Las salidas del CRISP-DM

“BIOCHEMICAL TOOLS”/“PHARMACOLOGICAL PROBES”

Until the mid-20th century, the discovery of bioactive entities from plants was largely the result of a trial and error process, without much under- standing of their mechanism of action in vertebrate physiological sys- tems. Opium, the dried latex of the capsules of Papaver somniferum L., for instance, has been used for its analgesic properties for several millen- nia, with the major active narcotic analgesic alkaloid morphine (13) being isolated at the beginning of 19th century.104 Nevertheless, it was only in 1973 that the opioid receptors were characterized, thus helping to explain the mode of action of opiates.73 In a similar fashion, plant secondary metabolites have helped define many receptor types and have served as invaluable tools to help elucidate biological processes. These types of com- pounds are known as “biochemical tools” or “pharmacological probes.” The structures of the compounds mentioned in this section of the chapter are in Figs. 1–3. Compounds are described in turn, in terms of their activ- ities on the CNS, DNA, proteins, enzymes, and miscellaneous molecular targets.

The isolation of pure plant natural products, such as, pilocarpine (15), physostigmine (14), (−)-hyoscyamine (2), and nicotine (11), and the fungal isolate, muscarine (53), have played a major role in the devel- opment of modern pharmacology through enabling the discovery of intricate receptor-ligand mechanisms, which could not be probed ear- lier. Natural products, including some traditionally known to have psy-

choactive properties, have provided many potent CNS-active agents.20

Based on the contribution of plant psychoactive compounds in the under- standing of neurochemical processes, in silico methods for screening the

(53) Muscarine O N OH H (54) Salvinorin A O O O O O O O O H H H (55) Cathinone O NH2 (56) d-Tubocurarine NO HO (57) Muscimol (59) Picrotoxinin O O _O OH O O (58) Bicuculline N O O O O O H O N N O O H H H H H

(60) Strychnine (61) (-)-Epigallocatechin-3-gallate (EGCG)

O O OH OH OH OH OH OH O HO OH (62) Curcumin O O HO MeO OH OMe (63) Quisqualic acid HN O N O O CO2H NH₂ (64) Genistein O O OH OH HO O H OH O OH OH O H H O O (65) 12-O-Tetradecanoylphorbol-13-acetate H N HO MeO O (66) Capsaicin O N OH OMe H O N H MeO OH (67) Resiniferatoxin O OH O H _OH H O O O O OH (68) (-)-Menthol H NH2 OMe

Fig. 3. Structures of “biological tools” (“pharmacological probes”) from plants.

receptorome for plant-derived compounds have been suggested to further elucidate and define molecular mechanisms, along with methods to make them more compatible with modern assay systems, such as fluorescent modifications.60,105

The cannabinoid-type psychoactive principles of marijuana (Cannabis sativa L.), have been of interest to pharmacologists for many years.106 The biological characterization of the major euphoriant principle, (−)-∆9_{-trans-tetrahydrocannabinol (THC) (20), has led to an understand-}

ing of the molecular mechanisms of these compounds, and ultimately enabled the characterization of the cannabinoid receptor (CB1).107This

finding was followed by the discovery of the endogenous ligands, and also another cannabinoid receptor subtype, CB2. The first selective antagonist-

inverse agonist at the CB1receptor, rimonabant, was recently approved in

the European Union for the treatment of obesity and metabolic disorder.108 Cocaine (8), from Erythroxylum coca Lam., besides causing euphoria by inhibiting the dopamine transport protein (DAT) responsible for its recre- ational and illegal use, exerts a local anesthetic activity through blocking sodium channels and is still used as a probe for this target.109

The discovery of a potent and selectiveκ-opioid receptor agonist com- pound, salvinorin A (54), a hallucinogenic neoclerodane diterpenoid from Salvia divinorum Epling and Jativa, has created particular interest in recent years, since it is the first nonnitrogenous compound found to demonstrate this type of activity.110,111

