2 310 Figure 4. Betulinic acid (13), Bevirimat (PA-457) (14) and Ganoderic acid β (15). In 2002, amrubicin hydrochloride (16), related to the anthracycline, doxorubicin (17) (Adriamycin®), was isolated from the fungus Streptomyces peucetius. Doxorubicin (17) is used to treat acute leukaemia, soft tissue and bone sarcomas, lung cancer, thyroid cancer and both Hodgkins and non-Hodgkins lymphomas (Figure 5) [5,26]. Torreyanic acid (18) was isolated from an endophyte from the endangered tree, Torreya taxifolia [53] and was tested in several cancer cell lines and found to display 5–10 times greater potentency/cytotoxicity in cell lines that are sensitive to protein kinase C causing cell death by apoptosis (Figure 5) [54]. Figure 5. Amrubicin hydrochloride (16), Doxorubicin (17) and Torreyanic acid (18). 2.2. Natural Products from Plants Plants have been well documented for their medicinal uses for thousands of years. They have evolved and adapted over millions of years to withstand bacteria, insects, fungi and weather to produce unique, structurally diverse secondary metabolites. Their ethnopharmacological properties have been used as a primary source of medicines for early drug discovery [55,56]. According to the World Health Organization (WHO), 80% of people still rely on plant-based traditional medicines for primary health care [57] and 80% of 122 plant derived drugs were related to their original ethnopharmacological purpose [58]. The knowledge associated with traditional medicine (complementary or alternative herbal products) has promoted further investigations of medicinal plants as potential medicines and has led to the isolation of many natural products that have become well known pharmaceuticals. The most widely used breast cancer drug is paclitaxel (Taxol®) (19), isolated from the bark of Taxus brevifolia (Pacific Yew). In 1962 the United States Department of Agriculture (USDA) first collected the bark as part of their exploratory plant screening program at the National Cancer Institute
Metabolites 2012.2 311 (NCI)(Figure 6)[59].The bark from about three mature 100 year old trees is required to provide I gram of 19 given that a course of treatment may need 2 grams of the drug.Current demand for 19 is in the region of 100-200 kg per annum (i.e.,50,000 treatments/year)and is now produced synthetically [26]. The first of several FDA approvals for various uses for Taxol was announced in 1992 [60].Taxol (19)is present in limited quantities from natural sources,its synthesis(though challenging and expensive)has been achieved [61].Baccatin III(20)present in much higher quantities and readily available from the needles of T.brevifolia and associated derivatives is an example of a structural analogue that can be efficiently transformed into 19(Figure 6)26. Figure 6.Paclitaxel (Taxol)(19)and baccatin III(20). 0 H 19 Other examples of antitumor compounds currently in clinical trials include ingenol 3--angelate (21)a derivative of the polyhydroxy diterpenoid ingenol isolated from the sap of Euphorbia peplus(known as "petty spurge"in England or"radium weed'in Australia)which is a potential chemotherapeutic agent for skin cancer is currently under clinical development by Peplin Biotech for the topical treatment of certain skin cancers (Figure 7)[62.63].PG490-88(22) (14-succinyl triptolide sodium salt),a semisynthetic analogue of triptolide is a diterpene-diepoxide isolated from which is used for autoimmune and inflammatory diseases in the People's Republic of China [64,65].Combretastatin A-4 phosphate(23)a stilbene derivative from the South African Bush Willow,Combretum caffrum acts as an anti-angiogenic agent causing vascular shutdowns in tumors(necrosis)and is currently in Phase II clinical trials(Figure 7)[66.67]. Figure 7.ingenol 3-0-angelate(21),PG490-88(22)and Combretastatin A-4 phosphate(23) 21 The Acquired Immune Deficiency Syndrome (AIDS)pandemic in the 1980s forced the National Cancer Institute(NCI)and other organizations to explore natural products as sources of potential drug
Metabolites 2012, 2 311 (NCI) (Figure 6) [59]. The bark from about three mature 100 year old trees is required to provide 1 gram of 19 given that a course of treatment may need 2 grams of the drug. Current demand for 19 is in the region of 100–200 kg per annum (i.e., 50,000 treatments/year) and is now produced synthetically [26]. The first of several FDA approvals for various uses for Taxol® was announced in 1992 [60]. Taxol® (19) is present in limited quantities from natural sources, its synthesis (though challenging and expensive) has been achieved [61]. Baccatin III (20) present in much higher quantities and readily available from the needles of T. brevifolia and associated derivatives is an example of a structural analogue that can be efficiently transformed into 19 (Figure 6) [26]. Figure 6. Paclitaxel (Taxol®) (19) and baccatin III (20). Other examples of antitumor compounds currently in clinical trials include ingenol 3-O-angelate (21) a derivative of the polyhydroxy diterpenoid ingenol isolated from the sap of