store-operated calcium (SOC) channel

When MRS1845 (10 μM) is applied, the impact of β-glycerophosphate on store-sorbed Ca2+ entry (SOCE) is almost completely eliminated [2]. After placental growth factor (PlGF) therapy, MRS1845 (10 μM) virtually completely removes the increase in SOCE and greatly lowers it [3].

Certain DHPs appeared to cause an incomplete blockade of SOC channel-dependent elevations of calcium, suggesting the presence of more than one class of such channels in HL-60 cells. N-Methylnitrendipine (IC(50) 2.6 microM, MRS 1844) and N-propargylnifrendipine (IC(50) 1.7 microM, MRS 1845) represent possible lead compounds for the development of selective SOC channel inhibitors.[1]

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Additional treatment with ORAI1 inhibitor MRS1845 or SGK1 inhibitor GSK650394 virtually disrupted the effects of β-glycerophosphate on SOCE. Moreover, the β-glycerophosphate-induced MSX2, CBFA1, SOX9, and ALPL mRNA expression and activity in HAoSMCs were suppressed in the presence of the ORAI1 inhibitor and upon ORAI1 silencing. In conclusion, enhanced phosphate upregulates ORAI1 and STIM1 expression and store-operated Ca2+-entry, which participate in the orchestration of osteo-/chondrogenic signaling of VSMCs. KEY MESSAGES: • In aortic SMC, phosphate donor ß-glycerophosphate upregulates Ca2+ channel ORAI1. • In aortic SMC, ß-glycerophosphate upregulates ORAI1-activator STIM1. • In aortic SMC, ß-glycerophosphate upregulates store-operated Ca2+-entry (SOCE). • The effect of ß-glycerophosphate on SOCE is disrupted by ORAI1 inhibitor MRS1845. • Stimulation of osteogenic signaling is disrupted by MRS1845 and ORAI1 silencing. [2]

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PlGF significantly increased store-operated Ca2+-entry following re-addition of extracellular Ca2+, an effect virtually abrogated by Orai1 inhibitor MRS1845 (10 μM). PlGF further increased HIF1α transcript and protein levels, an effect again significantly blunted by MRS1845 (10 μM). In conclusion, PlGF upregulates expression of both, Orai1 and STIM1 thus enhancing store-operated Ca2+-entry with subsequent upregulation of HIF1α.[3]

[1]. Dihydropyridines as inhibitors of capacitative calcium entry in leukemic HL-60 cells. Biochem Pharmacol. 2003 Feb 1;65(3):329-38.

[2]. Phosphate-induced ORAI1 expression and store-operated Ca2+ entry in aortic smooth muscle cells. J Mol Med (Berl). 2019 Oct;97(10):1465-1475.

[3]. Upregulation of Orai1 and STIM1 expression as well as store-operated Ca2+ entry in ovary carcinoma cells by placental growth factor. Biochem Biophys Res Commun. 2019 May 7;512(3):467-472.

A series of 1,4-dihydropyridines (DHPs) were investigated as inhibitors of capacitative calcium influx through store-operated calcium (SOC) channels. Such channels activate after ATP-elicited release of inositol trisphosphate (IP(3))-sensitive calcium stores in leukemia HL-60 cells. The most potent DHPs were those containing a 4-phenyl group with an electron-withdrawing substituent, such as m- or p-nitro- or m-trifluoromethyl (IC(50) values: 3-6 microM). Benzyl esters, corresponding to the usual ethyl/methyl esters of the DHPs developed as L-type calcium channel blockers, retained potency at SOC channels, as did N-substituted DHPs. N-Methylation reduced by orders of magnitude the potency at L-type channels resulting in DHPs nearly equipotent at SOC and L-type channels. DHPs with N-ethyl, N-allyl, and N-propargyl groups also had similar potencies at SOC and L-type channels. Replacement of the usual 6-methyl group of DHPs with larger groups, such as cyclobutyl or phenyl, eliminated activity at the SOC channels; such DHPs instead elicited formation of inositol phosphates and release of IP(3)-sensitive calcium stores. Other DHPs also caused a release of calcium stores, but usually at significantly higher concentrations than those required for the inhibition of capacitative calcium influx. Certain DHPs appeared to cause an incomplete blockade of SOC channel-dependent elevations of calcium, suggesting the presence of more than one class of such channels in HL-60 cells. N-Methylnitrendipine (IC(50) 2.6 microM, MRS 1844) and N-propargylnifrendipine (IC(50) 1.7 microM, MRS 1845) represent possible lead compounds for the development of selective SOC channel inhibitors.[1]

