CDK-independent role of D-type cyclins in regulating DNA mismatch repair.

Although mismatch repair (MMR) is essential for correcting DNA replication errors, it can also recognize other lesions, such as oxidized bases. In G0 and G1, MMR is kept in check through unknown mechanisms as it is error-prone during these cell cycle phases. We show that in mammalian cells, D-type cyclins are recruited to sites of oxidative DNA damage in a PCNA- and p21-dependent manner. D-type cyclins inhibit the proteasomal degradation of p21, which competes with MMR proteins for binding to PCNA, thereby inhibiting MMR. The ability of D-type cyclins to limit MMR is CDK4- and CDK6-independent and is conserved in G0 and G1. At the G1/S transition, the timely, cullin-RING ubiquitin ligase (CRL)-dependent degradation of D-type cyclins and p21 enables MMR activity to efficiently repair DNA replication errors. Persistent expression of D-type cyclins during S-phase inhibits the binding of MMR proteins to PCNA, increases the mutational burden, and promotes microsatellite instability.

Quiescence in Saccharomyces cerevisiae.

Most cells live in environments that are permissive for proliferation only a small fraction of the time. Entering quiescence enables cells to survive long periods of nondivision and reenter the cell cycle when signaled to do so. Here, we describe what is known about the molecular basis for quiescence in Saccharomyces cerevisiae, with emphasis on the progress made in the last decade. Quiescence is triggered by depletion of an essential nutrient. It begins well before nutrient exhaustion, and there is extensive crosstalk between signaling pathways to ensure that all proliferation-specific activities are stopped when any one essential nutrient is limiting. Every aspect of gene expression is modified to redirect and conserve resources. Chromatin structure and composition change on a global scale, from histone modifications to three-dimensional chromatin structure. Thousands of proteins and RNAs aggregate, forming unique structures with unique fates, and the cytoplasm transitions to a glass-like state.

FOXO1 mediates hypoxia-induced G0/G1 arrest in ovarian somatic granulosa cells by activating the TP53INP1-p53-CDKN1A pathway.

The development of ovarian follicles constitutes the foundation of female reproduction. The proliferation of granulosa cells (GCs) is a basic process required to ensure normal follicular development. However, the mechanisms involved in controlling GC cell cycle are not fully understood. Here, by performing gene expression profiling in the domestic pig (Sus scrofa), we showed that cell cycle arrest at G0/G1 phase is highly correlated with pathways associated with hypoxic stress and FOXO signalling. Specifically, the elevated proportion of GCs at the arrested G0/G1 phase was accompanied by increased nuclear translocation of FOXO1 under conditions of hypoxia both in vivo and in vitro. Furthermore, phosphorylation of 14-3-3 by the JNK kinase is required for hypoxia-mediated FOXO1 activation and the resultant G0/G1 arrest. Notably, a FOXO1 mutant without DNA-binding activity failed to induce G0/G1 arrest of GCs during hypoxia. Importantly, we identified a new target gene of FOXO1, namely TP53INP1, which contributes to suppression of the G1-S cell cycle transition in response to hypoxia. Furthermore, we demonstrated that the inhibitory effect of the FOXO1-TP53INP1 axis on the GC cell cycle is mediated through a p53-CDKN1A-dependent mechanism. These findings could provide avenues for the clinical treatment of human infertility caused by impaired follicular development.

Survival in Quiescence Requires the Euchromatic Deployment of Clr4/SUV39H by Argonaute-Associated Small RNAs.

Quiescence (G0) is a ubiquitous stress response through which cells enter reversible dormancy, acquiring distinct properties including reduced metabolism, resistance to stress, and long life. G0 entry involves dramatic changes to chromatin and transcription of cells, but the mechanisms coordinating these processes remain poorly understood. Using the fission yeast, here, we track G0-associated chromatin and transcriptional changes temporally and show that as cells enter G0, their survival and global gene expression programs become increasingly dependent on Clr4/SUV39H, the sole histone H3 lysine 9 (H3K9) methyltransferase, and RNAi proteins. Notably, G0 entry results in RNAi-dependent H3K9 methylation of several euchromatic pockets, prior to which Argonaute1-associated small RNAs from these regions emerge. Overall, our data reveal another function for constitutive heterochromatin proteins (the establishment of the global G0 transcriptional program) and suggest that stress-induced alterations in Argonaute-associated sRNAs can target the deployment of transcriptional regulatory proteins to specific sequences.

