Discovery of Novel Coumarin-quinolinium Derivatives as Pan-KRAS Translation Inhibitors by Targeting 5'-UTR RNA G-Quadruplexes.

Hyperactivated KRAS mutations fuel tumorigenesis and represent attractive targets for cancer treatment. While covalent inhibitors have shown clinical benefits against the KRASG12C mutant, advancements for non-G12C mutants remain limited, highlighting the urgent demand for pan-KRAS inhibitors. RNA G-quadruplexes (rG4s) in the 5'-untranslated region of KRAS mRNA can regulate KRAS translation, making them promising targets for pan-KRAS inhibitor development. Herein, we designed and synthesized 50 novel coumarin-quinolinium derivatives, leveraging our previously developed rG4-specific ligand, QUMA-1. Notably, several compounds exhibited potent antiproliferative activity against cancer cells as pan-KRAS translation inhibitors. Among them, 15a displayed exceptional capability in stabilizing KRAS rG4s, suppressing KRAS translation, and consequently modulating MAPK and PI3K-AKT pathways. 15a induced cell cycle arrest, prompted apoptosis in KRAS-driven cancer cells, and effectively inhibited tumor growth in a KRAS mutant xenograft model. These findings underscore the potential of 15a as a pan-KRAS translation inhibitor, offering a novel and promising approach to target various KRAS-driven cancers.

Design, synthesis and evaluation of 2-pyrimidinylindole derivatives as anti-obesity agents by regulating lipid metabolism.

Obesity, a global pandemic posing a growing threat to human health, necessitates the development of effective and safe anti-obesity agents. Our previous studies highlighted the lipid-lowering effects of indolylquinazoline Bouchardatine and its derivatives. In this study, we employed scaffold hopping and simplification strategies to design and synthesize two new series derivatives by modifying the D ring. Extensive discussions have been conducted regarding the structure-activity relationship between lipid-lowering activity and the new compounds. These discussions have resulted in the discovery of 2-pyrimidinylindole derivatives as a promising scaffold for anti-obesity treatment. The new 2-pyrimidinylindole derivatives exhibited comparable lipid-lowering activity to the previously reported indolylquinazoline derivatives, including SYSU-3d and R17, with reduced toxicity. The most potent compound, 5a, demonstrated a larger therapeutic index, improved aqueous solubility and oral bioavailability compared to the previous lead compounds. In vivo evaluation indicated that 5a effectively reduced lipid accumulation in adipose tissue, improved glucose tolerance, and mitigated insulin resistance and liver function damage caused by a high-fat and high-cholesterol diet. Mechanism studies indicated that 5a may regulate lipid metabolism through the modulation of the PPARγ signaling pathway. Overall, our study has identified a highly active compound 5a, and provided the basis for further development of 2-pyrimidinylindole as a promising scaffold for obesity treatment.

Design and Synthesis of Bouchardatine Derivatives as a Novel AMP-Activated Protein Kinase Activator for the Treatment of Colorectal Cancer.

Metabolic reprogramming is a crucial hallmark of tumorigenesis. Modulating the reprogrammed energy metabolism is an attractive anticancer therapeutic strategy. We previously found a natural product, bouchardatine, modulated aerobic metabolism and inhibited proliferation in the colorectal cancer cell (CRC). Herein, we designed and synthesized a new series of bouchardatine derivatives to discover more potential modulators. We applied the dual-parametric high-content screening (HCS) to evaluate their AMP-activated protein kinase (AMPK) modulation and CRC proliferation inhibition effect simultaneously. And we found their antiproliferation activities were highly correlated to AMPK activation. Among them, 18a was identified with nanomole-level antiproliferation activities against several CRCs. Interestingly, the evaluation found that 18a selectively upregulated oxidative phosphorylation (OXPHOS) and inhibited proliferation by modulating energy metabolism. Additionally, this compound effectively inhibited the RKO xenograft growth along with AMPK activation. In conclusion, our study identified 18a as a promising candidate for CRC treatment and suggested a novel anti-CRC strategy by AMPK activating and OXPHOS upregulating.

Study on Medication Rules of Traditional Chinese Medicine in Treating Constipation through Data Mining and Network Pharmacology.

