Arrest Of Mitosis Research Articles

Noscapine, a Non-addictive Opioid and Microtubule-Inhibitor in Potential Treatment of Glioblastoma.

Noscapine is a phthalide isoquinoline alkaloid that easily traverses the blood brain barrier and has been used for years as an antitussive agent with high safety. Despite binding opioid receptors, noscapine lacks significant hypnotic and euphoric effects rendering it safe in terms of addictive potential. In 1954, Hans Lettré first described noscapine as a mitotic poison. The drug was later tested for cancer treatment in the early 1960's, yet no effect was observed likely as a result of its short biological half-life and limited water solubility. Since 1998, it has regained interest thanks to studies from Emory University, which showed its anticancer activity in animal models with negligible toxicity. In contrast to other microtubule-inhibitors, noscapine does not affect the total intracellular tubulin polymer mass. Instead, it forces the microtubules to spend an increased amount of time in a paused state leading to arrest in mitosis and subsequently inducing mitotic slippage/mitotic catastrophe/apoptosis. In experimental models, noscapine does not induce peripheral neuropathy, which is common with other microtubule inhibitors. Noscapine also inhibits tumor growth and enhances cancer chemosensitivity via selective blockage of NF-κB, an important transcription factor in glioblastoma pathogenesis. Due to their anticancer activities and high penetration through the blood-brain barrier, noscapine analogues strongly deserve further study in various animal models of glioblastoma as potential candidates for future patient therapy.

The Biological Activity of the Novel Vinca Alkaloids 4-chlorochablastine and 4-chlorochacristine.

Vinca alkaloids are important cancer drugs belonging to the class of antimitotic agents. The most commonly used substances are vinblastine and vincristine, other compounds are vinorelbine and vinflunine. All of them are very effective drugs but their use is limited by severe side-effects including neurotoxicity and bone marrow depression. Therefore, it is very important to develop novel vinca alkaloids with similar efficacy but lower toxicity. Here, we analyzed two new compounds, 4-chlorochablastine and 4-chlorochacristine, with regard to their biological activity. These novel compounds were applied to a leukemia cell line at clinically relevant concentrations. For comparison, the established vinca alkaloids vinblastine, vincristine, vinorelbine, and vinflunine were also tested. Both novel substances decreased cellular proliferation. Apoptosis was found to be increased using two different methods reflecting early and late apoptosis. Cell cycle analysis revealed a clear decrease in G1-cells and an increase in G2/M-cells indicating an arrest in mitosis. In general, 4- chlorochablastine and 4-chlorochacristine caused these effects at concentrations higher than those needed for vinblastine, vincristine, and vinorelbine, but the potency was approximately in the range of vinflunine. Taken together, the results show first indications that these novel vinca alkaloids might be effective and that they warrant further analysis.

Cell cycle arrest in mitosis promotes interferon-induced necroptosis.

Resistance to apoptosis is a hallmark of cancer and deregulation of apoptosis often leads to chemoresistance. Therefore, new approaches to target apoptosis-resistant cancer cells are crucial for the development of directed cancer therapies. In the present study, we investigated the effect of cell cycle regulators on interferon (IFN)-induced necroptosis as an alternative cell death mechanism to overcome apoptosis resistance. Here, we report a novel combination treatment of IFNs with cell cycle arrest-inducing compounds that induce necroptosis in apoptosis-resistant cancer cells and elucidate the underlying molecular mechanisms. Combination treatment of IFNs (i.e. IFNβ) with inhibitors of the cell cycle (e.g. vinorelbine (VNR), nocodazole (Noc), polo-like kinase-1 (Plk-1) inhibitor BI 6727) co-operate to induce necroptotic cell death upon caspase inactivation. The mode of cell death was confirmed by pharmacological inhibition and siRNA-mediated downregulation of the key necroptotic factors receptor-interacting protein (RIP) kinase 3 (RIP3) and mixed-lineage kinase-like (MLKL) in various cell lines. Mechanistically, we show that necroptosis upon VNR/IFNβ/zVAD.fmk treatment is RIP1-independent but relies on IFNβ-induced gene expression of Z-DNA-binding protein 1 (ZBP1) as shown by quantitative RT-PCR and genetic knockdown experiments. Interestingly, we find that RIP3 is phosphorylated in response to compounds that trigger mitotic arrest, even in the absence of IFNβ signaling and necroptosis induction. Together, the identification of a novel combination treatment that triggers necroptosis has implications for the development of molecular-targeted therapies to circumvent apoptosis resistance and point to an underestimated role of cell cycle regulation in cell death signaling.

