Xiaoou Shan

Synthesis and Anti-inflammatory Evaluation of Novel Benzimidazole and Imidazopyridine Derivatives.

Sepsis, an acute inflammatory disease, remains the most common cause of death in intensive care units. A series of benzimidazole and imidazopyridine derivatives were synthesized and screened for anti-inflammatory activities, and the imidazopyridine series showed excellent inhibition of the expression of inflammatory cytokines in LPS-stimulated macrophages. Compounds X10, X12, X13, X14, and X15 inhibited TNF-α and IL-6 release in a dose-dependent manner, and X12 showed no cytotoxicity in hepatic cells. Furthermore, X12 exhibited a significant protection against LPS-induced septic death in mouse models. Together, these data present a series of new imidazopyridines with potential therapeutic effects in acute inflammatory diseases.

MD-2 as the target of a novel small molecule, L6H21, in the attenuation of LPS-induced inflammatory response and sepsis.

Myeloid differentiation 2 (MD‐2) recognizes LPS, which is required for TLR4 activation, and represents an attractive therapeutic target for severe inflammatory disorders. We previously found that a chalcone derivative, L6H21, could inhibit LPS‐induced overexpression of TNF‐α and IL‐6 in macrophages. Here, we performed a series of biochemical experiments to investigate whether L6H21 specifically targets MD‐2 and inhibits the interaction and signalling transduction of LPS‐TLR4/MD‐2.

Curcumin Analog L48H37 Prevents Lipopolysaccharide-Induced TLR4 Signaling Pathway Activation and Sepsis via Targeting MD2

Endotoxin-induced acute inflammatory diseases such as sepsis, mediated by excessive production of various proinflammatory cytokines, remain the leading cause of mortality in critically ill patients. Lipopolysaccharide (LPS), the characteristic endotoxin found in the outer membrane of Gram-negative bacteria, can induce the innate immunity system and through the myeloid differentiation protein 2 (MD2) and Toll-like receptor 4 (TLR4) complex, increase the production of inflammatory mediators. Our previous studies have found that a curcumin analog, L48H37 [1-ethyl-3,5-bis(3,4,5-trimethoxybenzylidene)piperidin-4-one], was able to inhibit LPS-induced inflammation, particularly tumor necrosis factor α and interleukin 6 production and gene expression in mouse macrophages. In this study, a series of biochemical experiments demonstrate L48H37 specifically targets MD2 and inhibits the interaction and signaling transduction of LPS-TLR4/MD2. L48H37 binds to the hydrophobic region of MD2 pocket and forms hydrogen bond interactions with Arg90 and Tyr102. Subsequently, L48H37 was shown to suppress LPS-induced mitogen-activated protein kinase phosphorylation and nuclear factor κB activation in macrophages; it also dose dependently inhibits the cytokine expression in macrophages and human peripheral blood mononuclear cells stimulated by LPS. In LPS-induced septic mice, both pretreatment and treatment with L48H37 significantly improved survival and protected lung injury. Taken together, this work identified a new MD2 specific inhibitor, L48H37, as a potential candidate in the treatment of sepsis.

Synthesis and evaluation of a series of novel asymmetrical curcumin analogs for the treatment of inflammation.

Curcumin has been reported to possess multiple bioactivities, such as antioxidant, anticancer, and anti-inflammatory properties, however the clinical application of curcumin has been significantly limited by its instability and poor metabolism. Modification of curcumin has led to discovery and development of lots of novel therapeutic candidates. In recent years acute and chronic inflammation has been the focus of numerous studies in various diseases. Here, we synthesized a series of asymmetrical curcumin analogs with high in vitro chemical stability, and their anti-inflammatory activity was evaluated in LPS-stimulated macrophages. According to the bio-screening results and QSAR analysis, these analogs exhibited potent activities against LPS-induced TNF-α and IL-6 release. Among the analogs of the potent anti-inflammatory activity, compounds 3b8 and 3b9 exhibited significant protection and possess enhanced anti-inflammatory activity thereby attenuated the LPS-induced septic death in mice.

