CyPPA

Cat No.:V19008 Purity: ≥98%

COA Product Data Instructions for use SDS

CyPPA is a positive modulator of hSK3 and hSK2 with EC50s of 14 μM and 5.6 μM, respectively.

CyPPA Chemical Structure CAS No.: 73029-73-9
Product category: New1

This product is for research use only, not for human use. We do not sell to patients.

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Product Description
Bioactivity & Protocols
Physicochemical Properties
Solubility Data
Calculators
Biological Data

Product Description

CyPPA is a positive modulator of hSK3 and hSK2 with EC50s of 14 μM and 5.6 μM, respectively. CyPPA is inactive against hSK1 and hIK channels.

Biological Activity I Assay Protocols (From Reference)

ln Vitro	The apparent Ca2+ sensitivity of channel activation is increased concentration-dependently by CyPPA, resulting in a change in the EC50 (Ca2+) from 429 nM to 59 nM [1].
ln Vivo	CyPPA is a positive regulator of small-conductance Ca2+-activated K+ channels that can delay premature delivery in mice and decrease phasic contractions of the uterus [2].
References	[1]. Neurons, Inhibits Dopamine Release, and Counteracts Hyperdopaminergic Behaviors Induced by Methylphenidate. Br J Pharmacol. 2007 Jul;151(5):655-65. [2]. CyPPA, a positive modulator of small-conductance Ca(2+)-activated K(+) channels, inhibits phasic uterine contractions and delays preterm birth in mice. Am J Physiol Cell Physiol. 2011 Nov;301(5):C1027-35.

These protocols are for reference only. InvivoChem does not independently validate these methods.

Physicochemical Properties

Molecular Formula	C16H23N5
Molecular Weight	285.38732
Exact Mass	285.195
CAS #	73029-73-9
PubChem CID	909822
Appearance	White to off-white solid powder
Density	1.2±0.1 g/cm3
Boiling Point	512.2±42.0 °C at 760 mmHg
Flash Point	263.6±27.9 °C
Vapour Pressure	0.0±1.3 mmHg at 25°C
Index of Refraction	1.648
LogP	2.13
Hydrogen Bond Donor Count	1
Hydrogen Bond Acceptor Count	4
Rotatable Bond Count	3
Heavy Atom Count	21
Complexity	331
Defined Atom Stereocenter Count	0
InChi Key	USEMRPYUFJNFQN-UHFFFAOYSA-N
InChi Code	InChI=1S/C16H23N5/c1-11-10-15(18-14-7-5-4-6-8-14)19-16(17-11)21-13(3)9-12(2)20-21/h9-10,14H,4-8H2,1-3H3,(H,17,18,19)
Chemical Name	N-cyclohexyl-2-(3,5-dimethylpyrazol-1-yl)-6-methylpyrimidin-4-amine
HS Tariff Code	2934.99.9001
Storage	Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month
Shipping Condition	Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Solubility Data

Solubility (In Vitro)	DMSO : ~100 mg/mL (~350.40 mM)
Solubility (In Vivo)	Solubility in Formulation 1: ≥ 2.5 mg/mL (8.76 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. Solubility in Formulation 2: ≥ 2.5 mg/mL (8.76 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. (Please use freshly prepared in vivo formulations for optimal results.)

Solubility (In Vitro)

DMSO : ~100 mg/mL (~350.40 mM)

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 2.5 mg/mL (8.76 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.

Solubility in Formulation 2: ≥ 2.5 mg/mL (8.76 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.

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

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	3.5040 mL	17.5199 mL	35.0398 mL
	5 mM	0.7008 mL	3.5040 mL	7.0080 mL
	10 mM	0.3504 mL	1.7520 mL	3.5040 mL

*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.

Calculator

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Molarity Calculator allows you to calculate the mass, volume, and/or concentration required for a solution, as detailed below:

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See Example

An example of molarity calculation using the molarity calculator is shown below:
What is the mass of compound required to make a 10 mM stock solution in 5 ml of DMSO given that the molecular weight of the compound is 350.26 g/mol?

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The answer of 17.513 mg appears in the Mass box. In a similar way, you may calculate the volume and concentration.

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Dilution Calculator allows you to calculate how to dilute a stock solution of known concentrations. For example, you may Enter C1, C2 & V2 to calculate V1, as detailed below:

What volume of a given 10 mM stock solution is required to make 25 ml of a 25 μM solution?
Using the equation C1V1 = C2V2, where C1=10 mM, C2=25 μM, V2=25 ml and V1 is the unknown:

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The answer of 62.5 μL (0.1 ml) appears in the Volume (Start) box

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Molecular Weight Calculator allows you to calculate the molar mass and elemental composition of a compound, as detailed below:

Note: Chemical formula is case sensitive: C12H18N3O4 c12h18n3o4
Instructions to calculate molar mass (molecular weight) of a chemical compound:

To calculate molar mass of a chemical compound, please enter the chemical/molecular formula and click the “Calculate’ button.

