The present study was designed to synthesize 2-Cyano-3, 12-dioxooleana-1, 9(11)-en-28-oate-13β, 28-olide (1), a lactone derivative of oleanolic acid (OA) and evaluate its anti-inflammatory activity. Compound 1 significantly diminished nitric oxide (NO) production and down-regulated the mRNA expression of iNOS, COX-2, IL-6, IL-1β, and TNF-α in lipopolysaccharide (LPS)-stimulated RAW264.7 cells. Further in vivo studies in murine model of LPS-induced acute lung injury (ALI) showed that 1 possessed more potent protective effects than the well-known anti-inflammatory drug dexamethasone by inhibiting myeloperoxidase (MPO) activity, reducing total cells and neutrophils, and suppressing inflammatory cytokines expression, and thus ameliorating the histopathological conditions of the injured lung tissue. In conclusion, compound 1 could be developed as a promising anti-inflammatory agent for intervention of LPS-induced ALI.

The increasing understanding of the role of nitric oxide (NO) in cancer biology has generated significant progress in the use of NO donor-based therapy to fight cancer. These advances strongly suggest the potential adoption of NO donor-based therapy in clinical practice, and this has been supported by several clinical studies in the past decade. In this review, we first highlight several types of important NO donors, including recently developed NO donors bearing a dinitroazetidine skeleton, represented by RRx-001, with potential utility in cancer therapy. Special emphasis is then given to the combination of NO donor(s) with other therapies to achieve synergy and to the hybridization of NO donor(s) with an anticancer drug/agent/fragment to enhance the activity or specificity or to reduce toxicity. In addition, we briefly describe inducible NO synthase gene therapy and nanotechnology, which have recently entered the field of NO donor therapy.

Hyperglycemia-induced inflammation and fibrosis have important roles in the pathogenesis of diabetic nephropathy and cardiomyopathy. With inflammatory cytokines and signaling pathways as important mediators, targeting inflammation may be an effective approach to new avenue for treating diabetic complications. J17, a molecule with structural similarities to curcumin, exhibited good anti-inflammatory activities by inhibiting LPS-induced inflammatory response in macrophages. However, its ability to alleviate hyperglycemia-induced injury via its anti-inflammatory actions remained unclear. Thus, we reported that J17 exerts significant inhibitory effects on hyperglycemia-induced inflammation and fibrosis in NRK-52E cells, H9C2 cells and a streptozotocin-induced diabetic mouse model. We also found that the anti-inflammatory and anti-fibrosis activities of J17 are associated with the inhibition of the P38 and AKT signal pathway, respectively. In vivo oral administration of J17 suppressed hyperglycemia-induced inflammation, hypertrophy and fibrosis, thereby reducing key markers for renal and cardiac dysfunction and improving in fibrosis and pathological changes in both renal and cardiac tissues of diabetic mice. The results of this study indicated that J17 can be potentially used as a cardio- and reno-protective agent and that targeting the P38 and AKT pathways may be an effective therapeutic strategy for diabetic complications.

Indoleamine 2,3-dioxygenase 1 (IDO1) is frequently hijacked by tumors to escape the host immune response, and the enzyme is now firmly established as an attractive target for cancer immunotherapy. To identify novel IDO1 inhibitors suitable for drug development, a scaffold-hopping strategy combined with the average electrostatic potentials calculation was ultilized to design novel benzoxazolinone derivatives. Among these, compounds 7e, 7f and 9c exhibited the inhibitory potency in the low micromolar range and displayed negligible level of cytotoxicity against HeLa cells. Treatment with these three compounds promoted the proliferation of T lymphocyte and led to the dramatic decrease of regulatory T cells in the B16F1 cells and naïve T cells co-culture system. Subsequent spectroscopic experiments suggested that these benzoxazolinones formed a coordinate bond with the heme iron to stabilize the complex. This study suggested that the benzoxazolinone was an interesting scaffold for discovering novel IDO1 inhibitors, and these compounds are attractive candidates for further development.

(R)- and (S)-1 were as potent as racemate 1 in antiplatelet aggregation, antioxidation, reduction of infarct size and brain-water content, as well as neurological deficit.

The elevation of oxidative stress preferentially in cancer cells by inhibiting thioredoxin reductase (TrxR) and/or enhancing reactive oxygen species (ROS) production has emerged as an effective strategy for selectively targeting cancer cells. In this study, we designed and synthesized 21 ligustrazine-curcumin hybrids (10a-u). Biological evaluation indicated that the most active compound 10d significantly inhibited the proliferation of drug-sensitive (A549, SPC-A-1, LTEP-G-2) and drug-resistant (A549/DDP) lung cancer cells but had little effect on nontumor lung epithelial-like cells (HBE). Furthermore, 10d suppressed the TrxR/Trx system and promoted intracellular ROS accumulation and cancer cell apoptosis. Additionally, 10d inhibited the NF-κB, AKT, and ERK signaling, P-gp-mediated efflux of rhodamine 123, P-gp ATPase activity, and P-gp expression in A549/DDP cells. Finally, 10d repressed the growth of implanted human drug-resistant lung cancer in mice. Together, 10d acts a novel TrxR inhibitor and may be a promising candidate for intervention of lung cancer.

