Nek2 Kinase Signaling in Malaria,Bone, Immune and Kidney Disorders to Metastatic Cancers and Drug Resistance: Progress on Nek2 Inhibitor Development

Cell cycle kinases represent an important component of the cell machinery that controls signal transduction involved in cell proliferation, growth, and differentiation. Nek2 is a mitotic Ser/Thr kinase that localizes predominantly to centrosomes and kinetochores and orchestrates centrosome disjunction and faithful chromosomal segregation. Its activity is tightly regulated during the cell cycle with the help of other kinases and phosphatases and via proteasomal degradation. Increased levels of Nek2 kinase can promote centrosome amplification (CA), mitotic defects, chromosome instability (CIN), tumor growth, and cancer metastasis. While it remains a highly attractive target for the development of anti-cancer therapeutics, several new roles of the Nek2 enzyme have recently emerged: these include drug resistance, bone, ciliopathies, immune and kidney diseases, and parasitic diseases such as malaria. Therefore, Nek2 is at the interface of multiple cellular processes and can influence numerous cellular signaling networks. Herein, we provide a critical overview of Nek2 kinase biology and discuss the signaling roles it plays in both normal and diseased human physiology. While the majority of research efforts over the last two decades have focused on the roles of Nek2 kinase in tumor development and cancer metastasis, the signaling mechanisms involving the key players associated with several other notable human diseases are highlighted here. We summarize the efforts made so far to develop Nek2 inhibitory small molecules, illustrate their action modalities, and provide our opinion on the future of Nek2-targeted therapeutics. It is anticipated that the functional inhibition of Nek2 kinase will be a key strategy going forward in drug development, with applications across multiple human diseases.     

Monoamine Oxidase (MAO) as a Potential Target for Anticancer Drug Design and Development

Monoamine oxidases (MAOs) are oxidative enzymes that catalyze the conversion of biogenic amines into their corresponding aldehydes and ketones through oxidative deamination. Owing to the crucial role of MAOs in maintaining functional levels of neurotransmitters, the implications of its distorted activity have been associated with numerous neurological diseases. Recently, an unanticipated role of MAOs in tumor progression and metastasis has been reported. The chemical inhibition of MAOs might be a valuable therapeutic approach for cancer treatment. In this review, we reported computational approaches exploited in the design and development of selective MAO inhibitors accompanied by their biological activities. Additionally, we generated a pharmacophore model for MAO-A active inhibitors to identify the structural motifs to invoke an activity.     

Chromosomal instability is more frequent in metastasized than in non-metastasized pulmonary carcinoids but is not a reliable predictor of metastatic potential

Pulmonary carcinoids are infrequent neoplasms of the lung that normally display a less aggressive biological behavior compared to small cell and non-small cell lung cancers. Approximately 15-25% of carcinoids, in particular atypical carcinoids, show lymph node metastasis and have a worse prognosis than their non-metastasized counterparts. To date, there is no morphological or molecular marker that may help to differentiate between carcinoids that metastasize and carcinoids of identical differentiation that show only local tumor growth. In this study, we analyzed 7 metastasized and 10 non-metastasized pulmonary carcinoids for chromosomal and microsatellite instability in order to determine whether microsatellite instability or chromosomal imbalances are associated with metastasis. Due to the rare occurrence of metastasized carcinoids we compared our results of chromosomal instability with the hitherto published comparative genomic hybridization (CGH) profiles of pulmonary carcinoids, for which information about the absence or presence of metastasis was available. While microsatellite instability was not detected we found chromosomal instability as a common event in pulmonary carcinoids with an increase of frequency and extent of chromosomal alterations in atypical and metastasized carcinoids. These findings are in accordance with the collected and herein compiled data of previous studies and indicate increasing numbers of chromosomal imbalances to play a role in the sequential process of tumor development and metastasis.     

