Phytochemistry Reviews (v.8, #2)

Preface by Virginia Lanzotti (311-311).

Nature: a vital source of leads for anticancer drug development by G. M. Cragg; D. J. Newman (313-331).
Over 60% of the current anticancer drugs have their origin in one way or another from natural sources. Nature continues to be the most prolific source of biologically active and diverse chemotypes, and it is becoming increasingly evident that associated microbes may often be the source of biologically active compounds originally isolated from host macro-organisms. While relatively few of the actual isolated compounds advance to become clinically effective drugs in their own right, these unique molecules may serve as models for the preparation of more efficacious analogs using chemical methodology such as total or combinatorial (parallel) synthesis, or manipulation of biosynthetic pathways. In addition, conjugation of toxic natural molecules to monoclonal antibodies or polymeric carriers specifically targeted to epitopes on tumors of interest can lead to the development of efficacious targeted therapies. The essential role played by natural products in the discovery and development of effective anticancer agents, and the importance of multidisciplinary collaboration in the generation and optimization of novel molecular leads from natural product sources is reviewed.
Keywords: Plants; Marine organisms; Microbes; Symbionts; Multidisciplinary collaboration

A precise regulation of redox balance is required for the cellular homeostatic control. Aberrant activation of redox-sensitive transcription factors, such as nuclear factor-kappaB (NF-κB), activator protein 1 (AP-1), cyclic adenosine monophosphate response element binding protein (CREB), and hypoxia inducible factor (HIF), contributes to carcinogenesis by promoting persistent inflammation, abnormal cell proliferation, evasion from apoptosis, angiogenesis, etc. A wide variety of dietary phytochemicals have been reported to exert cancer chemopreventive properties by suppressing the inappropriate activation of aforementioned transcription factors. On the other hand, transcription of genes involved in the activation of cellular antioxidant arsenal and carcinogen detoxification is largely regulated by another redox-sensitive transcription factor, i.e. NF-E2 related factor 2 (Nrf2), which plays a role in protecting cells/tissues from oxidative or electrophilic damage. Some food-derived phytochemicals have been shown to activate Nrf2, thereby augmenting cellular antioxidant capacity and inducing expression of phase-2 detoxification enzymes. Therefore, the modulation of cellular signaling mediated by redox-sensitive transcription factors in the right direction represents a promising approach to achieving molecular target-based chemoprevention with edible phytochemicals.
Keywords: Chemoprevention; Nrf2; NF-κB; AP-1; CREB; HIF; Phytochemicals

Phytochemical research has revealed that organic sulfur-containing compounds (OSCs) from Allium species exert biological effects, that might be beneficial in the treatment or prevention of a range of diseases, such as infections, cardiovascular and metabolic affections, cancers and related indispositions. Focusing physiological activities of these compounds in the context of cancer, it became clear from both epidemiological studies in men and experimental studies in diverse models, that the OSCs have a strong potential to prevent or to treat cancers even with selectivity against non-neoplastic cells. Though underlying mechanisms are not yet fully understood, several parts of their modes and mechanisms of action were elucidated: Pivotal molecular targets of as well chemoprevention as chemotherapy are metabolic, transporter or repair enzymes strongly affecting cell death, proliferation and formation of metastases. Accordingly effects are not restricted to the run of cell death programs, but they moreover comprise the strongly interdepending immune and inflammatory systems. Respectively, several hypotheses exist which are based on chemical properties of sulfur as the “pharmacophor” of the compounds appearing in up to ten different oxidation states (−2 to +6). Hence compounds can undergo redox-reactions and electrostatic interactions, making reactive oxygen species (ROS) a key feature of their mechanisms of action.
Keywords: Organic sulfur compounds from Allium species; Chemoprevention; Chemotherapy; Modes and mechanisms of action

Bioactive compounds are extra nutritional constituents found in small quantities in foods. We have recently shown that a bioactive compound, inositol pentakisphosphate (IP5), a naturally occurring substance that is present in most legumes, wheat bran and nuts, inhibits cell growth of ovarian, lung and breast cancer cells. We demonstrated that IP5 specifically blocks the activation of the critical phosphoinositide 3-kinase (PI3K) effector Akt, a serine/threonine kinase which plays a key role in different intracellular processes such as cell survival and proliferation. Due to its role in cancer development and progression, the PI3K/Akt pathway is an attractive target for therapeutic intervention. Interestingly, IP5 possesses anti tumour activity in mice to the same extent than cytotoxic drug cisplatin. Furthermore, IP5 enhances the effect of cytotoxic drugs in ovarian and lung cancer cells. These results support a role for IP5 as an anti-tumour agent that may sensitise cancer cells to the action of commonly used anti-cancer drugs. In addition we have recently observed that specific modifications of the IP5 structure may result in compounds with the same solubility and lack of toxicity in vivo but broader range of action and a higher activity compared to parental molecule indicating that IP5 may represents a promising molecule for further development of novel anticancer drugs. Therefore, our study reveals a new pharmacologically active nutrient (nutraceutical) as a potential chemopreventive agent and a lead compound for possible development of potent small molecule PI3K/Akt inhibitors.
Keywords: Bioactive compounds; Inositol polyphosphates; Nutraceutical; Phosphoinositide 3-kinase

