5-HT₆ Receptor ( pKi = 9.63 )

Intepirdine (SB742457) is a potent and selective 5-HT6 receptor antagonist that has been demonstrated to improve elderly rats' performance in a water maze task and to reverse a learning deficit caused by scopolamine in the novel object recognition test.[1]

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The present study aimed to examine the chronic effects of the selected APDs (haloperidol, risperidone, olanzapine), administered alone and in combination with a selective 5-HT6 agonist (WAY-181187) or antagonist (Intepirdine (SB742457)), on weight gain, food intake, serum lipid profile, glucose level, and a spectrum of hormones derived from adipose (leptin, adiponectin) and gastrointestinal (insulin, ghrelin) tissue in rats. SB-742457 inhibited increased weight gain and alleviated hyperglycemia induced by APDs more strongly than did WAY-181187, but also intensified dyslipidemia. WAY-181187 tended to improve the lipid profile, but increased the glucose level. The greatest benefits were obtained when WAY-181187 or Intepirdine (SB742457) were co-administered with haloperidol. It is difficult to assess whether the modification of the serum levels of insulin, leptin, ghrelin, and adiponectin depended on the treatment applied or other drug-independent factors; therefore, further research is needed. [2]

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The aim of the present study was to investigate and compare the effects of acute and chronic (21-day) administration of agonist (WAY-181187) and antagonist (Intepirdine (SB742457)) of the 5-hydroxytryptamine 6 receptor (5-HT6R) on MK-801-induced memory impairments in novel object recognition (NORT) and Y-maze continuous spontaneous alternation tests (Y-CAT). Further, the expression of the brain-derived neurotrophic factor (BDNF) in rat hippocampus was measured after 21-day administration to investigate BDNF participation in the pro-cognitive effects of 5-HT6R ligands. We found that acute administration of WAY-181187, as well as SB-742457, reversed the effects of MK-801 in NORT and Y-CAT, and that this influence persisted after prolonged application in NORT but not in Y-CAT. Both 5-HT6R ligands increased hippocampal BDNF protein expression, but WAY-181187 was much more potent than SB-742457 and alleviated the MK-801-induced inhibition of BDNF signaling pathways better, which seems to translate into a stronger WAY-181187 effect in behavioral tests. Collectively, both the 5-HT6R agonist and the antagonist, administered acutely and chronically, prevent memory impairments and alterations in BDNF signaling induced by MK-801 in rats. The present results confirm the pro-cognitive properties of both types of 5-HT6R ligands and suggest that BDNF pathways may be involved in their mechanism of action [3].

Group 2 (n = 6) was used for pharmacologic characterization of the regional binding of 11C-GSK215083. In addition to its affinity for 5HT6 receptors, the PET ligand 11C-GSK215083 also possesses an approximately 5-fold-lower affinity for the 5HT2A receptor (3). Therefore, we used ketanserin, a selective 5HT2A antagonist, to test for the existence of detectable binding to 5HT2A receptors and SIntepirdine (SB742457)to detect binding to 5HT6 receptors. Four subjects underwent a baseline 11C-GSK215083 scan followed by 2 further PET scans, after SB742457 (175 mg, orally, 5 h before 11C-GSK215083 administration) and after ketanserin (0.1 mg/kg, by slow intravenous injection, 2 h before 11C-GSK215083 administration), respectively, in a randomized fashion, 7 d apart. The remaining 2 subjects received a baseline scan, a second scan after SB742457 (175 mg), and a third scan, 7 d later (Fig. 1B).
Group 3 (n = 8) characterized the time and dose occupancy relationship of Intepirdine (SB742457) in the brain. Each subject underwent 3 PET scans. Scan 1 was a baseline scan with scans 2 and 3 following repeated once-daily dosing of SB742457 on 2 occasions between 7 and 28 d of dosing. Four subjects received a loading dose of SB742457 (175 mg) on day 1, followed by 35 mg daily for 21 or 28 d (2 subjects per group). Subjects were randomized to undergo 2 scans on days 7, 14, 21, and 28, such that 2 independent data points were collected for each time point (Fig. 1C). Similarly, 2 further subjects received SB742457 (70 mg) followed by 15 mg daily for 14 d, and the last 2 subjects received 15 mg followed by 3 mg daily for 14 d. All 4 subjects were scanned on days 7 and 14 (Fig. 1D). Scans were conducted approximately 5 h after SB742457 dosing.
Safety assessments included medical history, physical examination, adverse event reporting, clinical laboratory assessments and electrocardiogram, blood pressure, and pulse rate measurements (supplemental information). Blood samples were collected for pharmacokinetic analysis of SB742457 in plasma from subjects in groups 2 and 3 before and on completion of each scan for which Intepirdine (SB742457) was administered (supplemental information). https://pubmed.ncbi.nlm.nih.gov/26383152/

