Revaprazan Hydrochloride is a novel acid pump antagonist (APA) that reversibly inhibits gastric H+/K+-ATPase by competing with luminal K+ ions. In this study, its anti-inflammatory effects are mediated through the inhibition of signaling pathways leading to cyclooxygenase-2 (COX-2) expression. It does not have a defined IC50 or Ki value for these anti-inflammatory targets in this paper [1].

Inhibition of COX-2 Expression: In AGS human gastric epithelial cells infected with Helicobacter pylori (H. pylori, 100 MOI for 24h), pretreatment with revaprazan (20 μM for 2h) significantly reduces the H. pylori-induced up-regulation of COX-2 protein expression (p<0.05). This inhibitory effect is more pronounced at 24h than at 12h. This is associated with decreased levels of prostaglandin E2 (PGE2) [1].
Inhibition of NF-κB Activation: Revaprazan pretreatment (20 μM) inhibits H. pylori-induced NF-κB DNA-binding activity, as shown by electrophoretic mobility shift assay (EMSA). It does not affect AP-1 DNA-binding activity. The inhibition of NF-κB is mediated by preserving the cytosolic levels of its inhibitor, IκB-α, which is otherwise degraded upon H. pylori infection. Revaprazan prevents IκB-α degradation, thereby keeping NF-κB in an inactive state in the cytoplasm [1].
Inhibition of Akt and ERK Phosphorylation: Revaprazan pretreatment (5, 20 μM) significantly inhibits H. pylori-induced phosphorylation of Akt (at 24h) and ERK1/2, key kinases involved in inflammatory signaling and COX-2 regulation [1].
Cytoprotective Effect: Revaprazan pretreatment significantly rescues AGS cells from H. pylori-induced cytotoxicity, as measured by MTT assay [1].
Effect on Cell Migration (Wound Healing): In a live cell imaging wound healing assay, H. pylori infection delays the migration/re-epithelialization of AGS cells. Pretreatment with revaprazan at concentrations below 10 μM significantly accelerates wound healing and compensates for the H. pylori-induced delay. Concentrations of 20 μM or higher are less effective, possibly due to simultaneous induction of apoptosis [1].

Cell Culture and H. pylori Infection: AGS human gastric adenocarcinoma epithelial cells were cultured in RPMI 1640 medium with 10% FBS and antibiotics. H. pylori (ATCC 43504, CagA+, VacA+) was cultured on tryptic soy agar with sheep blood under microaerophilic conditions. For infection, AGS cells were co-cultured with H. pylori at a bacterium-to-cell ratio of 100:1 (100 MOI) for various time points (1h for EMSA, 12-24h for Western blot). For drug treatment, cells were pretreated with revaprazan (5, 20, or 50 μM) or vehicle (DMSO) for 2 hours before H. pylori infection [1].
Western Blot Analysis: Cells were lysed in RIPA buffer. Proteins (50 μg) were separated by SDS-PAGE and transferred to PVDF membranes. Membranes were probed with primary antibodies against COX-2, Akt, phospho-Akt, IκB-α, ERK1/2, and phospho-ERK1/2, followed by HRP-conjugated secondary antibodies. Protein bands were visualized using an ECL detection kit [1].
Electrophoretic Mobility Shift Assay (EMSA): Nuclear extracts were prepared from AGS cells. A double-stranded oligonucleotide probe containing the NF-κB consensus sequence was end-labeled with [γ-32P]ATP. Nuclear extracts (10 μg) were incubated with the labeled probe, and DNA-protein complexes were resolved on a 6% non-denaturing polyacrylamide gel. The gel was dried and exposed to X-ray film to visualize NF-κB DNA-binding activity [1].
MTT Cytotoxicity/Viability Assay: AGS cells were plated in 48-well plates. After treatment with H. pylori and/or various concentrations of revaprazan (5, 20, 50 μM), MTT solution (0.25 mg/mL) was added for 2h at 37°C. The resulting formazan crystals were dissolved in DMSO, and absorbance was measured at 570 nm. Cell viability was expressed as a percentage of the control (untreated) cells [1].
Live Cell Migration (Wound Healing) Assay: Confluent monolayers of AGS cells, infected with H. pylori (10 MOI) and pretreated with or without revaprazan (5, 10, 20 μM), were wounded with a pipette tip. Cell migration into the wound was monitored in real-time using a live cell imager for up to 24h, with images captured at 3-minute intervals. The mean velocity of cell growth/migration was calculated from the recorded data [1].

