S-phase Arrest Research Articles

Targeting the Homologous Recombination Pathway in Cancer With a Novel Class of RAD51 Inhibitors.

Targeting DNA damage response (DDR) pathway has been proposed as an approach for amplifying tumor-specific replicative lesions. RAD51 plays a central role in the DDR process, and thus represents a promising anti-tumor target. We here report the discovery of a series of next generation RAD51 inhibitors that can prevent RAD51 foci formation. The lead compounds dramatically impaired human cancer cell growth, induced cell cycle arrest in S-phase, and resulted in elevated γH2AX. Furthermore, cancer cells became sensitized to chemotherapy and other DDR inhibitors. Dosed either as a single agent or in combination with cisplatin, the compounds significantly inhibited tumor growth in vivo. By upregulating ATR-CHK1 signaling, the RAD51 inhibitors increased surface PD-L1 levels in various tumor cells, suggesting a potential combination of RAD51 inhibitors with PD-1/PD-L1 blockade. Overall, our findings provide the preclinical rationale to explore RAD51 inhibitors as monotherapy or in combination with chemotherapy, immunotherapy or DDR-targeting therapy in cancer treatment.

Radiosensitization to γ-Ray by Functional Inhibition of APOBEC3G.

The radiosensitization of tumor cells is one of the promising approaches for enhancing radiation damage to cancer cells and limiting radiation effects on normal tissue. In this study, we performed a comprehensive screening of radiosensitization targets in human lung cancer cell line A549 using an shRNA library and identified apolipoprotein B mRNA editing enzyme catalytic subunit 3G (APOBEC3G: A3G) as a candidate target. APOBEC3G is an innate restriction factor that inhibits HIV-1 infection as a cytidine deaminase. APOBEC3G knockdown with siRNA showed an increased radiosensitivity in several cancer cell lines, including pancreatic cancer MIAPaCa2 cells and lung cancer A549 cells. Cell cycle analysis revealed that APOBEC3G knockdown increased S-phase arrest in MIAPaCa2 and G2/M arrest in A549 cells after γ-irradiation. DNA double-strand break marker γH2AX level was increased in APOBEC3G-knocked-down MIAPaCa2 cells after γ-irradiation. Using a xenograft model of A549 in mice, enhanced radiosensitivity by a combination of X-ray irradiation and APOBEC3G knockdown was observed. These results suggest that the functional inhibition of APOBEC3G sensitizes cancer cells to radiation by attenuating the activation of the DNA repair pathway, suggesting that APOBEC3G could be useful as a target for the radiosensitization of cancer therapy.

The chemical structure impairs the intensity of genotoxic effects promoted by 1,2-unsaturated pyrrolizidine alkaloids in vitro

1,2-unsaturated pyrrolizidine alkaloids (PAs) represent a large group of secondary plant metabolites exhibiting hepatotoxic, genotoxic, and carcinogenic properties upon bioactivation.To examine how the degree of esterification affects the genotoxic profile of PA we investigated cytotoxicity, histone H2AX phosphorylation, DNA strand break induction, cell cycle perturbation, micronuclei formation, and aneugenic effects in different cell models.Analysis of cytotoxicity and phosphorylation of histone H2AX was structure- and concentration-dependent: diester-type PAs (except monocrotaline) showed more pronounced effects than monoester-type PAs. Cell cycle analysis identified that diester-type PAs induced a S-phase arrest and a decrease in the occurrence of cells in the G1-phase. The same structure-dependency was observed by flow-cytometric analysis of PA-induced micronuclei in CYP3A4-overexpressing V79 cells. Analysis of centromeres induced by lasiocarpine in the micronuclei by fluorescence in situ hybridization indicated an aneugenic effect in V79h3A4 cells. Comet assays revealed no significant induction of DNA strand breaks for all investigated PAs.Overall, diester-type PAs induced more pronounced effects than monoester-type PAs. Furthermore, our results indicate aneugenic effects upon exposure towards lasiocarpine in vitro. These data improve our understanding how structural features of PA influence the genotoxic profile. Especially, the monoester-type PAs seem to induce less severe effects than other PAs.

