Richard A. Glennon

Predicting new molecular targets for known drugs

Although drugs are intended to be selective, at least some bind to several physiological targets, explaining side effects and efficacy. Because many drug–target combinations exist, it would be useful to explore possible interactions computationally. Here we compared 3,665 US Food and Drug Administration (FDA)-approved and investigational drugs against hundreds of targets, defining each target by its ligands. Chemical similarities between drugs and ligand sets predicted thousands of unanticipated associations. Thirty were tested experimentally, including the antagonism of the β1 receptor by the transporter inhibitor Prozac, the inhibition of the 5-hydroxytryptamine (5-HT) transporter by the ion channel drug Vadilex, and antagonism of the histamine H4 receptor by the enzyme inhibitor Rescriptor. Overall, 23 new drug–target associations were confirmed, five of which were potent (<100 nM). The physiological relevance of one, the drug N,N-dimethyltryptamine (DMT) on serotonergic receptors, was confirmed in a knockout mouse. The chemical similarity approach is systematic and comprehensive, and may suggest side-effects and new indications for many drugs.

Evidence for 5-HT2 involvement in the mechanism of action of hallucinogenic agents

The affinities (Ki values) of twenty two psycho-active agents, including LSD, 5-OMe DMT and a series of phenalkylamine derivatives, for cortical 5-HT1 and 5-HT2 binding sites were compared with two measures of behavioral activity. It was found that a significant correlation (r = 0.938) exists between the 5-HT2 binding affinities of these agents and their ED50 values as determined in tests of stimulus generalization using 1-(2,5-dimethoxy-4-methylphenyl)-2-aminopropane (DOM) as the training drug. Furthermore, for fifteen of these agents where human data were available, a significant correlation (r = 0.924) also exists between 5-HT2 binding affinities and their human hallucinogenic potencies. The results of this study suggest that the mechanism of action of these agents involves 5-HT2-related events.

Radioligand binding evidence implicates the brain 5-HT2 receptor as a site of action for LSD and phenylisopropylamine hallucinogens

Alterations in brain serotonergic function have been implicated in the mechanism of action of LSD, mescaline, and other similarly acting hallucinogenic drugs of abuse such as STP (2,5-dimethoxyphenylisopropylamine; DOM). In order to test the hypothesis that the mechanism of action of LSD and phenylisopropylamine hallucinogens is through stimulation of a specific brain serotonin receptor sub-type, the affinities of these compounds for radiolabelled 5-HT2, 5-HT1A, 5-HT1B, and 5-HT1C receptors have been determined using recently developed in vitro radioligand binding methodologies. The 5-HT2 receptor was labelled with the agonist/hallucinogen radioligand 3H-DOB (4-bromo-2,5-dimethoxyphenylisopropylamine). The 5-HT1A, 5-HT1B, and 5-HT1C receptors were labelled with 3H-OH-DPAT, 3H-5-HT, and 3H-mesulergine, respectively. In general, the phenylisopropylamines displayed 10–100 fold higher affinities for the 5-HT2 receptor than for the 5-HT1C receptor and 100–1000 fold higher affinities for the 5-HT2 receptor than for the 5-HT1A or 5-HT1B receptor. There was a strong correlation between hallucinogenic potencies and 5-HT2 receptor affinities of the phenylisopropylamines (r=0.90); the correlation coefficients for the 5-HT1A, 5-HT1B, and 5-HT1C were 0.73, 0.85, and 0.78, respectively. Because there is no evidence that 5-HT1A-selective or 5-HT1B-selective agonists are hallucinogenic and because the phenylisopropylamines are potent hallucinogens, a 5-HT2 receptor interaction is implicated and supports our previous suggestions to this effect. A secondary role for 5-HT1C receptors cannot be discounted at this time. These results, when integrated with other receptor pharmacological information, indicate that an important component of the mechanism of action of LSD and the phenylisopropylamine hallucinogens is through stimulation of brain 5-HT2 receptors.

Do functional relationships exist between 5-HT1A and 5-HT2 receptors?

