Synthesis and Biological Evaluation of Novel Urea- and Guanidine-Based Derivatives for the Treatment of Obesity-Related Hepatic Steatosis

Leptin, the product of the obese gene, is an adipocyte-secreted protein hormone playing a key role in the progression of obesity and hepatic steatosis. In this study, 28 novel (thio)urea and guanidine-based analogues have been synthesized and N-(1-(4-(3-(2-chloroethyl)ureido)benzyl)piperidin-4-yl)-3-(trifluoromethyl) benzamide (7i) was found to be a potent regulator of leptin expression in 3T3-L1 adipocytes. Treatment with 7i at a dose of 50 mg/kg/day for 35 days reduced the body weight and liver weight of diet-induced obesity mice by 13.5% and 18.4%, respectively, while also improving the serum levels of triglyceride, total cholesterol, leptin, adiponectin, LDL-c, HDL-c. Hematoxylin-eosin (H&E) and Oil Red O staining also confirmed that 7i ameliorated fat deposition in liver tissue and restricted the size of adipocytes in obesity-related fatty liver disease

Studies on the anti-psoriasis effects and its mechanism of a dual JAK2/FLT3 inhibitor flonoltinib maleate

Psoriasis is a chronic, inflammatory autoimmune disease mediated by T cells, and characterized with abnormal proliferation and differentiation of keratinocytes, and inflammatory infiltration. The Janus kinase-signal transducer and activator of transcription (JAK-STAT) pathway has been identified to play essential roles in mediating various of biological processes, and is closely related to autoimmune diseases. Dendritic cells (DCs) are important antigen presenting cells and play an important regulatory role in T cells. The proliferation, differentiation and function of DCs are regulated by JAK and FMS-like tyrosine kinase 3 (FLT3) signal pathways. Flonoltinib maleate (FM), a high selectivity dual JAK2/FLT3 inhibitor with IC50 values of 0.8 nM and 15 nM for JAK2 and FLT3, respectively, was developed by our laboratory. Moreover, FM was a potent JAK2 inhibitor with 863-fold and 696-fold selectivity over JAK1 and JAK3, respectively. In this study, the anti-psoriasis activity of FM was evaluated both in vitro and in vivo. FM effectively inhibited the proliferation of HaCaT, the inflammatory keratinocyte induced by M5 and markedly suppressed the generation and differentiation of DCs from bone marrow (BM), and inhibited the expression of FLT3 in DCs in vitro. FM effectively inhibited the ear thickening and improved the pathological changes of the ear in interleukin (IL)-23-induced psoriasis-like acanthosis mouse model. Further in keratin 14-vascular endothelial growth factor (K14-VEGF) transgenic homozygous mice model, FM could obviously improve the psoriatic symptom and pathological changes, significantly inhibit the generations of Th1 and Th17 cells in the spleen, and the accumulations of DCs in the ears. FM could also significantly reduce the expression of various inflammatory factors both in C57BL/6 and K14-VEGF mice ears, and the serum of K14-VEGF mice. Mechanism revealed that FM effectively suppressed the phosphorylation of JAK2, STAT3 and STAT5 in inflammatory keratinocytes and the mice ears of C57BL/6 and K14-VEGF, as well as the phosphorylation of FLT3 in K14-VEGF mice ears. In conclusion, FM plays an excellent anti-psoriasis activity, including inhibiting keratinocyte proliferation and regulating inflammatory response through inhibiting JAK2 and FLT3 signaling pathway

Pharmacological Effects of the Water Fraction of Key Components in the Traditional Chinese Prescription Mai Tong Fang on 3T3-L1 Adipocytes and ob/ob Diabetic Mice

Mai Tong Fang (MTF), a Chinese herbal combination, has been used for the treatment of diabetic nephropathy in traditional medical clinics in China. However, the anti-adipogenic and anti-hyperglycemic effects of MTF have not been fully elucidated, so this study explored these pharmacological activities in 3T3-L1 adipocytes and ob/ob mice, respectively, of the water fraction of milkvetch root, salviae miltiorrhizae and mulberry as key components of MTF. MTF was found to inhibit adipogenesis and triglyceride accumulation in 3T3-L1 adipocytes. Oral administration of MTF in ob/ob mice for 8 weeks, exhibited positive controls on blood glucose and body weight, and further improved glucose tolerance according to an oral glucose tolerance test. Importantly, MTF extract alleviated fat deposition and ballooning degeneration in liver tissue and blocked the increase of adipocyte size in adipose tissue from treated ob/ob mice. These results indicated that the extract of key components in the traditional Chinese prescription MTF continue a potent anti-adipogenic and glucose-lowering agent

SKLB023 Blocks Joint Inflammation and Cartilage Destruction in Arthritis Models via Suppression of Nuclear Factor-Kappa B Activation in Macrophage

