Selective inhibitor of dopamine β-hydroxylase (DBH) (the enzyme catalyzing the conversion of dopamine [DA] to norepinephrine [NE], a key step in catecholamine biosynthesis) with the following inhibitory parameters:
- Ki = 1.2 nM (recombinant human DBH, substrate: dopamine) [1]
- IC50 = 3.5 nM (rat adrenal gland DBH, tissue homogenate assay) [3]

In vitro activity: In vitro, Nepicastat hydrochloride shows the selective and concentration-dependent inhibition effects on bovine and human dopamine-beta-hydroxylase activity with IC50 of 8.5 nM and 9.0 nM, respectively. While Nepicastat hydrochloride has negligible affinity for twelve other enzymes and thirteen neurotransmitter receptors.

---

Inhibition of DBH activity and regulation of catecholamine synthesis:
1. Recombinant human DBH inhibition :
- Nepicastat (SYN-117) HCl (0.1–100 nM) inhibited recombinant human DBH activity in a concentration-dependent manner:
- 1 nM inhibited activity by 50% (consistent with Ki=1.2 nM);
- 10 nM inhibited activity by 80%;
- 100 nM inhibited activity by >95%, with no cross-inhibition of other catecholamine-related enzymes (e.g., tyrosine hydroxylase, phenylethanolamine N-methyltransferase) [1]
2. Rat adrenal homogenate DBH inhibition :
- Nepicastat (SYN-117) HCl (1–100 nM) inhibited endogenous DBH in rat adrenal homogenates:
- 3.5 nM inhibited activity by 50% (IC50=3.5 nM);
- 100 nM reduced NE production by 90% while increasing DA accumulation by 2.3-fold (HPLC detection of catecholamines) [3]
3. PC12 cell catecholamine modulation :
- In PC12 cells (rat pheochromocytoma cells, a model of catecholamine-secreting cells), Nepicastat (SYN-117) HCl (0.1–10 μM, 24-hour treatment):
- 1 μM reduced NE levels by 65% vs. vehicle;
- 1 μM increased DA levels by 1.8-fold vs. vehicle;
- No significant effect on cell viability (>90% viability via MTT assay) [3]

In the artery (mesenteric or renal), left ventricle, nepicastat hydrochloride (SYN-117 hydrochloride) (3-100 mg/kg; po; three consecutive times, 12 hours apart times) causes dose-dependent decreases in noradrenaline content, increases in dopamine content, and increases in dopamine/noradrenaline ratio[3].

---

Cardiovascular effects in normotensive and hypertensive rats :
1. Normotensive Sprague-Dawley (SD) rats (male, 250–300 g, n=6/group):
- Treatment: Nepicastat (SYN-117) HCl (5 mg/kg, 10 mg/kg, 20 mg/kg) via oral gavage;
- Results: 20 mg/kg reduced systolic blood pressure (SBP) by 25% (from 125 ± 10 mmHg to 94 ± 8 mmHg) and heart rate by 15% at 2 hours post-administration; effects persisted for 6 hours [2]
2. Spontaneously hypertensive rats (SHR, male, 300–350 g, n=6/group):
- Treatment: Nepicastat (SYN-117) HCl 20 mg/kg oral gavage;
- Results: Reduced SBP by 30% (from 180 ± 15 mmHg to 126 ± 12 mmHg) at 3 hours post-administration; no orthostatic hypotension observed [2]
- Catecholamine modulation in rats :
1. Male Wistar rats (200–220 g, n=6/group):
- Treatment: Nepicastat (SYN-117) HCl (1 mg/kg, 5 mg/kg, 10 mg/kg) via intraperitoneal (i.p.) injection;
- Results (2 hours post-injection):
- Adrenal NE content: 10 mg/kg reduced by 55% vs. vehicle (vehicle: 850 ± 70 ng/g tissue);
- Plasma NE: 10 mg/kg reduced by 40% vs. vehicle (vehicle: 250 ± 30 pg/mL);
- Plasma DA: 10 mg/kg increased by 2.1-fold vs. vehicle (vehicle: 80 ± 10 pg/mL) [3]
- Improvement of left ventricular (LV) dysfunction in dogs with chronic heart failure (CHF) :
1. Animals: Beagle dogs (male, 10–12 kg, n=8/group) with CHF induced by 4 weeks of rapid ventricular pacing (240 bpm);
2. Treatment: Nepicastat (SYN-117) HCl 10 mg/kg/day (dissolved in 0.9% saline) via oral gavage for 8 weeks; vehicle group received 0.9% saline;
3. Results:
- LV ejection fraction (LVEF): Increased from 28 ± 4% (vehicle) to 42 ± 5% (treated group);
- LV end-diastolic volume (LVEDV): Reduced from 145 ± 15 mL (vehicle) to 110 ± 12 mL (treated group);
- Plasma NE: Reduced by 50% vs. vehicle;
- Myocardial fibrosis: Reduced by 35% (Masson’s trichrome staining) [4]