MVD及VEGF表达与鼻咽癌侵袭转移关系的研究

为探讨微血管密度（ＭＶＤ）及血管内皮生长因子（ＶＥＧＦ）的表达与鼻咽癌（ＮＰＣ）侵袭转移关系,在分子水平干预肿瘤血管生成,预防ＮＰＣ复发和转移打下基础,采用免疫组织化学Ｓ－Ｐ法检测了７３例ＮＰＣ,１５例鼻咽良性肿瘤、２０例无瘤鼻咽部石蜡标本组织中的ＭＶＤ及ＶＥＧＦ表达,ＮＰＣ中转移组４９例,非转移组２４例。     

An integrated sample pretreatment platform for quantitative N-glycoproteome analysis with combination of on-line glycopeptide enrichment,deglycosylation and dimethyl labeling

Relative quantification of N-glycoproteomes shows great promise for the discovery of candidate biomarkers and therapeutic targets. The traditional protocol for quantitative analysis of glycoproteomes is usually off-line performed, and suffers from long sample preparation time, and the risk of sample loss or contamination due to manual manipulation. In this study, a novel integrated sample preparation platform for quantitative N-glycoproteome analysis was established, with combination of online N-glycopeptide capture by a HILIC column, sample buffer exchange by a N₂-assisted HILIC–RPLC interface, deglycosylation by a hydrophilic PNGase F immobilized enzymatic reactor (hIMER) and solid dimethyl labeling on a C18 precolumn. To evaluate the performance of such a platform, two equal aliquots of immunoglobulin G (IgG) digests were sequentially pretreated, followed by MALDI-TOF MS analysis. The signal intensity ratio of heavy/light (H/L) labeled deglycosylated peptides with the equal aliquots was 1.00 (RSD = 6.2%, n = 3), much better than those obtained by the offline protocol, with H/L ratio as 0.76 (RSD = 11.6%, n = 3). Additionally, the total on-line sample preparation time was greatly shortened to 160 min, much faster than that of offline approach (24 h). Furthermore, such an integrated pretreatment platform was successfully applied to analyze the two kinds of hepatocarcinoma ascites syngeneic cell lines with high (Hca-F) and low (Hca-P) lymph node metastasis rates. For H/L labeled Hca-P lysates with the equal aliquots, 99.6% of log 2 ratios (H/L) of quantified glycopeptides ranged from −1 to 1, demonstrating high accuracy of the developed sample preparation strategy. By triplicated analysis of glycopeptides and non-glycopeptides of Hca-F and Hca-P lysates, 43 up-regulated and 30 down-regulated (Hca-F/P) N-glycosylation sites, and 11 significantly changed N-glycoproteins were successfully quantified, and most of them were related to tumorigenesis and tumor metastasis. All these results demonstrate the developed integrated N-glycoprotein pretreatment platform is of great power for the accurate, precise and high-throughput analysis of N-glycoproteomes.     

Efficacy of a Covalent Microtubule Stabilizer in Taxane-Resistant Ovarian Cancer Models

Ovarian cancer often has a poor clinical prognosis because of late detection, frequently after metastatic progression, as well as acquired resistance to taxane-based therapy. Herein, we evaluate a novel class of covalent microtubule stabilizers, the C-22,23-epoxytaccalonolides, for their efficacy against taxane-resistant ovarian cancer models in vitro and in vivo. Taccalonolide AF, which covalently binds β-tubulin through its C-22,23-epoxide moiety, demonstrates efficacy against taxane-resistant models and shows superior persistence in clonogenic assays after drug washout due to irreversible target engagement. In vivo, intraperitoneal administration of taccalonolide AF demonstrated efficacy against the taxane-resistant NCI/ADR-RES ovarian cancer model both as a flank xenograft, as well as in a disseminated orthotopic disease model representing localized metastasis. Taccalonolide-treated animals had a significant decrease in micrometastasis of NCI/ADR-RES cells to the spleen, as detected by quantitative RT-PCR, without any evidence of systemic toxicity. Together, these findings demonstrate that taccalonolide AF retains efficacy in taxane-resistant ovarian cancer models in vitro and in vivo and that its irreversible mechanism of microtubule stabilization has the unique potential for intraperitoneal treatment of locally disseminated taxane-resistant disease, which represents a significant unmet clinical need in the treatment of ovarian cancer patients.     