In the MTT assay, treatment of AGS cells with revaprazan alone at concentrations of 10, 30, 40, 50, 80, and 100 μM was performed to assess its effect on cell viability. The results showed a concentration-dependent effect, but specific numerical data on toxicity (e.g., LD50) are not provided in the text [1].

[1]. Revaprazan, a novel acid pump antagonist, exerts anti-inflammatory action against Helicobacter pylori-induced COX-2 expression by inactivating Akt signaling. J Clin Biochem Nutr. 2012 Sep;51(2):77-83.

Revaprazan hydrochloride is a lipophilic, weakly basic hydrochloride form with potassium-competitive acid-blocking (P-CAB) activity. Highly concentrated in the parietal tubules of the stomach, Revaprazan is immediately protonated upon entering an acidic environment and competitively and reversibly binds to the potassium-binding site of the proton pump potassium-hydrogen ATPase (H+/K+ ATPase), thereby inhibiting the pump's activity and the secretion of H+ ions from the parietal cells into the gastric lumen—the final step in gastric acid production.
See also: Revaprazan (note moved to).

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Revaprazan Hydrochloride is a clinically approved acid pump antagonist (APA) used to suppress gastric acid secretion by reversibly inhibiting the H+/K+-ATPase. This study demonstrates a significant additional pharmacological action: direct anti-inflammatory effects. It shows that revaprazan can attenuate H. pylori-induced gastric inflammation by inhibiting the expression of COX-2, a key pro-inflammatory enzyme linked to H. pylori-associated gastritis and gastric carcinogenesis. The mechanism involves the suppression of Akt and ERK phosphorylation and the blockade of NF-κB activation by preventing the degradation of its inhibitor, IκB-α. Furthermore, revaprazan exhibits a cytoprotective effect against H. pylori-induced cytotoxicity and promotes cell migration/wound healing at lower concentrations. These findings suggest that revaprazan's therapeutic benefit in gastritis may extend beyond acid suppression to include direct modulation of the inflammatory response to H. pylori infection [1].

398.167

178307-42-1

199463-33-7;178307-42-1 (HCl);

204103

White to off-white solid powder

205-208°

6.039

2

5

3

28

481

0

CC1=C(C)N=C(NC2=CC=C(C=C2)F)N=C1N3CCC4=CC=CC=C4C3C.Cl

MALPZYQJEDBIAK-UHFFFAOYSA-N

InChI=1S/C22H23FN4.ClH/c1-14-15(2)24-22(25-19-10-8-18(23)9-11-19)26-21(14)27-13-12-17-6-4-5-7-20(17)16(27)3;/h4-11,16H,12-13H2,1-3H3,(H,24,25,26);1H

N-(4-fluorophenyl)-4,5-dimethyl-6-(1-methyl-3,4-dihydro-1H-isoquinolin-2-yl)pyrimidin-2-amine;hydrochloride

YH 1885 YH-1885Revaprazan Hydrochloride YH1885

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment, avoid exposure to moisture.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~8.33 mg/mL (~20.88 mM)
H2O : < 0.1 mg/mL

Solubility in Formulation 1: ≥ 1.56 mg/mL (3.91 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 15.6 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: ≥ 1.56 mg/mL (3.91 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 15.6 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: ≥ 1.56 mg/mL (3.91 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 15.6 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.)