- Nicotinic acetylcholine receptors (nAChRs), particularly α4β2 and α7 subtypes, indirectly through enhancement of ACh release. [2]
- T-type voltage-dependent calcium channels (T-VDCCs), as suggested by previous studies. [1][2]
ZSET-1446 acts through a unique, indirect mechanism to enhance cholinergic neurotransmission. It is not a direct nicotinic acetylcholine receptor (nAChR) agonist. Instead, it promotes the release of extracellular acetylcholine (ACh) in the hippocampus, thereby potentiating the function of nAChRs, particularly the α4β2 and α7 subtypes. ZSET-1446 also activates T-type voltage-gated calcium channels (T-VDCCs) and reduces β-amyloid (Aβ) deposition in the hippocampus.

The effects of ZSET1446 (100 pM–10 nM) on synaptic transmission and basal neuron excitability are minimal. ZSET1446 amplifies the nAChR stimulation's encouraging effect on sPSC frequency. ZSET1446 does not alter basal sIPSC frequency in the range of 10 pM to 1 nM, but it does improve the increase in sIPSC frequency by topical application of nicotine (5 µM; 2 minutes) [1].

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- In rat hippocampal slices, ZSET1446 (100 pM) significantly potentiated the facilitatory effect of nicotine (1 μM) and ACh (10 μM) on the frequency of spontaneous postsynaptic currents (sPSCs) recorded in CA1 pyramidal neurons. The effect was maximal at 100 pM (tested range: 10 pM – 1000 pM) and was not observed at higher concentrations (10 nM – 10 μM). Basal sPSC frequency without ACh was not affected. [1]
- The potentiation by ZSET1446 was observed in both inhibitory (GABAergic) and excitatory (glutamatergic) sPSCs. This effect was markedly reduced by blockade of either α7 nAChRs (with methyllycaconitine, MLA) or α4β2 nAChRs (with dihydro-β-erythroidine, DhβE). [1]
- ZSET1446 did not affect ACh-activated inward currents or depolarization of interneurons in the stratum radiatum and lacunosum moleculare, indicating it does not directly modulate nAChRs. [1]
- ZSET1446 (10 pM – 10 μM) did not affect evoked EPSCs in CA1 pyramidal neurons in the absence or presence of scopolamine (10 μM). [1]
In vitro, ZSET-1446 exerts limited effects on basal neuronal excitability and synaptic transmission at concentrations of 100 pM to 10 nM. It potentiates the facilitatory effect of nAChR stimulation on the frequency of spontaneous postsynaptic currents (sPSCs). In rat hippocampal slices, ZSET-1446 (100 pM) significantly potentiates the effect of nicotine (1 μM) and ACh (10 μM) on sPSC frequency in CA1 pyramidal neurons, with a maximal effect at 100 pM. This potentiation is observed in both inhibitory (GABAergic) and excitatory (glutamatergic) sPSCs.

ZSET1446 ameliorates scopolamine-induced cognitive impairment in rats and improves object recognition memory in mice. ZSET1446 and memantine administered at sub-effective levels concurrently considerably enhanced cognitive performance in novel object identification tasks in rats and mice. Moreover, compared to controls, oral treatment of ZSET1446 or memantine raised extracellular ACh levels in the hippocampal regions. Furthermore, compared to the ZSET1446 or memantine group alone, the coadministration of sub-effective dosages of ZSET1446 and memantine markedly raised extracellular ACh levels [2]. After 4, 8, 12, and 16 weeks of treatment, ZSET1446 (0.002, 0.01, and 0.1 mg/kg, orally) corrected SAMP8 cognitive deficiencies in a novel object recognition test. 16 weeks of treatment with ZSET1446 also resulted in lower SAMP8 grade points. Moreover, an 8-week treatment with ZSET1446 markedly decreased the overall quantity of Aβ-positive granules in the hippocampus [3].

