Dopamine/serotonin receptor ligands. Part VIII[1]:the dopamine receptor antagonist LE300 - modelled and X-ray structure plus further pharmacological characterization, including serotonin receptor binding, biogenic amine transporter testing and in vivo testings
“…The idea behind the synthesis of the new heterocyclic system of 1 (Figure ) was to incorporate the substructures of tryptamine and β-phenylethylamine into a moderately constrained 10-membered azecine ring − 1 Structural variations of the lead compound 1 . R = CH 3 and other alkyl; R‘, R‘ ‘ = H, OCH 3 . …”
On the basis of the benz[d]indolo[2,3-g]azecine derivative 1 (LE300), structure-activity relations were investigated in order to identify the pharmacophore in this new class of ligands. Various structural modifications were performed and the inhibitory activities at human cloned D(1), D(2L), and D(5) receptors were measured by using a simple fluorescence microplate reader based calcium assay. Subsequently, the affinities of active compounds were estimated by radioligand binding experiments. Deleting one of the aromatic rings as well as replacing it by a phenyl moiety abolishes the inhibitory activities almost completely. Contraction of the 10-membered central ring decreases them significantly. The replacement of indole by thiophene or N-methylpyrrole reduces the inhibitory activity, whereas replacing the indole by benzene increases it. Finally, the hydroxylated dibenz[d,g]azecine derivative 11d (LE404) was found to be more active than the lead 1 in the functional calcium assay as well as in radioligand displacement experiments.
“…The idea behind the synthesis of the new heterocyclic system of 1 (Figure ) was to incorporate the substructures of tryptamine and β-phenylethylamine into a moderately constrained 10-membered azecine ring − 1 Structural variations of the lead compound 1 . R = CH 3 and other alkyl; R‘, R‘ ‘ = H, OCH 3 . …”
On the basis of the benz[d]indolo[2,3-g]azecine derivative 1 (LE300), structure-activity relations were investigated in order to identify the pharmacophore in this new class of ligands. Various structural modifications were performed and the inhibitory activities at human cloned D(1), D(2L), and D(5) receptors were measured by using a simple fluorescence microplate reader based calcium assay. Subsequently, the affinities of active compounds were estimated by radioligand binding experiments. Deleting one of the aromatic rings as well as replacing it by a phenyl moiety abolishes the inhibitory activities almost completely. Contraction of the 10-membered central ring decreases them significantly. The replacement of indole by thiophene or N-methylpyrrole reduces the inhibitory activity, whereas replacing the indole by benzene increases it. Finally, the hydroxylated dibenz[d,g]azecine derivative 11d (LE404) was found to be more active than the lead 1 in the functional calcium assay as well as in radioligand displacement experiments.
“…Our data suggest that transfected HEK-293 cells express 5-HT 2A R on the cell surface (fig 4B), and risperidone treatment has no significant effect on 5-HT 2A R immunoreactivity (data not shown). Further, ketanserin binding assay was conducted to analyze the effect of risperidone treatment on the binding of [Ethylene- 3 H]-labelled ketanserin hydrochloride, (Cat# R41 468, 25 μCi, 925 kBq) (PerkinElmer Life and Analytical Sciences, Inc, Waltham, MA) as described elsewhere (Decker et al, 2004). Briefly, HEK-293 cells were seeded on 100-mm plates and transfected with 12 μg 5-HT 2A R pDNA and 24 μL of LipofectamineTM 2000 transfection reagent (Invitrogen) according to the manufacturer’s instructions.…”
A recent report demonstrated that JCV employs serotonin receptor 2A (5HT 2A R) to infect the glial cells. To assess the ability of a potent 5HT 2A R blocker, risperidone, to inhibit JCV infection, we treated primary human fetal glial (PHFG) cells in-vitro with risperidone for 24 hr and inoculated with JCV(Mad1). There was no significant difference in JCV genome copies or mRNA transcripts and protein expression in treatment-naive and risperidone-treated PHFG cells. This data indicate that risperidone does not inhibit JCV(Mad1) attachment, internalization and replication in PHFG cells, and 5HT 2A R blockers may not be effective in treating progressive multifocal leukoencephalopathy (PML).
