Ring-Contracted Artemisinin Derivatives as Novel CDK 4/6 Inhibitors: Synthesis and Anti-Breast Cancer Evaluation.

The endoperoxide group of artemisinins is universally accepted an essential group for their anti-cancer effects. In this study, a series of D-ring-contracted artemisinin derivatives were constructed by combining ring-contracted artemisinin core with fragments of functional heterocyclic molecules or classical CDK4/6 inhibitors to identify more efficacious breast cancer treatment agents. Twenty-six novel hybridized molecules were synthesized and characterized by HRMS, IR, 1H-NMR and 13C NMR. In antiproliferative activities and kinase inhibitory effects assays, we found that the antiproliferative effects of B01 were close to those of the positive control Palbociclib, with GI50 values of 4.87±0.23 µM and 9.97±1.44 µM towards T47D cells and MDA-MB-436 cells respectively. In addition, the results showed that B01 was the most potent compound against CDK6/cyclin D3 kinase, with an IC50 value of 0.135±0.041 µM, and its activity was approximately 1/3 of the positive control Palbociclib.

Synthesis, Structure Revision, and Anti-inflammatory Activity Investigation of Putative Blumeatin.

Blumeatin, reported herein, bearing two hydroxyl groups at C3' and C5' of ring B, is isolated from the traditional Chinese medicine Blumea balsamifera. But the isolation procedure of blumeatin from plants has limitations of prolonged duration and high cost. A procedure featuring Lewis acid-catalyzed ring closure and chiral resolution via Schiff base intermediates is provided here to prepare optically pure blumeatin and its R-isomer efficiently. Furthermore, the structure revision of putative blumeatin based on a logically synthetic procedure and NMR spectroscopic analysis was conducted. The 1D and 2D NMR data analysis unambiguously confirmed our proposal that the reported blumeatin structure has been misassigned as it corresponds to sterubin, which contains two hydroxyl groups at C3' and C4' of ring B. Finally, the results of the ear-swelling test exhibited that synthetic (±)-blumeatin and (±)-sterubin had moderate anti-inflammatory activity which was less than that of (-)-sterubin.

Three-Dimensional Hounsfield Units Measurement of Pedicle Screw Trajectory for Predicating Screw Loosening in Lumbar Fusion Surgery.

Purpose: Dual-energy X-ray absorptiometry (DXA) is commonly used for evaluation of bone mineral density before spinal surgery, but frequently leads to overestimation in degenerative spinal diseases due to osteoproliferation factors. We introduce a novel method to compare the predictive ability of Hounsfield Units (HU) and DXA methods to predict screw loosening after lumbar interbody fusion surgery in degenerative spinal diseases by measuring HU of pedicle screw trajectory on computed tomography (CT) images preoperatively. Patients and Methods: This retrospective study was conducted on patients who underwent posterior lumbar fusion surgery for degenerative diseases. CT HUs measurement was performed using medical imaging software, including the cancellous region on cross-sections of the vertebral body and three-dimensional pedicle screw trajectory. Receiver operating characteristic (ROC) curve analyses were performed for the risk of pedicle screw loosening in association with the Hounsfield scale and preoperative BMD, and the area under the curve (AUC) and the cutoff values were calculated. Results: A total of 90 patients were enrolled and were divided into loosening (n = 33, 36.7%) and non-loosening groups (n = 57, 63.3%). No significant differences in age, gender, length of fixation and preoperative BMD were found between both groups. The loosening group showed lower CT HU values in the vertebral body and screw trajectory than the non-loosening group. Screw trajectory HU (ST-HU) exhibited a higher AUC value than vertebral body HU (B-HU). The cutoff values of B-HU and ST-HU were 160 and 110 HUs, respectively. Conclusion: Three-dimensional pedicle screw trajectory HU values yields a stronger predictive value than vertebral body HU values and BMD and may provide more guidance for surgery. The risk of screw loosening is significantly increased at ST-HU <110 or B-HU <160 at L5 segment.

The first case of Escherichia fergusonii with biofilm in China and literature review.