Design, Synthesis, and Biological Evaluation of Novel Quinazoline Derivatives as Anti-inflammatory Agents against Lipopolysaccharide-induced Acute Lung Injury in Rats.

Quinazoline has been reported to exhibit multiple bioactivities. The aim of this study was to discover new quinazoline derivatives with preventive effect on lipopolysaccharide‐induced acute lung injury via anti‐inflammatory actions. Thirty‐three 4‐amino quinazolin derivatives were synthesized and screened for anti‐inflammatory activities in lipopolysaccharide‐induced macrophages. The most potent four compounds, 6h, 6m, 6p, and 6q, were shown dose‐dependent inhibition against lipopolysaccharide‐induced TNF‐α and IL‐6 release. Then, the preliminary structure–activity relationship and quantitative structure–activity relationship analyses were conducted. To further determine the effects of quinazolines on acute lung injury treatment, lipopolysaccharide‐induced acute lung injury model was employed. Male Sprague Dawley rats were pretreated with 6m or 6q before instillation of lipopolysaccharide. The results showed that 6m and 6q, especially 6q, obviously alleviated lung histopathological changes, inflammatory cells infiltration, and cytokines mRNA expression initiated by lipopolysaccharide. Taken together, this work suggests that 6m and 6q suppressed the lipopolysaccharide‐induced acute lung injury through inhibition of the inflammatory response in vivo and in vitro, indicating that quinazolines might serve as potential agents for the treatment of acute lung injury and deserve the continuing drug development and research.

Discovery of new MD2 inhibitor from chalcone derivatives with anti-inflammatory effects in LPS-induced acute lung injury

Acute lung injury (ALI) is a life-threatening acute inflammatory disease with limited options available for therapy. Myeloid differentiation protein 2, a co-receptor of TLR4, is absolutely required for TLR4 sense LPS, and represents an attractive target for treating severe inflammatory diseases. In this study, we designed and synthesized 31 chalcone derivatives that contain the moiety of (E)-4-phenylbut-3-en-2-one, which we consider the core structure of current MD2 inhibitors. We first evaluated the anti-inflammatory activities of these compounds in MPMs. For the most active compound 20, we confirmed that it is a specific MD2 inhibitor through a series of biochemical experiments and elucidated that it binds to the hydrophobic pocket of MD2 via hydrogen bonds with Arg90 and Tyr102 residues. Compound 20 also blocked the LPS-induced activation of TLR4/MD2 -downstream pro-inflammatory MAPKs/NF-κB signaling pathways. In a rat model with ALI induced by intracheal LPS instillation, administration with compound 20 exhibited significant protective effect against ALI, accompanied by the inhibition of TLR4/MD2 complex formation in lung tissues. Taken together, the results of this study suggest the specific MD2 inhibitor from chalcone derivatives we identified is a potential candidate for treating acute inflammatory diseases.

Angiotensin II induces kidney inflammatory injury and fibrosis through binding to myeloid differentiation protein-2 (MD2)

Growing evidence indicates that angiotensin II (Ang II), a potent biologically active product of RAS, is a key regulator of renal inflammation and fibrosis. In this study, we tested the hypothesis that Ang II induces renal inflammatory injury and fibrosis through interaction with myeloid differentiation protein-2 (MD2), the accessory protein of toll-like receptor 4 (TLR4) of the immune system. Results indicated that in MD2−/− mice, the Ang II-induced renal fibrosis, inflammation and kidney dysfunction were significantly reduced compared to control Ang II-infused wild-type mice. Similarly, in the presence of small molecule MD2 specific inhibitor L6H21 or siRNA-MD2, the Ang II-induced increases of pro-fibrotic and pro-inflammatory molecules were prevented in tubular NRK-52E cells. MD2 blockade also inhibited activation of NF-κB and ERK. Moreover, MD2 blockade prevented the Ang II-stimulated formation of the MD2/TLR4/MyD88 signaling complex, as well as the increased surface binding of Ang II in NRK-52E cells. In addition, Ang II directly bound recombinant MD2 protein, rather than TLR4 protein. We conclude that MD2 is a significant contributor in the Ang II-induced kidney inflammatory injury in chronic renal diseases. Furthermore, MD2 inhibition could be a new and important therapeutic strategy for preventing progression of chronic renal diseases.

