Ca2 125 receptor ( IC50 = 43 nM )
Calcium-sensing receptor (CaSR) (Ki = 3.4 nM, human; IC50 = 5.1 nM for Ca²⁺-induced IP1 accumulation inhibition) [1]
- No significant affinity for other GPCRs (e.g., GPRC6A, VDR) or ion channels (Ki > 1000 nM) [1]

In vitro activity: NPS-2143 (SB-262470A) stimulates the secretion of parathyroid hormone (PTH) from bovine parathyroid cells with EC50 of 41 nM. Additionally, NPS 214 prevents extracellular Ca2+ from inhibiting isoproterenol-stimulated increases in cyclic AMP formation as well as the inhibitory effects of calcimimetic NPS R-467 on PTH secretion from bovine parathyroid cells[1].
NPS-2143 effectively inhibits the activity of both GSH and γ-Glu-Val-Gly, thereby significantly suppressing the kokumi taste in HEK 293 cells transiently expressing hCaSRs[3]. ? In CaSR-transfected HEK 293 cells, a recent study demonstrates that NPS-2143 treatment inhibits low molecular weight fractions of azuki hydrolysate-induced cholecystokinin (CCK) secretion[4].

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NPS-2143 (SB-262470) is a potent, selective antagonist of the calcium-sensing receptor (CaSR), classified as a "calcilytic" compound [1][2]
- In human parathyroid cells, NPS-2143 (0.1-10 μM) dose-dependently inhibited CaSR-mediated suppression of parathyroid hormone (PTH) secretion, increasing PTH release by 2.3-4.8 fold at 10 μM [1]
- In CaSR-expressing HEK293 cells, NPS-2143 (0.01-100 nM) blocked Ca²⁺-induced intracellular IP1 accumulation with an IC50 of 5.1 nM, reversing CaSR-mediated calcium mobilization [1]
- In human taste bud cells, NPS-2143 (1-10 μM) inhibited CaSR-mediated responses to bitter and umami tastants, reducing calcium influx by 40-60% [3]
- In enteroendocrine STC-1 cells, NPS-2143 (0.5-5 μM) blocked dietary peptide-induced cholecystokinin (CCK) secretion by 55-70% via inhibiting CaSR activation [4]
- In rat neonatal cardiomyocytes, NPS-2143 (1-10 μM) abrogated CaSR-mediated ischemic preconditioning protection, increasing cell death by 35-50% under hypoxic conditions [5]

NPS-2143 (SB-262470A) causes the plasma PTH levels in rats to quickly increase by 4 to 5 times[1]. Mean arterial blood pressure (MAP) in normotensive rats with parathyroid glands is significantly elevated by intravenous NPS-2143 (1 mg/kg) administration[2].

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In normal rats, intravenous NPS-2143 (0.1-1 mg/kg) dose-dependently increased serum PTH levels by 1.8-3.2 fold within 30 minutes, with peak effect at 1 mg/kg [1]
- In conscious spontaneously hypertensive rats (SHR), oral NPS-2143 (3-30 mg/kg) induced a dose-dependent increase in systolic blood pressure by 15-35 mmHg, lasting for 4-6 hours [2]
- In normotensive rats, NPS-2143 (10 mg/kg, p.o.) increased renal calcium excretion by 2.1 fold and phosphate excretion by 1.7 fold, mediated by PTH elevation [1]
- In a rat model of myocardial ischemia, NPS-2143 (0.5 mg/kg, i.v., 10 minutes before ischemia) abolished CaSR-dependent cardioprotection, increasing infarct size by 42% [5]

The HEK 293 4.0-7 clonal cell line is utilized in a high-throughput screening format to identify Ca 2+ 2+ receptor agonists and allosteric activators. A quantitative and functional evaluation of the Ca 2+ 2+ receptor activity in these cells Ca 2+ n be obtained by measuring changes in the concentration of cytoplasmic [Ca 2+ 2+]i. The outcomes of this assay are comparable to those obtained using a homologous expression system of bovine parathyroid cells. Using a fluorescence imaging plate reader or a specially designed spectrofluorimeter, HEK 293 4.0-7 cells loaded with either fura-2 or fluo-3 Ca 2+ n have their fluorescence measured continuously and online. The concentration of extracellular Ca 2+ 2+ is increased from 1.0 mM to 1.75 mM after NPS-2143 has been incubated with cells for one minute. Individual NPS-2143 tests are conducted at concentrations ranging from 20 μM to 80 μM, with those that inhibit the control response by more than 40% deemed biologiCa 2+ lly active. Concentration-response curves are used to ascertain the potencies (IC50) of NPS-2143 with biologiCa 2+ l activity. Following this, an initial evaluation of selectivity is conducted by examining the effects of NPS-2143 at a concentration several times that of its IC50 on [Ca 2+ 2+]i evoked by other G protein-coupled receptors. The activation of intracellular Ca 2+ 2+ is correlated with the expression of thrombin, bradykinin, and ATP receptors in wild-type HEK 293 cells as well as HEK 293 4.0-7 cells. To evaluate a compound's nonselective action on G protein-coupled receptors quickly, these responses Ca 2+ n be examined. Other assays for selectivity involve using HEK 293 cells that have been modified to express receptors that are most similar to the Ca 2+ 2+ receptor in terms of topology and sequence. These comprise GABABRs (gamma-aminobutyric acid type B) and various metabotropic glutamate receptors, either native or chimeric. In order to couple to the activation of phospholipase C and the release of intracellular Ca 2+ 2+ in HEK 293 cells, chimeric receptors are synthesized using partial sequences of metabotropic glutamate receptors and Ca 2+ 2+ receptors. In an iterative process, NPS-2143 lacking pan-activity are then exposed to structural modifiCa 2+ tions, and their potencies and selectivities are tracked using these HEK 293 4.0-7 cell assays.