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Compromised renal phosphate elimination in chronic kidney disease (CKD) leads to hyperphosphatemia, which in turn triggers osteo-/chondrogenic signaling in vascular smooth muscle cells (VSMCs) and vascular calcification. Osteo-/chondrogenic transdifferentiation of VSMCs leads to upregulation of the transcription factors MSX2, CBFA1, and SOX9 as well as tissue-nonspecific alkaline phosphatase (ALPL) which fosters calcification by degrading the calcification inhibitor pyrophosphate. Osteo-/chondrogenic signaling in VSMCs involves the serum- and glucocorticoid-inducible kinase SGK1. As shown in other cell types, SGK1 is a powerful stimulator of ORAI1, a Ca2+-channel accomplishing store-operated Ca2+-entry (SOCE). ORAI1 is stimulated following intracellular store depletion by the Ca2+ sensor STIM1. The present study explored whether phosphate regulates ORAI1 and/or STIM1 expression and, thus, SOCE in VSMCs. To this end, primary human aortic smooth muscle cells (HAoSMCs) were exposed to the phosphate donor β-glycerophosphate. Transcript levels were estimated by qRT-PCR, protein abundance by western blotting, ALPL activity by colorimetry, calcification by alizarin red S staining, cytosolic Ca2+-concentration ([Ca2+]i) by Fura-2-fluorescence, and SOCE from increase of [Ca2+]i following re-addition of extracellular Ca2+ after store depletion with thapsigargin. As a result, β-glycerophosphate treatment increased ORAI1 and STIM1 transcript levels and protein abundance as well as SOCE in HAoSMCs. Additional treatment with ORAI1 inhibitor MRS1845 or SGK1 inhibitor GSK650394 virtually disrupted the effects of β-glycerophosphate on SOCE. Moreover, the β-glycerophosphate-induced MSX2, CBFA1, SOX9, and ALPL mRNA expression and activity in HAoSMCs were suppressed in the presence of the ORAI1 inhibitor and upon ORAI1 silencing. In conclusion, enhanced phosphate upregulates ORAI1 and STIM1 expression and store-operated Ca2+-entry, which participate in the orchestration of osteo-/chondrogenic signaling of VSMCs. KEY MESSAGES: • In aortic SMC, phosphate donor ß-glycerophosphate upregulates Ca2+ channel ORAI1. • In aortic SMC, ß-glycerophosphate upregulates ORAI1-activator STIM1. • In aortic SMC, ß-glycerophosphate upregulates store-operated Ca2+-entry (SOCE). • The effect of ß-glycerophosphate on SOCE is disrupted by ORAI1 inhibitor MRS1845. • Stimulation of osteogenic signaling is disrupted by MRS1845 and ORAI1 silencing.[2]

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Placental growth factor (PlGF) is produced by tumor cells and stimulates tumor growth and metastasis in part by upregulation of hypoxia inducible factor HIF1α. Orchestration of tumor cell proliferation and migration involves oscillations of cytosolic Ca2+ activity ([Ca2+]i). The [Ca2+]i oscillations could be accomplished by triggering of intracellular Ca2+ release followed by store-operated Ca2+-entry (SOCE). Mechanisms accomplishing SOCE include the pore-forming ion channel unit Orai1 and its regulator STIM1. The present study explored whether PlGF influences the expression of Orai1 and STIM1, as well as SOCE and whether this effect impacts on HIF1α expression. To this end, ovary carcinoma cells were cultured for 24 h without and with PlGF (10 ng/ml). Orai1, STIM1 and HIF1α transcript levels were quantified utilizing RT-PCR and Orai1, STIM1 and HIF1α protein levels by Western blotting. [Ca2+]i was estimated from Fura-2-fluorescence and SOCE from increase of [Ca2+]i following Ca2+ re-addition after Ca2+-store depletion with extracellular Ca2+ removal and sarcoendoplasmatic Ca2+-ATPase (SERCA) inhibitor thapsigargin (1 μM). As a result, exposure of ovary carcinoma cells to PlGF was followed by a significant increase of Orai1 as well as STIM1 transcript and protein levels. PlGF significantly increased store-operated Ca2+-entry following re-addition of extracellular Ca2+, an effect virtually abrogated by Orai1 inhibitor MRS1845 (10 μM). PlGF further increased HIF1α transcript and protein levels, an effect again significantly blunted by MRS1845 (10 μM). In conclusion, PlGF upregulates expression of both, Orai1 and STIM1 thus enhancing store-operated Ca2+-entry with subsequent upregulation of HIF1α.[3]

C₂₁H₂₂N₂O₆

398.40918

398.148

C, 63.31; H, 5.57; N, 7.03; O, 24.09

544478-19-5

11538542

Typically exists as Off-white to light yellow solids at room temperature

1.318g/cm3

530.8ºC at 760 mmHg

274.8ºC

1.595

3.372

0

7

7

29

798

0

CCOC(C1=C(C)N(CC#C)C(C)=C(C(OC)=O)C1C1C=CC=C([N+]([O-])=O)C=1)=O

BITHABUTZRAUGT-UHFFFAOYSA-N

InChI=1S/C21H22N2O6/c1-6-11-22-13(3)17(20(24)28-5)19(18(14(22)4)21(25)29-7-2)15-9-8-10-16(12-15)23(26)27/h1,8-10,12,19H,7,11H2,2-5H3

3-ethyl 5-methyl 2,6-dimethyl-4-(3-nitrophenyl)-1-(prop-2-yn-1-yl)-1,4-dihydropyridine-3,5-dicarboxylate

MRS 1845; MRS1845; MRS-1845; N-Propargylnitrendipene; N-Propargylnitrendipene; MRS-1845; 5-O-ethyl 3-O-methyl 2,6-dimethyl-4-(3-nitrophenyl)-1-prop-2-ynyl-4H-pyridine-3,5-dicarboxylate; Lopac0_000763; MLS002172491;

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 : ≥ 83.33 mg/mL (~209.16 mM)

Solubility in Formulation 1: ≥ 5 mg/mL (12.55 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 50.0 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: ≥ 5 mg/mL (12.55 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 50.0 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: ≥ 5 mg/mL (12.55 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 50.0 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.)