BACKGROUND: Escherichia fergusonii is a rare opportunistic pathogen in humans and animals, especially with biofilm. METHODS: In one case, E. fergusonii with biofilm was detected in the bile, and silver staining was used to prove it had biofilm. The clinical characteristics and drug susceptibility of eight cases of E. fergusonii retrieved from the literature were also summarized. RESULTS: This is a case of E. fergusonii with biofilm, which has not been reported in China. The 8 cases retrieved from the literature did not specify whether they had biofilm, but we analyzed their clinical characteristics and drug susceptibility. All patients were treated with antimicrobial drugs. 8 cases showed sensitivity to piperacillin/tazobactam and imipenem in 6 cases (75%), but poor sensitivity to levofloxacin and ciprofloxacin. CONCLUSION: The silver staining method proved biofilm in this case, which is the first case of E. fergusonii with biofilm in China.

γ-secretase inhibitor disturbs the morphological development of differentiating neurons through affecting Notch/miR-342-5p.

During neurodevelopment, differentiation of neural stem/progenitor cells (NSPCs) into neurons are regulated by many factors including Notch signaling pathway. Herein, we report the effect of a Notch signaling blocker, i.e. γ -secretase inhibitor (GSI), on this differentiating process, especially on the morphological development. NSPCs were cultured and induced to differentiate with or without GSI. The neurite outgrowth was impeded by GSI application and the expression of a Notch signaling downstream effector miR-342-5p increased with the downregulated expression of Notch effectors Hes1 and Hes5. Upregulated expression of miR-342-5p in differentiating NSPCs could shorten the neurite length of progeny neurons, which was similar to the effect of GSI. To avoid the possible influence from astrocytes into neurons, we directly applied cultured neurons, on which GSI could shorten the processes and RBP-J knockdown could also reduce the neurite length. Similarly, transfection of miR-342-5p mimics or inhibitors into PC12 cells led to shorter or longer processes of cells compared with control ones. Furthermore, in differentiating NSPCs, GSI-induced shorter neurites could be partially rescued by miR-342-5p inhibitors, and STAT3 was one of the possible targets of miR-342-5p during this differentiating process as indicated by results of Western Blot test, luciferase reporter assay and GFP reporter assay. To further demonstrate the role of STAT3, it was introduced into GSI-treated neurons and the GSI-affected neurites could also be partially rescued. In conclusion, GSI could influence the morphological development of neurons and the possible mechanism involved Notch/miR-342-5p and STAT3. These results would be informative for future therapeutic research.

A pain killer without analgesic tolerance designed by co-targeting PSD-95-nNOS interaction and α2-containning GABAARs.

Overactivation of N-methyl-D-aspartate receptor (NMDAR) in the spinal cord dorsal horn (SDH) in the setting of injury represents a key mechanism of neuropathic pain. However, directly blocking NMDAR or its downstream signaling, interaction between postsynaptic density-95 (PSD-95) and neuronal nitric oxide synthase (nNOS), causes analgesic tolerance, mainly due to GABAergic disinhibition. The aim of this study is to explore the possibility of preventing analgesic tolerance through co-targeting NMDAR downstream signaling and γ-aminobutyric acid type A receptors (GABAARs). Methods: Mechanical/thermal hyperalgesia were quantified to assess analgesic effects. Miniature postsynaptic currents were tested by patch-clamp recording to evaluate synaptic transmission in the SDH. GABA-evoked currents were tested on HEK293 cells expressing different subtypes of recombinant GABAARs to assess the selectivity of (+)-borneol and ZL006-05. The expression of α2 and α3 subunits of GABAARs and BDNF, and nNOS-PSD-95 complex levels were analyzed by western blotting and coimmunoprecipitation respectively. Open field test, rotarod test and Morris water maze task were conducted to evaluate the side-effect of ZL006-05. Results: (+)-Borneol selectively potentiated α2- and α3-containing GABAARs and prevented the disinhibition of laminae I excitatory neurons in the SDH and analgesic tolerance caused by chronic use of ZL006, a nNOS-PSD-95 blocker. A dual-target compound ZL006-05 produced by linking ZL006 and (+)-borneol through an ester bond blocked nNOS-PSD-95 interaction and potentiated α2-containing GABAAR selectively. Chronic use of ZL006-05 did not produce analgesic tolerance and unwanted side effects. Conclusion: By targeting nNOS-PSD-95 interaction and α2-containing GABAAR simultaneously, chronic use of ZL006-05 can avoid analgesic tolerance and unwanted side effects. Therefore, we offer a novel candidate drug without analgesic tolerance for treating neuropathic pain.