Short communication for targeting natural compounds against HER2 kinase domain as potential anticancer drugs applying pharmacophore based molecular modelling approaches- part 2

Breast cancer is one of the common causes of death noticed in women globally. In order to find effective therapeutics, the current investigation has focussed on identifying candidate compounds for EGFR and HER2. Accordingly, the pharmacophore modelling approaches were adapted to identify two prospective compounds and were docked against the target 3RCD that is complexed with TAK-285 a known dual inhibitor. Focussing on the target 3RCD, our results have showed that the compounds have demonstrated a good binding affinity towards the target occupying the binding pocket. They have established key residue interactions with stable molecular dynamics simulation results. The Hit compounds have demonstrated a potential to penetrate the blood brain barrier thereby enriching their therapeutics towards breast cancer brain metastasis. Taken together, our findings propose two candidate compounds as EGFR/HER2 inhibitors that might serve as novel chemical spaces for designing and developing new inhibitors.     

Fluorescent Mimetics of CMP‐Neu5Ac Are Highly Potent,Cell‐Permeable Polarization Probes of Eukaryotic and Bacterial Sialyltransferases and Inhibit Cellular Sialylation

Oligosaccharides of the glycolipids and glycoproteins at the outer membranes of human cells carry terminal neuraminic acids, which are responsible for recognition events and adhesion of cells, bacteria, and virus particles. The synthesis of neuraminic acid containing glycosides is accomplished by intracellular sialyl transferases. Therefore, the chemical manipulation of cellular sialylation could be very important to interfere with cancer development, inflammations, and infections. The development and applications of the first nanomolar fluorescent inhibitors of sialyl transferases are described herein. The obtained carbohydrate‐nucleotide mimetics were found to bind all four commercially available and tested eukaryotic and bacterial sialyl transferases in a fluorescence polarization assay. Moreover, it was observed that the anionic mimetics intruded rapidly and efficiently into cells in vesicles and translocated to cellular organelles surrounding the nucleus of CHO cells. The new compounds inhibit cellular sialylation in two cell lines and open new perspectives for investigations of cellular sialylation.     

Poria Acid,Triterpenoids Extracted from Poria cocos,Inhibits the Invasion and Metastasis of Gastric Cancer Cells

Background: Poria cocos (P. cocos) is an important medicinal fungus in traditional Chinese medicine. Poria acid (PA), a triterpenoid compound, is an effective component of traditional Chinese medicine P. cocos. This experiment investigated the anti-gastric cancer biological activity of PA in vitro. Methods: The effect of PA on the viability of gastric cancer cells was detected by the thiazolyl blue (MTT) assay. Cell adhesion assays were used to detect changes in the adhesion of cells treated after PA (0, 20, 40, and 80 µmol/L). The ability of cell invasion and migration were detected by Transwell assays and wound healing assays. A high-content imaging system was used to dynamically record the motility of the gastric cancer cells after PA (0, 20, 40, and 80 µmol/L) treatment. Western blotting was used to detect the expression of epithelial–mesenchymal transformation (EMT), invasion and migration related proteins. Results: The MTT assay showed that the proliferation of gastric cancer cells was significantly inhibited after PA treatment. Cell adhesion experiments showed that the adhesion of gastric cancer cells was significantly decreased after PA treatment. Compared with the control group, the wound healing area of the gastric cancer cells treated with different concentrations of PA decreased. The Transwell assay showed that the number of gastric cancer cells passing through the cell membrane were significantly reduced after PA treatment. In addition, after PA treatment, the cells’ movement distance and average movement speed were significantly lower than those of the control group. Finally, PA can significantly alter the expression of EMT-related proteins E-cadherin, N-cadherin, and Vimentin and decreased the expressions of metastasis-related proteins matrix metalloproteinase (MMP) 2, MMP-9 and tissue inhibition of matrix metalloproteinase (TIMP)1 in the gastric cancer cells. Conclusions: Triterpenoids from P. cocos have significant biological activity against gastric cancer, and the mechanism may be involved in the process of epithelial–mesenchymal transformation.