Assessment of cytotoxicity and induction of apoptosis by cytolysin-A in MCF-7 human breast cancer cell line.

A protein called cytolysin A or ClyA, encoded by certain bacteria species, can cause cytotoxicity. Although the ClyA protein is not typically expressed at detectable levels in most E. coli strains, here it was successfully overproduced and purified by cloning the structural gene into a hns mutant strain. The cytotoxicity of the purified cytolysin was assessed on two MCF-7 cancer cell lines and HDF normal cell line using the MTT assay. Flow cytometry was employed to examine the cytolysin's ability to induce apoptosis in cancer cells. In addition, a Western blot analysis was carried out to evaluate the expression levels of P53, Bcl2, and Bax proteins. The results revealed that cytolysin exhibited dose-dependent and time-dependent toxicity towards cancer cells, while showing minimal toxicity against normal cells, indicating its selective action against cancer cells. Cytolysin had an IC50 value of 3.29 µg/ml against MCF-7 cancer cells and 12.6 µg/ml against HDF normal cells. Flow cytometry results further demonstrated that cytolysin induced apoptosis in cancer cells, evidenced by increased expression of p53 and BCL2, as well as decreased in Bax, in gene and protein levels. These findings underscore the potential of cytolysin as a targeted therapy for cancer, highlighting its selective cytotoxic effect on cancer cells. Keywords: Cytolysin-A, Breast cancer, Cytotoxicity, Flowcytometry, Western blotting

Benzyl isothiocyanate (BITC), a dietary cancer chemopreventive agent, causes apoptosis in MDA-MB-231 and MCF-7 human breast cancer cells, but the mechanism of cell death is not fully understood. We now demonstrate that the BITC-induced apoptosis in human breast cancer cells is initiated by reactive oxygen species (ROS) due to inhibition of complex III of the mitochondrial respiratory chain. The BITC-induced ROS production and apoptosis were significantly inhibited by overexpression of catalase and Cu,Zn-superoxide dismutase and pharmacological inhibition of the mitochondrial respiratory chain. The mitochondrial DNA-deficient Rho-0 variant of MDA-MB-231 cells was nearly completely resistant to BITC-mediated ROS generation and apoptosis. The Rho-0 MDA-MB-231 cells also resisted BITC-mediated mitochondrial translocation (activation) of Bax. Biochemical assays revealed inhibition of complex III activity in BITC-treated MDA-MB-231 cells as early as at 1 h of treatment. The BITC treatment caused activation of c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK), which function upstream of Bax activation in apoptotic response to various stimuli. Pharmacological inhibition of both JNK and p38 MAPK conferred partial yet significant protection against BITC-induced apoptosis. Activation of JNK and p38 MAPK resulting from BITC exposure was abolished by overexpression of catalase. The BITC-mediated conformational change of Bax was markedly suppressed by ectopic expression of catalytically inactive mutant of JNK kinase 2 (JNKK2(AA)). Interestingly, a normal human mammary epithelial cell line was resistant to BITC-mediated ROS generation, JNK/p38 MAPK activation, and apoptosis. In conclusion, the present study indicates that the BITC-induced apoptosis in human breast cancer cells is initiated by mitochondria-derived ROS.

Mitochondrial dysfunction of cancer cells includes increased aerobic glycolysis, elevated levels of ROS, decreased apoptosis, and resistance to chemotherapeutic agents. We hypothesized that the introduction of normal mitochondria into cancer cells might restore mitochondrial function and inhibit cancer cell growth, and reverse chemoresistance. First, in the present study, we tested if mitochondria of immortalized, untransformed mammary epithelial MCF-12A cells could enter into human cancer cell lines. Second, if introducing normal mitochondria into cancer cells would inhibit proliferation. And third, would the addition of normal mitochondria increase the sensitivity of human breast cancer MCF-7 cells to chemotherapy. We found that JC-1-stained mitochondria of immortalized, untransformed mammary epithelial MCF-12A cells can enter into the cancer cell lines MCF-7, MDA-MB-231, and NCI/ADR-Res, but cannot enter immortalized, untransformed MCF-12A cells. The normal mitochondria from immortalized, untransformed MCF-12A cells suppressed the proliferation of MCF-7 and NCI/ADR-Res cells in a dose-dependent pattern, but did not affect the proliferation of immortalized, untransformed MCF-12A cells. The normal mitochondria from immortalized, untransformed MCF-12A cells increased the sensitivity of human breast cancer MCF-7 cells to doxorubicin, Abraxane, and carboplatin. In conclusion, the introduction of normal mammary mitochondria into human breast cancer cells inhibits cancer cell proliferation and increases the sensitivity of the MCF-7 human breast cancer cell line to doxorubicin, Abraxane, and carboplatin. These results support the role of mitochondrial dysfunction in cancer and suggest the possible use of targeted mitochondria for cancer therapeutics.

