ABSTRACT
Aromatic iodination ortho to the phenolic hydroxyl reverts the activity of the ultrapotent vanilloid agonist resiniferatoxin (RTX, 1a), generating the ultrapotent antagonist 5'-iodoRTX (1b). To better understand the role of iodine in this remarkable switch of activity, a systematic investigation on the halogenation of vanillamides, a class of compounds structurally simpler than resiniferonoids, was carried out. The results showed that (a) the antagonistic activity depends on the site of halogenation and is maximal at C-6', (b) iodine is more efficient than chlorine and bromine at reverting the agonistic activity, and (c) iodine-carbon exchange decreases antagonist activity. Iodine-induced reversal of vanilloid activity was also observed in vanillamides more powerful than capsaicin, but a poor correlation was found between agonistic and antagonistic potencies, suggesting that differences exist in the way vanillamides and their 6'-iodo derivatives bind to TRPV1.' '
Subject(s)
Capsaicin/analogs & derivatives , Capsaicin/chemical synthesis , Iodine/chemistry , Ion Channels/antagonists & inhibitors , Bromine/chemistry , Calcium/metabolism , Capsaicin/chemistry , Capsaicin/pharmacology , Cell Line , Chlorine/chemistry , Diterpenes/chemistry , Diterpenes/pharmacology , Humans , Ion Channels/agonists , Structure-Activity Relationship , TRPV Cation ChannelsABSTRACT
Olvanil (N-9-Z-octadecenoyl-vanillamide) is an agonist of transient receptor potential vanilloid type 1 (TRPV1) channels that lack the pungency of capsaicin and was developed as an oral analgesic. Vanillamides are unmatched in terms of structural simplicity, straightforward synthesis, and safety compared with the more powerful TRPV1 agonists, like the structurally complex phorboid compound resiniferatoxin. We have modified the fatty acyl chain of olvanil to obtain ultra-potent analogs. The insertion of a hydroxyl group at C-12 yielded a compound named rinvanil, after ricinoleic acid, significantly less potent than olvanil (EC(50) = 6 versus 0.7 nM), but more versatile in terms of structural modifications because of the presence of an additional functional group. Acetylation and phenylacetylation of rinvanil re-established and dramatically enhanced, respectively, its potency at hTRPV1. With a two-digit picomolar EC(50) (90 pM), phenylacetylrinvanil (PhAR, IDN5890) is the most potent vanillamide ever described with potency comparable with that of resiniferatoxin (EC(50), 11 pM). Benzoyl- and phenylpropionylrinvanil were as potent and less potent than PhAR, respectively, whereas configurational inversion to ent-PhAR and cyclopropanation (but not hydrogenation or epoxidation) of the double bond were tolerated. Finally, iodination of the aromatic hydroxyl caused a dramatic switch in functional activity, generating compounds that behaved as TRPV1 antagonists rather than agonists. Since the potency of PhAR was maintained in rat dorsal root ganglion neurons and, particularly, in the rat urinary bladder, this compound was investigated in an in vivo rat model of urinary incontinence and proved as effective as resiniferatoxin at reducing bladder detrusor overactivity.