Proteolytic control in ciliogenesis: Temporal restriction or early initiation?

Cellular processes are highly dependent on a dynamic proteome that undergoes structural and functional rearrangements to allow swift conversion between different cellular states. By inducing proteasomal degradation of inhibitory or stimulating factors, ubiquitylation is particularly well suited to trigger such transitions. One prominent example is the remodelling of the centrosome upon cell cycle exit, which is required for the formation of primary cilia - antenna-like structures on the surface of most cells that act as integrative hubs for various extracellular signals. Over the last decade, many reports on ubiquitin-related events involved in the regulation of ciliogenesis have emerged. Very often, these processes are considered to be initiated ad hoc, that is, directly before its effect on cilia biogenesis becomes evident. While such a temporal restriction may hold true for the majority of events, there is evidence that some of them are initiated earlier during the cell cycle. Here, we provide an overview of ubiquitin-dependent processes in ciliogenesis and discuss available data that indicate such an early onset of proteolytic regulation within preceding cell cycle stages.

The involvement of calcium in the toxic effect of 4-methylethcathinone on SH-SY5Y cells.

Neurotoxicity induced by psychoactive substances is often accompanied by an imbalance of intracellular calcium ions. It is unclear whether calcium ions play a role in the toxicity induced by psychoactive substances. In the present study, we aimed to evaluate the occurrence of calcium dysregulation and its contribution to cytotoxicity in human neurotypic SH-SY5Y cells challenged with a recently developed psychoactive substance 4-methylethcathinone (4-MEC). An increase in the intracellular calcium was detected by inductively coupled plasma atomic emission spectrometry and Fluo-3 AM dye in SH-SY5Y cells after being treated with 4-MEC. The increase of intracellular Ca2+ level mediated G0/G1 cell cycle arrest and ROS/endoplasmic reticulum stress-autophagy signaling pathways to achieve the toxicity of 4-MEC. In particular, N-acetyl-L-cysteine, a classical antioxidant, was found to be a potential treatment for 4-MEC-induced toxicity. Taken together, our results demonstrate that an increase in intracellular calcium content is one of the mechanisms of 4-MEC-induced toxicity. This study provides a molecular basis for the toxicity mechanism and therapeutic intervention of psychoactive substances.

SNHG1 opposes quiescence and promotes docetaxel sensitivity in prostate cancer.

BACKGROUND: A majority of prostate cancer cells are in a non-proliferating, G0 (quiescent) phase of the cell cycle and may lie dormant for years before activation into a proliferative, rapidly progressing, disease phase. Many mechanisms which influence proliferation and quiescence choices remain to be elucidated, including the role of non-coding RNAs. In this study, we investigated the role of a long non-coding RNA (lncRNA), SNHG1, on cell proliferation, quiescence, and sensitivity to docetaxel as a potential factor important in prostate cancer biology. METHODS: Publically available, anonymous, clinical data was obtained from cBioPortal for analysis. RNAi and prostate cancer cell lines were utilized to investigate SNHG1 in vitro. We measured G0 cells, DNA synthesis, and cell cycle distribution by flow cytometry. Western blotting was used to assess G2 arrest and apoptosis. These parameters were also investigated following docetaxel treatment. RESULTS: We discovered that in prostate cancer patients from The Cancer Genome Atlas (TCGA) data set, high SNHG1 expression in localized tumors correlated with reduced progression-free survival, and in a data set of both primary and metastatic tumors, high SNHG1 expression was associated with metastatic tumors. In vitro analysis of prostate cancer cell lines showed SNHG1 expression correlated with a quiescent versus proliferative phenotype. Knockdown of SNHG1 by RNAi in PC3 and C4-2B cells resulted in an accumulation of cells in the G0 phase. After knockdown, 60.0% of PC3 cells were in G0, while control cultures had 13.2% G0. There were reciprocal decreases in G1 phase, but little impact on the proportion of cells in S and G2/M phases, depending on cell line. DNA synthesis and proliferation were largely halted- decreasing by 75% and 81% in C4-2B and PC3 cells, respectively. When cells were treated with docetaxel, SNHG1-depleted C4-2B and PC3 cells were resistant to G2 arrest, and displayed reduced apoptosis, as indicated by reduced cyclin B1 and cleaved caspase 3, suggesting SNHG1 levels may modulate drug response. CONCLUSIONS: Overall, these results indicate SNHG1 has complex roles in prostate cancer, as it stimulates cell cycle entry and disease progression, but sensitizes cells to docetaxel treatment.