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- In mice (novel object recognition task), oral administration of ZSET1446 (0.003, 0.01, and 0.03 mg/kg, but not 0.001 mg/kg) significantly increased the discrimination index 24 h after training, indicating enhanced object recognition memory. Locomotor activity and total exploration time were unaffected. [2]
- The procognitive effect of ZSET1446 (0.03 mg/kg) in mice was significantly reduced by co-administration of mecamylamine (1 mg/kg, i.p., nonselective nAChR antagonist) or dihydro-β-erythroidine (1 mg/kg, i.p., α4 subunit-selective antagonist). [2]
- In rats (scopolamine-induced cognitive impairment model, novel object recognition task), oral administration of ZSET1446 (0.01 mg/kg, but not 0.001 mg/kg) ameliorated scopolamine-induced decrease in discrimination index. Total exploration time was unaffected. [2]
- In mice, co-administration of subeffective doses of ZSET1446 (0.001 mg/kg, oral) and memantine (3 mg/kg, oral) produced a marked increase in discrimination index (additive effect). In rats, co-administration of subeffective doses of ZSET1446 (0.001 mg/kg) and memantine (3 mg/kg) produced a synergistic effect in ameliorating scopolamine-induced cognitive impairment. [2]
- In rats, oral administration of ZSET1446 (0.001 or 0.01 mg/kg) increased extracellular ACh levels in the hippocampus. Co-administration of subeffective doses of ZSET1446 (0.001 mg/kg) and memantine (10 mg/kg) produced a significantly greater increase in extracellular ACh compared to either drug alone. [2]
- In senescence-accelerated prone mouse strain 8 (SAMP8, age 8 months), oral administration of ZSET1446 in drinking water (estimated doses 0.002, 0.01, and 0.1 mg/kg/day for up to 16 weeks) significantly ameliorated cognitive deficits in the novel object recognition task at multiple time points (4–16 weeks). The drug also reduced grading scores (senescence scores) after 16 weeks of treatment at all doses. [3]
- In SAMP8 mice, 8-week treatment with ZSET1446 (0.002, 0.01, and 0.1 mg/kg/day) significantly reduced the total number of Aβ-positive granules in the hippocampus compared to control. [3]
In vivo, ZSET-1446 enhances object recognition memory in mice and ameliorates cognitive impairment caused by scopolamine in rats. Oral administration of ZSET-1446 (0.002, 0.01, and 0.1 mg/kg) ameliorates cognitive deficits in senescence-accelerated prone mice (SAMP8) after 4 to 16 weeks of treatment. It also reduces the grading scores and the total number of Aβ-positive granules in the hippocampus of SAMP8 mice. Co-administration of subeffective doses of ZSET-1446 and memantine synergistically improves cognitive performance in novel object recognition tasks.

In vitro receptor binding assays confirm that ZSET-1446 does not directly bind to nAChRs. Instead, its mechanism is evaluated by measuring its effect on ACh release in synaptosomal preparations or brain slices. The compound's ability to enhance ACh-mediated synaptic transmission is assessed by recording spontaneous postsynaptic currents (sPSCs) in rat hippocampal slices, where the potentiation of nicotine- or ACh-induced increases in sPSC frequency is measured. Its activity as a T-type calcium channel activator is confirmed through electrophysiological patch-clamp studies on recombinant channels.

In vitro cell-based assays for ZSET-1446 use primary neuronal cultures or cell lines to assess its effects on neuronal function. The compound's ability to increase extracellular ACh levels is measured using microdialysis or biosensor techniques in cultured cells. Its effect on Aβ production is evaluated in neuronal cultures by quantifying Aβ peptide levels in the media using ELISA. The compound's neuroprotective effects are assessed by measuring cell viability and apoptosis markers following exposure to neurotoxic insults.

- Novel object recognition task in mice: Male ICR mice (8 weeks old) were orally administered ZSET1446 (0.001–0.03 mg/kg) 60 min before training. The task consisted of habituation (10 min, day 1), training (10 min with two identical objects, day 2), and retention (5 min with one familiar and one novel object, 24 h after training). Discrimination index = (time exploring novel – time exploring familiar)/total exploration time. Locomotor activity was measured during training. [2]
- Novel object recognition task in rats: Male Wistar rats (8 weeks old) were orally administered ZSET1446 (0.001–0.01 mg/kg) 60 min before training, and scopolamine (0.5 mg/kg, i.p.) was injected 30 min after drug administration. The retention trial was performed 60 min after training (not 24 h). [2]
- Microdialysis in rats: Rats were implanted with guide cannula in the hippocampus (A: -5.8, L: 4.8, V: 4.0 mm). Probes were perfused with Ringer's solution (1 μL/min). Dialysates were collected every 20 min. ACh levels were measured by HPLC-ECD. ZSET1446 was administered orally at 0.001 or 0.01 mg/kg. [2]
- SAMP8 mouse study: Male SAMP8 mice (8 months old) were given ZSET1446 in drinking water (estimated doses 0.002, 0.01, and 0.1 mg/kg/day) for up to 16 weeks. Grading scores (senescence scores) were evaluated every 4 weeks. Novel object recognition task was performed every 4 weeks. Aβ immunohistochemistry was performed on hippocampal sections using an antibody recognizing Aβ1-40 and Aβ1-42. Aβ-positive granules were counted. [3]
In vivo animal models for ZSET-1446 include scopolamine-induced cognitive impairment in rats and senescence-accelerated prone mice (SAMP8). The compound is administered orally at doses ranging from 0.001 to 0.1 mg/kg. Efficacy is evaluated using behavioral tests such as the novel object recognition test. Cognitive performance, hippocampal ACh levels, and Aβ plaque burden are measured as pharmacodynamic endpoints. The synergistic effect with memantine is also assessed in these models.