Overexpression of Fbxo6 inactivates spindle checkpoint by interacting with Mad2 and BubR1

ABSTRACT The spindle assembly checkpoint prevents chromosome mis-segregation during mitosis by delaying sister chromatid separation. Several F-box protein members play critical roles in maintaining genome stability and regulating cell cycle progress via ubiquitin-mediated protein degradation. Here, we showed that Fbxo6 critically regulated spindle checkpoint and chromosome segregation. Fbxo6 was phosphorylated during mitosis. Overexpression of Fbxo6 lead to faster exit from nocodazole-induced mitosis arrest through premature sister chromatid separation. Moreover, we found substantially more binuclear and multilobed nuclei cells accompanied with impaired cell viability in Fbxo6-overexpressed HeLa cells. Mechanistically, Fbxo6 interacted with spindle checkpoint proteins including Mad2 and BubR1 leading to the premature exit from mitosis. Overall, we revealed a novel role of Fbxo6 in regulating spindle checkpoint, which may shed light on the regulation of genome instability of cancer cells.

Downregulation of NOP53 Ribosome Biogenesis Factor Leads to Abnormal Nuclear Division and Chromosomal Instability in Human Cervical Cancer Cells.

NOP53 ribosome biogenesis factor (NOP53) is a nucleolar protein involved in oncogenesis/tumor suppression, cell cycle regulation, and cell death. Here, we investigated the role of NOP53 in the maintenance of normal nuclear shape and chromosomal stability. Depletion of NOP53 by shRNA caused abnormal nuclear morphology, including large nucleus, irregular nucleus, and multinucleated cells, and chromosomal instability resulting in micronucleus or nuclear bud formation. The abnormal nuclear shape and chromosomal instability were restored by re-expression of NOP53. We further showed that NOP53 was involved in chromosome congression in metaphase. Downregulation of NOP53 induced aberrant chromosome congression and spindle checkpoint activation, resulting in delayed mitosis and mitotic arrest. Thus, our findings demonstrated that the nucleolar protein NOP53 participated in mitotic progression and that dysregulated NOP53 expression caused chromosomal instability in cancer cells.

JNK Inactivation Induces Polyploidy and Drug-Resistance in Coronarin D-Treated Osteosarcoma Cells.

Inhibition of proliferating cells is a critical strategy for cancer therapy. In this study, we demonstrated that coronarin D, a natural component extracted from the rhizomes of Hedychium coronarium, significantly suppressed the proliferation of osteosarcoma cells. The treatment with coronarin D resulted in the activation of caspase-3 and apoptosis. This treatment induced the accumulation of cyclin B1 and DNA condensation indicating the treated osteosarcoma cells were arrested in mitotic phase. Furthermore, the treatment with coronarin D increased the levels of phosphorylated c-Jun NH2-terminal kinase (JNK) in human osteosarcoma cells. Pretreatment with JNK inhibitor blocked the accumulation of cyclin B1 and DNA condensation, resulting the accumulation of tetraploid cells in coronarin D-treated osteosarcoma HOS cells, indicating JNK inactivation blocked the mitotic entry and arrested cells in the 4 N state. After adaptation, the arrested tetraploid cells continued to duplicate their DNA resulting in polyploidy. Interestingly, when the arrested mitotic cells induced by coronarin D were treated with JNK inhibitor, the accumulated cyclin B1 and DNA condensation were immediately eliminated. These arrested 4 N cells loss the ability to undergo cytokinesis, and ultimately continued to duplicate DNA upon prolonged arrest resulting in the production of polyploid populations. JNK inactivation, either by the pretreatment with JNK inhibitor or the treatment with JNK inhibitor in coronarin D-induced mitotic cells, both caused resistance to coronarin D-induced cell death. Taken together, our findings indicate that coronarin D induces the apoptosis and mitosis arrest in human osteosarcoma cells. JNK has a crucial role in coronarin D-induced mitosis arrest and apoptosis. We hypothesize that functional evaluation of JNK may produce more specific and effective therapies in coronarin D-related trail for treatment of human osteosarcoma.