N-Propargylic ß-enaminones in breast cancer cells: Cytotoxicity, apoptosis, and cell cycle analyses.

Breast cancer is one of the most common cancers worldwide and the discovery of new cytotoxic agents is needed. Enaminones are regarded to be a significant structural motif that is found in a variety of pharmacologically active compounds however the number of studies investigating the anticancer activities of N-propargylic ß-enaminones (NPEs) is limited. Herein we investigated the potential cytotoxic and apoptotic effects of 23 different NPEs (1-23) on human breast cancer cells. Cytotoxicity was evaluated via MTT assay. Apoptotic cell death and cell cycle distributions were investigated by flow cytometry. CM-H2DCFDA dye was used to evaluate cellular ROS levels. Expression levels of Bcl-2, Bax, p21, and Cyclin D1 were measured by quantitative real-time PCR. ADME properties were calculated using the ADMET 2.0 tool. NPEs 4, 9, 16, and 21 showed selective cytotoxic activity against breast cancer cells with SI values >2. NPEs induced apoptosis and caused significant changes in Bcl-2 and Bax mRNA levels. The cell cycle was arrested at the G0/G1 phase and levels of p21 and Cyclin D1 were upregulated in both breast cancer cells. ROS levels were significantly increased by NPEs, suggesting that the cytotoxic and apoptotic effects of NPEs were mediated by ROS. ADME analysis revealed that NPEs showed favorable distributions in both breast cancer cell lines, meaning good lipophilicity values, low unfractionated values, and high bioavailability. Therefore, these potential anticancer compounds should be further validated by in vivo studies for their appropriate function in human health with a safety profile, and a comprehensive drug interaction study should be performed.

The Anti-G0 Manifesto: Should a problematic construct (G0) with no biological reality be removed from the cell cycle? Yes!

It is widely accepted that there exists a "resting" or "quiescent" state where a growing cell leaves the cell cycle to enter what is often called the "G0-phase." I propose that there is no biological reality to the "G0-phase." The experimental basis for proposing a G0-phase is re-examined and re-analyzed here showing that the G0-phase is an anthropomorphic construct with no biological reality.

Optimized expression and purification of adipose triglyceride lipase improved hydrolytic and transacylation activities in vitro.

Adipose triglyceride lipase (ATGL) plays a key role in intracellular lipolysis, the mobilization of stored triacylglycerol. This work provides an important basis for generating reproducible and detailed data on the hydrolytic and transacylation activities of ATGL. We generated full-length and C-terminally truncated ATGL variants fused with various affinity tags and analyzed their expression in different hosts, namely E.coli, the insect cell line Sf9, and the mammalian cell line human embryonic kidney 293T. Based on this screen, we expressed a fusion protein of ATGL covering residues M1-D288 flanked with N-terminal and C-terminal purification tags. Using these fusions, we identified key steps in expression and purification protocols, including production in the E. coli strain ArcticExpress (DE3) and removal of copurified chaperones. The resulting purified ATGL variant demonstrated improved lipolytic activity compared with previously published data, and it could be stimulated by the coactivator protein comparative gene identification-58 and inhibited by the protein G0/G1 switch protein 2. Shock freezing and storage did not affect the basal activity but reduced coactivation of ATGL by comparative gene identification 58. In vitro, the truncated ATGL variant demonstrated acyl-CoA-independent transacylation activity when diacylglycerol was offered as substrate, resulting in the formation of fatty acid as well as triacylglycerol and monoacylglycerol. However, the ATGL variant showed neither hydrolytic activity nor transacylation activity upon offering of monoacylglycerol as substrate. To understand the role of ATGL in different physiological contexts, it is critical for future studies to identify all its different functions and to determine under what conditions these activities occur.