Phytoestrogens as natural prodrugs in cancer prevention: dietary flavonoids by Randolph R. J. Arroo; Vasilis Androutsopoulos; Kenneth Beresford; Ketan Ruparelia; Somchaiya Surichan; Nicola Wilsher; Gerry A. Potter (375-386).
There are many reasons why vegetables and fruits may protect against cancer. As well as containing vitamins and minerals, which help keep the body healthy and strengthen the immune system, they are also good sources of biologically active compounds, which can help to protect cells in the body from damage that can lead to cancer. Notably, dietary flavonoids and other polyphenols are thought to have an important role as chemopreventive agents. Most studies on the possible mechanism of the chemopreventive action of dietary compounds have assumed that free hydroxyl groups of flavonoids and other polyphenols are necessary for their biological effects. However, in the human body dietary polyphenols are rapidly conjugated by glucuronosyltransferases and sulfotransferases, two enzymes that are abundantly present in the small intestine and liver, through which all of the oral dose must pass. Thus, most polyphenols that have been studied, e.g. quercetin, kaempferol, diosmetin, and resveratrol, would not be expected to reach internal organs beyond sites directly along the gastrointestinal tract. When the hydroxyl groups in polyphenols are methylated, the resulting compounds are much less prone to glucuronidation and sulfation. Thus methoxylated compounds are more metabolically stable, increasing their bioavailablity. The peel of various Citrus species can contain high concentrations of polymethoxyflavones, whereas the juice mainly contains hydroxylated flavones. At present, very little is known about the mechanisms by which methoxylated flavones may affect growth and development of tumour cells. Recently, it was shown that tumour specific enzymes can catalyze the O-demethylation of methoxylated flavones, resulting in the formation of flavones with free hydroxyl groups. We propose that demethylation of methoxylated flavones is another example of bioactivation of naturally occurring prodrugs.
Keywords: Cancer prevention; CYP1; Flavonoids; Metabolism

Recent developments in the rapid analysis of plants and tracking their bioactive constituents by Teris A. van Beek; Kishore K. R. Tetala; Irina I. Koleva; Airidas Dapkevicius; Vassiliki Exarchou; Suzanne M. F. Jeurissen; Frank W. Claassen; Elbert J. C. van der Klift (387-399).
Natural products chemistry has witnessed many new developments in the last 5 years like extractions with subcritical water and ionic liquids, LC/HRMS and LC/SPE/cryo-NMR, UHPLC, TLC/MS, MS-based preparative HPLC, comprehensive chromatography (GC × GC, LC × LC), high-throughput screening, introduction of monolithic columns, miniaturisation, and automated structure identification. Nevertheless identifying bioactive constituents in complex plant extracts remains a tedious process. The classical approach of bioassay guided fractionation is time-consuming while off-line screening of extracts does not provide information on individual compounds and sometimes suffers from false positives or negatives. One way out of this is by coupling chromatography with chemical or biochemical assays, so called high resolution screening. An example is the development of HPLC on-line assays for antioxidants. By the post-column addition of a relatively stable coloured radical like DPPH or ABTS•+, radical scavengers are detected as negative peaks because in a reaction coil they reduce the model radical to its reduced, non-coloured form. When combined with LC/DAD/MS and LC/SPE/NMR, reliable identification of active constituents becomes possible without the necessity of ever isolating them in a classical sense. Also for finding leads for new drugs, combining HPLC with biochemical assays is interesting but technically more difficult. Most enzymes do not work at the organic modifier concentrations commonly encountered in RP-HPLC and the reaction time is often longer requiring dilution and lengthy coils respectively. Therefore, new techniques have to be implemented to gain the required sensitivity for on-line enzyme assays. For stable analytes, high temperature LC offers a solution to the organic modifier problem. When enzymes are highly expensive, like those used in the screening for Cytochrome P450 inhibitors, miniaturisation to chip format may offer a way out. Microreactors (chips) are not only useful for miniaturising larger assays but also offer completely new prospects in phytochemical analysis. One such application is in the sample clean-up of acids and bases like alkaloids. In a lay-out of three parallel channels of 100 μm width with the middle one containing organic phase and the two outer ones water of high pH (feed phase) and low pH (trapping phase) such a chip replaces two classical LLE steps but is much faster and requires less solvents and less manpower input.
Keywords: Modern phytochemistry; On-line HPLC; High resolution screening; Radical scavenging; Liquid–liquid extraction chip