Definitions of molecular mass, molecular weight, molar mass and molar weight:

Molecular mass (or molecular weight) is the mass of one molecule of a substance and is expressed in the unified atomic mass units (u). (1 u is equal to 1/12 the mass of one atom of carbon-12)
Molar mass (molar weight) is the mass of one mole of a substance and is expressed in g/mol.

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Reconstitution Calculator allows you to calculate the volume of solvent required to reconstitute your vial.

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The answer appears in the Volume (to add to vial) box

In vivo Formulation Calculator (Clear solution)

Step 1: Enter information below (Recommended: An additional animal to make allowance for loss during the experiment)

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Step 2: Enter in vivo formulation (This is only a calculator, not the exact formulation for a specific product. Please contact us first if there is no in vivo formulation in the solubility section.)

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Calculation results

Working concentration： mg/mL；

Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.

Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH₂O，mix and clarify.

Note： (1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.

Biological Data

CyPPA is a positive modulator of hSK3 but not hIK channels. (a) Chemical structures of the subtype-selective SK channel activator CyPPA and the pan-selective compound NS309. (b) The left panel shows whole-cell current–voltage (I-V) relationships from a HEK293 cell expressing hSK3. The currents were recorded upon application of 200 ms long voltage ramps (−80 to +80 mV) elicited every 5 s from a holding potential of 0 mV. The free [Ca2+] in the pipette solution was buffered at 0.3 μm and the [K+] was 154 mm in the intra- as well as extracellular solutions. The traces were obtained before (Ctrl) and during application of 40 nm NS309 or 300 nm CyPPA as indicated. The right panel shows the current at −75 mV as a function of time. The first 7 min after obtaining the whole-cell configuration are not shown. During this period the hSK3 current increased as a result of Ca2+ equilibration between the pipette solution and the cell; also 0.1% DMSO was shown to be without effect on the currents. NS309 (40 nm), BMB (30 μm) and CyPPA (300 nm) were present in the bath solution as indicated by the bars. BMB inhibited the current by 75 and 77% in the absence and presence of CyPPA, respectively. (c) Same experimental conditions as in (b) but using a hIK-expressing HEK293 cell. The hIK current was activated by 40 nm NS309 and inhibited by 30 nm ChTX. The hIK current was inhibited by 54 and 51% before and during application of 10 μm CyPPA.Br J Pharmacol. 2007 Jul;151(5):655-65.

SK subtype-dependent effects of CyPPA on the Ca2+ activation curves. In the left panels, currents at −75 mV are depicted as a function of time. Currents were measured from inside-out patches obtained from HEK293 cells stably expressing hSK3 (a), hSK2 (b) or hSK1 (c). The patches were exposed to a bath/intracellular [Ca2+] of 0.01, 0.2 or 10 μm as indicated, and CyPPA (10 μm) was applied at the different [Ca2+] as shown by the bars. In the right panels, the Ca2+ concentration–response relationships for hSK3, hSK2 and hSK1 are depicted in the absence or presence of 10 μm CyPPA. Currents from individual patches were normalized with respect to the effect of 10 μm Ca2+ (in the absence of CyPPA). The solid lines are the fit of the Hill equation to mean data. The following EC50(Ca2+) and Hill coefficients were estimated from separate experiments: hSK3, 0.43±0.01 μm and 6.3±0.4 (control, n=14), 0.12±0.02 μm and 3.4±0.2 (+10 μm CyPPA, n=3); hSK2, 0.42±0.02 μm and 5.2±0.1 (control, n=8), 0.20±0.01 and 3.9±0.3 (+10 μm CyPPA, efficacy: 91±4%, n=3); hSK1, 0.42±0.01 μm and 4.5±0.2 (control, n=8), 0.38±0.03 μm and 3.5±0.2 (+10 μm CyPPA, efficacy: 84±7%, n=3); ***P<0.001 compared to corresponding control values (Bonferroni post hoc comparisons).Br J Pharmacol. 2007 Jul;151(5):655-65.

CyPPA activates the endogenous SK channels in TE671 cells but not the endogenous IK channels in HeLa cells. Whole cell I-V relationships measured in a TE671 (a) or HeLa (b) cell upon application of 200 ms long voltage ramps (−80 to +80 mV) elicited every 5 s from a holding potential of 0 mV; [Ca2+] in the pipette solution was 0.4 μm. Whole-cell I-V relationships were measured in the absence of compound (Ctrl) and in the presence of NS309 (1 μm), or CyPPA (10 μm). BMB (100 μm) was applied as an inhibitor of SK channel activity in TE671 cells (a) and clotrimazole (clot, 1 μm) as an inhibitor of IK channels in HeLa cells (b). Membrane potential measured under current-clamp conditions in a TE671 cells (c) or a HeLa cell (d). The cells were exposed to a near-physiological K+ gradient at an intracellular [Ca2+] of 0.1 μm. NS309 (1 μm) and CyPPA (10 μm) were applied to the extracellular solution as indicated and experiments terminated by addition of BMB (100 μm, C) or clot (1 μm, D); n=3–5.Br J Pharmacol. 2007 Jul;151(5):655-65..