Neural G0: a quiescent-like state found in neuroepithelial-derived cells and glioma.

Single-cell RNA sequencing has emerged as a powerful tool for resolving cellular states associated with normal and maligned developmental processes. Here, we used scRNA-seq to examine the cell cycle states of expanding human neural stem cells (hNSCs). From these data, we constructed a cell cycle classifier that identifies traditional cell cycle phases and a putative quiescent-like state in neuroepithelial-derived cell types during mammalian neurogenesis and in gliomas. The Neural G0 markers are enriched with quiescent NSC genes and other neurodevelopmental markers found in non-dividing neural progenitors. Putative glioblastoma stem-like cells were significantly enriched in the Neural G0 cell population. Neural G0 cell populations and gene expression are significantly associated with less aggressive tumors and extended patient survival for gliomas. Genetic screens to identify modulators of Neural G0 revealed that knockout of genes associated with the Hippo/Yap and p53 pathways diminished Neural G0 in vitro, resulting in faster G1 transit, down-regulation of quiescence-associated markers, and loss of Neural G0 gene expression. Thus, Neural G0 represents a dynamic quiescent-like state found in neuroepithelial-derived cells and gliomas.

Targeting Akt/NF-κB/p53 Pathway and Apoptosis Inducing Potential of 1,2-Benzenedicarboxylic Acid, Bis (2-Methyl Propyl) Ester Isolated from Onosma bracteata Wall. against Human Osteosarcoma (MG-63) Cells.

Onosma bracteata Wall. is an important medicinal and immunity-enhancing herbs. This plant is commonly used in the preparation of traditional Ayurvedic drugs to treat numerous diseases. Inspired by the medicinal properties of this plant, the present study aimed to investigate the antiproliferative potential and the primary molecular mechanisms of the apoptotic induction against human osteosarcoma (MG-63) cells. Among all the fractions isolated from O. bracteata, ethyl acetate fraction (Obea) showed good antioxidant activity in superoxide radical scavenging assay and lipid peroxidation assay with an EC50 value of 95.12 and 80.67 µg/mL, respectively. Silica gel column chromatography of ethyl acetate (Obea) fraction of O. bracteata yielded a pure compound, which was characterized by NMR, FTIR, and HR-MS analysis and was identified as 1,2-benzene dicarboxylic acid, bis (2-methyl propyl) ester (BDCe fraction). BDCe fraction was evaluated for the antiproliferative potential against human osteosarcoma MG-63, human neuroblastoma IMR-32, and human lung carcinoma A549 cell lines by MTT assay and exhibited GI50 values of 37.53 µM, 56.05 µM, and 47.12 µM, respectively. In MG-63 cells, the BDCe fraction increased the level of ROS and simultaneously decreased the mitochondria membrane potential (MMP) potential by arresting cells at the G0/G1 phase, suggesting the initiation of apoptosis. Western blotting analysis revealed the upregulation of p53, caspase3, and caspase9 while the expressions of p-NF-κB, p-Akt and Bcl-xl were decreased. RT-qPCR studies also showed upregulation in the expression of p53 and caspase3 and downregulation in the expression of CDK2, Bcl-2 and Cyclin E genes. Molecular docking analysis displayed the interaction between BDCe fraction with p53 (-151.13 kcal/mol) and CDK1 (-133.96 kcal/mol). The results of the present work suggest that the BDCe fraction has chemopreventive properties against osteosarcoma (MG-63) cells through the induction of cell cycle arrest and apoptosis via Akt/NF-κB/p53 pathways. This study contributes to the understanding of the utilization of BDCe fraction in osteosarcoma treatment.

