64907-22-8

Basic information

• Product Name: 7-alpha,25-Dihydroxycholesterol
• Synonyms: 7-alpha,25-Dihydroxycholesterol;3β,7α,25-Trihydroxycholest-5-ene;5-Cholesten-3β,7α,25-triol;25-dihydroxycholesterol
• CAS NO: 64907-22-8
• Molecular Formula: C₂₇H₄₆O₃
• Molecular Weight: 418.65
• Mol File: 64907-22-8.mol
• Article Data: 2

Chemical Properties

• Appearance: white to beige/
• Storage Temp.: 2-8°C
• Solubility: DMSO: soluble5mg/mL (clear solution)
• CAS DataBase Reference: 7-alpha,25-Dihydroxycholesterol(CAS DataBase Reference)
• NIST Chemistry Reference: 7-alpha,25-Dihydroxycholesterol(64907-22-8)
• EPA Substance Registry System: 7-alpha,25-Dihydroxycholesterol(64907-22-8)

Safety Data

• WGK Germany: 3
• Hazardous Substances Data: 64907-22-8(Hazardous Substances Data)

Usage

Description

7-alpha,25-Dihydroxycholesterol, also known as 3β,7α,25-Trihydroxycholest-5-ene, is a potent agonist of GPR183, an orphan G protein-coupled receptor (GPCR) that plays a crucial role in humoral immune responses, including B cell migration and T cell-dependent antibody production. It is a cholesterol derivative with a unique chemical structure that allows it to specifically activate the GPR183 receptor with an EC50 value of 50 nM (Kd = 450 pM). At concentrations below 3 μM, it does not show any appreciable binding activity when tested against other nuclear hormone receptors and G protein-coupled receptors.

Uses

Used in Immunology Research:
7-alpha,25-Dihydroxycholesterol is used as a G-protein-coupled receptor 183 (GPR183) ligand in various immunological research applications. It is employed in chemotaxis assays, intracellular cytokine staining methods, and fluorescence-activated cell sorting (FACS) analysis of natural killer cells. These applications help researchers understand the role of GPR183 in immune cell migration and activation.
Used in Oxysterol-based Assays:
7-alpha,25-Dihydroxycholesterol is used in oxysterol-based calcium mobilization assays in Chinese hamster ovary (CHO) cells. These assays are crucial for studying the effects of oxysterols on cellular signaling pathways and their potential role in various physiological and pathological processes.
Used in Radioligand Binding Assays:
7-alpha,25-Dihydroxycholesterol is also used in competitive radioligand binding assays of Epstein-Barr virus-induced gene 2 (EBI2) expressing COS-7 cells. These assays help in determining the binding affinity and specificity of 7-alpha,25-Dihydroxycholesterol for the GPR183 receptor, providing valuable information for drug discovery and development efforts targeting this receptor.

Biological Activity

7α,25-dihydroxy cholesterol (7α,25-dhc, 7α,25-ohc) is a potent and selective gpr183 agonist [1].epstein-barr virus (ebv)-induced gene 2 (ebi2, also known as gpr183) is an orphan g protein-coupled receptor that is highly expressed in spleen and up-regulated upon epstein–barr-virus infection. gpr183 is required for humoral immune responses and polymorphisms in the receptor is associated with inflammatory autoimmune diseases [1][2].7α,25-dihydroxy cholesterol is a potent and selective gpr183 (ebi2) agonist and most potent ebi2 ligand with ec50 and kd values of 140 pm and 450 pm, respectively. in a competition binding study, 7α,25-ohc competed for binding with ic50 value of 242 pm. in ebi2-expressing cells, 7α,25-ohc dose-dependently inhibited forskolin-induced camp with ic50 value of 2 nm. 7α,25-ohc was probably the endogenous ligand for ebi2. in ebi2-expressing mouse b and t cells, 7α,25-dihydroxy cholesterol stimulated cell migration with ic50 value of around 500 pm, but had no effect on ebi2-deficient cells [1].in ebi2-deficient mice, unseparated splenocytes, b cells and t cells were completely unresponsive to 7α,25-ohc stimulation [1]. mice deficient in ch25h, a key enzyme for the generation of 7α, 25-ohc, failed to position activated b cells within the spleen to the outer follicle [2].

Biochem/physiol Actions

7α,25-dihydroxycholesterol (7α,25-OHC) serves as an endogenous ligand for G-protein-coupled receptor 183 (GPR183) or Epstein-Barr virus-induced gene 2 (EBI2). It plays a key role in the migration of type 3 innate lymphoid cells (ILC3) in the small intestine and colon. 7α,25-OHC mediates immune cell migration functionality by their chemoattractant property. Inhibition of 7α,25-OHC synthesis leads to impairment in the migration of B cells.

references

[1]. liu c, yang xv, wu j, et al. oxysterols direct b-cell migration through ebi2. nature. 2011 jul 27;475(7357):519-23.[2]. hannedouche s, zhang j, yi t, et al. oxysterols direct immune cell migration via ebi2. nature. 2011 jul 27;475(7357):524-7.