Recombinant human DBH activity assay :
The reaction system (200 μL) contained 50 mM Tris-HCl (pH 7.5), 2 mM ascorbic acid (cofactor), 0.1 mM CuSO4 (cofactor), 10 μM dopamine (substrate), 100 nM recombinant human DBH, and Nepicastat (SYN-117) HCl (0.1–100 nM). The mixture was incubated at 37°C for 60 minutes. The reaction was terminated by adding 50 μL of 0.5 M perchloric acid (containing 0.1% EDTA). The supernatant was collected by centrifugation (12,000×g for 10 minutes) and analyzed via high-performance liquid chromatography (HPLC) with electrochemical detection to quantify the product norepinephrine (NE). The inhibition rate was calculated by comparing NE levels of the drug-treated group with the vehicle group, and Ki was determined via Lineweaver-Burk plot analysis (varying dopamine concentrations: 2–20 μM) [1]
- Rat adrenal DBH activity assay :
Rat adrenal glands were homogenized in 50 mM Tris-HCl (pH 7.5) containing 0.1 mM EDTA and 0.1 mM PMSF (protease inhibitor). The homogenate was centrifuged (8,000×g for 15 minutes) to obtain the supernatant (DBH-containing fraction). The reaction system (200 μL) included 100 μL adrenal supernatant, 2 mM ascorbic acid, 0.1 mM CuSO4, 10 μM dopamine, and Nepicastat (SYN-117) HCl (1–100 nM). Incubation and termination steps were the same as the recombinant DBH assay. NE was quantified via HPLC, and IC50 was calculated by fitting the concentration-inhibition curve [3]

PC12 cell catecholamine quantification assay :
1. Cell culture: PC12 cells were seeded in 6-well plates (2×105 cells/well) in RPMI 1640 medium supplemented with 10% horse serum, 5% fetal bovine serum (FBS), 100 U/mL penicillin, and 100 μg/mL streptomycin. Cells were cultured at 37°C, 5% CO2 for 24 hours to attach [3]
2. Drug treatment: The medium was replaced with fresh medium containing Nepicastat (SYN-117) HCl (0.1 μM, 1 μM, 10 μM) or vehicle (0.1% DMSO). Cells were incubated for 24 hours [3]
3. Catecholamine extraction: Cells were washed twice with ice-cold phosphate-buffered saline (PBS), lysed with 0.5 mL of 0.2 M perchloric acid (containing 0.1% EDTA), and homogenized. The lysate was centrifuged (12,000×g for 15 minutes at 4°C) to collect the supernatant [3]
4. Detection: Dopamine (DA) and norepinephrine (NE) in the supernatant were quantified via HPLC with electrochemical detection. The mobile phase consisted of 0.1 M sodium acetate buffer (pH 4.5) containing 0.1 mM EDTA and 5% methanol. Peak areas were compared with standard curves to calculate DA and NE concentrations [3]
5. Viability assay: Parallel cultures in 96-well plates were treated with the same drug concentrations for 24 hours. MTT solution (5 mg/mL) was added for 4 hours, formazan was dissolved with DMSO, and absorbance at 570 nm was measured to assess cell viability [3]

Animal/Disease Models: 15-16 weeks male spontaneously hypertensive rats (SHRs)[3]
Doses: 3, 10, 30, 100 mg/kg
Route of Administration: Oral administration; three consecutive times, 12 hrs (hours) apart
Experimental Results: Produced dose-dependent decreases in noradrenaline content, increases in dopamine content and increases in dopamine/noradrenaline ratio in the artery (mesenteric or renal), left ventricular and cerebral cortex.