Background: To explore the rules of TCM medication in the treatment of constipation in network pharmacology. Methods: Collect and screen the clinical intervention literature on TCM for constipation from China's national knowledge infrastructure, Wanfang and VIP databases established a database of TCM for constipation, applied R software (3.3.1) to analyze the pattern of prescriptions for TCM for constipation, and summarized the core prescription. The effective active compounds and action targets in the core prescription were screened by Traditional Chinese Medicine Systems Pharmacology (TCMSP) and Traditional Chinese Medicine Integrated Databases (TCMID), constipation-related targets were derived from the DisGeNET and GeneCards databases, Protein-protein interaction network (PPI) was drawn by STRING database, and enrichment analysis was conducted by the Clusterprofiler package in R software (3.3.1). Finally, molecular docking was used to validate the binding ability of candidate compounds to potential targets. Results: Two hundred sixteen target prescriptions were screened through data mining, involving 226 herbs. Association rule analysis results suggested that the "Angelicae sinensis-Radix-dried rehmanniae-Cistanche deserticola-Atractylodes macrocephala-Astragali Radix" was a strong affinity for medicine. Network pharmacology analysis of the core prescription resulted in the screening of 115 candidate compounds, such as quercetin, kaempferol, mangostin, eugenol A, and beta-sitosterol; 131 potential targets, such as PTGS2, PTGS1, and CHRM3; and 160 signaling pathways, such as lipid and atherosclerosis, proteoglycans in cancer, hepatitis B, Kaposi's sarcoma-associated herpesvirus infection, and PI3K/AKT pathways. Molecular docking showed that PTGS1-formononetin, PTGS2-kaempferol, and CHRM3-kaempferol were all well bound and well matched. Conclusions: This study provides a new method and ideas for clinical applications of integrated Chinese and western medicine in treating constipation.

[The effects of vitamin E and dexamethasone on inflammation of acute lung injury and expression of myosin light chain kinase].

OBJECTIVE: To observe the effect of vitamin E (VitE) combined with dexamethasone (DXM) on inflammation of acute lung injury and expression of myosin light chain kinase. METHODS: Forty female Balb/c mice were randomly divided into 4 groups, a saline control group (1.5 ml/kg), a LPS group (1 mg/kg), a VitE and DXM group (VitE 50 mg/kg, DXM 1 mg/kg), and a VitE group (50 mg/kg). Lung tissue histopathological changes were observed. Immunohistochemistry assays (SABC) were used to determine the myosin light chain kinase (MLCK) immunoreactive cells in the lung tissues, and the MLCK mRNA and the MLCK protein was assayed by RT-PCR and by Western blot, respectively. Means were compared with analysis of variance and Student-Newman-Keuls were used to compare 2 means. RESULTS: Histological examination showed that extensive lung inflammation were seen in the LPS group, which manifested by accumulation of significant numbers of neutrophils, accompanied by marked pulmonary edema and hemorrhage. The inflammation and hemorrhage in the 2 treatment groups were significantly improved. Immunoreactive cells of MLCK numbers in BALF in the control group, the LPS group, the VitE and DXM group, and the VitE group was (1.1 +/- 0.4), (5.6 +/- 2.1), (4.0 +/- 1.0), (4.2 +/- 1.3) x 10(9)/L respectively. Compared with other groups, the difference of LPS group was significant (F = 14. 53, all P < 0.05). Immunoreactive cells of MLCK located airway epithelial and endothelia in the LPS group were more than which in the control group, decreased immunoreactive cells of MLCK in two treatment groups. Compared with LPS group, the difference of MLCK mRNA expression (A) of lung tissue in two treatment groups was no significant (F = 2.76, all P > 0.05). Compared with LPS group, the difference of A values of MLCK protein of lung tissue in two treatment groups was statistical significance (F = 12.06, all P < 0.01). CONCLUSIONS: Vitamin E combined with low dose of DXM could effectively inhibit inflammation and expression of MLCK protein in acute lung injury induced by LPS lung. It is suggested that, inhibition of MLCK activation leads to stabilize vascular barrier function and attenuation of pulmonary edema and inflammation, which also suggests a possible role of MLCK in the pathogenesis of acute lung injury.

[Protective effect of myosin light-chain kinase inhibitor on acute lung injury].