To investigate the possible functional relationship between 5-HT1 and 5-HT2 receptors, we studied the effects of a nonselective 5-HT agonist (5-MeO DMT), a 5-HT1A-selective (8-OH-DPAT) and a 5-HT1B/5-HT1C-selective (TFMPP) agonist on the head-twitch behavior induced by the putative 5-HT2-selective receptor agonist (+/-)-DOI. In the mouse (+/-)-DOI produced the head-twitch response in a dose-dependent manner and (-)-DOI was twice as potent as the (+) isomer. Selective 5-HT2 antagonists, ketanserin and spiperone, dose-dependently inhibited the (+/-)-DOI-induced head-twitch response. The nonselective and the 5-HT1A-selective agonists also dose-dependently reduced the behavior, whereas 5-HT1B/5-HT1C-selective agonist (TFMPP) failed to affect the (+/-)-DOI-induced response. Taken together with previously published literature data, we propose a 5-HT1A inhibitory action on the 5-HT2 receptor-mediated response when induced by its selective agonist (+/-)-DOI.

Serotonin receptors: Clinical implications

Over the past decade, a variety of serotonin (5-hydroxytryptamine, 5-HT) receptor/binding sites have been identified. These include 5-HT1, 5-HT2, and 5-HT3 sites. The 5-HT1 sites have been further divided into 5-HT1A, 5-HT1B, 5-HT1C, 5-HT1D and 5-HT1E sites. It would be of interest to identify those pharmacological effects that are specifically mediated by a particular population of 5-HT sites and, indeed, attempts have been made to do this almost since the initial discovery of multiple populations of sites. Unfortunately, much of the early work made use of serotonergic agents that are now known to be somewhat less selective than originally suspected. Nevertheless, there is ample information in the literature suggesting that site-selective serotonergic agents may ultimately lead (and, in some cases, has already led) to the development of therapeutically-useful agents. The present review examines the pharmacological effects that are thought to be related to the individual types of 5-HT sites and provides some clinical implications for agents that act at these sites.

NAN-190: an arylpiperazine analog that antagonizes the stimulus effects of the 5-HT1A agonist 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT)

In order to determine whether NAN 190 might serve as a 5-HT 1A antagonist, it was evaluated in rats trained to discriminate 8-OH-DPAT HBr from saline in a 2-lever operant procedure

Antagonism of the effects of the hallucinogen dom and the purported 5-HT agonist quipazine by 5-HT2 antagonists

Rats trained to discriminate 1.0 mg/kg of 1-(2,5-dimethoxy-4-methylphenyl)-2-aminopropane (DOM) from saline in a two-lever operant choice task were administered doses of mescaline, LSD, 5-methoxy-N,N-dimethyltryptamine (5-OMe DMT), quipazine, TFMPP and RU-24969. The DOM-stimulus generalized to the three hallucinogenic agents and to quipazine, but not to the purported serotonin agonists TFMPP or RU-24969. Pretreatment of the animals with the 5-HT2 antagonists ketanserin and pirenperone antagonized the effect produced by DOM. Pirenperone also blocked DOM-stimulus generalization to mescaline, LSD, 5-OMe DMT and quipazine. The results of this study suggest that the discriminative stimulus effects of DOM, the three hallucinogenic agents to which DOM-stimulus generalization occurred, and quipazine, may involve those sub-populations of serotonin receptors that are labeled by tritiated ketanserin (i.e. 5-HT2 sites).

Binding of β-carbolines and related agents at serotonin (5-HT2 and 5-HT1A), dopamine (D2) and benzodiazepine receptors

A large series of β-carbolines was examined for their ability to bind at [3H]agonist-labeled 5-HT2A serotonin receptors. Selected β-carbolines were also examined at 5-HT2C serotonin receptors, 5-HT1A serotonin receptors, dopamine D2 receptors, and benzodiazepine receptors. Indolealkylamines and phenylisopropylamines were also evaluated in some of these binding assays. The β-carbolines were found to bind with modest affinity at 5-HT2A receptors, and affinity was highly dependent upon the presence of ring substituents and ring saturation. The β-carbolines displayed little to no affinity for 5-HT1A serotonin receptors, dopamine D2 receptors and, with the exception of β-CCM, for benzodiazepine receptors. Examples of β-carbolines, indolealkylamines (i.e. N,N-dimethyltryptamine analogs), and phenylisopropylamines have been previously shown to produce common stimulus effects in animals trained to discriminate the phenylisopropylamine hallucinogen DOM (i.e. 1-(2,5-dimethoxy-4-methylphenyl)-2-aminopropane) from vehicle. Although the only common receptor population that might account for this action is 5-HT2A, on the basis of a lack of enhanced affinity for agonist-labeled 5-HT2A receptors, as well as on their lack of agonist action in the PI hydrolysis assay, it is difficult to conclude that the β-carbolines behave in a manner consistent with that of other classical hallucinogens.