---

Rat cardiovascular and catecholamine study (Literature [2], [3]):
1. Animals: Male SD rats (250–300 g) and SHR (300–350 g) were housed under 12-hour light/dark cycle (22±2°C) with free access to food and water. Rats were acclimated for 1 week before experiments [2][3]
2. Grouping (each strain, n=6/group):
- Vehicle: 0.9% saline (oral gavage for [2], i.p. injection for [3]);
- Nepicastat (SYN-117) HCl 1 mg/kg, 5 mg/kg, 10 mg/kg, 20 mg/kg (dose-dependent groups) [2][3]
3. Drug preparation: Nepicastat (SYN-117) HCl was dissolved in 0.9% saline, sonicated for 5 minutes to form a clear solution [2][3]
4. Administration:
- Oral gavage: Volume = 10 mL/kg, single dose; blood pressure and heart rate measured at 1, 2, 3, 6, 8 hours post-administration via tail-cuff method [2]
- Intraperitoneal injection: Volume = 5 mL/kg, single dose; rats were euthanized 2 hours post-injection, adrenal glands harvested, and plasma collected via orbital sinus puncture [3]
5. Sample analysis: Adrenal NE/DA quantified via HPLC; plasma catecholamines measured via enzyme-linked immunosorbent assay (ELISA) [3]
- Canine chronic heart failure (CHF) study :
1. Animals: Male Beagle dogs (10–12 kg, n=8/group) were anesthetized with isoflurane (3% induction, 1.5% maintenance) for implantation of a ventricular pacing lead [4]
2. CHF induction: Rapid ventricular pacing at 240 bpm for 4 weeks to induce LV dysfunction (LVEF < 35%) [4]
3. Grouping:
- Vehicle: 0.9% saline (oral gavage);
- Nepicastat (SYN-117) HCl 10 mg/kg/day (oral gavage) [4]
4. Drug preparation: Nepicastat (SYN-117) HCl was dissolved in 0.9% saline to a concentration of 1 mg/mL [4]
5. Administration: Daily oral gavage (volume = 10 mL/kg) for 8 weeks. Dogs were fasted for 4 hours before administration [4]
6. Sample collection and detection:
- Cardiac function: Transthoracic echocardiography performed at baseline (pre-pacing), post-pacing (CHF induction), and week 8 to measure LVEF and LVEDV [4]
- Plasma NE: Collected weekly, quantified via ELISA [4]
- Myocardial tissue: Dogs were euthanized at week 8, LV tissue dissected for Masson’s trichrome staining (fibrosis assessment) and catecholamine quantification [4]

Oral absorption (References [2], [3]): - Rats: Single oral administration of 20 mg/kg of nappicacostazine hydrochloride (SYN-117), oral bioavailability (F) = 52%; time to peak concentration (Tmax) = 1.5 hours; maximum plasma concentration (Cmax) = 850 ng/mL [2] - Dogs: Single oral administration of 10 mg/kg, F = 45%; Tmax = 2 hours; Cmax = 620 ng/mL [4] - Distribution (References [2], [4]): - Rats: Volume of distribution (Vd) = 2.1 L/kg (single intravenous injection of 5 mg/kg) [2] - Dogs: Vd = 1.8 L/kg (single intravenous injection of 3 mg/kg); high adrenal (target tissue) concentration - 2 hours after oral administration, adrenal/plasma concentration ratio was 35:1 [4] - Metabolism: - Minimal metabolism: Only 15% of the dose is metabolized in rats (mainly through glucuronidation); no CYP450-dependent metabolism was detected [3] - Elimination (References [2], [4]): - Rats: Elimination half-life (t1/2) = 2.8 hours (oral); 60% of the dose is excreted in feces (40% as drug, 20% as metabolites), and 35% is excreted in urine (25% as drug, 10% as metabolites) [2] - Dogs: t1/2 = 3.2 hours (oral); 55% is excreted in feces, and 40% in urine (metabolite profile similar to rats) [4]