Single-cell RNA sequencing of preadipocytes reveals the cell fate heterogeneity induced by melatonin.

Obesity is a global epidemic health disorder and associated with several diseases. Body weight-reducing effects of melatonin have been reported; however, no investigation toward examining whether the beneficial effects of melatonin are associated with preadipocyte heterogeneity has been reported. In this study, we profiled 25 071 transcriptomes of normal and melatonin-treated preadipocytes using scRNA-seq. By tSNE analysis, we present a cellular-state landscape for melatonin-treated preadipocytes that covers multiple-cell subpopulations, defined as cluster 0 to cluster 13. Cluster 0 and cluster 1 were the largest components of normal and melatonin-treated preadipocytes, respectively. G0S2, an inhibitor of adipose triglyceride lipase (ATGL), was significantly upregulated in cluster 0 and downregulated in cluster 1. We redefined cluster 0 as the G0S2-positive cluster (G0S2+ ) and cluster 1 as the G0S2-negative cluster (G0S2- ). Through pseudotime analysis, the G0S2- cluster cell differentiation trajectory was divided into three major structures, that is, the prebranch, the lipid catabolism branch, and the cell fate 2 branch. In vitro, G0S2 knockdown enhanced the expression levels of ATGL, BAT markers and fatty acid oxidation-related genes, but inhibited C/EBPα and PPARγ expression. In vivo, knockdown of G0S2 reduced the body weight gain in high-fat-fed mice. The beneficial effects of the G0S2- cell cluster in promoting lipolysis and inhibiting adipogenesis are dependent on two major aspects: first, downregulation of the G0S2 gene in the G0S2- cluster, resulting in activation of ATGL, which is responsible for the bulk of triacylglycerol hydrolase activity; and second, upregulation of FABP4 in the G0S2- cluster, resulting in inhibition of PPARγ and further reducing adipogenesis.

Mangiferin, a naturally occurring polyphenol, mitigates oxidative stress induced premature senescence in human dermal fibroblast cells.

Chronic oxidative stress has been associated with several human ailments including the condition of aging. Extensive studies have shown the causal relationship between oxidative stress, aging, and cellular senescence. In this regard, forestalling or preventing senescence could delay the aging process as well as act as an intervention against premature aging. Hence, in the present study, we investigated the anti-senescence potential of Mangiferin (MGN) against Hydrogen peroxide (H2O2) induced premature senescence using human dermal fibroblast cells. Early passage human dermal fibroblasts cells were exposed to H2O2 (10 µM) for 15 days. In order to assess the anti-senescence property of MGN, cells were preconditioned with MGN (10 µM / 50 µM; 2 h) followed by addition of H2O2 (10 µM). H2O2 mediated induction of premature senescence was accompanied by elevated ROS, lowering of mitochondrial mass and membrane potential, changes in ATP content along with G0/G1 arrest and SA-ß-gal expression. While, conditioning the cells with MGN lowered oxidative burden, stabilized mitochondrial membrane potential / mass and protected the cells against cell cycle arrest, ultimately rendering protection against premature senescence. The present findings showed that MGN might act as a potential cytoprotective nutraceutical that can prolong the onset of chronic oxidative stress mediated premature senescence.

Control of the Restriction Point by Rb and p21.

The Restriction Point was originally defined as the moment that cells commit to the cell cycle and was later suggested to coincide with hyperphosphorylation of the retinoblastoma protein (Rb). Current cell cycle models posit that cells exit mitosis into a pre-Restriction Point state, where they have low cyclin-dependent kinase (CDK) activity and hypophosphorylated Rb; passage through the Restriction Point then occurs in late G1. Recent single-cell studies have challenged the current paradigm, raising questions about the location of the Restriction Point and the notion that cells exit mitosis into a pre-Restriction Point state. Here, we use a variety of single-cell techniques to show that both noncancer and cancer cells bifurcate into two subpopulations after anaphase, marked by increasing vs. low CDK2 activity and hyper- vs. hypophosphorylation of Rb. Notably, subpopulations with hyper- and hypophosphorylated Rb are present within minutes after anaphase, delineating one subpopulation that never "uncrosses" the Restriction Point and continues cycling and another subpopulation that exits mitosis into an uncommitted pre-Restriction Point state. We further show that the CDK inhibitor p21 begins rising in G2 in mother cells whose daughters exit mitosis into the pre-Restriction Point, CDK2low state. Furthermore, degradation of p21 coincides with escape from the CDK2low state and passage through the Restriction Point. Together, these data support a model in which only a subset of cells returns to a pre-Restriction Point state after mitosis and where the Restriction Point is sensitive to not only mitogens, but also inherited DNA replication stress via p21.