5-HT1 and 5-HT2 binding properties of derivatives of the hallucinugen 1-(2,5-dimethoxyphenyl)-2-aminopropane (2,5-DMA)

The affinities of a series of 1-(2,5-dimethoxyphenyl)-2-aminopropane (2,5-DMA) derivatives, most of which are hallucinogenic in man, and several related agents were determined for rat cortical serotonin (5-HT) binding sites. Competition assays were performed in which these agents were competed for the 5-HT2 binding of [3H]ketanserin, or the 5-HT1 binding of [3H]LSD (in the presence of ketanserin). The R(-)-isomers of DOI, DOM and DON (i.e. the 4-iodo, -methyl and -nitro derivatives of 2,5-DMA) were found to be more potent than their racemates and demonstrated selectivity for 5-HT2 sites. These same agents in competing for [3H]ketanserin binding resulted in Hill coefficients significantly less than unity; computer-assisted analysis indicated a two-state model better fit the data. In the presence of 10(-4) M Gpp(NH)p the competition curve for R(-)-DOI produced a Hill coefficient close to unity. These results are consistent with the hypothesis that certain derivatives of 2,5-DMA, in particular R(-)-DOI, may be potent and selective agonists at 5-HT2 binding sites, sites that may constitute a serotonin receptor that is regulated by a guanine nucleotide regulatory protein. Conversely, the interactions of these agents at 5-HT1 sites was with a lower affinity and a lack of stereoselectivity. Although DOI and DOM are amongst the most potent of these agents as hallucinogens, it is still too premature to draw any conclusions regarding a possible relationship between 5-HT binding and hallucinogenic potency.

Selectivity of serotonergic drugs for multiple brain serotonin receptors: Role of [3H]-4-bromo-2,5-dimethoxyphenylisopropylamine ([3H]DOB), A 5-HT2 agonist radioligand

The affinities of putative serotonin receptor agonists and antagonists for 5-HT1A, 5-HT1B, 5-HT1C, and 5-HT2 receptors were assayed using radioligand binding assays. The 5-HT1 sites were labeled with the agonist radioligands [3H]-8-hydroxy-2-(di-n-propylamino)-tetralin [3H]-8-OH-DPAT, [3H]-5-HT, and [3H]mesulergine. The 5-HT2 receptor was labeled with the antagonist radioligand [3H]ketanserin or the agonist radioligand [3H]-4-bromo-2,5-dimethoxyphenylisopropylamine ([3H]DOB). The apparent 5-HT1 receptor selectivity of agonist compounds was found to be 50- to 100-fold higher when the 5-HT2 receptor affinity was determined using the antagonist radioligand [3H]ketanserin than when the agonist radioligand [3H]DOB was used. Quipazine, a putative specific 5-HT2 agonist, appeared to be only 3-fold more potent at 5-HT2 than at 5-HT1A receptors when [3H]ketanserin was used as the 5-HT2 radioligand. When [3H]DOB was used as the 5-HT2 radioligand, quipazine was determined to be 100-fold more potent at 5-HT2 receptors than at 5-HT1A receptors. 1-(3-trifluoromethylphenyl)piperazine (TFMPP), a putative specific 5-HT1B receptor agonist was apparently 10-fold more potent at 5-HT1B receptors than at 5-HT2 receptors when [3H]ketanserin was used as the 5-HT2 radioligand. When [3H]DOB was used as the 5-HT2 radioligand, TFMPP was found to be equipotent at 5-HT1B and 5-HT2 receptors. Using the 5-HT2 antagonist radioligand [3H]ketanserin, a similar pattern of underestimating 5-HT2 receptor selectivity and/or overestimating 5-HT1A or 5-HT1B receptor selectivity was observed for a series of serotonin receptor agonists. Antagonist receptor selectivity was not affected significantly by the nature of the 5-HT2 receptor assay used. These data indicate that, by using an antagonist radioligand to label 5-HT2 receptors and agonist radioligands to label 5-HT1 receptors, the 5-HT1 receptor selectivity may be overestimated. This may be an especially severe problem in serotonin drug development as drugs that interact potently with 5-HT2 receptors have been reported to be psychoactive and/or hallucinogenic.