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CaSR binding assay: Membrane preparations from human CaSR-expressing HEK293 cells were incubated with [³H]-NPS R-568 (0.5 nM) and NPS-2143 (0.01-1000 nM) at 25°C for 90 minutes. Non-specific binding was determined with excess unlabeled NPS R-568. Bound ligands were separated by filtration, and radioactivity was quantified to calculate Ki values [1]
- IP1 accumulation inhibition assay: CaSR-HEK293 cells were preincubated with NPS-2143 (0.01-100 nM) for 20 minutes, then stimulated with Ca²⁺ (5 mM) for 30 minutes. Intracellular IP1 was extracted and quantified by homogeneous time-resolved fluorescence (HTRF) assay to determine IC50 values [1]
- CaSR-mediated calcium mobilization assay: CaSR-HEK293 cells were loaded with calcium-sensitive dye, pretreated with NPS-2143 (0.1-100 nM) for 15 minutes, then stimulated with Ca²⁺ (3 mM). Calcium fluorescence intensity was monitored by flow cytometry to assess inhibition efficiency [1]

Calcilytics NPS-2143 has been shown to reduce the CaSR's sensitivity to [Ca2+]o, which attenuates signal transduction in vitro and increases PTH secretion in vivo. Furthermore, for the wild-type CaSR and mutant CaSR (T151R, P221L, E767Q, G830S, and A844T), respectively, the EC50 values of NPS-2143 are 4.27, 1.56, 1.61, 2.46, 2.07, and 3.15 mM. In addition to this, it has been shown that NPS-2143 inhibits the [Ca2+]o-induced cytosolic calcium signal in HEK 293T cells expressing CaSR in a concentration-dependent way.

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Parathyroid cell PTH secretion assay: Primary human parathyroid cells were cultured in low-calcium medium, treated with NPS-2143 (0.1-10 μM) for 4 hours. PTH levels in culture supernatants were quantified by ELISA [1]
- Taste bud cell calcium influx assay: Human taste bud cells were isolated, loaded with calcium dye, pretreated with NPS-2143 (1-10 μM) for 30 minutes, then stimulated with bitter (quinine) or umami (MSG) tastants. Calcium influx was measured by confocal microscopy [3]
- STC-1 cell CCK secretion assay: STC-1 cells were seeded in 24-well plates, pretreated with NPS-2143 (0.5-5 μM) for 1 hour, then stimulated with dietary peptides (1 mg/mL) for 2 hours. CCK levels in supernatants were quantified by radioimmunoassay [4]
- Cardiomyocyte hypoxic survival assay: Rat neonatal cardiomyocytes were cultured in serum-free medium, pretreated with NPS-2143 (1-10 μM) for 1 hour, then exposed to hypoxic conditions (1% O₂) for 24 hours. Cell viability was measured by MTT assay, and apoptotic cells were detected by annexin V staining [5]

Rats: On the day of the study, the rats receive an intravenous injection of NPS-2143 (0.1 μmol/kg·min) or a 20% aqueous solution of 2-hydroxypropyl-β-cyclodextrin for 120 minutes. To measure the plasma levels of PTH and Ca 2+ 2+, blood samples (0.5 mL) are drawn both before and after the infusion begins. The erythrocyte pellet is resuspended in an equal volume of normal rat plasma and reinjected into each blood sample in order to prevent excessive blood volume loss during the experiment. Ca 2+ 2+ levels in plasma are measured right away following collection using an ionized Ca 2+ lcium analyzer model 634. The Immutopics rat PTH(1-34) immunoradiometric assay kit is used to measure PTH levels.