Quercetin attenuates the proliferation of arsenic-related lung cancer cells via a caspase-dependent DNA damage signaling.

Exposure to arsenic (As) mainly through contaminated drinking water enhances the lung tumor progression, invasion, and metastasis. The carcinogenic effect of As is due to the generation of reactive oxygen species (ROS) and DNA damage, and interference with DNA repair machinery. Herein, we investigated the potential therapeutic function of quercetin on As-treated lung cancer cells. Quercetin is a natural product with antioxidative, anti-inflammatory, and antiproliferative properties. We showed that quercetin induced cell death in the As-exposed lung cancer cells in a dose-dependent manner. Quercetin was able to significantly inhibit the proliferation of the As-treated cells over a period of 5 weeks. In addition, quercetin induced ROS-mediated DNA double-strand breaks in the As-treated lung cancer cells. We also showed that ROS generation induced by quercetin activated caspase-3 to a sufficient level to induce DNA damage but insufficient to induce death in As-treated lung cancer cells. Moreover, transient activation of caspase-2 was detected in quercetin- and As-cotreated cells. The flow cytometry-based cell cycle analysis showed that the antiproliferative function of quercetin was mediated by S-phase cell cycle arrest, which was associated with upregulation of the Ataxia Telangiectasia-mutated (ATM), but not ATM and RAD3-related. In conclusion, quercetin synergized the As-driven ROS generation and DNA damage, and induced the S-phase arrest, thus inhibiting the proliferation of As-exposed lung cancer cells. This data suggested that quercetin is an alternative reagent to chemo-drugs to prevent the growth of As-exposed lung cancer cells.

The radiosensitizing effect of β-Thujaplicin, a tropolone derivative inducing S-phase cell cycle arrest, in head and neck squamous cell carcinoma-derived cell lines

BackgroundResistance to radiotherapy is a common cause of treatment failure in advanced head and neck squamous cell carcinoma (HNSCC). ß-Thujaplicin, a natural tropolone derivative, acts as an anti-cancer agent and has recently been shown to radiosensitize non-HNSCC cancer cells. However, no data is currently available on its radiosensitizing potential in HNSCC.MethodsTo investigate the effect of ß-Thujaplicin and irradiation in HNSCC cell lines CAL27 and FADU, we performed a cell viability assay, colony forming assay, flow cytometry for cell cycle analysis and a wound healing assay. Drug-irradiation interaction was analyzed using a zero-interaction potency model.ResultsTreatment with ß-Thujaplicin led to a dose-dependent decrease in cell viability and enhanced the effect of irradiation. Clonogenic survival was inhibited with synergistic drug-irradiation interaction. ß-Thujaplicin further led to S-phase arrest and increased the sub-G1 population. Moreover, combined ß-Thujaplicin and irradiation treatment had a higher anti-migratory effect compared to irradiation alone.Conclusionsß-Thujaplicin acts as a radiosensitizer in HNSCC cell lines. Further evaluation of its use in HNSCC therapy is warranted.

Phosphatidylserine-Specific Phospholipase A1 Limits Aggressiveness of Lung Adenocarcinoma by Lysophosphatidylserine and Protein Kinase A–Dependent Pathway