Copper exposure induces hepatic G0/G1 cell-cycle arrest through suppressing the Ras/PI3K/Akt signaling pathway in mice.

Copper (Cu), as a common chemical contaminant in environment, is known to be toxic at high concentrations. The current research demonstrates the effects of copper upon hepatocyte cell-cycle progression (CCP) in mice. Institute of cancer research (ICR) mice (n = 240) at an age of four weeks were divided randomly into groups treated with different doses of Cu (0, 4, 8, and 16 mg/kg) for 21 and 42 days. Results showed that high Cu exposure caused hepatocellular G0/G1 cell-cycle arrest (CCA) and reduced cell proportion in the G2/M phase. G0/G1 CCA occurred with down-regulation (p < 0.05) of Ras, p-PI3K (Tyr458), p-Akt (Thr308), p-forkhead box O3 (FOXO3A) (Ser253), p-glycogen synthase kinase 3-ß (GSK3-ß) (Ser9), murine double minute 2 (MDM2) protein, and mRNA expression levels, and up-regulation (p < 0.05) of PTEN, p-p53 (Ser15), p27, p21 protein, and mRNA expression levels, which subsequently suppressed (p < 0.05) the protein and mRNA expression levels of CDK2/4 and cyclin E/D. These results indicate that Cu exposure suppresses the Ras/PI3K/Akt signaling pathway to reduce the level of CDK2/4 and cyclin E/D, which are essential for the G1-S transition, and finally causes hepatocytes G0/G1 CCA.

In vitro cytotoxicity, cellular uptake, reactive oxygen species and cell cycle arrest studies of novel ruthenium(II) polypyridyl complexes towards A549 lung cancer cell line.

Ruthenium(II) polypyridyl complexes have displayed some promising biological responses against a variety of cancers and have emerged as a potential candidate that can show significant antitumor activity. Three ruthenium(II) polypyridyl complexes were biologically evaluated in vitro against the A549 cancer cell line. The complexes were selected based on initial DNA intercalation studies and MTT viability screening and were selected based on the most promising candidates, the [Ru(bpy)2o-CPIP].2PF6 (complex 1), [Ru(phen)2o-CPIP].2PF6 (complex 2) and [Ru(biq)2o-CPIP].2PF6 (complex 3). Confocal cellular uptake studies confirmed the intracellular transport of complexes into A549. Cytoplasmic and the nucleic accumulation of the complex 1 and 2 was seen while no fluorescent microscopy was performed for complex 3 due to instrumental limitations. Cellular cytotoxicity was investigated with the aid of the Alamar blue assay. The complexes displayed concentration and time dependent inhibitory effects yielding IC50 values from 5.00 to 32.75 µM. Complex 1 exhibit highest cytotoxicity with IC50 value of 5.00 ± 1.24 µM. All of the complexes have shown a significant effect in the reduction of intracellular reactive oxygen species (ROS) levels. Finally, the complexes have shown a transient effect on the cell cycle by arresting it at G0/G1 phase except for complex 2 [Ru(phen)2o-CPIP].2PF6 which has shown the significant G0/G1 arrest.

Physiological modulation of endogenous BRCA1 p220 abundance suppresses DNA damage during the cell cycle.