OBJECTIVE: To investigate the influence of inhibitor of myosin light-chain kinase (MLCK) on the human pulmonary arterial endothelial cell (HPAEC) challenged with lipopolysaccharide (LPS) and LPS induced of acute lung injury (ALI) in mice. METHODS: HPAECs were cultured in ECM medium and its passages 4-6 were used. After treatment with inhibitor of MLCK (ML-7) for 60 minutes, the HPAECs were incubated in LPS for another 60 minutes, and then cell viability was measured by the methyl thiazolyl tetrazolium (MTT) assay. Immunofluorescence microscope was used to detect phosphorylated-MLCK (p-MLCK) immunoreactive cells. Twenty female BALB/c mice were randomly divided into two groups. The mice of LPS group were exposed to LPS (1 microg/g) through nasal instillation, and the mice of ML-7 group were pretreated with ML-7 before intranasal instillation of LPS. Wet/dry weight (W/D) ratio of lung, bronchoalveolar lavage fluid (BALF) protein content, myeloperoxidase (MPO) activity and histopathological changes of lung tissue were observed. Immunohistochemistry assays were used to determine the status of MLCK and CD11b immunoreactive cells in lung tissue, and expression of MLCK mRNA in lung tissue was assessed by reverse transcription-polymerase chain reaction (RT-PCR). Expression of MLCK protein in lungs was assayed by Western blotting. RESULTS: Compared with LPS group, increased absorbance (A) value of HPAEC was found in ML-7 group (P<0.01). Immunoreactive cells of p-MLCK were more reduced in the ML-7 group (P<0.05), and W/D ratio of lung, MPO activity and BALF protein content of lung tissue were decreased in ML-7 group (P<0.05 or P<0.01). Histological examination showed that an extensive lung inflammation was seen in mice of LPS group, with an accumulation of a large number of neutrophils, marked pulmonary edema and hemorrhage, but the inflammation and parenchymal hemorrhage was significantly alleviated in ML-7 group. Both MLCK immunoreactive cells located in endothelium and CD11b in infiltrated inflammatory cells were decreased in ML-7 group compared with those in LPS group. Compared with LPS group, MLCK mRNA and protein expressions (A) in ML-7 group were significantly decreased (both P<0.05). CONCLUSION: ML-7, an MLCK inhibitor, enhances activity of HPAEC induced by LPS and reduces expression of p-MLCK. It also reduces the LPS-induced infiltration of neutrophils in lung tissues, pulmonary edema and expression of MLCK and CD11b protein and MLCK mRNA in lung tissues, demonstrating that inhibition of activation of MLCK, leading to an abatement of phosphorylation of myosin light chain or MLCK, resulting in stabilization of vascular barrier function. The results suggest that MLCK has a crucial role in the pathogenesis of ALI.

[Effect of hypoxia on expression of iNOS mRNA in cultured rat astrocytes].

AIM: To explore the effects of hypoxia on expression of inducible nitric oxide synthase (iNOS) mRNA in cultured rat astrocytes. METHODS: Cultured rat astrocytes were randomly divided into 4 groups: glutamate group (G), hypoxic group (H), hypoxia + glutamate group (H + G) and the control (C). Cells of control group were exposed to normoxic (95% air, 5% CO2) condition, and cells of G and H + G were incubated with 100 micromol/L L-glutamate, cells of H and H + G exposed to hypoxic conditions (5% CO2, 95% N2) at 37 degrees C. Each group had five timepoints which included 0 h, 3 h, 6 h, 12 h, 24 h, respectively. Expression of mRNAs of iNOS were detected with reverse transcription polymerase chain reaction (RT-PCR). RESULTS: Expression of iNOS mRNA was not detectable in G and C, while it increased dramatically and continuously from 6 h to 24 h in H and G + H. Expression of iNOS mRNA was significantly higher in H than both in G and C at 6 h, 12 h and 24 h, and expression of iNOS mRNA was the highest of all groups in G + H. CONCLUSION: Hypoxia upregulates the expression of iNOS mRNA in cultured astrocytes. Glutamate does not induce the expression of iNOS mRNA but enhance the effect of hypoxia, which is maybe one of the adaptive mechanisms of hypoxia-induced cerebral dilation.