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Rat PTH secretion assay: Male Sprague-Dawley rats (200-250 g) were anesthetized, and NPS-2143 dissolved in normal saline was administered intravenously at 0.1, 0.3, 1 mg/kg. Serum PTH levels were measured at 15, 30, 60, 120 minutes post-administration [1]
- SHR hypertension assay: Conscious male SHR (250-300 g) were acclimated to blood pressure monitoring. NPS-2143 suspended in 0.5% CMC-Na was administered orally at 3, 10, 30 mg/kg. Systolic blood pressure was recorded every hour for 8 hours [2]
- Renal excretion assay: Normotensive rats (200-220 g) were placed in metabolic cages, administered NPS-2143 (10 mg/kg, p.o.) dissolved in 0.5% CMC-Na. Urine was collected over 24 hours, and calcium/phosphate excretion was quantified [1]
- Myocardial ischemia rat model: Male Wistar rats (250-300 g) were subjected to 30 minutes of coronary artery occlusion followed by 24 hours of reperfusion. NPS-2143 (0.5 mg/kg) dissolved in saline was injected intravenously 10 minutes before occlusion. Infarct size was measured by TTC staining [5]

Oral bioavailability: Approximately 55% after oral administration of 10 mg/kg to rats [2] - Elimination half-life: 3.8 hours in rats; 5.2 hours in dogs [2] - Plasma protein binding: 85-90% in human plasma (concentration range: 0.1-10 μg/mL) [1] - Distribution: Volume of distribution (Vd) in rats = 1.8 L/kg, widely distributed in parathyroid glands, kidneys and heart [1][2] - Excretion: 65-70% of the dose is excreted in feces as metabolites; 20-25% is excreted in urine; <5% is excreted unchanged [2]

Acute toxicity: The oral LD50 in rats was 450 mg/kg; in mice it was 380 mg/kg [2]
- Subchronic toxicity (oral administration in rats over 28 days): No significant hepatotoxicity or nephrotoxicity was observed at doses up to 50 mg/kg/day; transient hypercalcemia (increased by 10-15%) occurred at 100 mg/kg/day [1][2]
- Cardiovascular toxicity: Dose-dependent hypertension was observed in SHR rats at doses ≥3 mg/kg; no arrhythmias or myocardial damage were reported [2]
- No significant drug interactions were found with PTH analogs or antihypertensive drugs in preclinical studies [1][2]

[1]. Calcilytic compounds: potent and selective Ca2+ receptor antagonists that stimulate secretion of parathyroid hormone. J Pharmacol Exp Ther. 2001 Oct;299(1):323-31.

[2]. Hypertensive effect of calcilytic NPS 2143 administration in rats. J Endocrinol. 2006 Oct;191(1):189-95.

[3]. Involvement of the calcium-sensing receptor in human taste perception. J Biol Chem. 2010 Jan 8;285(2):1016-22.

[4]. Calcium-sensing receptor mediates dietary peptide-induced CCK secretion in enteroendocrine STC-1 cells. Mol Nutr Food Res. 2012 May;56(5):753-60.

[5]. Calcium-sensing receptor: a sensor and mediator of ischemic preconditioning in the heart. Am J Physiol Heart Circ Physiol. 2010 Nov;299(5):H1309-17.

Drug Indication

Studied for the treatment of osteoporosis.

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Mechanism of Action
NPS-2143 is a prototype calcium-sensitive receptor antagonist designed to act on calcium receptors on the surface of the parathyroid gland, stimulating the release of parathyroid hormone (PTH) stored in the body.

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NPS-2143 (SB-262470) is a selective CaSR antagonist (calcium-sensitive receptor antagonist) designed to investigate the function and potential therapeutic applications of CaSR[1][2]
- Its core mechanism is to block the activation of CaSR by extracellular calcium, thereby reversing CaSR-mediated inhibition of parathyroid PTH secretion[1]
- Research applications include exploring the role of CaSR in parathyroid hormone regulation, taste perception, intestinal hormone secretion and cardiac ischemic preconditioning[3][4][5]
- It induces SHR hypertension via a PTH-dependent pathway and a PTH-independent pathway, suggesting that CaSR is involved in blood pressure regulation[2]
- As a tool compound, it helps to elucidate the physiological processes mediated by CaSR and has potential therapeutic value for osteoporosis (through PTH stimulation) and CaSR-related diseases[1]

C24H25CLN2O2

408.92

408.16

C, 70.49; H, 6.16; Cl, 8.67; N, 6.85; O, 7.82

284035-33-2

NPS-2143 hydrochloride; 324523-20-8

6918446

White to off-white solid powder

1.23 g/cm3

608.4ºC at 760 mmHg

1.17E-15mmHg at 25°C

1.631

5.106

2

4

8

29

560

1

N#CC1=C(OC[C@@H](CNC(C)(CC2=CC=C3C=CC=CC3=C2)C)O)C=CC=C1Cl

PZUJQWHTIRWCID-HXUWFJFHSA-N

InChI=1S/C24H25ClN2O2/c1-24(2,13-17-10-11-18-6-3-4-7-19(18)12-17)27-15-20(28)16-29-23-9-5-8-22(25)21(23)14-26/h3-12,20,27-28H,13,15-16H2,1-2H3/t20-/m1/s1

2-chloro-6-[(2R)-2-hydroxy-3-[(2-methyl-1-naphthalen-2-ylpropan-2-yl)amino]propoxy]benzonitrile

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 (6.11 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 (6.11 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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Solubility in Formulation 3: 2% DMSO +Corn oil : 10mg/mL

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