Subject(s)
Capsaicin/analogs & derivatives , Capsaicin/therapeutic use , Ion Channels/agonists , Amidohydrolases/metabolism , Animals , Animals, Newborn , Anti-Inflammatory Agents, Non-Steroidal/chemistry , Anti-Inflammatory Agents, Non-Steroidal/pharmacology , Arachidonic Acids/metabolism , Capsaicin/chemical synthesis , Capsaicin/chemistry , Capsaicin/pharmacology , Carrier Proteins/metabolism , Cell Line, Tumor , Endocannabinoids , Female , Ganglia, Spinal/cytology , Ganglia, Spinal/drug effects , Humans , In Vitro Techniques , Indicators and Reagents , Neurons/drug effects , Polyunsaturated Alkamides , Rats , Rats, Sprague-Dawley , Receptor, Cannabinoid, CB1/drug effects , Receptor, Cannabinoid, CB2/drug effects , Structure-Activity Relationship , TRPV Cation Channels , Urinary Bladder/drug effects , Urinary Incontinence/drug therapyABSTRACT
1. The C-5 halogenation of the vanillyl moiety of resiniferatoxin, an ultrapotent agonist of vanilloid TRPV1 receptors, results in a potent antagonist for these receptors. Here, we have synthesized a series of halogenated derivatives of 'synthetic capsaicin' (nonanoyl vanillamide=nordihydrocapsaicin) differing for the nature (iodine, bromine-chlorine) and the regiochemistry (C-5, C-6) of the halogenation. 2. The activity of these compounds was investigated on recombinant human TRPV1 receptors overexpressed in HEK-293 cells. None of the six compounds exerted any significant agonist activity, as assessed by measuring their effect on TRPV1-mediated calcium mobilization. Instead, all compounds antagonized, to various extents, the effect of capsaicin in this assay. 3. All 6-halo-nordihydrocapsaicins behaved as competitive antagonists against human TRPV1 according to the corresponding Schild's plots, and were more potent than the corresponding 5-halogenated analogues. The iodo-derivatives were more potent than the bromo- and chloro-derivatives. 4. Using human recombinant TRPV1, 6-iodo-nordihydrocapsaicin (IC(50)=10 nM against 100 nM capsaicin) was about four times more potent than the prototypical TRPV1 antagonist, capsazepine, and was tested against capsaicin also on native TRPV1 in: (i) rat dorsal root ganglion neurons in culture; (ii) guinea-pig urinary bladder; and (iii) guinea-pig bronchi. In all cases, except for the guinea-pig bronchi, the compound was significantly more potent than capsazepine as a TRPV1 antagonist. 5. In conclusion, 6-iodo-nordihydrocapsaicin, a stable and easily prepared compound, is a potent TRPV1 antagonist and a convenient replacement for capsazepine in most of the in vitro preparations currently used to assess the activity of putative vanilloid receptor agonists.
Subject(s)
Capsaicin/analogs & derivatives , Capsaicin/pharmacology , Hydrocarbons, Halogenated/pharmacology , Receptors, Drug/antagonists & inhibitors , Animals , Calcium/metabolism , Capsaicin/chemistry , Cell Line , Dose-Response Relationship, Drug , Ganglia, Spinal/cytology , Guinea Pigs , Humans , Hydrocarbons, Halogenated/chemistry , Kidney/cytology , Kidney/embryology , Muscle Contraction/drug effects , Muscle, Smooth/drug effects , Neurons/drug effects , Neurons/metabolism , Rats , Recombinant Proteins/antagonists & inhibitors , Structure-Activity Relationship , Trachea/drug effects , Transfection , Urinary Bladder/drug effectsABSTRACT
N-Arachidonoyldopamine (NADA) was recently identified as an endogenous ligand for the vanilloid type 1 receptor (VR1). Further analysis of the bovine striatal extract from which NADA was isolated indicated the existence of substances corresponding in molecular mass to N-oleoyldopamine (OLDA), N-palmitoyldopamine (PALDA), and N-stearoyldopamine (STEARDA). Quadrupole time-of-flight mass spectrometric analysis of bovine striatal extracts revealed the existence of OLDA, PALDA, and STEARDA as endogenous compounds in the mammalian brain. PALDA and STEARDA failed to affect calcium influx in VR1-transfected human embryonic kidney (HEK) 293 cells or paw withdrawal latencies from a radiant heat source, and there was no evidence of spontaneous pain behavior. By contrast, OLDA induced calcium influx (EC(50) = 36 nm), reduced the latency of paw withdrawal from a radiant heat source in a dose-dependent manner (EC(50) = 0.72 microg), and produced nocifensive behavior. These effects were blocked by co-administration of the VR1 antagonist iodo-resiniferatoxin (10 nm for HEK cells and 1 microg/50 micro;l for pain behavior). These findings demonstrate the existence of an endogenous compound in the brain that is similar to capsaicin and NADA in its chemical structure and activity on VR1. Unlike NADA, OLDA was only a weak ligand for rat CB1 receptors; but like NADA, it was recognized by the anandamide membrane transporter while being a poor substrate for fatty-acid amide hydrolase. Analysis of the activity of six additional synthetic and potentially endogenous N-acyldopamine indicated the requirement of a long unsaturated fatty acid chain for an optimal functional interaction with VR1 receptors.