Euphorbia peplus (known as “petty spurge” in England or “radium weed” in Australia) which is a potential chemotherapeutic agent for skin cancer is currently under clinical development by Peplin Biotech for the topical treatment of certain skin cancers (Figure 7) [62,63]. PG490-88 (22) (14-succinyl triptolide sodium salt), a semisynthetic analogue of triptolide is a diterpene-diepoxide isolated from Tripterygium wilfordii which is used for autoimmune and inflammatory diseases in the People’s Republic of China [64,65]. Combretastatin A-4 phosphate (23) a stilbene derivative from the South African Bush Willow, Combretum caffrum acts as an anti-angiogenic agent causing vascular shutdowns in tumors (necrosis) and is currently in Phase II clinical trials (Figure 7) [66,67]. Figure 7. ingenol 3-O-angelate (21), PG490-88 (22) and Combretastatin A-4 phosphate (23). HO O O OH O OH 21 O O O O OPO3 23 O O O O O H O O O O Na 22 The Acquired Immune Deficiency Syndrome (AIDS) pandemic in the 1980s forced the National Cancer Institute (NCI) and other organizations to explore natural products as sources of potential drug
Metabolites 2012.2 312 candidates for the treatment of AIDS.Over 60,000 extracts of plants and marine organisms were tested against lymphoblastic cells infected with HIV-1.The most important result of these tests is the class of compounds known as the calanolides.In particular the isolation of calanolide a (24)and calanolide b (25)from the Calonphyllum species,along with prostratin (26)from Homalanthus mtans,have now progressed into clinical and preclinical development (Figure 8)[68-70].Calanolide A (24)was licensed and evaluated to Phase II clinical trials by Sarawak Medichem Pharmaceuticals,however there has been no subsequent announcement for further drug development.In 2010.Phase I human clinical trials of prostratin (26)were carried out by the AIDS ReSearch Alliance in Los Angeles, California(Figure 8). Figure 8.Calanolide A(24),Calanolide B(25)and Prostratin(26). HO Arteether (27),introduced in 2000,as Artemotil is derived from artemisinin(28)(introduced in 1987 as Artemisin)which was first isolated from the plant Artemisia annua and are both approved antimalarial drugs (Figure 9)[47].The plant was originally used in traditional Chinese medicine as a remedy for chills and fevers.Other derivatives of artemisinin (28)are in various stages of clinical development as antimalarial drugs in Europe [3,26].To date,a synthetic trioxolane modeled on the 28 pharmacophore,is being assessed in combination with piperaquine (a synthetic bisquinoline antimalarial drug)in an effort to treat malaria(Figure 9)[71]. Figure 9.Arteether(27)and artemisinin(28) H H H Grandisines A (29)and B(30)are two indole alkaloids which were isolated from the leaves of the Australian rainforest tree,Elaeocarpus grandis (Figure 10).Grandisine A (29)contains a unique tetracyclic skeleton,while Grandisine B(30)possesses an unusual combination of isoquinuclidinone and indolizidine ring systems.Both 29 and 30 exhibit binding affinity for the human 6-opioid receptor
Metabolites 2012, 2 312 candidates for the treatment of AIDS. Over 60,000 extracts of plants and marine organisms were tested against lymphoblastic cells infected with HIV-1. The most important result of these tests is the class of compounds known as the calanolides. In particular the isolation of calanolide A (24) and calanolide B (25) from the Calonphyllum species, along with prostratin (26) from Homalanthus nutans, have now progressed into clinical and preclinical development (Figure 8) [68–70]. Calanolide A (24) was licensed and evaluated to Phase II clinical trials by Sarawak Medichem Pharmaceuticals, however there has been no subsequent announcement for further drug development. In 2010, Phase I human clinical trials of prostratin (26) were carried out by the AIDS ReSearch Alliance in Los Angeles, California (Figure 8). Figure 8. Calanolide A (24), Calanolide B (25) and Prostratin (26). Arteether (27), introduced in 2000, as Artemotil is derived from artemisinin (28) (introduced in 1987 as Artemisin) which was first isolated from the plant Artemisia annua and are both approved antimalarial drugs (Figure 9) [47]. The plant was originally used in traditional Chinese medicine as a remedy for chills and fevers. Other derivatives of artemisinin (28) are in various stages of clinical development as antimalarial drugs in Europe [3,26]. To date, a synthetic trioxolane modeled on the 28 pharmacophore, is being assessed in combination with piperaquine (a synthetic bisquinoline antimalarial drug) in an effort to treat malaria (Figure 9) [71]. Figure 9. Arteether (27) and artemisinin (28). Grandisines A (29) and B (30) are two indole alkaloids which were isolated from the leaves of the Australian rainforest tree, Elaeocarpus grandis (Figure 10). Grandisine A (29) contains a unique tetracyclic skeleton, while Grandisine B (30) possesses an unusual combination of isoquinuclidinone and indolizidine ring systems. Both 29 and 30 exhibit binding affinity for the human δ-opioid receptor