Design, synthesis and biological activity of novel asymmetric C66 analogs as anti-inflammatory agents for the treatment of acute lung injury

Acute lung injury (ALI) is a leading cause of morbidity and mortality in critically-ill patients. Previously, we reported that a symmetric mono-carbonyl analog of curcumin, (C66), exhibits enhanced stability and was found to have efficacy and be involved in potential cytokines inhibition. In the present study, a series of novel asymmetric C66 analogs were designed and synthesized. A majority of them effectively inhibited the LPS-induced expression of TNF-α and IL-6. Significantly, compound 4b2 was found to effectively reduce LPS-induced pulmonary inflammation, as reflected by reductions in concentration of total protein, inflammatory cell count as well as the lung W/D ratio in bronchoalveolar lavage (BAL) fluid. Furthermore, in vivo administration of 4b2 resulted in remarkable improvement in histopathological changes of lung in rats.

Targeting myeloid differentiation protein 2 by the new chalcone L2H21 protects LPS-induced acute lung injury

Acute inflammatory diseases are the leading causes of mortality in intensive care units. Myeloid differentiation 2 (MD‐2) is required for recognizing lipopolysaccharide (LPS) by toll‐like receptor 4 (TLR4), and represents an attractive therapeutic target for LPS‐induced inflammatory diseases. In this study, we report a chalcone derivative, L2H21, as a new MD2 inhibitor, which could inhibit LPS‐induced inflammation both in vitro and in vivo. We identify that L2H21 as a direct inhibitor of MD‐2 by binding to Arg90 and Tyr102 residues in MD‐2 hydrophobic pocket using a series of biochemical experiments, including surface plasmon response, molecular docking and amino acid mutation. L2H21 dose dependently inhibited LPS‐induced inflammatory cytokine expression in primary macrophages. In mice with LPS intratracheal instillation, L2H21 significantly decreased LPS‐induced pulmonary oedema, pathological changes in lung tissue, protein concentration increase in bronchoalveolar lavage fluid, inflammatory cells infiltration and inflammatory gene expression, accompanied with the decrease in pulmonary TLR4/MD‐2 complex. Meanwhile, administration with L2H21 protects mice from LPS‐induced mortality at a degree of 100%. Taken together, this study identifies a new MD2 inhibitor L2H21 as a promising candidate for the treatment of acute lung injury (ALI) and sepsis, and validates that inhibition of MD‐2 is a potential therapeutic strategy for ALI.

Synthesis and biological evaluation of novel indole-2-one and 7-aza-2-oxindole derivatives as anti-inflammatory agents

Sepsis, a typically acute inflammatory disease, is the biggest cause of death in ICU (intensive care unit). Novel anti-inflammatory alternatives are still in urgent need. In this study, we designed and synthesized 30 indole-2-one and 7-aza-2-oxindole derivatives based on the skeleton of tenidap, and their anti-inflammatory activity was determined by evaluating the inhibitory potency against lipopolysaccharide (LPS)-stimulated tumor necrosis factor (TNF)-α and interleukin (IL)-6 release in RAW264.7 macrophages. Quantitative SAR (structure–activity relationship) analysis revealed that a high molecular polarizability and low lipid/water partition coefficient (ALogP) in indole-2-one are beneficial for anti-inflammatory activity. Moreover, compounds 7i and 8e inhibited the expression of TNF-α, IL-6, COX-2, PGES, and iNOS in LPS-stimulated macrophages, and 7i exhibited a significant protection from LPS-induced septic death in mouse models. These data present a series of new indole-2-one compounds with potential therapeutic effects in acute inflammatory diseases.