Aim: Compound 10b is a hybrid molecule of edaravone and a ring-opening derivative of 3- n -butylphthalide (NBP). The aim of this study was to examine the effects of compound 10b on brain damage in rats after focal cerebral ischemia. Methods: SD rats were subjected to 2-h-middle cerebral artery occlusion (MCAO). At the onset of reperfusion, the rats were orally treated with NBP (60 mg/kg), edaravone (3 mg/kg), NBP (60 mg/kg)+edaravone (3 mg/kg), or compound 10b (70, 140 mg/kg). The infarct volume, motor behavior deficits, brain water content, histopathological alterations, and activity of GSH, SOD, and MDA were analyzed 24 h after reperfusion. The levels of relevant proteins in the ipsilateral striatum were examined using immunoblotting. Results: Administration of compound 10b (70 or 140 mg/kg) significantly reduced the infarct volume and neurological deficits in MCAO rats. The neuroprotective effects of compound 10b were more pronounced compared to NBP, edaravone or NBP+edaravone. Furthermore, compound 10b significantly upregulated the protein levels of the cytoprotective molecules Bcl-2, HO-1, Nrf2, Trx, P-NF-κB p65, and IκB-α, while decreasing the expression of Bax, caspase 3, caspase 9, Txnip, NF-κB p65, and P-IκB-α. Conclusion: Oral administration of compound 10b effectively attenuates rat cerebral ischemia injury.

The NEDD8-activating enzyme (NAE) is an emerging target for cancer therapy, which regulates the degradation and turnover of a variety of cancer-related proteins by activating the cullin-RING E3 ubiquitin ligases. Among a limited number of known NAE inhibitors, the covalent inhibitors have demonstrated the most potent efficacy through their covalently linked adducts with NEDD8. Inspired by this unique mechanism, in this study, a novel combined strategy of virtual screening (VS) was adopted with the aim to identify diverse covalent inhibitors of NAE. To be specific, a docking-enabled pharmacophore model was first built from the possible active conformations of chosen covalent inhibitors. Meanwhile, a dynamic structure-based phamacophore was also established based on the snapshots derived from molecular dynamic simulation. Subsequent screening of a focused ZINC database using these pharmacophore models combined with covalent docking discovered three novel active compounds. Among them, compound LZ3 exhibited the most potent NAE inhibitory activity with an IC50 value of 1.06 ± 0.18 μM. Furthermore, a cell-based washout experiment proved the proposed covalent binding mechanism for compound LZ3, which confirmed the successful application of our combined VS strategy, indicating it may provide a viable solution to systematically discover novel covalent ligands.

Hybrid 5f significantly inhibited both Ras-related signaling and phosphorylated NF-κB, which may synergistically contribute to its apoptosis induction and tumor growth inhibition in vitro and in vivo.

Fifteen novel derivatives of glycyrrhetinic acid (GA) were synthesized and evaluated for anti-inflammatory activities. It was found that the introduction of 1-en-3-one and 9(11),12-diene and 2,20-dinitrile functionalities into the scaffold of GA led to the discovery of potent compound 19 for inhibition of LPS-induced NO production. Furthermore, 19 effectively inhibited the protein and mRNA expression of inducible NO synthase (iNOS) and the mRNA expression of TNF-α, IL-6, and IL-1β in LPS-stimulated RAW 264.7 macrophages. Mechanistically, 19 exerted inhibitory effects on the activation of the three main MAPKs and phosphorylation and degradation of IκB-α, as well as the ratio of nuclear/cytosolic content of p65. Importantly, 19 significantly decreased the mortality rate in the mouse model of LPS-induced sepsis shock. It is noteworthy that inhibitory effect of 19 on NO production was not blocked by the glucocorticoid receptor antagonist mifepristone, indicating that it does not act through the glucocorticoid receptor.

A series of hybrids (12a-k) from (phenylsulfonyl)furoxan and anilinopyrimidine were synthesized and biologically evaluated as epidermal growth factor receptor (EGFR) inhibitors for intervention of non-small-cell lung cancer (NSCLC). Compound 12k exhibited strong and selective EGFR L858R/T790M inhibitory activity (IC50 = 0.047 μM) and displayed antiproliferative effects on EGFR mutation NSCLC cell lines HCC827 (del E746_A750) and H1975 (L858R/T790M) with IC50 values of 0.007 and 0.029 μM, respectively. Additionally, 12k released high levels of NO in H1975 cells but not in normal human cells, and its activity was diminished by pretreatment with a NO scavenger. Furthermore, 12k induced apoptosis of H1975 and HCC827 cells more strongly than WZ4002 (1), inhibited EGFR downstream signaling in H1975 cells, and suppressed the nuclear factor-κB activation in H1975 cells, while 1 had no significant effects under the same conditions. Finally, 12k substantially inhibited tumor growth in an H1975 xenograft mouse model. Overall, 12k might be a promising candidate for the treatment of NSCLC.

