Nordamnacanthal, an anthraquinone extracted from the root of Morinda elliptica from Rubiaceae family has cytotoxic properties towards cancer cell lines and antitumor promoting activities. This study was conducted to determine the cytotoxic effect of nordamnacanthal towards MOLT-4 and MCF-7 cell lines. Nordamnacanthal was found to be more cytotoxic towards MOLT-4 than MCF-7 with the IC50 of 3.8 μg/ml and 54 μg/ml, respectively, as detected by using the trypan blue dye exclusion test. The nordamnacanthal-treated cells showed characteristics of apoptosis such as membrane blebbing, chromatin condensation and formation of apoptotic bodies as observed under an inverted light microscope. Fluorescence analysis of cell death using acridine orange and propidium iodide staining showed that the population of MOLT-4 and MCF-7 cells underwent apoptosis at the IC50 value was 32% and 30.4%, respectively. Cell cycle analysis by flow cytometry indicated that nordamnacanthal did arrest MCF-7 cells at the G2/M phase. For MOLT-4, no cell cycle arrest was observed. Bcl-2 and Bax were downregulated in nordamnacanthal-treated MCF-7 cells. On the other hand, expression of the proteins in MOLT-4 was not significantly different from the control. In conclusion, nordamnacanthal was more cytotoxic towards MOLT-4 than MCF-7 cell line. The compound induced apoptosis in both cell lines, but with G2/M arrest and the involvement of Bcl-2 and Bax only in MCF-7.
- Keywords
- cytotoxicita,
- MeSH
- Adenocarcinoma pathology MeSH
- Aldehydes chemistry therapeutic use MeSH
- Anthraquinones * chemistry therapeutic use MeSH
- Apoptosis * drug effects MeSH
- Cell Cycle drug effects MeSH
- Cytotoxins MeSH
- Down-Regulation MeSH
- Enzyme-Linked Immunosorbent Assay statistics & numerical data MeSH
- Fluorescent Dyes MeSH
- Microscopy, Fluorescence statistics & numerical data MeSH
- Inhibitory Concentration 50 MeSH
- G2 Phase Cell Cycle Checkpoints drug effects MeSH
- Plant Roots MeSH
- Cells, Cultured MeSH
- Humans MeSH
- Precursor T-Cell Lymphoblastic Leukemia-Lymphoma pathology MeSH
- MCF-7 Cells cytology drug effects MeSH
- Morinda MeSH
- Cell Line, Tumor * cytology drug effects MeSH
- Breast Neoplasms * pathology MeSH
- bcl-2-Associated X Protein MeSH
- Flow Cytometry statistics & numerical data MeSH
- Plant Extracts MeSH
- Statistics as Topic MeSH
- In Vitro Techniques MeSH
- Check Tag
- Humans MeSH
Microtubule dynamic is exceptionally sensitive to modulation by small-molecule ligands. Our previous work presented the preparation of microtubule-targeting estradiol dimer (ED) with anticancer activity. In the present study, we explore the effect of selected linkers on the biological activity of the dimer. The linkers were designed as five-atom chains with carbon, nitrogen or oxygen in their centre. In addition, the central nitrogen was modified by a benzyl group with hydroxy or methoxy substituents and one derivative possessed an extended linker length. Thirteen new dimers were subjected to cytotoxicity assay and cell cycle profiling. Dimers containing linker with benzyl moiety substituted with one or more methoxy groups and longer branched ones were found inactive, whereas other structures had comparable efficacy as the original ED (e.g. D1 with IC50 = 1.53 μM). Cell cycle analysis and immunofluorescence proved the interference of dimers with microtubule assembly and mitosis. The proposed in silico model and calculated binding free energy by the MM-PBSA method were closely correlated with in vitro tubulin assembly assay.
- MeSH
- Apoptosis MeSH
- Ethinyl Estradiol * chemistry pharmacology MeSH
- G2 Phase Cell Cycle Checkpoints drug effects MeSH
- Microtubules MeSH
- Tubulin Modulators * chemistry pharmacology MeSH
- Cell Line, Tumor MeSH
- Antineoplastic Agents * chemistry pharmacology MeSH
- Triazoles * chemistry pharmacology MeSH
- Tubulin * metabolism MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Glioblastoma multiforme (GBM) represents approximately 60% of all brain tumors in adults. This malignancy shows a high biological and genetic heterogeneity associated with exceptional aggressiveness, leading to a poor survival of patients. This review provides a summary of the basic biology of GBM cells with emphasis on cell cycle and cytoskeletal apparatus of these cells, in particular microtubules. Their involvement in the important oncosuppressive process called mitotic catastrophe will next be discussed along with select examples of microtubule-targeting agents, which are currently explored in this respect such as benzimidazole carbamate compounds. Select microtubule-targeting agents, in particular benzimidazole carbamates, induce G2/M cell cycle arrest and mitotic catastrophe in tumor cells including GBM, resulting in phenotypically variable cell fates such as mitotic death or mitotic slippage with subsequent cell demise or permanent arrest leading to senescence. Their effect is coupled with low toxicity in normal cells and not developed chemoresistance. Given the lack of efficient cytostatics or modern molecular target-specific compounds in the treatment of GBM, drugs inducing mitotic catastrophe might offer a new, efficient alternative to the existing clinical management of this at present incurable malignancy.