Lipid metabolic abnormalities in cancer cells are increasingly being studied. Several studies have reported that phosphatidylserine-specific phospholipase A1 (PLA1A) might be involved in the pathogenesis of cancers. Nevertheless, the function and mechanistic details of PLA1A in lung adenocarcinoma (LUAD) progression remain largely undefined. In the present study, low PLA1A expression was correlated with poor prognosis in patients with LUAD. Results from invitro and invivo animal studies showed that overexpressed PLA1A suppressed the proliferation of LUAD cells invitro and tumor growth invivo through regulation of cyclin abundance, thereby inducing S-phase arrest. Meanwhile, PLA1A overexpression attenuated migration and invasion of LUAD cells, including by inhibiting the epithelial-mesenchymal transition. Mechanistically, PLA1A overexpression inhibited aggressiveness of LUAD cells through elevated lysophosphatidylserine, which acts via G-protein-coupled receptor 174, further activating cAMP/protein kinase A pathway. Activating G-protein-coupled receptor 174/protein kinase A pathway may involve effects on cell cycle regulators and transcription factors-regulated epithelial-mesenchymal transition. The study uncovered the mechanism through which PLA1A regulates LUAD proliferation, invasion, and migration. These results demonstrate the potential use of PLA1A as a biomarker for diagnosing LUAD, which may therefore potentially serve as a therapeutic target for LUAD.

MicroRNA-150 targets major epigenetic repressors and inhibits cell proliferation

The Polycomb Repressive Complex (PRC) proteins, EZH2 and EZH1 regulate many biological processes by generating the repressive H3K27me3 modifications in the chromatin. However, the factors that regulate the EZH1/EZH2 functions are poorly studied. We identify that the 3′UTRs of EZH2 and EZH1 mRNAs contain the binding sites for the miRNA, miR-150. MicroRNA-150 (miR-150) controls numerous biological processes including cell proliferation, differentiation and pathogenesis of a variety of diseases including cancer. We find that miR-150 regulates the levels of EZH1 and EZH2 through various experimental investigations. Since EZH2 is known to form a repressive complex with other epigenetic repressors especially DNMT3A and DNMT3B, we investigated whether miR-150 also regulates the DNMT3A and DNMT3B levels. We report that miR-150 regulates DNMT3A and DNMT3B levels through direct and indirect mechanisms respectively. Since these epigenetic repressors promote cell proliferation, we investigated the effect of miR-150 perturbation on HEK293 cell proliferation. We found that miR-150 inhibits cell proliferation and induces S-phase arrest by increasing the levels of tumor suppressors and decreasing the cell cycle regulators. Collectively, our study shows that miR-150 act as a tumor suppressor by down-regulating the oncogenic major epigenetic repressors and controls cell proliferation.

Arene Ru(II) Complexes with Difluorinated Ligands Act as Potential Inducers of S-Phase Arrest via the Stabilization of c-myc G-Quadruplex DNA.

Here, a series of half-sandwich arene Ru(II) complexes with difluorinated ligands [Ru(η6-arene)(L)Cl] (L1 = 2-(2,3-difluorophenyl)imidazole[4,5f][1,10]-phenanthroline; L2 = 2-(2,4-difluorophenyl)imidazole[4,5f][1,10]-phenanthroline; arene = benzene, toluene, and p-cymene) were synthesized and characterized. Molecular docking analysis showed that these complexes bind to c-myc G-quadruplex DNA through either groove binding or π–π stacking, and the relative difluorinated site in the main ligand plays a role in regulating the binding mode. The binding behavior of these complexes with c-myc G-quadruplex DNA was evaluated using ultraviolet–visible spectroscopy, fluorescence intercalator displacement assay, fluorescence resonance energy transfer melting assay, and polymerase chain reaction. The comprehensive analysis indicated that complex 1 exhibited a better affinity and stability in relation to c-myc G-quadruplex DNA with a DC50 of 6.6 μM and ΔTm values of 13.09 °C, than other molecules. Further activity evaluation results displayed that this class of complexes can also inhibit the growth of various tumor cells, especially complexes 3 and 6, which exhibited a better inhibitory effect against human U87 glioblastoma cells (51.61 and 23.75 μM) than other complexes, even superior to cisplatin (32.59 μM). Owing to a befitting lipophilicity associated with the high intake of drugs by tumor cells, complexes 3 and 6 had favorable lipid-water partition coefficients of −0.6615 and −0.8077, respectively. Moreover, it was found that complex 6 suppressed the proliferation of U87 cells mainly through an induced obvious S phase arrest and slight apoptosis, which may have resulted from the stabilization of c-myc G-quadruplex DNA to block the transcription and expression of c-myc. In brief, these types of arene Ru(II) complexes with difluorinated ligands can be developed as potential inducers of S-phase arrest and apoptosis through the binding and stabilization of c-myc G-quadruplex DNA, and could be used in clinical applications in the future.