To search for potent anti-Alzheimer's disease (AD) agents with multifunctional effects, 12 NO-donating tacrine-flurbiprofen hybrid compounds (2a-l) were synthesized and biologically evaluated. It was found that all the new target compounds showed selective butyrylcholinesterase (BuChE) inhibitory activity in vitro comparable or higher than tacrine and the tacrine-flurbiprofen hybrid compounds 1a-c, and released moderate amount of NO in vitro. The kinetic study suggests that one of the most active and highest BuChE selective compounds 2d may not only compete with the substrate for the same catalytic active site (CAS) but also interact with a second binding site. Furthermore, 2d and 2l exhibited significant vascular relaxation effect, which is beneficial for the treatment of AD. All the results suggest that 2d and 2l might be promising lead compounds for further research.

A series of novel nitric oxide releasing derivatives of 6-amino-3-n-butylphthalide were designed, synthesized and evaluated as potential antiplatelet agents. Compound 10b significantly inhibited the adenosine diphosphate (ADP)-induced platelet aggregation in vitro, superior to 6-amino-3-n-butylphthalide, 3-n-butylphthalide (NBP) and ticlopidine. Meanwhile 10b released moderate levels of NO, which could be beneficial for improving cardiovascular and cerebral circulation. Furthermore, 10b had an enhanced aqueous solubility relative to NBP. These findings may provide new insights into the development of novel antiplatelet agents for the treatment of thrombosis-related ischemic stroke.

To search for potent nitric oxide (NO) donating epidermal growth factor receptor (EGFR) inhibitors, a series of phenylsulfonylfuroxan-based anilinopyrimidines 10a-h were synthesized and biologically evaluated. Compounds 10f-h exhibited potent inhibitory activity against EGFR L858R/T790M and were as potent as WZ4002 in inhibition of H1975 cells harboring EGFR L858R/T790M. Additionally, 10h produced high levels of NO in H1975 cells but not in normal human cells, and its antiproliferative activity was diminished by hemoglobin, an NO scavenger. Furthermore, 10h inhibited EGFR activation and downstream signaling in H1975 cells. These results suggest that the strong antiproliferative activity of 10h could be attributed to the synergic effects of high levels of NO production and inhibition of EGFR and downstream signaling in the cancer cells.

A series of O(2)-glycosylated diazeniumdiolate-based derivatives of oleanolic acid (4-19) were synthesized and their anti-human hepatocellular carcinoma (HCC) activities were evaluated. Compound 6 selectively inhibited HCC, but not non-tumor liver cell proliferation. This inhibition was attributed to high levels of nitric oxide (NO) released in HCC cells. Importantly, 6 exhibited low acute toxicity (LD(50) = 173.3 mg kg(-1)) and potent inhibition of HCC tumor growth in mice (3 mg kg(-1) iv). Furthermore, 6 induced HCC cell apoptosis, which was accompanied by lower mitochondrial membrane potentials and Bcl2 expression, but with higher cytochrome C release, Bax, caspase 3 and 9 expression activities in HCC cells. Collectively, 6 may be a promising candidate drug for the intervention of HCC.

Nitric oxide (NO) and its reduced form nitroxyl (HNO), effective vasodilation agents that can inhibit platelet aggregation and adhesion, could suppress adverse cardiovascular effects associated with the use of selective COX-2 inhibitors. In this regard, a sulfohydroxamic acid (SO(2)NHOH) substituent, that can act as a dual NO/HNO donor moiety, was inserted at the para-position of the C2 phenyl ring of acyclic 2-alkyl-1,1,2-triaryl olefins previously shown to be potent and highly selective COX-2 inhibitors. Although this new group of 1,1-diaryl-2-(4-hydroxyaminosulfonylphenyl)alk-1-enes exhibited weak inhibition of the constitutive cyclooxygenase-1 (COX-1) and inducible COX-2 isozymes, in vivo studies showed anti-inflammatory potencies that were generally intermediate between that of the reference drugs aspirin and ibuprofen. All compounds released NO (5.6-13.5% range) upon incubation with phosphate buffer which was increased further (8.3-25.6% range) in the presence of the oxidant K(3)(FeCN(6)).The low release of HNO in MeOH-buffer (< 2% at 24 h incubation) was much higher at alkaline pH (11-37% range). The concept of designing better anti-inflammatory drugs possessing either an effective HNO, or dual NO/HNO, donor moiety that are devoid of adverse ulcerogenic and/or cardiovascular side effects warrants further investigation.

A series of nitric oxide (NO)-releasing glycosyl derivatives () of oleanolic acid were synthesized to improve the aqueous solubility and cytotoxicity of the parent compound . Derivative exhibited better solubility and strong cytotoxicity against human hepatocellular carcinoma (HCC) in vitro. Furthermore, displayed low acute toxicity to mice and significantly inhibited the growth of HCC tumors in vivo, indicating that may be a promising candidate for the treatment of human HCC.