Synthetic route

cholest-5-ene-3β,7α,25-triol 3β,25-dibenzoate (306288-44-8) → [3H]-7alpha, 25-Dihydroxycholesterol (64907-22-8)

Conditions	Yield
With methanol; potassium hydroxide for 4h; Reflux; Inert atmosphere;	81%
With potassium hydroxide In methanol for 1h; Heating;	51%

25-hydroxychlolesterol (2140-46-7) → [3H]-7alpha, 25-Dihydroxycholesterol (64907-22-8)

Conditions	Yield
Multi-step reaction with 4 steps 1: 98 percent / pyridine / 3 h / 20 °C 2: 87 percent / PCC; 3A molecular sieves / benzene / 24 h / Heating 3: 71 percent / L-Selectride / tetrahydrofuran / 5 h / -78 °C 4: 51 percent / KOH / methanol / 1 h / Heating View Scheme
Multi-step reaction with 4 steps 1.1: pyridine / dichloromethane / 42 h / 20 °C / Inert atmosphere 2.1: tert.-butylhydroperoxide / ethyl acetate / 0.5 h / Molecular sieve; Inert atmosphere 2.2: 48 h / 20 °C / Molecular sieve; Inert atmosphere 3.1: L-Selectride / tetrahydrofuran / 3 h / -78 °C / Inert atmosphere 4.1: potassium hydroxide; methanol / 4 h / Reflux; Inert atmosphere View Scheme

cholest-5-ene-3β,25-diol 3β,25-dibenzoate (64164-61-0) → [3H]-7alpha, 25-Dihydroxycholesterol (64907-22-8)

Conditions	Yield
Multi-step reaction with 3 steps 1: 87 percent / PCC; 3A molecular sieves / benzene / 24 h / Heating 2: 71 percent / L-Selectride / tetrahydrofuran / 5 h / -78 °C 3: 51 percent / KOH / methanol / 1 h / Heating View Scheme
Multi-step reaction with 3 steps 1.1: tert.-butylhydroperoxide / ethyl acetate / 0.5 h / Molecular sieve; Inert atmosphere 1.2: 48 h / 20 °C / Molecular sieve; Inert atmosphere 2.1: L-Selectride / tetrahydrofuran / 3 h / -78 °C / Inert atmosphere 3.1: potassium hydroxide; methanol / 4 h / Reflux; Inert atmosphere View Scheme

3β,25-dihydroxycholest-5-en-7-one 3β,25-dibenzoate (306288-41-5) → [3H]-7alpha, 25-Dihydroxycholesterol (64907-22-8)

Conditions	Yield
Multi-step reaction with 2 steps 1: 71 percent / L-Selectride / tetrahydrofuran / 5 h / -78 °C 2: 51 percent / KOH / methanol / 1 h / Heating View Scheme
Multi-step reaction with 2 steps 1: L-Selectride / tetrahydrofuran / 3 h / -78 °C / Inert atmosphere 2: potassium hydroxide; methanol / 4 h / Reflux; Inert atmosphere View Scheme

[3H]-7alpha, 25-Dihydroxycholesterol (64907-22-8) → 7α,25-dihydroxy-4-cholesten-3-one (98774-41-5)

Conditions	Yield
With recombinant Brevibacterium cholesterol oxidase In phosphate buffer; isopropyl alcohol at 25℃; for 12h; pH=7;

[3H]-7alpha, 25-Dihydroxycholesterol (64907-22-8) → 1-[(7R,8S,9S,10R,13R,14S,17R)-7-Hydroxy-17-((R)-5-hydroxy-1,5-dimethyl-hexyl)-10,13-dimethyl-1,2,6,7,8,9,10,11,12,13,14,15,16,17-tetradecahydro-cyclopenta[a]phenanthren-(3Z)-ylidene-hydrazinocarbonylmethyl]-pyridinium; chloride

Conditions	Yield
Multi-step reaction with 2 steps 1: recombinant Brevibacterium cholesterol oxidase / propan-2-ol; aq. phosphate buffer / 12 h / 25 °C / pH 7 2: methanol; H2O; acetic acid / 20 °C View Scheme

Upstream product

• 306288-41-5 3β,25-dihydroxycholest-5-en-7-one 3β,25-dibenzoate 
• 64164-61-0 cholest-5-ene-3β,25-diol 3β,25-dibenzoate 
• 2140-46-7 25-hydroxychlolesterol 
• 306288-44-8 cholest-5-ene-3β,7α,25-triol 3β,25-dibenzoate 

Relevant articles and documents

Fluoro analogs of bioactive oxy-sterols: Synthesis of an EBI2 agonist with enhanced metabolic stability

Synthesis of several 7-hydroxy oxysterols and their potential roles as signaling molecules in the innate and adaptive immune responses is discussed. Discovery of a new, fluorinated, synthetic analog of the 7α,25-dihydroxycholesterol—the endogenous ligand of GPR 183 (EBI2), a G-protein coupled receptor highly expressed upon Epstein–Barr virus infection is described. Fluoro oxysterol 12 showed good metabolic stability while maintaining excellent EBI2 agonist activity.