Subject(s)
Capsaicin/chemistry , Dopamine/analogs & derivatives , Dopamine/pharmacology , Hyperalgesia/etiology , Animals , Arachidonic Acids/pharmacology , Brain/metabolism , Calcium/metabolism , Calcium Channel Blockers/pharmacology , Capsaicin/pharmacology , Cattle , Cell Line , Cell Membrane/metabolism , Cytosol/metabolism , Diterpenes/pharmacology , Dopamine/chemistry , Dose-Response Relationship, Drug , Endocannabinoids , Humans , Inhibitory Concentration 50 , Ions , Lipids , Male , Mass Spectrometry , Models, Chemical , Polyunsaturated Alkamides , Protein Binding , Rats , Rats, Sprague-Dawley , Receptors, Drug/antagonists & inhibitors , TRPV Cation Channels , Temperature , Time FactorsABSTRACT
BACKGROUND: Capsiate, the non-pungent ester isoster of capsaicin, and its dihydroderivative are the major capsaicinoids of sweet peppers. The remarkable difference between the sensory properties of capsaicin vs capsiate is solely due to the way the vanillyl and the acyl moieties of this basic structural motif are linked, via an amide bond in capsaicin-type compounds and via an ester bond in capsiate-type compounds. AIM OF THE STUDY: Since capsaicin induces apoptosis in tumoral cells by a vanilloid receptor type 1(VR1)-independent pathway, we examined the effects of capsiates derived from sweet peppers in the ROS generation and induction of apoptosis in tumoral cells and if these are mediated independently from VR1. METHODS: We have developed an expeditious synthesis of capsiates based on the esterification of vanillol with the Mitsunobu protocol. Capsiate-induction of apoptosis, generation of reactive oxygen species and disruption of the mitochondria transmembrane potential in tumoral cell lines were measured by flow cytometry. Chemopreventive activity was studied in a two-stage mouse skin carcinogenesis assay. RESULTS: Capsiates induce apoptosis that was preceded by an increase in the production of reactive oxygen species and by a subsequent loss of mitochondria transmembrane potential (DeltaPsi(m)). These properties were retained in simplified synthetic analogues of natural capsiates, one of which (nor-dihydrocapsiate) showed powerful chemopreventive activity. CONCLUSIONS: These results suggest that capsiates and related synthetic analogues target a variety of pathways involved in cancer development and inflammation, and have considerable potential for dietary health benefits as well as for pharmaceutical development.
Subject(s)
Apoptosis/drug effects , Capsaicin/analogs & derivatives , Capsaicin/pharmacology , Capsicum/chemistry , 9,10-Dimethyl-1,2-benzanthracene/toxicity , Animals , Anticarcinogenic Agents/chemistry , Anticarcinogenic Agents/pharmacology , Anticarcinogenic Agents/therapeutic use , Capsaicin/chemistry , Capsaicin/therapeutic use , Carcinogens/toxicity , Cell Line , Chemoprevention , Flow Cytometry , Humans , Jurkat Cells , Membrane Potentials/drug effects , Mice , Papilloma/chemically induced , Papilloma/prevention & control , Reactive Oxygen Species/metabolism , Receptors, Drug/drug effects , Receptors, Drug/metabolism , Skin Neoplasms/chemically induced , Skin Neoplasms/prevention & control , Tumor Cells, CulturedABSTRACT
Two regioisomers and 13 analogues of the putative endocannabinoid noladin ether (2-arachidonyl glyceryl ether, 2-AGE, 1) were synthesized and tested for their interaction with CB(1) receptors in rat brain membranes. The results showed that a C-20 tetra-unsaturated moiety is necessary for high affinity, and that a series of alkyl glyceryl ethers of potential occurrence in brain tissues have less affinity than 2-AGE for CB(1) receptors.
Subject(s)
Fatty Acids, Unsaturated/chemical synthesis , Glycerides/chemistry , Receptors, Drug/chemistry , Animals , Brain/ultrastructure , Cannabinoid Receptor Modulators , Cell Membrane/chemistry , Endocannabinoids , Fatty Acids, Unsaturated/chemistry , Fatty Acids, Unsaturated/metabolism , Glycerides/metabolism , Protein Binding , Radioligand Assay , Rats , Receptors, Cannabinoid , Receptors, Drug/metabolism , Stereoisomerism , Structure-Activity RelationshipABSTRACT
[formula: see text] The Mitsunobu reaction can distinguish between alcohol and phenol hydroxyls in esterification reactions, providing an expeditious and broadly applicable entry into various phenolics and polyphenolics of biomedical and nutritional relevance.