Western blot analysis [3]
Hippocampus samples were homogenized using T-PER mammalian protein extraction reagent with protease and phosphatase inhibitors. Protein concentrations was determined using the Bradford reaction. Aliquots (40 μg) were solubilise in Laemmli buffer with 2% 2-mercaptoethanol, and were subject to 10% SDS-polyacrylamide gel electrophoresis. The anti-brain derived neurotrophic factor (BDNF, 15 kDa) diluted 1:500 and anti-β-actin diluted 1:1000 were used. The secondary antibody was anti rabbit IgG (HRP) diluted 1:2000.

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Quantitative real-time PCR [3]
RNA was extracted from tissues using RNA Isolation kit. After quantity and quality evaluation, RNA concentration was normalized to 15 ng/µL. Reverse transcription was done with a High-Capacity Reverse Transcription Kit. qPCR 96-wells reaction plate was performed with TaqMan primers and probes for BDNF (Rn02531967_s1) according to the manufacturer’s protocol on The Applied Biosystems® 7500 Fast Real-Time PCR Instrument. Endogenous control genes Gapdh (Rn01775763_g1) and Tbp (Rn01455646_m1) were selected on the basis of the pilot experiment. Relative expression was calculated using ΔΔCq method.

A total of 140 rats were used in the study, and each treatment group consisted of 10 randomly selected animals. Due to the large number of animals and limited laboratory space, the experiment was carried out in three turns: the first included 4 treatment groups: vehicle (1% Tween 80), haloperidol, risperidone, and olanzapine; the second consisted of 5 treatment groups: vehicle (1% Tween 80), WAY-181187, haloperidol + WAY-181187, risperidone + WAY-181187, and olanzapine + WAY-181187; and the third consisted of 5 treatment groups: vehicle (1% Tween 80), Intepirdine (SB742457), haloperidol + SB-742457, risperidone + SB-742457, and olanzapine + SB-742457. One animal died during the administration of the compounds; therefore, one experimental group (i.e., risperidone + WAY-181187-treated group) eventually consisted of only 9 animals. 24 hours after the last drug administration, trained personnel sacrificed the rats by dislocating the cervical spinal cord.[2]

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Haloperidol risperidone, olanzapine, WAY-181187 (oxalate), and Intepirdine (SB742457) were used in the experiment. Doses of APDs (haloperidol 0.5 mg/kg, risperidone 0.5 mg/kg, and olanzapine 5 mg/kg) and 5-HT6 ligands (WAY-181187 3 mg/kg and SB-742457 3 mg/kg) were selected for the experiments, based on literature review and our previous studies which presented their separate and combined behavioral effects. The compounds were suspended in a 1% solution of Tween 80 (Sigma Aldrich, St. Louis, MO, USA) immediately before administration and injected intraperitoneally (ip) in a volume of 2 mL/kg. The compounds were dispensed to the rats once daily between 10:00 and 11:00 a.m. for 28 days. The last injection was given 24 h before sacrifice. The control rats received 1% Tween 80, on the same dosing regimen.[2]

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The following drugs were used: WAY-181187 (oxalate), Intepirdine (SB742457), (+)-MK-801 (hydrogen maleate). All the compounds, except for MK-801 which was dissolved in distilled water, were suspended in 1% solution of Tween 80 immediately before administration, and were injected i.p. in a volume of 2 ml/kg. In acute experiments 5-HT6R ligands were injected 60 min before testing, while in chronic tests once a day during consecutive 21 days, with the last injection 24 h before the test. MK-801 was administered only once, 30 min before the tests. Control rats received vehicle according to the same schedule. The doses of drugs refer to their salt forms. [3]

[1]. 5-HT6 receptor antagonists as novel cognitive enhancing agents for Alzheimer's disease. Neurotherapeutics. 2008 Jul;5(3):458-69.