KeywordsJCV; human polyomavirus; PML; serotonin receptor; risperidone; mirtazapine Progressive multifocal leukoencephalopathy (PML), a fatal, subacute demyelinating disease of the central nervous system (CNS), is caused by human polyomavirus JC (JCV) (Padgett et al, 1971). Although, some AIDS-associated PML patients had survival benefit when treated with highly active antiretroviral therapy (HAART) (Antinori et al, 2003;Lima et al, 2007), in the post-HAART era incidence of PML has not significantly changed , and PML currently is the second most frequently diagnosed neurological disorder among AIDS patients . Few reports have demonstrated beneficial effect of cytosine arabinoside (Aksamit, 2001;De Luca et al, 1999;Elphick et al, 2004) and cidofovir in treating PML patients, however studies have concluded that cytosine arabinoside (Enting and Portegies, 2000) and cidofovir (Marra et al, 2002;Wyen et al, 2004) therapy have no significant therapeutic benefit. Although, clinical outcome among PML patients treated with interferon is controversial (Colosimo et al, 1992;Geschwind et al, 2001;Huang et al, 1998;Nath et al, 2006), IFN is effective in in-vitro inhibition of JCV replication and intrathecal infusion of IFN may be beneficial as an adjunct therapy for PML (Co et al, 2007). It is important to note that there is currently no proven therapy or vaccine available for treatment and prevention of the fatal disease, PML.A recent study demonstrated that the chimera polyomavirus JC (Mad-1/SVEΔ) consisting of JCV-SV40 promoter enhancer sequences in the backbone of JCV coding region sequences employs the 5HT 2A R to infect SVG-A cells, a subclone of transformed human fetal glial cells by an origin-defective SV40 mutant (Elphick et al, 2004). Altschuler and Kast (2005), have suggested that newer safer antipsychotic drugs such as risperidone, ziprasidone and olanzapine with significantly more potent 5HT 2A R receptor antagonist activity, may be useful in treating or preventing PML. Since, half of the healthy adult population shed large amounts of JCV in the urine (Agostini et al, 2001;Agostini et al, 1997;Agostini et al, 1996;Shah et al, 1997), this provides an unique opportunity to test safe and easy-to-tolerate drugs such as, risperidone and mirtazapine for anti-JCV activity, using urinary tract clearance of JCV as an additional su...
“…LE‐300, is 7‐methyl‐6,7,8,9,14,15‐hexahydro‐5H‐benz‐[d]indolo[2,3‐g]azecine (Fig. ), with nanomolar affinities to the hD receptor family, suppresses in vivo spontaneous locomotor activity and attenuates locomotor activity induced by cocaine.…”
Section: Introductionmentioning
confidence: 99%
“…LE‐300, is 7‐methyl‐6,7,8,9,14,15‐hexahydro‐5H‐benz‐[d]indolo[2,3‐g]azecine (Fig. ), with nanomolar affinities to the hD receptor family, suppresses in vivo spontaneous locomotor activity and attenuates locomotor activity induced by cocaine. LE‐300 was the most potent compound with a 10‐ to 20‐fold selectivity for D1 over D2L receptors compared with three of its azecine‐N‐substituted congeners combining structural elements of serotonin and dopamine .…”
A simple, rapid and highly sensitive spectrofluorimetric method was developed for determination of a novel type of dopamine receptor antagonist LE300 in mouse plasma. The method is based on measuring the native fluorescence of LE-300 in methanol at 343 nm after excitation at 280 nm. The fluorescence concentration plot was rectilinear over the range of 3.5-100 ng/mL with a lower detection limit of 1.0 ng/mL and quantification limit of 3.5 ng/mL. The method was statistically validated for linearity, accuracy, precision and selectivity according to the International Conference on Harmonization guidelines. The accuracy and precision results was expressed as % recovery and relative standard deviation (RSD). The accuracy for LE-300 was in the range 95.5-103.6% and RSD values were in the range of 0.21-1.55% of the theoretical value. The method was successfully applied to the analysis of LE-300 in mice plasma. The results were compared statistically with those obtained by the reported method and were found to be in good agreement, which could be applied in a pharmacokinetic study.
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