We investigated the effect of UVA-activated 8-methoxypsoralen (PUVA) on the cell line Karpas 299 derived from anaplastic large-cell lymphoma (ALCL) expressing chimeric fusion protein nucleophosmin-anaplastic lymphoma kinase (NPM/ALK). NPM/ALK activates phosphatidylinositol 3 kinase (PI3K)/Akt pathway responsible for the cell protection from apoptosis. We found that PUVA treatment first induced G2/M cell cycle arrest resulting in a decrease in the cell proliferation rate. The mitochondrial apoptosis was triggered immediately following PUVA treatment, as we judged from the unmasking of mitochondrial membrane antigen 7A6. However, the mitochondrial membrane depolarization was not observed and caspase-3 was only slightly activated. The late apoptotic events were lacking: neither translocation of phosphatidylserine to the outer side of plasma membrane nor DNA fragmentation occurred. We revealed that PUVA enhanced the expression of peroxiredoxin, stress protein endoplasmin and galectin-3. Galectin-3 has been shown to protect mitochondrial membrane integrity and prevent cytochrome c release thereby blocking the effector stage of apoptosis. We suggest that the elevated level of this protein following PUVA treatment acts in synergy with the constitutively expressed chimeric kinase NPM/ALK to block the apoptosis.
- MeSH
- Cell Line MeSH
- Cytochromes c metabolism MeSH
- Financing, Organized MeSH
- Phosphatidylinositol 3-Kinases metabolism MeSH
- G2 Phase drug effects MeSH
- Galectin 3 metabolism MeSH
- Caspase 3 metabolism MeSH
- Lymphoma, T-Cell metabolism pathology MeSH
- Membrane Glycoproteins metabolism MeSH
- Membrane Proteins metabolism MeSH
- Methoxsalen pharmacology MeSH
- Mitochondria metabolism MeSH
- Peroxidases metabolism MeSH
- Peroxiredoxins MeSH
- Cell Cycle Proteins drug effects MeSH
- Heat-Shock Proteins metabolism MeSH
- PUVA Therapy adverse effects MeSH
- Protein-Tyrosine Kinases metabolism MeSH
- Ultraviolet Rays MeSH
A total of 41 new triterpenoids were prepared from allobetulone, methyl betulonate, methyl oleanonate, and oleanonic acid to study their influence on cancer cells. Each 3-oxotriterpene was brominated at C2 and substituted with thiocyanate; subsequent cyclization with the appropriate ammonium salts gave N-substituted thiazoles. All compounds were tested for their in vitro cytotoxic activity on eight cancer cell lines and two non-cancer fibroblasts. 2-Bromoallobetulone (2 b) methyl 2-bromobetulonate (3 b), 2-bromooleanonic acid (5 b), and 2-thiocyanooleanonic acid (5 c) were best, with IC50 values less than 10 μm against CCRF-CEM cells (e.g., 3 b: IC50 =2.9 μm) as well as 2'-(diethylamino)olean-12(13)-eno[2,3-d]thiazole-28-oic acid (5 f, IC50 =9.7 μm) and 2'-(N-methylpiperazino)olean-12(13)-eno[2,3-d]thiazole-28-oic acid (5 k, IC50 =11.4 μm). Compound 5 c leads to the accumulation of cells in the G2 phase of the cell cycle and inhibits RNA and DNA synthesis significantly at 1×IC50 . The G2 /M cell-cycle arrest probably corresponds to the inhibition of DNA/RNA synthesis, similar to the mechanism of action of actinomycin D. Compound 5 c is new, active, and nontoxic; it is therefore the most promising compound in this series for future drug development. Methyl 2-bromobetulonate (3 b) and methyl 2-thiocyanometulonate (3 c) were found to inhibit nucleic acid synthesis only at 5×IC50 . We assume that in 3 b and 3 c (unlike in 5 c), DNA/RNA inhibition is a nonspecific event, and an unknown primary cytotoxic target is activated at 1×IC50 or lower concentration.