Novel nicotinoyl pyrazoline derivates bearing N-methyl indole moiety as antitumor agents: Design, synthesis and evaluation

In the present work, twenty-five nicotinoyl pyrazoline derivates bearing N-methyl indole moiety have been designed and synthesized. The biological evaluation of these compounds as tubulin assembly inhibitors revealed that most of them were potential antitumor agents. Among them, compound 28 exhibited most potency against cancer cell line panels (GI50 = 29–90 nM for HeLa, HepG2 and MCF-7 cells) without toxicity to non-tumor cells (CC50 > 300 μM for 293 T cell), bound to the colchicine site of tubulin and displayed excellent inhibitory activity in tubulin assembly assay (IC50 = 1.6 μM, better than CA-4). Molecular dynamics simulation was carried out to validate the docking pose of compound 28 with tubulin crystalline. Further investigation on HepG2 and HeLa cells demonstrated that compound 28 could cause mitosis arrest to G2/M phase, and subsequently induced cell apoptosis. The efficiency in vivo of compound 28 was also evaluated on HeLa-Xenograft nude mice, and the relative tumor inhibition ration was up to 61.52% without noticeable weight loss and tissue damage (examined by H&E staining), which was comparable to CA-4 (inhibited 59.92%). In brief, compound 28 is a promising candidate for tumor therapy as tubulin assembly inhibitor.

New 6- and 7-heterocyclyl-1H-indole derivatives as potent tubulin assembly and cancer cell growth inhibitors

We designed new 3-arylthio- and 3-aroyl-1H-indole derivatives 3–22 bearing a heterocyclic ring at position 5, 6 or 7 of the indole nucleus. The 6- and 7-heterocyclyl-1H-indoles showed potent inhibition of tubulin polymerization, binding of colchicine to tubulin and growth of MCF-7 cancer cells. Compounds 13 and 19 inhibited a panel of cancer cells and the NCI/ADR-RES multidrug resistant cell line at low nanomolar concentrations. Compound 13 at 50 nM induced 77% G2/M in HeLa cells, and at 20 nM caused 50% stable arrest of mitosis. As an inhibitor of HepG2 cells (IC50 = 20 nM), 13 was 4-fold superior to 19. Compound 13 was a potent inhibitor of the human U87MG glioblastoma cells at nanomolar concentrations, being nearly one order of magnitude superior to previously reported arylthioindoles. The present results highlight 13 as a robust scaffold for the design of new anticancer agents.

Synthesis and anti-mitotic activity of 2,4- or 2,6-disubstituted- and 2,4,6-trisubstituted-2H-pyrazolo[4,3-c]pyridines

An efficient synthetic route for the synthesis of 2H-pyrazolo[4,3-c]pyridines, primarily varying by the substituents at the 2-, 4- and 6-positions, is described here. A Sonogashira-type cross-coupling reaction was employed to yield 3-alkynyl-1H-pyrazole-4-carbaldehydes, ethanones and propanones from the corresponding 1H-pyrazol-3-yl trifluoromethanesulfonates. Subsequent treatment of the coupling products with dry ammonia afforded a versatile library of 2H-pyrazolo[4,3-c]pyridines, which were then evaluated for their cytotoxicity against K562 and MCF-7 cancer cell lines. The most potent of these compounds displayed low micromolar GI50 values in both cell lines. Active compounds induced dose-dependent cell-cycle arrest in mitosis, as shown by flow cytometric analysis of DNA content and phosphorylation of histone H3 at serine-10. Moreover, biochemical assays revealed increased activities of caspases-3/7 in treated cells, specific fragmentation of PARP-1, and phosphorylation of Bcl-2, collectively confirming apoptosis as the mechanism of cell death.

Abstract A070: Uncovering unknown Aurora kinase signaling in adrenal tumor cell line, NCI-H295