Sessile marine animals like sponges, tunicates, and bryozoans are a rich source of bioactive natural products, many of which exhibit potent anticancer activities. However, most of these substances are available in very limited amounts only, which has prohibited further drug development. Recent evidence suggests that symbiotic bacteria might be the true producers of many animal-derived metabolites. In addition to revealing fascinating perspectives for research in marine chemical ecology, these findings suggest new solutions to the supply problem. Although most symbionts remain uncultivated, bacterial production systems might be created by isolating biosynthetic genes from marine metagenomes, and expressing them in culturable bacterial hosts. This review discusses cell-sorting, natural product visualization, and phylogenetic approaches to identify symbiotic producers. In addition, strategies to isolate genes and gene clusters from marine species consortia are described. These techniques have provided insights into the bacterial origin and biosynthesis of polyketides like the onnamides, swinholides, and bryostatins, of peptides including the patellamides, chlorinated dipeptides, and theopalauamide as well as of brominated biphenylethers.
Keywords: Marine natural products; Metagenomics; Symbiosis; Uncultivated bacteria; Gene cloning; Sponges; Tunicates; Bryozoans; Biosynthesis

Marine natural products have become a major source of new chemical entities in the discovery of potential anticancer agents that potently suppress various antitumor molecular targets. As a consequence of insufficient vascularization, hypoxic regions form within rapidly growing solid tumor masses. Specific alterations of gene expression in these hypoxic tumor cells help facilitate the survival and metastatic spread of solid tumors. The transcriptional response to cellular hypoxia is primarily mediated by the transcription factor hypoxia-inducible factor-1 (HIF-1) that regulates the expression of more than 100 genes involved in cellular adaptation and survival under hypoxic stress. Clinical studies in cancer patients indicate that HIF-1 activation is directly correlated with advanced disease stages and treatment resistance. HIF-1 has emerged as an important tumor-selective molecular target for anticancer drug discovery. As a result, natural product-based inhibitors of HIF-1 activation have been identified from plants and microorganisms. Recently, structurally unique natural products from marine sponges, crinoids, and algae have been identified as HIF-1 activation inhibitors. The US National Cancer Institute’s Open Repository of marine invertebrate and algae extracts has proven to be a valuable source of natural product HIF-1 inhibitors. Among the active compounds identified, certain marine natural products have also been shown to suppress the hypoxic induction of HIF-1 target genes such as vascular endothelial growth factor (VEGF). Some of these marine HIF-1 inhibitors act by interfering with the generation of mitochondrial signaling molecules in hypoxic cells. However, the precise mechanisms of action for many newly identified marine natural product HIF-1 inhibitors remain unresolved.
Keywords: Cellular signaling; Crinoids; Gene expression; HIF-1 inhibitors; Hypoxia-inducible factor-1; Marine natural products; Molecular-targeted antitumor agents; Sponges; Transcription factor; Tumor hypoxia; Tunicates

Jatrophane diterpenes from Euphorbia spp. as modulators of multidrug resistance in cancer therapy by G. Corea; A. Di Pietro; C. Dumontet; E. Fattorusso; V. Lanzotti (431-447).
A phytochemical investigation of the aerial parts of Euphorbia spp., considered one of the most common elements of Mediterranean landascape, led to the isolation of a large number of bioactive plant metabolites, belonging to the diterpenes family. Above all, over seventy jatrophane, modified jatrophane, segetane, pepluane, and paraliane diterpenoids, fifty of them reported for the first time, were extracted, purified and characterized from Euphorbia dendroides, Euphorbia characias, Euphorbia peplus, Euphorbia paralias and Euphorbia helioscopia. These compounds showed interesting pharmacological activities. In particular, jatrophanes, modified jatrophanes and lathyranes exhibited a potent inhibitory activity against P-glycoprotein (Pgp), a membrane protein that confers cellular ability to resist lethal doses of certain cytotoxic drugs by pumping them to the outside, thus resulting in a reduced cytotoxicity. Among the others, our chemical survey led to isolation of the most powerful inhibitors of daunomycin-efflux activity isolated to date for this class of inhibitors, named euphodendroidin D and pepluanin A. Their efficiency was found to be at least two-fold higher than the conventional modulator cyclosporin A, taken as a reference. In addition, the isolation of a high number of natural structurally-related analogues allowed us to perform Structure Activity Relationship (SAR) studies, without any chemical modification, which gave information on the key pharmacophoric elements of these new class of promising drugs. A further set of diterpene analogues, very recently isolated from sun spurge, E. helioscopia, individually investigated for their Pgp- and BCRP-inhibiting properties, appeared to be specific inhibitors of Pgp since they showed no significant activity against BCRP, thus resembling to the third-generation class of specific MDR inhibitors.
Keywords: Euphorbia ; Diterpenes; Jatrophanes; Modified jatrophanes; Lathyranes; Multidrug resistance; Pgp; BCRP; Third generation inhibitors