Chiral protoalkaloids, such as cathinone (55), a psychotropic con- stituent of Catha edulis Forssk. (khat), have provided probes for studying the mechanistic properties of biogenic amine transporters, and afforded information regarding the effect of stereochemistry at the transportation level of these nitrogenous compounds.112

d -Tubocurarine (56), isolated from a South American plant used as an arrow poison, Chondodendron tomentosum Ruiz and Pav. (curare), while no longer employed as a skeletal muscle relaxant drug, is still used in pharma- cological studies as a competitive agonist of nicotinic acetylcholine recep- tors (nAChR) and also as an agonist of the 5-hydroxytryptamine (5-HT3)

receptor.48 Muscimol (57), a centrally acting principle of the mushroom Amanita muscaria (fly agaric), is known to be a potent GABAA receptor

agonist and led to the development of a bicyclic analogue, gaboxadol, a drug currently in phase III clinical trials for the treatment of sleep disor- ders. Bicuculline (58) (Dicentra cucullaria Bernh.) and picrotoxinin (59) (Cocculus indicus Royle), on the other hand, serve as potent antagonists

at GABAA receptors.48 Strychnine (60), the toxic principle of Strychnos

nux-vomica L., has proven to be useful in the studies on glycine (Gly) receptors as a competitive antagonist of this receptor type at nM levels.48 Members of the chalcone subtype of flavonoids have provided useful tools for probing the p53/MDM2 system through the inhibition of ubiquitin ligase.50Quisqualic acid (63) (Quisqualis indica L.) is a potent agonist for the ionotopic glutamate (iGlu) receptors.48

Epigenetic modulation is important for the treatment of cancer and the understanding of cancer cell dynamics. DNA-methylation is regulated by the DNA-methyl transferase (DNMT) family of enzymes which constitute potential targets with antiproliferative outcome in cancer chemotherapy. (−)-Epigallocatechin-3-gallate (EGCG) (61) (Camellia sinensis Kuntze), and other flavan derivatives have been shown to be DNMT1 inhibitors atµM levels.49 Curcumin (62) (Curcuma longa L.) has been found to act as a histone acetyl transferase (HAT) inhibitor, leading to transcriptional silencing.50

Podophyllotoxin (38) (Podophyllum peltatum L.), colchicine (9) (Colchicum autumnale L.), vinblastine (4), and vincristine (5) [Catharan- thus roseus (L.) G. Don] are standard microtubule-destabilizing agents used in cancer research.80,113Paclitaxel (21), from Taxus brevifolia Nutt., acts as a promoter of stabilization of microtubules and causes mitotic arrest in an unusual fashion.77,114

Genistein (64), an isoflavone found in plants of the family Legu- minosae, is an inhibitor of several protein tyrosine kinases (PTK) and is currently in phase II clinical trials for its potential as an angiogene- sis inhibitor.23,115 Genistein also has been utilized as a probe to identify binding sites for PTKs by observing the effect it demonstrates on cyclic- nucleotide-gated channels.116

Long-chain ester derivatives of phorbol, a tetracyclic diterpene from the seed oil of Croton tiglium L., including its most abundant representa- tive, 12-O-tetradecanoylphorbol-13-acetate (65), are potent activators of protein kinase C (PKC) and are used as standard tumor promoters for the study of experimental carcinogenesis in animal models.117

Investigations on capsaicin (66), the vanillyl-group containing “hot” principle in chili peppers (Capsicum spp.), as well as the irritant compound

resiniferatoxin (67) (Euphorbia resinifera Berg.), have led to the character- ization of transient receptor potential (TRP) channels and the vanilloid receptors (TRPV1).118These receptors show the possibility of serving as new targets to assist with diseases that involve chronic pain.119 While ligands acting on TRPV1 cause a burning sensation, compounds with a cooling effect, such as (−)-menthol (68) (Mentha spicata L.), have led to the characterization of cold sensors, most importantly the TRPM8 receptor, which also plays a role in the nociceptive process.22

2.6 STRATEGIES FOR MEDICINAL PLANT