Abstract Adrenocortical carcinomas (ACC) are rare tumors with no effective therapy for patients with advanced or metastatic disease. Therefore, efforts have been made to develop new therapeutic approaches for effective ACC treatment. Studies suggest that the dysregulation of Aurora kinases might be strongly associated with tumorigenesis and that Aurora kinases are involved in the promotion of the epithelial-mesenchymal transition (EMT) and invasion in different cancer cells. Molecular markers for EMT include decreased expression of E-cadherin, nuclear localization of β-catenin, and upregulation of genes related to the Wnt/β-catenin pathway, such as Cyclin D1 and c-Myc. Thus, the aim of this study was to investigate whether Aurora kinases are involved in the promotion of EMT in ACC cells. NCI-H295 cells were treated with 50 nM of the pan-Aurora kinase inhibitor, AMG 900, for 6, 24, and 48 hours. Multiple Reaction Monitoring proteomic approach was performed to screen proteins that are known to be involved in EMT. Total and enriched membrane, nucleus, and cytosol proteins were obtained by different centrifugation grades and detergents. Western blotting was performed for protein expression and subcellular localization. Cell cycle, proliferation, and migration assays were performed to study the effect of AMG 900 in NCI-H295 cells. MRM studies showed increased expression of E-cadherin 24 and 48 hours after AMG 900 treatment and decreased expression of N-cadherin after 48 hours of treatment. Conversely, our data showed increased of β-catenin expression and Vimentin expression after 48 hours of treatment. Moreover, our results showed higher expression of β-catenin in the nucleus when compared to control. Western blotting analysis confirmed these findings and gene expression analysis showed increased of c-Myc, one of the main products of β-catenin nuclear translocation, after 24 hours of treatment. We performed functional assays in order to check whether the studied pathways would affect NCI-H295 cell proliferation and migration, and our results showed that inhibition of Aurora kinases leads to cell cycle arrest in mitosis and decreases cell proliferation and migration. Further studies need to be performed combining Aurora kinases and β-catenin inhibitors; however, our results suggest that Aurora kinases might regulate proteins involved in EMT process during ACC progression. Citation Format: Andrea G. Maria, Carolina H. Thomé, Kleiton S. Borges, Mariana L. Grassi, Augusto F. Andrade, Regia C.P. Lira, Fabio R. Faucz, Constantine A. Stratakis, Luiz G. Tone, Vitor M. Faca. Uncovering unknown Aurora kinase signaling in adrenal tumor cell line, NCI-H295 [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr A070.

Protocols for the Study of Taxanes Chemosensitivity in Prostate Cancer.

Prostate cancer is major cause of cancer-related death among men in Western countries. Locally advanced prostate cancers are treated with castration therapy, which is initially effective, but after months the disease progresses to a hormone-refractory state whose treatment is chemotherapy based on taxanes. Although taxanes improve the survival of patients with castration-resistant prostate cancers, these patients often develop chemotherapy resistance, and new therapeutic strategies are necessary. Taxanes exert their action through interaction with β-tubulin which triggers cell cycle arrest in mitosis and the subsequent induction of the intrinsic apoptotic pathway. Since taxanes are widely used for the treatment of advanced prostate cancers, we present in this chapter protocols that allow the study of the prostate cancer sensitivity as well as determine the mechanisms of resistance to these chemotherapeutic agents.

1674P - Exploratory study of CK-M30 and pHH3 expression in Circulating Tumor Cells (CTCs) as biomarkers of docetaxel (DOC) efficacy in metastatic castration resistant prostate cancer (mCRPC)

Background: A drop in CTCs counts as early as 4 weeks following treatment initiation have been suggested as an indicator of overall survival (OS) benefit. DOC remains a pivotal treatment in mCRPC for which there are no early pharmacodynamic (PD) markers of its activity in patients (pts). CTCs may be used as surrogate tumor tissue to study PD markers of apoptosis (CK-M30) or mitosis arrest (pHH3) in mCRPC pts receiving DOC. Methods: We conducted a prospective 2-cohort multicenter exploratory study in mCRPC pts progressing by PCWG2 criteria who were eligible for DOC 75 mg/m^2. Pts were prescreened using the CellSearch system and selected if CTCs≥5/7.5mLs of blood (Basal 1). A 2nd blood sample (Basal 2) was drawn in eligible pts within 0-7 days prior to C1D1 of DOC and further samples were collected at 8h, 24h, 7d and 21d. Directly conjugated mAb against pHH3 and CK-M30 were used in combination with CellSearch. Statistical analyses were performed to evaluate the baseline and post-treatment variability. Increases in % of biomarker CTC+ greater than the median baseline variability were correlated with achieving a 50% PSA response (PSA50) and OS from DOC start using chi-square and long-rank test, respectively. Results: 60 mCRPC pts (CK-M30 = 30; pHH3 = 30), 95% ECOG 0-1, 95% and 17% have bone and visceral metastases respectively, received a median of 7 cycles (range 2-10) of DOC. Biomarker results are summarised in Table.Table1674P VariabilityBasal 1 (N = 60) Median (Range)Basal 2 (N = 57) Median (Range)%marker Corr. CoeffPost-24h (n = 59) median (Range)P-value Basal vs post-24hCohort CK-M30CTC/7.5mL9 (5-1266)8 (3-863)0.54 p = 0.7639 (4-1129)NSCK-M30+48% (0-76%)35% (0-100%)51% (0-100%)Cohort pHH3CTC/7.5mL10 (5-567)12 (4-623)0.98 p < 0.0019 (3-584)<0.001pHH3+0% (0-10%)0% (0-8%)7% (0-67%)ResponseNPSA50p-valueOS median (CI95%)p-valueCK-M30+ change 24hCKM30 + >50%52NS24 m (–)NSCKM30 + <50%251121 m (12-30)pHH3+ change 24hpHH3 + >10%1380.04717 m (12-22)NSpHH3 + <10%16412 m (6-18) Open table in a new tab Conclusions: pHH3 in CTC+ may be a potential PD biomarker of a favourable response to DOC treatment. Legal entity responsible for the study: Spanish National Cancer Research Centre Funding: None Disclosure: All authors have declared no conflicts of interest.