Plants of the Amaryllidaceae family have been under intense scrutiny for the presence of the specific metabolites responsible for the medicinal properties associated with them. The study began in 1877 with the isolation of alkaloid lycorine from Narcissus pseudonarcissus and since then more than 100 alkaloids, exhibiting diverse biological activities, have been isolated from the Amaryllidaceae plants. Based on the present scientific evidence, it is likely that isocarbostyril constituents of the Amaryllidaceae, such as narciclasine, pancratistatin and their congeners, are the most important metabolites responsible for the therapeutic benefits of these plant species in the folk medical treatment of cancer. Notably, Narcissus poeticus L., used by the ancient Greek physicians, is now known to contain about 0.12 g of narciclasine per kg of fresh bulbs. The focus of the present research work is the chemistry and biology of these compounds as specifically relevant to their potential use in medicine. In particular, the anticancer evaluation of lycorine, narciclasine as well as of other Amaryllidaceae alkaloids and their synthetic derivatives are presented in this paper. The structure–activity relationships among some groups of Amaryllidaceae alkaloids will be discussed.
Keywords: Medicinal plants; Amaryllidaceae; Alkaloids; Lycorine; Narciclasine; Anticancer activity

Ascidians, invertebrates belonging to the subphylum Urochordata (Tunicata), are renowned for their great chemical diversity, and during the last 25 years, they have been shown to produce an array of cytotoxic molecules. Among the first six marine-derived compounds that have reached clinical trials as antitumor agents, three are derived from ascidians, as evidence of the high potential of these organisms as a new source of antitumor compounds. Reported in this communication are some recent results on the chemistry of Mediterranean ascidians; a number of new molecules with different structural features but all endowed with antiproliferative or cytotoxic activity are discussed. These results strongly evidence the highly significant role that Mediterranean ascidians natural products could play in anticancer drug discovery and development process.
Keywords: Marine natural products; Tunicates; Ascidians; Cytotoxicity; Anticancer activity

Biological activities and chemistry of saponins from Chenopodium quinoa Willd. by Tiwatt Kuljanabhagavad; Michael Wink (473-490).
Chenopodium quinoa Willd. is a valuable food source which has gained importance in many countries of the world. The plant contains various bitter-tasting saponins which present an important antinutritional factor. Various triterpene saponins have been reported in C. quinoa including both monodesmosidic and bidesmosidic triterpene saponins of oleanolic acid, hederagenin, phytolaccagenic acid, and serjanic acid as the major aglycones and other aglycones as 3β-hydroxy-23-oxo-olean-12-en-28-oic acid, 3β-hydroxy-27-oxo-olean-12-en-28-oic acid, and 3β, 23α, 30β-trihydroxy-olean-12-en-28-oic acid. A tridesmosidic saponin of hederagenin has also been reported. Here we review the occurrence, analysis, chemical structures, and biological activity of triterpene saponins of C. quinoa. In particular, the mode of action of the mono- and bidesmosidic triterpene saponins and aglycones are discussed.
Keywords: Quinoa; Oleanane triterpene; Phytolaccagenane triterpene; Triterpene saponins; Biological activity

Terminalia arjuna a sacred medicinal plant: phytochemical and pharmacological profile by Sunyana Jain; Prem Prakash Yadav; Vikrant Gill; Neeru Vasudeva; Neelam Singla (491-502).
Terminalia arjuna Wight & Arn. (Combretaceae) is a tree having an extensive medicinal potential. The plant is used traditionally in the treatment of various aliments. T. arjuna is a very good hypocholsteremic, hypolipidemic, anticoagulant, antihypertensive, antithrombotic, antiviral, antifungal and antibacterial agent Various parts of plant have been investigated for the presence of phytoconstituents and pharmacological activities. Many useful phytoconstituents have been isolated from T. arjuna. Triterpenoids are mainly responsible for cardiovascular properties. Tannins and flavonoids are responsible for its anticancer properties. The present review summarizes the ethnic use, pharmacological activities of the extracts and phytoconstituents of T. arjuna for last 90 years.
Keywords: Cardio protective; Phytoconstituents; Terminalia arjuna