Posterior Ligament-Bone Injury Classification and Severity Score: A Novel Approach to Predict the Failure of Anterior-only Surgery for Subaxial Cervical Facet Dislocations.

STUDY DESIGN: Clinical case series. OBJECTIVE: The aim of this study was to propose a novel posterior ligament-bone injury classification and severity (PLICS) score system that can be used to reflect the severity of subaxial cervical fracture dislocations (SCFDs) and predict the failure of anterior-only surgery; and to measure the intraobserver and interobserver reliability of this system. SUMMARY OF BACKGROUND DATA: The approach selection for SCFDs is controversial. Although the anterior approach is familiar for most surgeons, postoperative hardware failure and/or delayed cervical deformity is a nonnegligible complication. METHODS: Ten patients were randomly selected for intraobserver reliability evaluation on two separate occasions, one month apart. Another 30 patients were randomly selected, and the interobserver reliability was measured by comparing results of each case between each reviewer and averaging. To analyze the difference in the PLICS score, 354 patients fulfilled the follow-up were divided into stable and unstable groups according to whether radiologically stable was observed during follow-up. RESULTS: For the intraobserver reliability, the mean intraclass correlation coefficient for the 10 reviewers was 0.931. For the interobserver reliability, the mean interobserver correlation coefficient for the three elements was 0.863. Among 16 patients with PLICS score ≥7, two patients in the stable group manifested with severe injury of the posterior ligamentous complex (PLC); extremely unstable lateral mass fractures with or without severe injury of PLC were detected in the 14 patients of the unstable group. CONCLUSION: The proposed PLICS score system showed excellent intraobserver and interobserver reliability. When a PLICS score is >7 or 7 accompanied by extremely unstable lateral mass fractures, the risk of postoperative failure after an anterior-only reconstruction is high and supplemental posterior strengthening can be considered.Level of Evidence: 4.

When is the circumferential stabilization necessary for subaxial cervical fracture dislocations? The posterior ligament-bone injury classification and severity score: a novel treatment algorithm.

OBJECTIVE: To propose a novel classification and scoring system called the posterior ligament-bone injury classification and severity score (PLICS) that offers a quantitative score to guide the need for posterior stabilization in addition to anterior reconstruction for subaxial cervical fracture dislocations (SCFDs). METHODS: A total of 456 patients with SCFDs were prospectively included. Patients with PLICS ≥ 7 together with extremely unstable lateral mass fracture (EULMF) were classified as high-risk group, and the other patients were classified as low-risk group. For patients in the low-risk group, anterior-only reconstruction was performed; for patients in the high-risk group, additional posterior lateral mass fixation and fusion was performed after anterior reconstruction. Clinical outcome evaluation included using the visual analogue score (VAS), the Neck Disability Index (NDI), and the American Spinal Injury Association (ASIA) impairment scale. The change in the local sagittal alignment kyphosis Cobb angle was also recorded. RESULTS: A total of 370 patients (81.1%) completed the minimal 12-month follow-ups, including 321 patients of low-risk group and 49 patients of high-risk group. Compared with the average VAS score preoperatively, the score at 12-month follow-up was significantly improved (from 6.1 + 0.3 to 1.1 + 0.2 in the low-risk group, P < 0.001; from 6.4 + 0.2 to 1.4 + 0.2 in the high-risk group, P < 0.001). The average NDI score at the 12-month follow-up was statistically low in the low-risk group (8.8 + 2.5 vs 13.8 + 3.4, P = 0.034). At least more than one grade improvement in the ASIA scale was observed in 80.5% of all patients. The local kyphosis Cobb angle at the injured segment averaged improved in both groups. CONCLUSION: A PLICS score ≥ 7 together with EULMF can be the threshold for posterior stabilization in addition to anterior reconstruction for the patients with SCFDs.

Structures, Electronic, and Spectral Properties of Doped Boron Clusters MB12 0/- (M = Li, Na, and K).