- MeSH
- Apoptosis drug effects MeSH
- G2 Phase Cell Cycle Checkpoints drug effects MeSH
- M Phase Cell Cycle Checkpoints drug effects MeSH
- Oleanolic Acid analogs & derivatives chemistry MeSH
- Humans MeSH
- Cell Line, Tumor MeSH
- Antineoplastic Agents chemical synthesis chemistry toxicity MeSH
- Drug Screening Assays, Antitumor MeSH
- Thiazoles chemical synthesis chemistry toxicity MeSH
- Triterpenes chemistry MeSH
- Structure-Activity Relationship MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
Oplopanax elatus (Nakai) Nakai has a long history of use as an ethnomedicine by the people living in eastern Asia. However, its bioactive constituents and cancer chemopreventive mechanisms are largely unknown. The aim of this study was to prepare O. elatus extracts, fractions, and single compounds and to investigate the herb's antiproliferative effects on colon cancer cells and the involved mechanisms of action. Two polyyne compounds were isolated from O. elatus, falcarindiol and oplopandiol. Based on our HPLC analysis, falcarindiol and oplopandiol are major constituents in the dichloromethane (CH2Cl2) fraction. For the HCT-116 cell line, the dichloromethane fraction showed significant effects. Furthermore, the IC50 for falcarindiol and oplopandiol was 1.7 µM and 15.5 µM, respectively. In the mechanistic study, after treatment with 5 µg/ml for 48 h, dichloromethane fraction induced cancer cell apoptosis by 36.5% (p < 0.01% vs. control of 3.9%). Under the same treatment condition, dichloromethane fraction caused cell cycle arrest at the G2/M phase by 32.6% (p < 0.01% vs. control of 23.4%), supported by upregulation of key cell cycle regulator cyclin A to 21.6% (p < 0.01% vs. control of 8.6%). Similar trends were observed by using cell line HT-29. Data from this study filled the gap between phytochemical components and the cancer chemoprevention of O. elatus. The dichloromethane fraction is a bioactive fraction, and falcarindiol is identified as an active constituent. The mechanisms involved in cancer chemoprevention by O. elatus were apoptosis induction and G2/M cell cycle arrest mediated by a key cell cycle regulator cyclin A.
AIMS: Chalcones, naturally occurring open-chain polyphenols abundant in plants, have demonstrated antiproliferative activity in several cancer cell lines. In the present study, the potential anticancer activity of two synthetic analogues named Ch1 and Ch2 in colon cancer cell line was investigated. MAIN METHODS: Antiproliferative activities of both synthetic analogues were assessed by Growth Inhibition Assay (MTT) and xCELLigence cell analysis. Apoptosis was assessed by annexin V/PI staining (early stage) or by DNA fragmentation (final stage). To study the cell death mechanism induced by tested substances, we assessed a series of assays including measurements of the caspase 3 activity, membrane mitochondrial potential (MMP) changes, reactive oxygen species (ROS) production by flow cytometry and expression of important apoptosis-related genes by realtime PCR. KEY FINDINGS: We found concentration and time-dependent cytotoxicity, inhibition of proliferation of Caco-2 cells after Ch1 and Ch2 treatment in parallel with G2/M phase cell cycle arrest and increased cell proportion in subG0/G1 population with annexin V positivity. We demonstrated that both Ch1 and Ch2 induced caspase-dependent cell death associated with increased ROS production, suppressed Bcl-2 and Bcl-xL and enhanced Bax expression. Treatment of Ch1 also suppressed α-, α1- and β5-tubulins, on the other hand Ch2 only suppressed α-tubulin expression. SIGNIFICANCE: Presented chalcones induce apoptosis by intrinsic pathways, and therefore may be an interesting strategy for cancer therapy.
- MeSH
- Apoptosis drug effects MeSH
- Cell Death drug effects MeSH
- Cell Division drug effects MeSH
- Caco-2 Cells MeSH
- Chalcones pharmacology MeSH
- DNA Fragmentation drug effects MeSH
- G2 Phase drug effects MeSH
- Caspase 3 metabolism MeSH
- Humans MeSH
- Membrane Potential, Mitochondrial drug effects MeSH
- Cell Line, Tumor MeSH
- Colonic Neoplasms pathology MeSH
- Antineoplastic Agents pharmacology MeSH
- Reactive Oxygen Species metabolism MeSH
- Gene Expression Regulation, Neoplastic drug effects MeSH
- Tubulin biosynthesis MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
BACKGROUND: Although it is clear that functional p53 is not required for radiation-induced G2 block, certain experimental findings suggest a role for p53 in this context. For instance, as we also confirm here, the maximum accumulation in the G2 compartment after X-ray exposure occurs much later in p53 mutants than in wild types. It remains to be seen, however, whether this difference is due to a longer block in the G2 phase itself. MATERIAL AND METHODS: We observed the movement of BrdU-labeled cells through G2 and M into G1. From an analysis of the fraction of labeled cells that entered the second posttreatment cell cycle, we were able to determine the absolute duration of the G2 and M phases in unirradiated and irradiated cells. RESULTS: Our experiments with four cell lines, two melanomas and two squamous carcinomas, showed that the radiation-induced delay of transition through the G2 and M phases did not correlate with p53 status. CONCLUSION: We conclude that looking at the accumulation of cells in the G2 compartment alone is misleading when differences in the G2 block are investigated and that the G2 block itself is indeed independent of functional p53.