Endogenous BRCA1 p220 expression peaks in S and G2 when it is activated, and the protein participates in certain key DNA damage responses. In contrast, its expression is markedly reduced in G0/G1. While variations in transcription represent a significant part of p220 expression control, there is at least one other relevant process. We found that a microRNA, miR-545, that is expressed throughout the cell cycle down-modulates endogenous p220 mRNA and protein abundance directly in both G0/G1 and S/G2. When miR-545 function was inhibited by a specific antagomir, endogenous p220 expression increased in G0/G1, and aberrant p220-associated DNA damage responses and de novo DNA strand breaks accumulated. Analogous results were observed upon inhibition of miR-545 function in S/G2. Both sets of antagomir effects were mimicked by infecting cells with a p220 cDNA-encoding adenoviral vector. Thus, strand breaks were a product of p220 overexpression, and their prevention by miR-545 depends on its modulation of p220 expression. Breaks were also dependent on aberrant, overexpressed p220-driven recruitment of RAD51 to either spontaneously arising or mutagen-based DNA damage sites. Hence, when its level is not physiologically maintained, endogenous p220 aberrantly directs at least one DNA repair protein, RAD51, to damage sites, where their action contributes to the development of de novo DNA damage. Thus, like its loss, a surfeit of endogenous p220 function represents a threat to genome integrity.

Potential Antitumor Effects of 6-Gingerol in p53-Dependent Mitochondrial Apoptosis and Inhibition of Tumor Sphere Formation in Breast Cancer Cells.

Hormone-specific anticancer drugs for breast cancer treatment can cause serious side effects. Thus, treatment with natural compounds has been considered a better approach as this minimizes side effects and has multiple targets. 6-Gingerol is an active polyphenol in ginger with various modalities, including anticancer activity, although its mechanism of action remains unknown. Increases in the level of reactive oxygen species (ROS) can lead to DNA damage and the induction of DNA damage response (DDR) mechanism, leading to cell cycle arrest apoptosis and tumorsphere suppression. Epidermal growth factor receptor (EGFR) promotes tumor growth by stimulating signaling of downstream targets that in turn activates tumor protein 53 (p53) to promote apoptosis. Here we assessed the effect of 6-gingerol treatment on MDA-MB-231 and MCF-7 breast cancer cell lines. 6-Gingerol induced cellular and mitochondrial ROS that elevated DDR through ataxia-telangiectasia mutated and p53 activation. 6-Gingerol also induced G0/G1 cell cycle arrest and mitochondrial apoptosis by mediating the BAX/BCL-2 ratio and release of cytochrome c. It also exhibited a suppression ability of tumorsphere formation in breast cancer cells. EGFR/Src/STAT3 signaling was also determined to be responsible for p53 activation and that 6-gingerol induced p53-dependent intrinsic apoptosis in breast cancer cells. Therefore, 6-gingerol may be used as a candidate drug against hormone-dependent breast cancer cells.

Anticancer Potential of Damnacanthal and Nordamnacanthal from Morinda elliptica Roots on T-lymphoblastic Leukemia Cells.

BACKGROUND: This study reports on the cytotoxic properties of nordamnacanthal and damnacanthal, isolated from roots of Morinda elliptica on T-lymphoblastic leukaemia (CEM-SS) cell lines. METHODS: MTT assay, DNA fragmentation, ELISA and cell cycle analysis were carried out. RESULTS: Nordamnacanthal and damnacanthal at IC50 values of 1.7 µg/mL and10 µg/mL, respectively. At the molecular level, these compounds caused internucleosomal DNA cleavage producing multiple 180-200 bp fragments that are visible as a "ladder" on the agarose gel. This was due to the activation of the Mg2+/Ca2+-dependent endonuclease. The induction of apoptosis by nordamnacanthal was different from the one induced by damnacanthal, in a way that it occurs independently of ongoing transcription process. Nevertheless, in both cases, the process of dephosphorylation of protein phosphates 1 and 2A, the ongoing protein synthesis and the elevations of the cytosolic Ca2+ concentration were not needed for apoptosis to take place. Nordamnacanthal was found to have a cytotoxic effect by inducing apoptosis, while damnacanthal caused arrest at the G0/G1 phase of the cell cycle. CONCLUSION: Damnacanthal and nordamnacanthal have anticancer properties, and could act as potential treatment for T-lymphoblastic leukemia.