Isoelectric point-based fractionation by HiRIEF coupled to LC-MS allows for in-depth quantitative analysis of the phosphoproteome

Protein phosphorylation is involved in the regulation of most eukaryotic cells functions and mass spectrometry-based analysis has made major contributions to our understanding of this regulation. However, low abundance of phosphorylated species presents a major challenge in achieving comprehensive phosphoproteome coverage and robust quantification. In this study, we developed a workflow employing titanium dioxide phospho-enrichment coupled with isobaric labeling by Tandem Mass Tags (TMT) and high-resolution isoelectric focusing (HiRIEF) fractionation to perform in-depth quantitative phosphoproteomics starting with a low sample quantity. To benchmark the workflow, we analyzed HeLa cells upon pervanadate treatment or cell cycle arrest in mitosis. Analyzing 300 µg of peptides per sample, we identified 22,712 phosphorylation sites, of which 19,075 were localized with high confidence and 1,203 are phosphorylated tyrosine residues, representing 6.3% of all detected phospho-sites. HiRIEF fractions with the most acidic isoelectric points are enriched in multiply phosphorylated peptides, which represent 18% of all the phospho-peptides detected in the pH range 2.5–3.7. Cross-referencing with the PhosphoSitePlus database reveals 1,264 phosphorylation sites that have not been previously reported and kinase association analysis suggests that a subset of these may be functional during the mitotic phase.

Prolonged mitotic arrest induced by Wee1 inhibition sensitizes breast cancer cells to paclitaxel.

Wee1 kinase is a crucial negative regulator of Cdk1/cyclin B1 activity and is required for normal entry into and exit from mitosis. Wee1 activity can be chemically inhibited by the small molecule MK-1775, which is currently being tested in phase I/II clinical trials in combination with other anti-cancer drugs. MK-1775 promotes cancer cells to bypass the cell-cycle checkpoints and prematurely enter mitosis. In our study, we show premature mitotic cells that arise from MK-1775 treatment exhibited centromere fragmentation, a morphological feature of mitotic catastrophe that is characterized by centromeres and kinetochore proteins that co-cluster away from the condensed chromosomes. In addition to stimulating early mitotic entry, MK-1775 treatment also delayed mitotic exit. Specifically, cells treated with MK-1775 following release from G1/S or prometaphase arrested in mitosis. MK-1775 induced arrest occurred at metaphase and thus, cells required 12 times longer to transition into anaphase compared to controls. Consistent with an arrest in mitosis, MK-1775 treated prometaphase cells maintained high cyclin B1 and low phospho-tyrosine 15 Cdk1. Importantly, MK-1775 induced mitotic arrest resulted in cell death regardless the of cell-cycle phase prior to treatment suggesting that Wee1 inhibitors are also anti-mitotic agents. We found that paclitaxel enhances MK-1775 mediated cell killing. HeLa and different breast cancer cell lines (T-47D, MCF7, MDA-MB-468 and MDA-MB-231) treated with different concentrations of MK-1775 and low dose paclitaxel exhibited reduced cell survival compared to mono-treatments. Our data highlight a new potential strategy for enhancing MK-1775 mediated cell killing in breast cancer cells.

