ML221

Catalog No.S8695 Batch:S869501

Technical Data

Formula

C₁₇H₁₁N₃O₆S

Molecular Weight

385.35

CAS No.

877636-42-5

Solubility (25°C)*

In vitro

DMSO

9 mg/mL (23.35 mM)

Water

Insoluble

Ethanol

Insoluble

In Vivo (Add solvents to the product individually and in order.)

Homogeneous suspension	CMC-NA	≥5mg/ml	Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.
Clear solution	5%DMSO 1 40%PEG300 2 5%Tween80 3 50%ddH2O Validated by Selleck labs. Should you need adjustments to this formulation, contact our sales team for custom testing.	0.450mg/ml (1.17mM)	Taking the 1 mL working solution as an example, add 50 μL of 9 mg/ml clarified DMSO stock solution to 400 μL of PEG300, mix evenly to clarify it; add 50 μL of Tween80 to the above system, mix evenly to clarify; then continue to add 500 μL of ddH2O to adjust the volume to 1 mL. The mixed solution should be used immediately for optimal results.
Clear solution	5% DMSO 1 95% Corn oil Validated by Selleck labs. Should you need adjustments to this formulation, contact our sales team for custom testing.	0.112mg/ml (0.29mM)	Taking the 1 mL working solution as an example, add 50 μL of 2.24 mg/ml clear DMSO stock solution to 950 μL of corn oil and mix evenly. The mixed solution should be used immediately for optimal results.

* <1 mg/ml means slightly soluble or insoluble.
* Please note that Selleck tests the solubility of all compounds in-house, and the actual solubility may differ slightly from published values. This is normal and is due to slight batch-to-batch variations.
* Room temperature shipping (Stability testing shows this product can be shipped without any cooling measures.)

Preparing Stock Solutions

Biological Activity

Description

ML221 is a potent Apelin receptor (APJ) functional antagonist in cell-based assays that is >37-fold selective over the closely related Angiotensin Receptor type 1 (AT1) receptor. The IC50 values of ML221 are 0.70 and 1.75 μM in a cAMP assay and β-Arrestin assay, respectively.

Targets

apelin receptor
(β-arrestin assay)

1.75 μM

In vitro

In a PAMPA permeability assay, ML221 exhibits moderate permeability. It displays moderate plasma and poor microsomal stability, as it is rapidly metabolised in both human and mouse liver homogenates (4.2% and 4.9% remaining at 60 min). It shows no toxicity (>50 μM) toward human hepatocytes. This compound displays limited cross reactivity against a range of GPCRs. It inhibits endothelial cell proliferation by blocking apelin-APJ signalling without affecting the expression of VEGF and VEGFR2.

In Vivo

Intraperitoneal administration of ML221 inhibits pathological angiogenesis but enhances the recovery of normal vessels into the ischaemic regions in the retina of the OIR model mice. A single application of this compound alleviates mechanical allodynia and heat hyperalgesia 7 days following chronic constriction injury (CCI), in a dose‑dependent manner. Intraspinal delivery of this chemical, at the onset of and in fully‑established neuropathic pain, persistently attenuates CCI‑induced pain hypersensitivity, indicating that the apelin‑APJ system is involved in initiating and maintaining pain. Intrathecal ML221 downregulates phosphorylated extracellular signal‑related kinase (ERK) in the rat spinal cord dorsal horn, suggesting that the effect of apelin on neuropathic pain may be mediated via ERK signalling.