<div><p>Rheumatoid arthritis (RA) is the most common arthritis and is mainly characterized by symmetric polyarticular joint disorders. Our previous study demonstrated a novel small molecule compound (Z)-N-(3-Chlorophenyl)-2-(4-((2,4-dioxothiazolidin-5-ylidene) methyl) phenoxy) acet-amide (SKLB023) showed potently anti-arthritic effects in a rat arthritis model, however, the underlying mechanisms for this are largely unknown. Both NF-<i>κ</i>B and macrophages were reported to play important roles in the pathologic processes of RA. The purposes of this study were to indicate whether NF-<i>κ</i>B and macrophages contributed to anti-arthritic effects of SKLB023 in two experimental arthritis models. Our results showed that SKLB023 could significantly improve joint inflammation and cartilage destruction both in adjuvant induced arthritis (AIA) and collagen-induced arthritis (CIA) models. We further found that the binding activation of NF-<i>κ</i>B to DNA in joint tissues and RAW264.7 macrophages were suppressed by SKLB023. SKLB023 also inhibited the NF-<i>κ</i>B activity in peritoneal macrophages by luciferase assay. Furthermore, the number of macrophages in synovial tissues was decreased after the treatment of different doses of SKLB023. The levels of TNF-α, IL-1β, and IL-6 in plasma, and the levels of TNF-α, NO, and IL-1β in peritoneal macrophages were down-regulated by SKLB023. Finally, SKLB023 attenuated the expression of iNOS and COX-2 in vivo and suppressed the phosphorylations of components of the mitogen-activated protein kinases (MAPKs). These observations identify a novel function for SKLB023 as an inhibitor of NF-<i>κ</i>B in macrophages of RA, highlighting that SKLB023 was a potential therapeutic strategy for RA.</p> </div

SNX17 Recruits USP9X to Antagonize MIB1-Mediated Ubiquitination and Degradation of PCM1 during Serum-Starvation-Induced Ciliogenesis

Centriolar satellites are non-membrane cytoplasmic granules that deliver proteins to centrosome during centrosome biogenesis and ciliogenesis. Centriolar satellites are highly dynamic during cell cycle or ciliogenesis and how they are regulated remains largely unknown. We report here that sorting nexin 17 (SNX17) regulates the homeostasis of a subset of centriolar satellite proteins including PCM1, CEP131, and OFD1 during serum-starvation-induced ciliogenesis. Mechanistically, SNX17 recruits the deubiquitinating enzyme USP9X to antagonize the mindbomb 1 (MIB1)-induced ubiquitination and degradation of PCM1. SNX17 deficiency leads to enhanced degradation of USP9X as well as PCM1 and disrupts ciliogenesis upon serum starvation. On the other hand, SNX17 is dispensable for the homeostasis of PCM1 and USP9X in serum-containing media. These findings reveal a SNX17/USP9X mediated pathway essential for the homeostasis of centriolar satellites under serum starvation, and provide insight into the mechanism of USP9X in ciliogenesis, which may lead to a better understating of USP9X-deficiency-related human diseases such as X-linked mental retardation and neurodegenerative diseases

SKLB023 improved histological changes and arthritis scores in Lewis rats.

<p>(A) Representative photographs of tarsotibial joint swelling of the hind paws in rats treated with or without SKLB023 in AIA (n = 8). (B) Representative histopathologies of the ankle joints stained with hematoxylin and eosin (H&E) in AIA. (C) Histological examinations of a total of 8 rats for each pair were scored from 0 to 4 as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0056349#s2" target="_blank">Materials and Methods</a>. (D) Arthritis scores during the progression of AIA. Error bars represented SEM, *P<0.05, **P<0.01, and ***P<0.001 indicated significant differences from the control group.</p

SKLB023 improved histological changes and arthritis scores in DBA/1J mice.

<p>(A) Representative photographs of tarsotibial joint swelling of the hind paws in mice treated with or without SKLB023 (n = 8). (B) Representative histopathologies of the ankle joints stained with hematoxylin and eosin (H&E) in CIA. (C) Histological examinations of a total of 8 mice for each pair were scored from 0 to 4 as described in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0056349#s2" target="_blank">Methods</a>. (D) Arthritis scores during the progression of CIA. Error bars represented SEM, *P<0.05, **P<0.01, and ***P<0.001 indicated significant differences from the control group.</p

SKLB023 inhibited the phosphorylation of MAPKs.

<p>(A) Chemical structure of SKLB023. (B) iNOS and COX-2 expression in the ankle joints of AIA rats untreated or treated with various doses of SKLB023. (C) RAW264.7 cells were stimulated with LPS (1 µg/ml) and incubated with SKLB023 (20 µM) for the indicated time. Cell lysates were prepared and blotted with total or phosphospecific antibodies to ERK1/2 (Thr202/Tyr204), p38 MAPK (Thr180/Tyr182), and JNK (Thr183/Tyr185).</p

SKLB023 improved cartilage destruction and macrophages infiltration in AIA model.

<p>(A) Representative histopathologies of the ankle joints stained with Safranin-O in AIA. (B) SKLB023 inhibiting the infiltration of CD68 positive cells in synovial tissue of AIA model. Synovial tissue sections were analyzed for the expression of CD68 using immunohistochemical method as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0056349#s2" target="_blank">Materials and Methods</a>. (C) Histologic analysis of cartilage in AIA rats after SKLB023 therapy. Sections were scored in a blinded manner on a 4-point scale as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0056349#s2" target="_blank">Materials and Methods</a>. Significantly less loss of safranin O staining was observed in the animals treated with SKLB023 than in controls, indicating inhibition of cartilage destruction. (D) CD68 expression was measured by immunohistochemistry in rats with AIA. It was scored in a blinded manner on a scale of 0 to 4 as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0056349#s2" target="_blank">Materials and Methods</a>. A statistically significant reduction of CD68 expression was observed in the rats treated with different doses of SKLB023. Error bars represented SEM, **P<0.01, and ***P<0.001 indicated significant differences from the control group.</p