Urushiol V Suppresses Cell Proliferation and Enhances Antitumor Activity of 5-FU in Human Colon Cancer Cells by Downregulating FoxM1.

Colorectal cancer (CRC) is one of the most common malignant tumor. 5-FU is commonly used for the treatment of CRC. However, the development of drug resistance in tumor chemotherapy can seriously reduce therapeutic efficacy of 5-FU. Recent data show that FoxM1 is associated with 5-FU resistance in CRC. FoxM1 plays a critical role in the carcinogenesis and drug resistance of several malignancies. It has been reported that urushiol V isolated from the cortex of Rhus verniciflua Stokes is cytotoxic to several types of cancer cells. However, the underlying molecular mechanisms for its antitumor activity and its potential to attenuate the chemotherapeutic resistance in CRC cells remain unknown. Here, we found that urushiol V could inhibit the cell proliferation and induced S-phase arrest of SW480 colon cancer cells. It inhibited protein expression level of FoxM1 through activation of AMPK. We also investigated the combined effect of urushiol V and 5-FU. The combination treatment reduced FoxM1 expression and consequently reduced cell growth and colony formation in 5-FU resistant colon cancer cells (SW480/5-FUR). Taken together, these result suggest that urushiol V from Rhus verniciflua Stokes can suppress cell proliferation by inhibiting FoxM1 and enhance the antitumor capacity of 5-FU. Therefore, urushiol V may be a potential bioactive compound for CRC therapy.

Ribonucleotide reductase M2 subunit silencing suppresses tumorigenesis in pancreatic cancer via inactivation of PI3K/AKT/mTOR pathway

Background/objectivesRibonucleotide Reductase M2 subunit (RRM2) is elevated in pancreatic cancer and involved in DNA synthesis and cell proliferation. But its specific mechanism including genetic differences and upstream regulatory pathways remains unclear. MethodsWe analyzed RRM2 expression of 178 pancreatic cancer patients in Gene Expression Profiling Interactive Analysis (GEPIA) database. Besides, more pancreatic cancer specimens were collected and detected RRM2 expression by immunohistochemistry. RRM2 knockdown by shRNA was applied for functional and mechanism analysis in vitro. Xenograft tumor growth was significantly slower by RRM2 silencing in vivo. ResultsIt showed that high RRM2 expression had a poorer overall survival and disease free survival. RRM2 expression was higher in tumor grade 2 and 3 than grade 1. Immunohistochemistry data validated that high RRM2 expression predicted worse survival. RRM2 knockdown significantly reduced cell proliferation, inhibited colony formation and suppressed cell cycle progress. Further mechanism assay showed silencing RRM2 lead to inactivation of PI3K/AKT/mTOR pathway and inhibition of mutant p53, which induce S phase arrest and/or apoptosis. In panc-1 cells, S-phase arrest mediated by mutant p53 inhibition, p21 increase and cell cycle related proteins change. While in miapaca-2 cells, induction of apoptosis and S-phase arrest mediated by CDK1 played a coordinated role. ConclusionTaken together, high RRM2 expression was associated with worse prognosis. Importantly, RRM2 knockdown deactivated PI3K/AKT/mTOR pathway, resulting in cell cycle arrest and/or apoptosis. This study shed light on the molecular mechanism of RRM2 in pancreatic tumor progression and is expected to provide a new theoretical basis for pancreatic cancer treatment.

