S1P3 ( IC50 = 4.6 μM ); CAY10444 (also known as BML-241) was initially developed as a potential S1P3 receptor antagonist through rational drug design. However, subsequent studies have demonstrated that it lacks selectivity for S1P3 receptors and exhibits significant off-target effects [1].
In a β-arrestin recruitment assay, the IC50 of CAY10444 for inhibiting the S1P3 receptor response was determined to be 4.6 μM [1].
S1P₃ receptor (IC₅₀ = 4.6 μM in a β-arrestin recruitment assay using Tango EDG3 cell lines) [1]

BML-241 blocks P2 receptor or α1A-adrenoceptor stimulation-induced increases in intracellular Ca²⁺ concentration as well as α1A-adrenoceptor-mediated contraction of the rat mesenteric artery. However, it has no effect on S1P3-mediated reduction of Forskolin-induced cyclic AMP accumulation[1].

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In HeLa cells expressing S1P3 receptors, CAY10444 at 10 μM inhibited S1P-induced increases in intracellular calcium concentration by 37%. At the same concentration, it inhibited S1P-induced calcium increases in cells expressing S1P1 receptors by only about 7% [1].
In transfected CHO cells, CAY10444 (10 μM) was shown to lack selectivity, as it inhibited calcium increases induced not only by S1P but also by purinergic P2 receptor or α1A-adrenoceptor stimulation [1].
In a β-arrestin recruitment assay using Tango EDG3 cell lines, the S1P3 receptor response was inhibited by CAY10444 with an IC50 of 4.6 μM, with 100 μM achieving approximately 78% inhibition [1].
In multiple other cell systems, including fibroblasts, keratinocytes, cardiomyocytes, and various cancer cell lines, CAY10444 (tested at concentrations ranging from 0.1 to 100 μM) was used to assess S1P3 involvement, with many studies showing no effect or concluding against a role for S1P3 [1].

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In HeLa cells expressing S1P₃ or S1P₁ receptors, CAY10444 (BML-241) at 10 μM inhibited S1P-induced [Ca²⁺]i increases by 37% (S1P₃) and 7% (S1P₁), respectively. [1]
- In transfected CHO cells, CAY10444 (BML-241) at 10 μM failed to block S1P₃-mediated decreases in forskolin-induced cAMP accumulation, but it inhibited [Ca²⁺]i increases mediated by purinergic P₂ receptors or α₁A-adrenoceptor stimulation, as well as α₁A-adrenoceptor-mediated contraction of rat mesenteric artery. [1]
- In human keratinocytes, CAY10444 (BML-241) at 50 μM blocked S1P-induced [Ca²⁺]i increase, an effect also reproduced by the mixed S1P₁/S1P₃ antagonist VPC23019, while the S1P₁ agonist SEW2871 had no effect. [1]
- In a Tango β-arrestin recruitment assay with EDG3 (S1P₃) cell lines, CAY10444 (BML-241) inhibited the S1P-induced response to 78% of the receptor response at 100 μM, with an IC₅₀ of 4.6 μM. [1]
- In immature B lymphocyte chemotaxis assays, CAY10444 (BML-241) at 100 μM only slightly and non-significantly inhibited S1P-induced migration; however, B cells from S1P₃ knockout mice were unable to migrate to S1P at all concentrations tested. [1]

β-Arrestin Recruitment Assay: A Tango EDG3 cell line system was used to assess S1P3 receptor activity. Cells were treated with S1P in the presence or absence of varying concentrations of CAY10444. The β-arrestin recruitment response was measured, and the IC50 value for CAY10444 was calculated to be 4.6 μM [1].

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In a β-arrestin recruitment assay (Tango technology), cells expressing the EDG3 (S1P₃) receptor were stimulated with S1P in the presence of varying concentrations of CAY10444 (BML-241). The compound inhibited the receptor response in a concentration-dependent manner, with an IC₅₀ value of 4.6 μM; at 100 μM, the response was reduced to 78% of the control. [1]
- In a calcium mobilization assay using HeLa cells transiently or stably expressing S1P₁ or S1P₃ receptors, CAY10444 (BML-241) was tested at a single concentration of 10 μM. The compound inhibited S1P-evoked [Ca²⁺]i increases by 37% in S1P₃-expressing cells and by approximately 7% in S1P₁-expressing cells. [1]
- In a cAMP accumulation assay in CHO cells expressing S1P₃ receptors, cells were pre-incubated with forskolin to elevate cAMP levels. Treatment with S1P decreased forskolin-induced cAMP accumulation. CAY10444 (BML-241) at 10 μM did not block this S1P₃-mediated effect. In contrast, in the same cell system, the compound inhibited [Ca²⁺]i increases triggered by purinergic P₂ receptor or α₁A-adrenoceptor stimulation. [1]