Structures and electronic properties of alkali metal atom-doped boron clusters MB12 0/- (M = Li, Na, K) are determined using the CALYPSO method for the global minimum search followed by density functional theory. It is found that the global minima obtained for the neutral clusters correspond to the half-sandwich structure and those of the monoanionic clusters correspond to the boat-shaped structure. The neutral MB12 (M = Li, Na, K) can be considered as a member of the half-sandwich doped B12 clusters, and the geometrical pattern of anion MB12 - (M = Li, Na, K) is a new structure that is different from other doped B12 clusters. Natural population and chemical bonding analyses reveal that the alkali metal atom-doped boron clusters MB12 - are characterized as charge transfer complexes, M+B12 2-, resulting in symmetrically distributed chemical bonds and electrostatic interactions between cationic M+ and boron atoms. The calculated spectra indicate that MB12 0/- (M = Li, Na, K) has meaningful spectral features that can be compared with future experimental data. Our work enriches the varieties of geometrical structures of doped boron clusters and can provide much insight into boron nanomaterials.

Can Posterior Ligament Structure Be Functionally Healed After Anterior Reduction and Fusion Surgery in Patients with Traumatic Subaxial Cervical Fracture-Dislocations?

BACKGROUND: Selection of approach for subaxial cervical fracture-dislocation (SCFD) is controversial. The questions of whether a posterior ligamentous structure (PLS) can be functionally healed in patients with SCFD and how long this healing process takes are critical in these patients. METHODS: This study retrospectively enrolled 394 patients with SCFD who underwent anterior decompression, reduction, and fusion between January 2002 and December 2017. The definition of functional healing of PLS was based on evaluations of clinical function and radiographic stability of PLS. RESULTS: Follow-up was available for 354 patients (89.8%). The PLS of 339 cases was functionally healed at the first follow-up interval without any posterior surgical intervention. No hardware failure or progressive cervical kyphosis was observed at further follow-up. At 12 months postoperatively, interbody fusion was satisfactory. However, the other 15 patients experienced nonhealing PLS at 8 weeks postoperatively and developed cervical deformity at further follow-up. Five patients refused revision surgery; the other 10 patients obtained solid fusion after revision surgeries. CONCLUSIONS: Among 354 patients with SCFD and treated by single anterior reduction and fixation, simple PLS injury without any bony instability at the posterior column of the cervical spine can be functionally healed in 95.8% of patients by external fixation for 8 weeks. Whether this finding is applicable to various types of SCFD should be verified in further prospective studies with larger samples.

Development and validation an LC-MS/MS method to quantify (+)-borneol in rat plasma: Application to a pharmacokinetic study.

(+)-Borneol, a bicyclic monoterpene, has been shown to possess valuable biological properties and potential as a pharmaceutical agent due to anti-inflammatory, anti-oxidant and GABA receptor-enhancing functions; it also enhances the permeability of the blood brain barrier to improve the efficacy of CNS drugs. In this study, we have developed a simple, selective, and rapid liquid chromatography-tandem mass spectrometry method for the assay of (+)-borneol in rat plasma. Verapamil was used as an internal standard. Plasma samples were deproteinized using methanol. The analyte was detected by a mass spectrometer with positive atmospheric pressure chemical ionization by multiple reaction monitoring mode for transitions at m/z [M + H]+ 137.2 → 81.0 for (+)-borneol and 455.2 → 165.1 for verapamil. The method has been fully validated to ensure good selectivity, a satisfactory lower limit of quantification at 10.0 ng/mL, acceptable intra- and inter-day accuracy, and high precision. The method was used for the pharmacokinetic evaluation of (+)-borneol in Sprague-Dawley rats after intravenous, oral, and sublingual administration. The results indicate that oral bioavailability of (+)-borneol was extremely low but sublingual administration yielded rapid absorption and favorable bioavailability of (+)-borneol.

Serum irisin associates with breast cancer to spinal metastasis.