6-Methoxyflavone induces S-phase arrest through the CCNA2/CDK2/p21CIP1 signaling pathway in HeLa cells

ABSTRACT This study aimed to elucidate the specific anticancer mechanism of 6-methoxyflavone in HeLa cells. A total of 178 putative targets of 6-methoxyflavone were obtained from the PharmMapper database. Microarray analyses, transcriptome sequencing analyses, functional enrichment analyses, and gene set enrichment analyses were performed to preliminarily explore the roles and mechanisms of the 178 targets in cervical cancer. Cell counting kit-8, cell cycle assays, polymerase chain reactions, and western blotting were used to clarify the mechanism of action of 6-methoxyflavone. Molecular docking and noncovalent interaction analyses were performed to further confirm the mechanism of action in three-dimensional structures. Functional enrichment analyses and gene set enrichment analyses indicated that high mRNA expression of cyclin A2 (CCNA2) and cyclin-dependent kinase 2 (CDK2) stimulated cell cycle progression in cervical cancer. Cell proliferation and cycle assays, transcriptome sequencing, polymerase chain reactions, and western blotting revealed that 6-methoxyflavone inhibited HeLa cell proliferation and induced S-phase arrest via the CCNA2/CDK2/ cyclin-dependent kinase inhibitor 1A (p21CIP1) pathway. Molecular docking and noncovalent interaction analyses showed that 6-methoxyflavone had the strongest affinity toward, inhibitory effect on, and noncovalent interactions with CDK2, and that the combination of CDK2 and CCNA2 enhanced these effects. An analysis of clinical characteristics showed that 6-methoxyflavone might be related to six clinicopathological parameters of cervical cancer patients. 6-Methoxyflavone induces S-phase arrest in HeLa cells via the CCNA2/CDK2/p21CIP1 pathway.

Adapalene Inhibits Prostate Cancer Cell Proliferation In Vitro and In Vivo by Inducing DNA Damage, S-phase Cell Cycle Arrest, and Apoptosis

Aims: Prostate cancer is a well-known aggressive malignant tumor in men with a high metastasis rate and poor prognosis. Adapalene (ADA) is a third-generation synthetic retinoid with anticancer properties. We investigated the anti-tumor activity and molecular mechanisms of ADA in the RM-1 prostate cancer cell line in vivo and in vitro. Methods: The effects of ADA on cell proliferation were estimated using the CCK-8 and colony formation assays. The wound-healing assay and the Transwell assay were employed to examine the migratory capacity and invasiveness of the cells. Flow cytometry was utilized to evaluate the cell cycle and apoptosis, and Western blotting analysis was used to assess the expression of the associated proteins. Micro-CT, histomorphological, and immunohistochemical staining were used to assess the effects of ADA on bone tissue structure and tumor growth in a mouse model of prostate cancer bone metastasis. Result: ADA dramatically inhibited cell proliferation, migration, invasiveness, and induced S-phase arrest and apoptosis. ADA also regulated the expression of S-phase associated proteins and elevated the levels of DNA damage markers, p53, and p21 after ADA treatment, suggesting that the anti-tumor effect of ADA manifests through the DNA damage/p53 pathway. Furthermore, we observed that ADA could effectively inhibited tumor growth and bone destruction in mice. Conclusion: ADA inhibited prostate cancer cell proliferation, elicited apoptosis, and arrested the cell cycle in the S-phase. ADA also slowed the rate of tumor growth and bone destruction in vitro. Overall, our results suggest that ADA may be a potential treatment against prostate cancer.

Modulation of lncRNA H19 enhances resveratrol-inhibited cancer cell proliferation and migration by regulating endoplasmic reticulum stress.