Calcium Mobilization Assay (HeLa cells): HeLa cells expressing S1P3 or S1P1 receptors were used. Cells were loaded with a calcium-sensitive dye, and changes in intracellular calcium concentration ([Ca²⁺]i) were measured after stimulation with S1P. CAY10444 was tested at 10 μM to assess its ability to block S1P-induced calcium responses [1].
Calcium Mobilization Assay (CHO cells): Transfected CHO cells expressing various receptors (including S1P3, P2 receptors, and α1A-adrenoceptors) were used. Cells were stimulated with S1P, UTP, or phenylephrine in the presence or absence of CAY10444 (10 μM), and changes in [Ca²⁺]i were measured to assess the selectivity of the compound [1].
β-Arrestin Recruitment Assay: Tango EDG3 cell lines were used. Cells were incubated with S1P and varying concentrations of CAY10444. The recruitment of β-arrestin to the activated S1P3 receptor was measured to determine antagonist potency [1].

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HeLa cells expressing S1P₃ or S1P₁ receptors were loaded with a calcium indicator, and changes in [Ca²⁺]i were measured upon S1P stimulation. Cells were pretreated with CAY10444 (BML-241) at 10 μM for a specified period before agonist addition. The percentage inhibition of the S1P-induced calcium signal was calculated relative to control. [1]
- CHO cells transfected with S1P₃ receptors were used for cAMP accumulation assays. Cells were incubated with forskolin to stimulate cAMP production, then treated with S1P in the presence or absence of CAY10444 (BML-241) (10 μM). Cellular cAMP levels were quantified. Separately, CHO cells were used to measure [Ca²⁺]i increases induced by purinergic P₂ receptor or α₁A-adrenoceptor agonists, with or without CAY10444 (BML-241) pretreatment. [1]
- Human keratinocytes were loaded with a calcium-sensitive dye, and [Ca²⁺]i responses to S1P were recorded. Cells were pre-incubated with CAY10444 (BML-241) at 50 μM, and the inhibition of the calcium signal was assessed. Parallel experiments were performed with the S1P₁ agonist SEW2871 and the mixed S1P₁/S1P₃ antagonist VPC23019 for comparison. [1]
- Immature B lymphocyte migration toward S1P was evaluated in transwell chambers. Cells were pre-treated with CAY10444 (BML-241) at 100 μM, and the number of migrated cells was counted. The effect of the compound was compared to the migration ability of B cells from S1P₃ knockout mice. [1]

[1]. Selectivity and specificity of sphingosine-1-phosphate receptor ligands: caveats and critical thinking in characterizing receptor-mediated effects. Front Pharmacol. 2011 Feb 22;2:9.

4-Thiazolidinic acid, 2-undecyl-,(4r)- is an L-α-amino acid.

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CAY10444 (BML-241) is commonly used as a putative S1P₃ receptor antagonist despite reports of its lack of selectivity. It has been shown to inhibit responses via purinergic P₂ receptors and α₁A-adrenoceptors, while failing to block S1P₃-mediated cAMP decrease. The compound exhibits low affinity for S1P₃ receptors (IC₅₀ 4.6 μM), and at concentrations of 1–10 μM it often shows no effect, leading some authors to incorrectly conclude the absence of S1P₃ involvement in their paradigms. Higher concentrations (50–100 μM) are required to observe antagonism, but even at 100 μM, the inhibition may be modest or non-significant. These observations highlight the caveats of relying on CAY10444 (BML-241) as a pharmacological tool without confirmatory genetic (knockout/knockdown) evidence. [1]

C15H29NO2S

287.46

287.191

C, 62.67; H, 10.17; N, 4.87; O, 11.13; S, 11.15

298186-80-8

2727678

White to off-white solid powder

1.0±0.1 g/cm3

439.2±40.0 °C at 760 mmHg

219.4±27.3 °C

0.0±2.3 mmHg at 25°C

1.495

5.42

2

4

11

19

248

1

CCCCCCCCCCCC1N[C@@H](CS1)C(=O)O

FNBSOIBCKUUVJJ-LSLKUGRBSA-N

InChI=1S/C15H29NO2S/c1-2-3-4-5-6-7-8-9-10-11-14-16-13(12-19-14)15(17)18/h13-14,16H,2-12H2,1H3,(H,17,18)/t13-,14?/m0/s1

(4R)-2-undecyl-1,3-thiazolidine-4-carboxylic acid

CAY-10444; BML-241; CAY10444; CAY10,444; 298186-80-8; (4R)-2-undecyl-1,3-thiazolidine-4-carboxylic acid; (4R)-2-Undecyl-4-thiazolidinecarboxylic acid; 2-Undecyl-thiazolidine-4-carboxylic acid; BML241; CAY 10444; BML 241

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: 14~41.7 mg/mL (48.7~145 mM)

Solubility in Formulation 1: 2.5 mg/mL (8.70 mM) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), suspension solution; with sonication.
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 (8.70 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.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (8.70 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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 (Please use freshly prepared in vivo formulations for optimal results.)