Novel Glycyrrhetin Ureas: Design,Synthesis And The Therapeutic Evaluation Of Acute Kidney Injury

Cisplatin,an inorganic platinum chemotherapeutic drug widely used in the treatment of various solid tumors,but its adverse reactions greatly limit its clinical use,among which the nephrotoxicity is more significant.Among patients treated with cisplatin,26%of patients developed acute kidney injury（AKI）,50%of patients still had clinical manifestations of elevated serum creatinine after 15 days of treatment.At present,the treatment of AKI is mainly supportive,and there is no effective treatment method.Acute kidney injury is associated with a strong inflammatory response,and inhibiting this response can effectively prevent or improve AKI.Glycyrrhetinic Acid（GA）is a triterpene aglycon component of natural product glycyrrhetinic acid,which is high in licorice root.GA has a variety of significant pharmacological activities,including anti-inflammatory,anti-viral,anti-tumor and hepatoprotective effects.The small structural changes of triterpenoids can cause extensive changes in the chemical and biological activities of medicines.Most of the structural modifications of GA on the 3-hydroxy（or 3-oxo）or/and 30-carboxyl group show better activity than GA.The treatment and application of amino acids in renal failure,respiratory tract,heart disease,neurological diseases and congenital defects have attracted many people’s attention.In addition,amino acid drugs have higher bioavailability,lower toxicity,and good osmotic pressure and pharmacokinetic properties.Because urea can form multiple stable hydrogen bonds with proteins and receptor targets,urea stents are often incorporated to adjust drug efficacy and selectivity,and improve drug design and drug performance in medicinal chemistry.The soluble epoxide hydrolase（s EH）inhibitor AR9281 exhibits anti-inflammatory activity through the urea moiety bound to the catalytic residue of s HE.The protein-ligand complex between vascular endothelial growth factor receptor 2（VEGFR2）and lovatinib highlights the importance of urea function,so we further confirmed that introducing the urea component into the structure of GA can improve the potency and drug-like properties of GA.In order to develop new type for the prevention and treatment of acute kidney injury anti-inflammatory drugs,glycyrrhetinic acid as the mother nucleus structure,design and synthesis of a series of compounds containing urea unit,and explore new glycyrrhetinic acid urea compounds in vivo experiments to LPS stimulation of RAW264.7 cells inhibit inflammation,and its protective effect of AKI in mice induced by cisplatin.First,we add all the compounds at 20mM to the medium containing RAW264.7cells and incubated for 24 h.The MTT method was used to detect the effect of the compounds on cell viability.The RAW264.7 cells were stimulated with 0.5mg/mL lipopolysaccharide solution and cultured in the incubator for 24 hours.The NO content in the supernatant was detected with a nitric oxide kit to evaluate the compound’s ability to inhibit NO release.Compound 5r with final concentrations of 20mM,10mM and 5mM was added to LPS-stimulated RAW264.7 cells,incubated for 24 h,and the content of inflammatory factors in the supernatant was detected by enzyme-linked immunosorbent assay.Thirty male C57BL/6 mice weighing about 18-25 g was randomly divided into 3 groups,the blank control group,the model group and the 5r group.The mice in the 5r group were given 30 mg/kg of compound 5r by intragastric administration,and the other two groups were given the same amount of normal saline.Mice in the model group and 5r group were intraperitoneally injected with 20 mg/kg cisplatin solution to induce acute kidney injury,and all mice in the blank control group were given the same dose of normal saline,to detect the level of inflammation in vivo and the degree of kidney injury in each group.The experimental results indicated that the cell viability percentages of all compounds are above 80%,and glycyrrhetinic urea agents have no obvious toxic effect on RAW264.7 cells.The results of NO detection experiments showed that most glycyrrhetinic acid urea agents have a fine inhibitory effect on NO.When 3-OH was acetylated,the inhibitory effect of the compound on NO was reduced.For cyclic amines or aliphatic amines at the R2 position,the smaller substituents showed better NO inhibition,and the compounds 5r-5u with acidic or neutral amino acids showed significant NO inhibition(IC₅₀=2-3μM).The compound 5r has the best anti-inflammatory activity,therefore,it was selected as a further mechanism study.The results of enzyme-linked immunosorbent assay showed that the compound 5r inhibited the release of TNF-aand IL-6 in a dose-dependent manner.At the same time,the compound 5r significantly reduced the content of inflammatory factors TNF-aand IL-6 in AKI mice induced by cisplatin,and inhibited the development of inflammation in AKI mice.Novel glycyrrhetin urea agents partly introduced urea into the structure of GA,thereby improving the efficacy and drug-like properties of GA.Some of the new glycyrrhetinic urea agents have obvious ability to inhibit the production of NO.Compound 5r can not only inhibit the production of pro-inflammatory factors TNF-αand IL-6 at the cellular and animal levels,reduce the damage caused by excessive metabolic reactions,but also protect the kidney damage caused by cisplatin toxicity and maintain metabolic balance.