The phytoalexin resveratrol exhibits anti‐tumour activity in many types of cancer. In this study, we showed that resveratrol suppressed the survival of gastric tumour cells both in vivo and in vitro. Resveratrol promoted apoptosis, autophagy and endoplasmic reticulum (ER) stress in a dose‐dependent manner. RNA‐seq analysis showed that multiple cell death signalling pathways were activated after resveratrol treatment, while the use of ER stress activators (tunicamycin and thapsigargin) in combinatorial with resveratrol led to further inhibition of cancer cell survival. Results also showed that resveratrol altered the expression of several long non‐coding RNAs (lncRNAs), including MEG3, PTTG3P, GAS5, BISPR, MALAT1 and H19. Knockdown of H19 in resveratrol‐treated cells further enhanced the effects of resveratrol on apoptosis, ER stress and cell cycle S‐phase arrest. Furthermore, the migratory ability of resveratrol‐treated cells was dramatically decreased after H19 knockdown. In conclusion, resveratrol inhibited cancer cell survival, while knockdown of lncRNA H19 resulted in increased sensitivity to resveratrol therapy.

Targeting HNRNPU to overcome cisplatin resistance in bladder cancer

PurposeThe overall response of cisplatin-based chemotherapy in bladder urothelial carcinoma (BUC) remains unsatisfactory due to the complex pathological subtypes, genomic difference, and drug resistance. The genes that associated with cisplatin resistance remain unclear. Herein, we aimed to identify the cisplatin resistance associated genes in BUC.Experimental designThe cytotoxicity of cisplatin was evaluated in six bladder cancer cell lines to compare their responses to cisplatin. The T24 cancer cells exhibited the lowest sensitivity to cisplatin and was therefore selected to explore the mechanisms of drug resistance. We performed genome-wide CRISPR screening in T24 cancer cells in vitro, and identified that the gene heterogeneous nuclear ribonucleoprotein U (HNRNPU) was the top candidate gene related to cisplatin resistance. Epigenetic and transcriptional profiles of HNRNPU-depleted cells after cisplatin treatment were analyzed to investigate the relationship between HNRNPU and cisplatin resistance. In vivo experiments were also performed to demonstrate the function of HNRNPU depletion in cisplatin sensitivity.ResultsSignificant correlation was found between HNRNPU expression level and sensitivity to cisplatin in bladder cancer cell lines. In the high HNRNPU expressing T24 cancer cells, knockout of HNRNPU inhibited cell proliferation, invasion, and migration. In addition, loss of HNRNPU promoted apoptosis and S-phase arrest in the T24 cells treated with cisplatin. Data from The Cancer Genome Atlas (TCGA) demonstrated that HNRNPU expression was significantly higher in tumor tissues than in normal tissues. High HNRNPU level was negatively correlated with patient survival. Transcriptomic profiling analysis showed that knockout of HNRNPU enhanced cisplatin sensitivity by regulating DNA damage repair genes. Furthermore, it was found that HNRNPU regulates chemosensitivity by affecting the expression of neurofibromin 1 (NF1).ConclusionsOur study demonstrated that HNRNPU expression is associated with cisplatin sensitivity in bladder urothelial carcinoma cells. Inhibition of HNRNPU could be a potential therapy for cisplatin-resistant bladder cancer.

Pharmacological targeting of MTHFD2 suppresses acute myeloid leukemia by inducing thymidine depletion and replication stress

The folate metabolism enzyme MTHFD2 (methylenetetrahydrofolate dehydrogenase/cyclohydrolase) is consistently overexpressed in cancer but its roles are not fully characterized, and current candidate inhibitors have limited potency for clinical development. In the present study, we demonstrate a role for MTHFD2 in DNA replication and genomic stability in cancer cells, and perform a drug screen to identify potent and selective nanomolar MTHFD2 inhibitors; protein cocrystal structures demonstrated binding to the active site of MTHFD2 and target engagement. MTHFD2 inhibitors reduced replication fork speed and induced replication stress followed by S-phase arrest and apoptosis of acute myeloid leukemia cells in vitro and in vivo, with a therapeutic window spanning four orders of magnitude compared with nontumorigenic cells. Mechanistically, MTHFD2 inhibitors prevented thymidine production leading to misincorporation of uracil into DNA and replication stress. Overall, these results demonstrate a functional link between MTHFD2-dependent cancer metabolism and replication stress that can be exploited therapeutically with this new class of inhibitors.