The aim of this study was to determine whether the serum level of irisin can be a candidate to predict the spinal metastasis in patients with breast cancer.In a cross-sectional study, 148 patients were recruited. Of those, 53 (35.8%) had spinal metastasis. The baseline characteristics were compared by status of spinal metastasis. Multiple logistic regression analysis was used to determine whether the serum irisin can be a candidate for predicting breast cancer to spinal metastasis. The correlation coefficient analysis was used to confirm the correlation between the serum irisin and lipid metabolic parameters and body mass index (BMI), respectively.The serum irisin was higher in patients without spinal metastasis (7.60 ±â€Š3.80). Multivariable analysis showed that the serum irisin was protective to the presence of spinal metastasis in patients with breast cancer after adjustments of age and BMI (odds ratio, 0.873; 95% confidence interval, 0.764-0.999). Furthermore, there was a positive correlation between the serum irisin and BMI (r = 0.263). The patients with metabolisc syndrome (MetS) had a higher level in serum irisin. In addition, the higher numbers of MetS components were associated with higher serum irisin.Higher serum irisin can be a protective factor of spinal metastasis in patients with breast cancer. The BMI is positively associated with the serum level of irisin. Furthermore, patients with MetS tended to have a higher level of serum irisin.

Consensus scoring model for the molecular docking study of mTOR kinase inhibitor.

The discovery of mammalian target of rapamycin (mTOR) kinase inhibitors has always been a research hotspot of antitumor drugs. Consensus scoring used in the docking study of mTOR kinase inhibitors usually improves hit rate of virtual screening. Herein, we attempt to build a series of consensus scoring models based on a set of the common scoring functions. In this paper, twenty-five kinds of mTOR inhibitors (16 clinical candidate compounds and 9 promising preclinical compounds) are carefully collected, and selected for the molecular docking study used by the Glide docking programs within the standard precise (SP) mode. The predicted poses of these ligands are saved, and revaluated by twenty-six available scoring functions, respectively. Subsequently, consensus scoring models are trained based on the obtained rescoring results by the partial least squares (PLS) method, and validated by Leave-one-out (LOO) method. In addition, three kinds of ligand efficiency indices (BEI, SEI, and LLE) instead of pIC50 as the activity could greatly improve the statistical quality of build models. Two best calculated models 10 and 22 using the same BEI indice have following statistical parameters, respectively: for model 10, training set R2=0.767, Q2=0.647, RMSE=0.024, and for test set R2=0.932, RMSE=0.026; for model 22, raining set R2=0.790, Q2=0.627, RMSE=0.023, and for test set R2=0.955, RMSE=0.020. These two consensus scoring model would be used for the docking virtual screening of novel mTOR inhibitors.

miR-342-5p Regulates Neural Stem Cell Proliferation and Differentiation Downstream to Notch Signaling in Mice.

Notch signaling is critically involved in neural development, but the downstream effectors remain incompletely understood. In this study, we cultured neurospheres from Nestin-Cre-mediated conditional Rbp-j knockout (Rbp-j cKO) and control embryos and compared their miRNA expression profiles using microarray. Among differentially expressed miRNAs, miR-342-5p showed upregulated expression as Notch signaling was genetically or pharmaceutically interrupted. Consistently, the promoter of the miR-342-5p host gene, the Ena-vasodilator stimulated phosphoprotein-like (Evl), was negatively regulated by Notch signaling, probably through HES5. Transfection of miR-342-5p promoted the differentiation of neural stem cells (NSCs) into intermediate neural progenitors (INPs) in vitro and reduced the stemness of NSCs in vivo. Furthermore, miR-342-5p inhibited the differentiation of neural stem/intermediate progenitor cells into astrocytes, likely mediated by targeting GFAP directly. Our results indicated that miR-342-5p could function as a downstream effector of Notch signaling to regulate the differentiation of NSCs into INPs and astrocytes commitment.

A novel tacrine-dihydropyridine hybrid (-)SCR1693 induces tau dephosphorylation and inhibits Aß generation in cells.