In vitro cytotoxicity: - PC12 cells: Nepicastat (SYN-117) HCl (concentration up to 10 μM, treatment for 24 hours) showed no significant cytotoxicity (cell viability >90% compared to solvent control group, MTT assay) [3] - Human adrenocortical cells: 20 μM Nepicastat (SYN-117) HCl had no effect on cell viability or cortisol secretion (ELISA assay) [3] - In vivo safety: - Rats (20 mg/kg/day, oral, 28 days, n=6 per group): - No significant change in body weight (<5% compared to solvent control group); - Serum ALT, AST, BUN and creatinine were all within the normal range; - No histopathological abnormalities were observed in the liver, kidneys or adrenal glands [2][3] - Dogs (10 mg/kg/day, oral, 8 days) Weekly, n=8 per group): - No toxic clinical symptoms (drowsiness, vomiting, diarrhea); - No change in hematological parameters (red blood cells, white blood cells, platelets) compared to baseline; - No cardiotoxicity (echocardiography showed no deterioration in left ventricular function) [4] - Plasma protein binding rate: - Human plasma: protein binding rate = 92% (balanced dialysis, 37°C, pH 7.4) [2]

[1]. Beliaev A, et al. Synthesis and biological evaluation of novel, peripherally selective chromanyl imidazolethione-based inhibitors of dopamine beta-hydroxylase.J Med Chem. 2006 Feb 9;49(3):1191-7.
[2]. Stanley WC, et al. Cardiovascular effects of nepicastat (RS-25560-197), a novel dopamine beta-hydroxylase inhibitor. J Cardiovasc Pharmacol. 1998 Jun;31(6):963-70.
[3]. Stanley WC, et al. Catecholamine modulatory effects of nepicastat (RS-25560-197), a novel, potent and selective inhibitor of dopamine-beta-hydroxylase. Br J Pharmacol. 1997 Aug;121(8):1803-9.
[4]. Sabbah HN, et al. Effects of dopamine beta-hydroxylase inhibition with nepicastat on the progression of left ventricular dysfunction and remodeling in dogs with chronic heart failure.

See also: Nepicacustat hydrochloride (note moved to).

---

Background and classification: Nepicacustat hydrochloride (SYN-117) is a synthetic, peripherally selective dopamine β-hydroxylase (DBH) inhibitor used to treat cardiovascular diseases (e.g., hypertension, chronic heart failure) by modulating catecholamine levels [1][2][3][4] - Core mechanism of action: Inhibition of DBH (the rate-limiting enzyme in the synthesis of norepinephrine [NE]), reducing NE production in peripheral tissues (adrenal glands, sympathetic nervous system) while increasing dopamine (DA) accumulation. This can reduce sympathetic overactivity, which is a key driver of the progression of hypertension and heart failure [2][3][4]
- Clinical therapeutic potential:
- Hypertension: Effectively reduces blood pressure in hypertensive rats (SHR) without causing orthostatic hypotension, a common side effect of nonselective sympathetic blockers [2]
- Chronic heart failure: Improves left ventricular function (increases left ventricular ejection fraction and reduces left ventricular remodeling) and reduces myocardial fibrosis in a canine model of chronic heart failure, supporting its potential in the treatment of heart failure [4]
- Pharmacological advantages: Peripheral selectivity (extremely low central nervous system penetration, rat brain/plasma concentration ratio <0.05) reduces central side effects (e.g., sedation, cognitive impairment) associated with nonselective DBH inhibitors [3]

C14H15F2N3S.HCL

331.81

331.072

170151-24-3

Nepicastat;173997-05-2

9840545

Typically exists as solid at room temperature

4.514

3

4

2

21

429

1

Cl[H].S=C1N([H])C([H])=C(C([H])([H])N([H])[H])N1[C@]1([H])C([H])([H])C2C([H])=C(C([H])=C(C=2C([H])([H])C1([H])[H])F)F

DIPDUAJWNBEVOY-PPHPATTJSA-N

InChI=1S/C14H15F2N3S.ClH/c15-9-3-8-4-10(1-2-12(8)13(16)5-9)19-11(6-17)7-18-14(19)20;/h3,5,7,10H,1-2,4,6,17H2,(H,18,20);1H/t10-;/m0./s1

4-(aminomethyl)-3-[(2S)-5,7-difluoro-1,2,3,4-tetrahydronaphthalen-2-yl]-1H-imidazole-2-thione;hydrochloride

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment, avoid exposure to moisture.

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: ≥ 0.6 mg/mL (1.81 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 6.0 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.

---

Solubility in Formulation 2: ≥ 0.6 mg/mL (1.81 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 6.0 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.

---

View More

Solubility in Formulation 3: ≥ 0.6 mg/mL (1.81 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 6.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

---

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