Ceftazidime and cefepime antagonize 5-fluorouracil\u2019s effect in colon cancer cells

BackgroundDrug-drug interaction (DDI), which can occur at the pharmacokinetics and/or the pharmacodynamics (PD) levels, can increase or decrease the therapeutic or adverse response of a drug itself or a combination of drugs. Cancer patients often receive, along their antineoplastic agents, antibiotics such as ß-lactams to treat or prevent infection. Despite the narrow therapeutic indices of antibiotics and antineoplastic agents, data about their potential interaction are insufficient. 5-fluorouracil (5-FU), widely used against colon cancer, is known for its toxicity and large intra- and inter- individual variability. Therefore, knowledge about its interaction with antibiotics is crucial.MethodsIn this study, we evaluated at the PD levels, against HCT-116 colon cancer cells, DDI between 5-FU and several ß-lactams (ampicillin, benzypenicillin, piperacillin, meropenem, flucloxacillin, ceftazidime (CFT), and cefepime (CFP)), widely used in intensive care units. All drugs were tested at clinically achieved concentrations. MTT assay was used to measure the metabolic activity of the cells. Cell cycle profile and apoptosis induction were monitored, in HCT-116 and DLD-1 cells, using propidium iodide staining and Caspase-3/7 activity assay. The uptake of CFT and CFP by the cells was measured using LC-MS/MS method.ResultsOur data indicate that despite their limited uptake by the cells, CFT and CFP (two cephalosporins) antagonized significantly 5-FU-induced S-phase arrest (DLD-1 cells) and apoptosis induction (HCT-116 cells). Remarkably, while CFP did not affect the proliferation of colon cancer cells, CFT inhibited, at clinically relevant concentrations, the proliferation of DLD-1 cells via apoptosis induction, as evidenced by an increase in caspase 3/7 activation. Unexpectedly, 5-FU also antagonized CFT’s induced cell death in DLD-1 cells.ConclusionThis study shows that CFP and CFT have adverse effects on 5-FU’s action while CFT is a potent anticancer agent that inhibits DLD-1 cells by inducing apoptotic cell death. Further studies are needed to decipher the mechanism(s) responsible for CFT’s effects against colon cancer as well as the observed antagonism between CFT, CFP, and 5-FU with the ultimate aim of translating the findings to the clinical settings.

In-vitro Evaluation of Anti-microbial and Cytotoxic Activity of Artemisia judaica Leaves and Stem Extracts via Induction of Caspase Dependent Apoptosis

Abstract: Medicinal plants and herbs are commonly used in the world to treat various human disorders. Artemisia judaica is one of these herbal species that is commonly used in medicine due to its contents of many bioactive compounds such as; flavonoids, lactones, essential oil and sesqui-terpenoids. It was used in many traditional medicines as an anthelmintic, antispasmodic, anti-rheumatic, and antibacterial agent. In recent years, anti-bacterial and anti-cancer activity of medicinal herbs are highly investigated. The present study is focused on the anti-microbial and cytotoxic effect of methanolic extract of Artemisia judaica leaves and stem. The antimicrobial assay was done on different gram-positive and gram-negative bacteria and it was revealed that leaves and stem extracts possess high and moderate activity against Staphylococcus aureus (MIC 312.5μg/ml and 625μg/ml) and Proteus vulgaris (MIC 312.5μg/ml and 1250μg/ml) for leaves and stem, respectively. Extracts were screened against HepG2, HCT-116, MCF-7, A-549 and MRC-5 cancer cells and it was found that both extracts were active against all cell lines with highest selectivity and cytotoxic activity observed against HepG2 cells (IC50 = 3.38 and 6.84μg/ml, for leaves and stem respectively). Further mechanistic studies on HepG2 cells showed that both extracts resulted in S-phase arrest and induced apoptosis via activation of caspase-3, p53 and Bax Key words: Artemisia judaica, Antimicrobial, Anticancer, Apoptosis, Mechanism.