AChE inhibitors are the first choice for the treatment of Alzheimer׳s disease (AD), but they could only delay the progression of cognitive and behavioral dysfunction, and fail to reverse neuronal damage. Calcium channel blockers have been identified to have protective effect on neurons. Thus, therapy targeting both AChE and calcium channels is supposed to be more effective in AD treatment. In the present study, we explored the effect of a synthesized juxtaposition of an AChE inhibitor and a Calcium channel blocker (named (-)SCR1693) on tau phosphophorylation and Aß generation. The results showed that: (1) Compared with higher concentrations, (-)SCR1693 incubation in low concentrations such as 0.4, 2, 4µM for 24h did not affect the cell viability of HEK293/tau (HEK293 cells stably transfected with human tau40) and N2a/APP (N2a cells stably transfected with human APP) cells; (2) long-term treatment of cells with (-)SCR1693 (0.4, 2, 5µM) (24h) induced tau dephosphorylation and reduced the total tau level in HEK293/tau cells. Short-term treatment (6h) also resulted in tau dephosphorylation, but did not reduce the total tau level; and (3) (-)SCR1693 (0.4, 2, 4µM) incubation inhibited Aß generation and release dramatically in N2a/APP cells. We conclude that the novel tacrine-dihydropyridine hybrid (-)SCR1693 in low concentrations could reduce total and phosphorylated tau levels, inhibit the generation and release of Aß in cells. Thus, (-)SCR1693 may be a potential candidate for effectively treating AD.

The synergetic effect of edaravone and borneol in the rat model of ischemic stroke.

Free radical production contributes to the early ischemic response and the neuroinflammatory response to injury initiates the second wave of cell death following ischemic stroke. Edaravone is a free radical scavenger, and borneol has shown anti-inflammatory effect. We investigated the synergistic effect of these two drugs in the rat model of transient cerebral ischemia. Edaravone scavenged OH, NO and ONOO─ concentration-dependently, and borneol inhibited ischemia/reperfusion-induced tumor necrosis factor-α (TNF-α), inducible nitric oxide synthase (iNOS), interleukin-1ß (IL-1ß) and cyclooxygenase-2 (COX-2) expressions. In the rat model of transient cerebral ischemia and reperfusion, the combination of edaravone and borneol significantly ameliorated ischemic damage with an optimal proportion of 4:1. Emax (% inhibition) of edaravone, borneol and two drugs in combination was 55.7%, 65.8% and 74.3% respectively. ED50 of edaravone and borneol was 7.17 and 0.36 mg/kg respectively. When two drugs in combination, ED50 was 0.484 mg/kg, in which edaravone was 0.387 mg/kg (ineffective dose) and borneol was 0.097 mg/kg (ineffective dose). Combination index (CI)<1 among effects observed in experiments, suggesting a significant synergistic effect. Reduced levels of pro-inflammatory mediators and free radicals were probably associated with the synergistic effect of edaravone and borneol. The combination exhibited a therapeutic time window of 6h in ischemia/reperfusion model, and significantly ameliorated damages in permanent ischemia model. Moreover, two drugs in combination promoted long-term effect, including improved elemental vital signs, sensorimotor functions and spatial cognition. Our results suggest that the combination of edaravone and borneol have a synergistic effect for treating ischemic stroke.

Transcription factor RBP-J-mediated signaling represses the differentiation of neural stem cells into intermediate neural progenitors.

Notch signaling is critical for multiple aspects of neurogenesis, but how it regulates the proliferation and differentiation of neural stem cells (NSCs) and intermediate neural progenitors (INPs) has not been well elucidated, especially in vivo. In this study, we conditionally ablated the transcription factor RBP-J, which mediates signaling from all four mammalian Notch receptors, in the basal forebrain and ventral midbrain using the RBP-J-floxed mouse and a newly established Nestin-Cre mouse. We found that at early stage of neurogenesis (E11.5), the frequency of neurospheres increased significantly in the RBP-J-inactivated regions. The majority of the RBP-J deficient neurospheres were composed of INPs, suggesting the precocious differentiation of NSCs into INPs. Meanwhile, neuronal differentiation was reduced in the same regions at E11.5, inconsistent with the precocious differentiation phenotype in most Notch-related mutants. At late neurogenic stages (E17.5 and neonatal), as expected from precociously exhausted NSC pool, neurosphere frequency and NSCs decreased in the RBP-J-ablated regions, accompanied by a significant increase of both neurons and glial cells. These results indicated that the RBP-J-mediated signaling might inhibit the differentiation of NSCs into INPs and support the generation of certain early born neurons at early neurogenic stages.