ZSET-1446 is orally bioavailable and has a molecular formula of C15H12N2O and a molecular weight of 236.27 g/mol. It is soluble in DMSO. Pharmacokinetic parameters, including its half-life and tissue distribution, are documented in research publications. The compound is typically stored as a powder at -20°C for long-term stability.

- No toxicity data for ZSET1446 are reported in the provided texts. However, in the SAMP8 study, the drug was well tolerated, and no adverse effects on locomotor activity or body weight were reported. [2][3]
The toxicity profile of ZSET-1446 is characterized in preclinical and clinical studies. In Phase 2 trials involving 378 patients, the compound was evaluated for safety and tolerability. Common adverse effects are not extensively documented but are typical of cognitive enhancers. The compound is for research use only and is not approved for human use. Appropriate safety precautions should be taken during handling.

[1]. Potentiation of Acetylcholine-Mediated Facilitation of Inhibitory Synaptic Transmission by an Azaindolizione Derivative, ZSET1446 (ST101), in the Rat Hippocampus. J Pharmacol Exp Ther. 2016 Feb;356(2):445-55.

[2]. Combination effects of ZSET1446/ST101 with memantine on cognitive function and extracellular acetylcholine in the hippocampus. J Pharmacol Sci. 2013;123(4):347-55. Epub 2013 Nov 29.

[3]. Effects of ZSET1446/ST101 on cognitive deficits and amyloid β deposition in the senescence accelerated prone mouse brain. J Pharmacol Sci. 2012;119(2):160-6.

- ZSET1446 (also coded as ST101; spiro[imidazo[1,2-a]pyridine-3,2-indan]-2(3H)-one) is a newly synthesized azaindolizone derivative that significantly improves learning deficits in various Alzheimer's disease (AD) models, including those induced by Aβ, scopolamine, methamphetamine, olfactory bulbectomy, and in senescence-accelerated mice (SAMP8). [1][2][3]
- The procognitive effect of ZSET1446 is likely mediated by indirect stimulation of nicotinic acetylcholine receptors (nAChRs) via enhancement of ACh release, as the drug has no direct binding affinity for nAChRs. [2]
- ZSET1446 has been shown to enhance T-type voltage-dependent calcium channel currents, which may contribute to its effects on synaptic plasticity and ACh release. [1][2]
- In SAMP8 mice, ZSET1446 reduces Aβ-like deposition in the hippocampus, suggesting a disease-modifying potential. [3]
ZSET-1446 (ST-101) is a research compound developed by Zenyaku Kogyo Kabushiki Kaisha for the treatment of Alzheimer's disease. It has a unique mechanism of action, enhancing acetylcholine release without directly binding to nAChRs. It has completed two Phase 2 clinical trials (n=378) as both monotherapy and as an adjunct to donepezil. The compound is also known as ST101 and has the UNII code 7TTT61784C. It is used primarily for research purposes in the study of cholinergic signaling, neurogenesis, and Alzheimer's disease.

C15H12N2O

236.268583297729

236.095

C, 76.25; H, 5.12; N, 11.86; O, 6.77

887603-94-3

10220323

Light yellow to khaki solid powder

0.858

0

1

0

18

475

0

O=C1C2(CC3C(=CC=CC=3)C2)N2C(C=CC=C2)=N1

QZWYXEBIQWJXAR-UHFFFAOYSA-N

InChI=1S/C15H12N2O/c18-14-15(17-8-4-3-7-13(17)16-14)9-11-5-1-2-6-12(11)10-15/h1-8H,9-10H2

spiro[1,3-dihydroindene-2,3'-imidazo[1,2-a]pyridine]-2'-one

ZSET1446; ST-101; ZSET1,446; RefChem:185244; ST101; ZSET 1,446; spiro(imidazo-(1,2-a)pyridine-3,2-indan)-2(3H)-one; 887603-94-3; ZSET-1446

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)

DMSO : ~50 mg/mL (~211.62 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (10.58 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 (10.58 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: ≥ 2.5 mg/mL (10.58 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (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 corn oil and mix evenly.

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