- MeSH
- Radiation Dosage MeSH
- Neoplasms, Experimental metabolism pathology radiotherapy MeSH
- G2 Phase Cell Cycle Checkpoints radiation effects MeSH
- Humans MeSH
- Cell Line, Tumor MeSH
- Tumor Suppressor Protein p53 metabolism MeSH
- Cell Survival radiation effects MeSH
- Dose-Response Relationship, Radiation MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
While p53-dependent apoptosis is triggered by combination of methyltransferase inhibitor decitabine (DAC) and histone deacetylase inhibitor suberoylanilide hydroxamic acid (SAHA) in leukemic cell line CML-T1, reactive oxygen species (ROS) generation as well as survivin and Bcl-2 deregulation participated in DAC + SAHA-induced apoptosis in p53-deficient HL-60 cell line. Moreover, decrease of survivin expression level is accompanied by its delocalization from centromere-related position in mitotic cells suggesting that both antiapoptotic and cell cycle regulation roles of survivin are affected by DAC + SAHA action. Addition of subtoxic concentration of all-trans-retinoic acid (ATRA) increases the efficiency of DAC + SAHA combination on viability, apoptosis induction, and ROS generation in HL-60 cells but has no effect in CML-T1 cell line. Peripheral blood lymphocytes from healthy donors showed no damage induced by DAC + SAHA + ATRA combination. Therefore, combination of ATRA with DAC and SAHA represents promising tool for therapy of leukemic disease with nonfunctional p53 signalization.
- MeSH
- Apoptosis drug effects MeSH
- Azacitidine analogs & derivatives toxicity MeSH
- Down-Regulation drug effects MeSH
- HL-60 Cells MeSH
- Inhibitor of Apoptosis Proteins metabolism MeSH
- G2 Phase Cell Cycle Checkpoints drug effects MeSH
- M Phase Cell Cycle Checkpoints drug effects MeSH
- Hydroxamic Acids toxicity MeSH
- Humans MeSH
- Lymphocytes cytology drug effects immunology MeSH
- Cell Line, Tumor MeSH
- Tumor Suppressor Protein p53 deficiency genetics MeSH
- Antimetabolites, Antineoplastic pharmacology MeSH
- Reactive Oxygen Species metabolism MeSH
- Drug Synergism MeSH
- Tretinoin pharmacology MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
The phenolic compounds of methanolic extracts of Salvia pomifera and Salvia fruticosa were identified by liquid chromatography tandem mass spectrometry. Carnosic acid and its metabolite carnosol were the most abundant terpene phenolic compounds of S. fruticosa, while they were completely absent in S. pomifera. The main terpene phenolic constituent of S. pomifera was 12-O-methylcarnosic acid and its mass/mass fragmentation pathway was explained. The detailed mechanism of carnosic acid oxidation to carnosol was suggested. The effects of Salvia extracts and/or carnosic acid, the main diterpene phenolic component of S. fruticosa, on the proliferation and cell cycle of two melanoma cell lines (A375, Mel JuSo) and human fibroblast cell line (HFF) were investigated by MTT assay, PI-exclusion assay and flow cytometry cell cycle analysis. Extract of S. fruticosa more efficiently than S. pomifera extract reduced the proliferation of the human melanoma cells. Carnosic acid showed the most significant effect. The first evidence that carnosic acid affects microtubule dynamics and arrests the cell cycle in the G2/M phase was provided. Collectively, our results demonstrate that these two Salvia species are plants of medicinal interest with perspective for further investigation. Carnosic acid could be the compound responsible for the biological activities of S. fruticosa extracts.
- MeSH
- Cell Line MeSH
- Abietanes chemistry isolation & purification pharmacology MeSH
- Epithelial Cells drug effects pathology MeSH
- Phenols chemistry isolation & purification pharmacology MeSH
- Fibroblasts cytology drug effects MeSH
- Inhibitory Concentration 50 MeSH
- G2 Phase Cell Cycle Checkpoints drug effects MeSH
- Humans MeSH
- Methanol chemistry MeSH
- Cell Line, Tumor MeSH
- Plant Components, Aerial chemistry MeSH
- Oxidation-Reduction MeSH
- Cell Proliferation drug effects MeSH
- Antineoplastic Agents chemistry isolation & purification pharmacology MeSH
- Plant Extracts chemistry MeSH
- Solvents chemistry MeSH
- Salvia chemistry MeSH
- Cell Survival drug effects MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