Mollugin induced oxidative DNA damage via up-regulating ROS that caused cell cycle arrest in hepatoma cells

Mollugin has been proven to have anti-tumor activity. However, its potential anti-tumor mechanism remains to be fully elaborated. Herein, we investigated the growth inhibition of HepG2 cells, as well as the anti-tumor effect of mollugin and its molecular mechanism on H22-tumor bearing mice. In vitro, mollugin was shown to have a strong inhibitory effect on HepG2 cells in a concentration-dependent manner. Mollugin induced S-phase arrest of HepG2 cells, and increased intracellular reactive oxygen species (ROS) levels. Comet assay demonstrated that mollugin induced DNA damage in HepG2 cells, as well as an increase in the expression of p-H2AX. In addition, mollugin induced changes in cyclin A2 and CDK2. However, the addition of antioxidant glutathione (GSH) was able to reverse the effect of mollugin. In vivo, mollugin significantly inhibited tumor growth and reduced the tendency of tumor volume growth in mice. The tumor cell density was found to be decreased in the administration group, and the content of ROS in the tumor tissue significantly increased. The expression of p-H2AX, cyclin A2 and CDK2 were consistent with in vitro results. Mollugin demonstrated anti-hepatocellular carcinoma activity in vitro and in vivo, and its anti-hepatocellular carcinoma activity was found to be related to DNA damage and cell cycle arrest induced by excessive ROS production in cells.

Reprogramming of nucleotide metabolism by interferon confers dependence on the replication stress response pathway in pancreatic cancer cells.

We determine that type I interferon (IFN) response biomarkers are enriched in a subset of pancreatic ductal adenocarcinoma (PDAC) tumors; however, actionable vulnerabilities associated with IFN signaling have not been systematically defined. Integration of a phosphoproteomic analysis and a chemical genomics synergy screen reveals that IFN activates the replication stress response kinase ataxia telangiectasia and Rad3-related protein (ATR) in PDAC cells and sensitizes them to ATR inhibitors. IFN triggers cell-cycle arrest in S-phase, which is accompanied by nucleotide pool insufficiency and nucleoside efflux. In combination with IFN, ATR inhibitors induce lethal DNA damage and downregulate nucleotide biosynthesis. ATR inhibition limits the growth of PDAC tumors in which IFN signaling is driven by stimulator of interferon genes (STING). These results identify a cross talk between IFN, DNA replication stress response networks, and nucleotide metabolism while providing the rationale for targeted therapeutic interventions that leverage IFN signaling in tumors.

Targeting the LSD1-G9a-ER Stress Pathway as a Novel Therapeutic Strategy for Esophageal Squamous Cell Carcinoma.

Despite recent advances in the management and treatment of esophageal squamous cell carcinoma (ESCC), the prognosis remains extremely poor, and current nonsurgical treatment options are limited. To identify new therapeutic targets, we screened a curated library of epigenetic compounds using a panel of cancer cell lines and found that coinhibiting the histone demethylase LSD1 and the histone methyltransferase G9a potently suppresses cell growth; similar results were obtained by knocking down both LSD1 and G9a expression. Importantly, we also found that inhibiting LSD1 and G9a significantly decreased tumor growth in a xenograft mouse model with ESCC cell lines. To examine the clinical relevance of these findings, we performed immunohistochemical analyses of microarray profiling data obtained from human esophageal squamous cancer tissues and found that both LSD1 and G9a are upregulated in cancer tissues compared to healthy tissues, and this increased expression was significantly correlated with poor prognosis. Mechanistically, we discovered that inhibiting LSD1 and G9a induces cell death via S-phase arrest and apoptosis, and cotargeting ER stress pathways increased this effect both in vitro and in vivo. Taken together, these findings provide compelling evidence that targeting LSD1, G9a, and ER stress-related pathways may serve as a viable therapeutic strategy for ESCC.