The Zebrafish curly fry Is Required for Proper Centrosome and Mitotic Spindle Assembly.

The zebrafish curly fry (cfy) mutation leads to a dramatic increase in mitotic index and cell death starting during neural tube formation. The mutant phenotype is cell autonomous and does not result from defects in apical/basal polarity within the neuroepithelium. The increase in mitotic index could be due to increased proliferation or cell cycle arrest in mitosis. cfy embryos were analyzed to examine these two possibilities. By labeling embryos with a pulse of BrdU and anti-phospho-histone 3 and examining the DNA content by fluorescence activated cell sorting, we show that cfy mutants exhibit no increase in proliferation, but a significant increase in the number of cells arrested in mitosis. Furthermore, time-lapse microscopy in vivo confirmed that a great majority of dividing cells arrest during mitosis and that these mitotically arrested cells die in cfy embryos. Finally, immunostaining and confocal microscopy in cfy mutant embryos revealed that mitotic cells in mutants contain aberrant centrosomes and often exhibit monopolar spindles, thereby leading to mitotic cell cycle arrest. Our results suggest that the cfy gene is required for proper centrosome assembly and mitotic spindle formation, therefore critical for normal cell division.

Single-Cell RNA-Seq Analysis Maps Development of Human Germline Cells and Gonadal Niche Interactions

Human fetal germ cells (FGCs) are precursors to sperm and eggs and are crucial for maintenance of the species. However, the developmental trajectories and heterogeneity of human FGCs remain largely unknown. Here we performed single-cell RNA-seq analysis of over 2,000 FGCs and their gonadal niche cells in female and male human embryos spanning several developmental stages. We found that female FGCs undergo four distinct sequential phases characterized by mitosis, retinoic acid signaling, meiotic prophase, and oogenesis. Male FGCs develop through stages of migration, mitosis, and cell-cycle arrest. Individual embryos of both sexes simultaneously contain several subpopulations, highlighting the asynchronous and heterogeneous nature of FGC development. Moreover, we observed reciprocal signaling interactions between FGCs and their gonadal niche cells, including activation of the bone morphogenic protein (BMP) and Notch signaling pathways. Our work provides key insights into the crucial features of human FGCs during their highly ordered mitotic, meiotic, and gametogenetic processes invivo.

Allelopathy and Anti-mitotic Effects of Cuscuta campestris and Cuscuta monogyna Extracts on Plant Cell Division

Some natural compounds of plants including phenols and alkaloids induce mitosis depressive that are blocked cell division. Field dodder (Cuscuta campestris Yunck.) and eastern dodder (Cuscuta monogyna Vahl.) are the most widespread obligatory parasitic species. In the present study, end of summer 2015, field dodder and eastern dodder are collected from Alhagi maurorum Medik. host and Vine (Vitis sylvestris C.C.Gmel.) host respectively from Markazi province of Iran and identified by department of botany of Islamic Azad University of Saveh. The aerial parts of dodders were used for aqueous extract. Dodders alkaloid and poly phenol were visualized by TLC and Dragendorff (for alkaloid) and ferric salt (for poly phenol) reagents. Total phenols and alkaloid were measured by using the calibration and spectrophotometry methods. Root length and roots tip meristem model of onion bulbs and barley seedlings were utilized to allelopathy and antimitotic effects of dodders aqueous extracts. Standard cytotoxic 8-Hydroxyquinoline served as controls. The bulbs and barley seedlings were treated at various concentrations: 0.05, 0.1 and 0.2 mgml-1 (dw) of and aqueous extract of dodders and 8-Hydroxyquinoline for 6 and 12 h. The inhibitory effect of dodders extract were evaluated on the growth and mitotic activity (Mitotic index) of barley seedlings and onion root meristems and the effect was compared with standard 8-Hydroxyquinoline. The One Way ANOVA test was used for statistical analysis. The poly phenols of field dodder was condensed tannins (include flavonoids) but for eastern dodder hydrolysable tannins. Like 8-Hydroxyquinoline, both field and eastern dodders extracts significantly inhibited the growth of roots and mitotic activity in incubation time and dose-dependent manner. However, C. campestris extract was more allelopathy and antimitotic potent in this regard and produced root decay and mitosis arrest. The extract of dodders had inhibitory and mitosis depressive effect on root tip meristem cells. In the present study we found, the dodders aqueous extracts can been used to produce desirable effects as it pertains to chromosome condensation and spread, and though the roles of these chemicals in spindle fiber inhibition have been elucidated, as 8-Hydroxyquinoline.