[2]. Selective 5-HT6 Receptor Ligands (Agonist and Antagonist) Show Different Effects on Antipsychotic Drug-Induced Metabolic Dysfunctions in Rats. Pharmaceuticals (Basel). 2023 Jan 20;16(2):154.

[3]. 5-HT6 receptor agonist and antagonist improve memory impairments and hippocampal BDNF signaling alterations induced by MK-801. Brain Res. 2019 Nov 1:1722:146375.

Intepirdine has been used in clinical trials for the treatment of Alzheimer's disease. Intepirdine is a small molecule drug that has completed up to three Phase III clinical trials (covering all indications) and has two investigational indications. In summary, the results clearly indicate whether adding a selective 5-HT6 receptor agonist or antagonist can provide further benefit for metabolic disorders following APD. The greatest benefit was observed when a 5-HT6 receptor ligand was used in combination with haloperidol. Unlike risperidone (Ki = 420 nM) and olanzapine (Ki = 2.5 nM), haloperidol has no affinity for the 5-HT6 receptor (Ki > 5000 nM). WAY-181187 normalized serum changes in appetite and metabolic activity induced by haloperidol and reduced weight gain and food intake to some extent; while Intepirdine (SB742457) significantly reduced weight gain and food intake and was unlikely to alter hormone levels. Overall, SB-742457 is more effective at inhibiting weight gain and alleviating hyperglycemia caused by antipsychotic drugs, but it should be noted that it can also exacerbate dyslipidemia. On the other hand, WAY-181187 tends to improve the lipid profile, but it can increase blood glucose levels. In addition, it is difficult to assess whether changes in serum insulin, leptin, ghrelin and adiponectin levels are dependent on the treatment method used or on other factors unrelated to the drug (e.g., weight gain, daily activity level, adipose tissue content). Therefore, further research is needed. [2] MK-801 is a non-competitive NMDA receptor antagonist (Wong et al., 1986) that can induce cognitive impairments similar to those associated with dementia (Ellison, 1995) and schizophrenia (Bubeníková-Valešová et al., 2008). MK-801-induced memory deficit models have been widely used in preclinical cognitive studies (van der Staay et al., 2011). Numerous animal studies demonstrating MK-801 sensitivity, including the NORT and Y-CAT tests, have been conducted, reflecting the natural exploratory behavior of rodents (Dix and Aggleton, 1999; Lalonde, 2002). In this study, rats exhibited memory impairment in the NORT and Y-CAT behavioral tests following acute injection of MK-801. These tests reflect episodic memory and spatial working memory processes, respectively. Subsequently, we investigated the effects of acute and long-term (21 days) intraperitoneal injections of the selective 5-HT6R agonist WAY-181187 and the antagonist SB-745427 on these MK-801-induced cognitive impairments. In the acute experiments, both the 5-HT6R agonist (3 mg/kg) and the antagonist (1 and 3 mg/kg) prevented MK-801-induced cognitive impairment in the NORT test. WAY-181187 showed significantly stronger memory-enhancing effects than Intepirdine (SB742457), but the mean difference was not statistically significant (F(1, 13) = 1,9383, ns). Both 5-HT6R ligands enhanced performance on the Y-CAT spatial memory task after a single dose, significantly improving alternating memory ability in rats; WAY-181187 showed significant effects at both 1 and 3 mg/kg doses, while SB-742457 showed a significant effect only at a 3 mg/kg dose. Considering the efficacy of single-dose administration of the two compounds at the same dose, some subtle differences can be observed in different memory function models. In the NORT task, WAY-181187 was more effective than SB-742457 at high doses, but showed no activity at a 1 mg/kg dose. Furthermore, the DI value of WAY-181187 was significantly higher than that of the control group, while the DI value of SB-742457 showed no statistically significant difference (a Bonferroni post-hoc test after one-way ANOVA showed p = 0.08, compared with the carrier control group). This result suggests that the memory-enhancing mechanism of WAY-181187 may involve other factors, but this phenomenon cannot be explained at the current stage of research. In the Y-CAT model, the situation was reversed; both doses of WAY-181187 and the higher dose of SB-742457 had similar cognitive-enhancing effects, while the lower dose of SB-742457 had no activity. In both tests, the active doses of WAY-181187 and SB-742457 did not change the exploratory behavior of rats, therefore the observed effects can be considered specific. Long-term use of two selective 5-HT6R ligands significantly improved cognitive processes interfered with by MK-801, with WAY-181187 showing a slightly stronger effect in the NORT model. However, the beneficial effects of WAY-181187 and SB-742457 in the Y-CAT model were no longer observed. At this stage of research, it is difficult to explain the loss of Y-CAT model activity after long-term use of these two 5-HT6R ligands, although their beneficial effects in the NORT model persist. The beneficial effects of 5-HT6R ligands on memory function have been reported multiple times in the literature. Studies of 5-HT6R antagonists in animal models of cognitive impairment have yielded more consistent results. 5-HT6R antagonists have been shown to be effective in episodic memory (NORT tasks), spatial working memory (maze or spontaneous alternation tasks), social cognition, and executive function (transitional or reverse learning tasks) paradigms, and to prevent memory impairment induced by scopolamine, phencyclidine (PCP), MK-801, ketamine, streptozotocin, and age-related memory disorders (reviews see Bokare et al. (2018), de Bruin and Kruse (2015), de Jong and Mørk (2017), Fone (2008), and Upton et al. (2008)). However, reports on the activity of SB-742457 in animal cognitive models are scarce. (de Bruin et al.) A 2011 study showed that intraperitoneal injection of 3 and 10 mg/kg doses of Intepirdine (SB742457) improved scopolamine-induced object recognition impairment; oral administration of 10 mg/kg dose of Intepirdine (SB742457) alleviated scopolamine-induced object localization task impairment. The same research team also reported that subchronic (5 consecutive days) administration of SB-742457 (0.63 mg/kg) alleviated PCP-induced reversal learning impairment in rats during a two-bar manipulator task (de Bruin et al., 2013); however, similar experiments by Idris et al. showed that subcutaneous injection of 2.5 and 5 mg/kg doses of SB-742457 produced activity (Idris et al., 2010). Callaghan et al. showed that oral administration of the compound at 3 mg/kg for 7 consecutive days reversed age-related cognitive deficits in middle-aged (13-month-old) rats in a delayed non-matched sample task (Callaghan et al., 2012). Therefore, the results of this study on the activity of SB-742457 are consistent with existing preclinical data and further enrich our understanding of the memory-enhancing potential of this compound in rats (especially episodic memory models). [3]