Protocol (from reference)

Cell Assay:^{^[2]}	Cell Lines bEnd.3 cells Concentrations 0–30 μM Incubation Time 24 h Method The proliferation of bEnd.3 cells following 24 h incubation with ML221 (0-30 μM) are assessed by the MTT assay and the BrdU incorporation assay. Mouse endothelial cell line bEnd.3 cells are maintained in Dulbecco's modified Eagle's medium supplemented with 10% heat-inactivated fetal bovine serum. The cells are plated at a density of 2.5 × 10⁴/well in 24 well culture plates for the MTT assay and the BrdU incorporation assay. To perform the BrdU incorporation assay, 10 μM BrdU is added to the culture medium. After 2 h, the cells are fixed with 4% paraformaldehyde for 10 minutes. After removing the 4% paraformaldehyde, the cells are incubated with 2 M hydrochloric acid for 10 min at 37℃ for DNA hydrolysis, followed by neutralization with 0.1 M Sodium Borate buffer (pH 8.5) for 30 min at room temperature. Intranuclear BrdU is labeled with mouse anti-BrdU antibodies. The primary antibody is visualized with biotinylated goat anti-mouse IgG antibodies plus streptavidin fluorescein isothiocyanate. Nuclei are detected with Hoechst33342. The BrdU incorporation rate is determined by calculating the numbers of BrdU positive cells per Hoechst positive cells.
Animal Study:^{^[2]}	Animal Models An ischemic retinopathy mouse model (C57BL/6 N mice) Dosages 10 mg/kg Administration i.p.

Cell Assay:^{^[2]}

Cell Lines
bEnd.3 cells
Concentrations
0–30 μM
Incubation Time
24 h
Method
The proliferation of bEnd.3 cells following 24 h incubation with ML221 (0-30 μM) are assessed by the MTT assay and the BrdU incorporation assay. Mouse endothelial cell line bEnd.3 cells are maintained in Dulbecco's modified Eagle's medium supplemented with 10% heat-inactivated fetal bovine serum. The cells are plated at a density of 2.5 × 10⁴/well in 24 well culture plates for the MTT assay and the BrdU incorporation assay. To perform the BrdU incorporation assay, 10 μM BrdU is added to the culture medium. After 2 h, the cells are fixed with 4% paraformaldehyde for 10 minutes. After removing the 4% paraformaldehyde, the cells are incubated with 2 M hydrochloric acid for 10 min at 37℃ for DNA hydrolysis, followed by neutralization with 0.1 M Sodium Borate buffer (pH 8.5) for 30 min at room temperature. Intranuclear BrdU is labeled with mouse anti-BrdU antibodies. The primary antibody is visualized with biotinylated goat anti-mouse IgG antibodies plus streptavidin fluorescein isothiocyanate. Nuclei are detected with Hoechst33342. The BrdU incorporation rate is determined by calculating the numbers of BrdU positive cells per Hoechst positive cells.

Animal Study:^{^[2]}

Animal Models
An ischemic retinopathy mouse model (C57BL/6 N mice)
Dosages
10 mg/kg
Administration
i.p.

Sellecks ML221 Has Been Cited by 3 Publications

A temperature-regulated circuit for feeding behavior [ Nat Commun, 2022, 13(1):4229]	PubMed: 35869064
MiR-185-5p regulates the development of myocardial fibrosis [ J Mol Cell Cardiol, 2021, S0022-2828(21)00246-7]	PubMed: 34973276
A temperature-regulated circuit for feeding behavior [ Research Square, 2021, 10.21203/rs.3.rs-916525/v1]	PubMed: None

RETURN POLICY
Selleck Chemical’s Unconditional Return Policy ensures a smooth online shopping experience for our customers. If you are in any way unsatisfied with your purchase, you may return any item(s) within 7 days of receiving it. In the event of product quality issues, either protocol related or product related problems, you may return any item(s) within 365 days from the original purchase date. Please follow the instructions below when returning products.

SHIPPING AND STORAGE
Selleck products are transported at room temperature. If you receive the product at room temperature, please rest assured, the Selleck Quality Inspection Department has conducted experiments to verify that the normal temperature placement of one month will not affect the biological activity of powder products. After collecting, please store the product according to the requirements described in the datasheet. Most Selleck products are stable under the recommended conditions.

NOT FOR HUMAN, VETERINARY DIAGNOSTIC OR THERAPEUTIC USE.