Effect of SP600125 on the mitotic spindle in HeLa Cells, leading to mitotic arrest, endoreduplication and apoptosis.

BackgroundThe JNK inhibitor SP600125 strongly inhibits cell proliferation in many human cancer cells by blocking mitosis progression and inducing cell death. Despite, all this study, the mechanism by which SP600125 inhibits mitosis-related effects in human cervical cells (HeLa cells) remains unclear. In this study, we investigated the effects of SP600125 on the cell viability, cell cycle, and on the spindle assembly during mitosis in HeLa cells.MethodsTo explore this approach, we used a viability test, an immunofluorescence microscopy to detect Histone phosphorylation and mitotic spindle aberrations. Apoptosis was characterised using Western Blotting.ResultsTreatment of HeLa cells with varying concentrations of SP600125 induces significant G2/M cell cycle arrest with elevated phosphorylation of histone H3 within 48 h, and endoreduplication after 48 h. SP600125 also induces significant abnormal mitotic spindle. High concentrations of SP600125 (20 μM) induce disturbing microtubule assembly in vitro. Additionally, SP600125- induced delayed apoptosis and cell death was accompanied by significant poly ADP-ribose polymerase (PARP) cleavage and caspase-3 activation in the late phase (at 72 h).ConclusionOur results confirmed that SP600125 induce mitosis arrest in G2/M, endoreduplication, mitotic spindle aberrations and apoptosis.

Inactivation of transforming growth factor-β-activated kinase 1 promotes taxol efficacy in ovarian cancer cells.

Resistance to taxol represents a major obstacle for long-term remission in ovarian cancer. Transforming Growth Factor-β-Activated Kinase 1 (TAK1) is a critical component in immune response pathway. However, the role of TAK1 in the development of chemoresistance in ovarian cancer remains unknown. Here, we showed that in vitro, taxol-resistant cells expressed higher TAK1, and the ratio of p-TAK1/TAK1 positively associated with taxol resistance in ovarian cancer cells. Inactivation of TAK1 by inhibitor 5Z-7-oxozeaenol or gene knockdown sensitized taxol cytotoxicity in vitro, promoting cell apoptosis and mitosis arrest. Moreover, resistant cells were much more sensitive to the combined TAK1 inhibitor and taxol treatment than their parental counterparts. Using xenograft mouse model, we found that 5Z-7-oxozeaenol significantly enhanced taxol efficacy in vivo. Thus, targeting TAK1 pathway is a promising strategy to enhance taxol response in ovarian cancer treatment.

Investigation on the isoform selectivity of novel kinesin-like protein 1 (KIF11) inhibitor using chemical feature based pharmacophore, molecular docking, and quantum mechanical studies

Kinesin-like protein (KIF11) is a molecular motor protein that is essential in mitosis. Removal of KIF11 prevents centrosome migration and causes cell arrest in mitosis. KIF11 defects are linked to the disease of microcephaly, lymph edema or mental retardation. The human KIF11 protein has been actively studied for its role in mitosis and its potential as a therapeutic target for cancer treatment. Pharmacophore modeling, molecular docking and density functional theory approaches was employed to reveal the structural, chemical and electronic features essential for the development of small molecule inhibitor for KIF11. Hence we have developed chemical feature based pharmacophore models using Discovery Studio v 2.5 (DS). The best hypothesis (Hypo1) consisting of four chemical features (two hydrogen bond acceptor, one hydrophobic and one ring aromatic) has exhibited high correlation co-efficient of 0.9521, cost difference of 70.63 and low RMS value of 0.9475. This Hypo1 is cross validated by Cat Scramble method; test set and decoy set to prove its robustness, statistical significance and predictability respectively. The well validated Hypo1 was used as 3Dquery to perform virtual screening. The hits obtained from the virtual screening were subjected to various scrupulous drug-like filters such as Lipinski’s rule of five and ADMET properties. Finally, six hit compounds were identified based on the molecular interaction and its electronic properties. Our final lead compound could serve as a powerful tool for the discovery of potent inhibitor as KIF11 agonists.