C19H19N3O2S

353.44

353.119

C, 64.57; H, 5.42; N, 11.89; O, 9.05; S, 9.07

607742-69-8

11256720

Light yellow to yellow solid powder

1.3±0.1 g/cm3

608.3±45.0 °C at 760 mmHg

321.7±28.7 °C

0.0±1.7 mmHg at 25°C

1.649

2.1

1

5

3

25

535

0

S(C1C([H])=C([H])C([H])=C([H])C=1[H])(C1=C([H])N=C2C(C([H])=C([H])C([H])=C2N2C([H])([H])C([H])([H])N([H])C([H])([H])C2([H])[H])=C1[H])(=O)=O

JJZFWROHYSMCMU-UHFFFAOYSA-N

InChI=1S/C19H19N3O2S/c23-25(24,16-6-2-1-3-7-16)17-13-15-5-4-8-18(19(15)21-14-17)22-11-9-20-10-12-22/h1-8,13-14,20H,9-12H2

3-(benzenesulfonyl)-8-piperazin-1-ylquinoline

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Solubility in Formulation 1: ≥ 2.5 mg/mL (7.07 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.5 mg/mL (7.07 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 3: 2% DMSO , 48% PEG300, 2% Tween 80 and 48% water: 5mg/mL

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 (Please use freshly prepared in vivo formulations for optimal results.)