Objective: To study the effects of Akt inhibitor MK2206 on the proliferation and apoptosis of human breast cancer cell lines T47D and MDA-MB-231, and to elucidate the possible mechanism of such effects. Methods: Human breast cancer T47D and MDA-MB-231 cells were treated with different concentrations of MK2206, respectively. The inhibitory effect of MK2206 on cell proliferation was examined by MTT assay. The apoptosis rates of T47D and MDA-MB-231 cells induced by MK2206 were detected by flow cytometry (FCM). The expression levels of caspase-3, poly (ADP-ribose) polymerase (PARP), bcl-2, bax, phosphorate-AKT (p-AKT) and total Akt (T-Akt) proteins were detected by Western blotting. Results: The proliferation of T47D and MDA-MB-231 cells were inhibited after treatment with MK2206 at 0.1, 1, 10 and 100 nmol/L for 24 h, respectively. The apoptosis rates of T47D and MDA-MB-231 cells were increased induced by MK2206. The expression levels of caspase-3, PARP and bax proteins were up-regulated, while the expression levels of bcl-2 and p-Akt proteins were down-regulated. All of these effects occurred in a dose-dependent manner. MK2206 had no significant effect on the expression of T-Akt. Conclusion: MK2206 can inhibit the proliferation and induce the apoptosis in human breast cancer cell lines T47D and MDA-MB-231. These effects may be related with the down-regulation of p-AKT and the inhibition of PI3K-Akt signaling pathway. DOI:10.3781/j.issn.1000-7431.2011.02.008

The aim of this research was to evaluate the anticancer properties of methanolic extracts from inner bark and wood of jabon (Anthocephalus cadamba). The extracts were investigated in vitro bioassay for its possible antiproliferative activities on human MCF7 breast cancer cell line and HeLa cervical adenocarcinoma cell lines. The cell viability were assessed using microculture tetrazolium technique (MTT) colorimetric assay. The results showed that inner bark extract exhibited higher antiproliferative activity on MCF7 cancer cell line (IC50 91 μg ml-1) than wood extract (IC50 312 μg ml-1). But, antiproliferative activity of inner bark extract on HeLa cell lines was higher (IC50 5 μg ml-1). The inner bark extract is potential to be developed as anti cancer agent in cervical adenocarcinoma cancer therapy because moresecure against Vero normal cells (IC50 288 μg ml-1). Whereas compounds such as phenolic and fatty acid contribute to high antiproliferative activities of inner bark extract. The qualitative analysis detect the extracts containing flavonoids, triterpenoids, saphonin which are thought to contribute to the high antiproliferative activities of this extract. Keywords : Anthocephalus cadamba, antiproliferative activity, human MCF7 breast cancer cell line, HeLa cervical adenocarcinoma cell lines, Vero normal cell lines

Breast cancer is the most prevalent cancer and the leading cause of cancer-associated mortalities among women worldwide today. Accumulating evidence suggested that miR-372 may serve important roles in the initiation and development of various human cancers. However, the role of miR-372 in breast cancer remains unknown. The present study demonstrated that the expression level of miR-372 in human breast cancer tissues and cell lines is significantly reduced compared with normal breast tissues cell lines. Furthermore, results of functional assays indicated that miR-372 inhibits cell proliferation and induces apoptosis in the MCF-7 human breast cancer cell line. E2F1 was identified as a direct functional target of miR-372 in breast cancer. In conclusion, the findings revealed that miR-372 may have the potential to act as a novel molecule for the diagnosis and therapy of patients with breast cancer.

The present paper focused on antioxidant and cytotoxicity assessment of crude and total saponin fraction of Chlorophytum borivilianum as an important medicinal plant. In this study, three different antioxidant activities (2,2-diphenyl-1-picrylhydrazyl radical scavenging (DPPH), ferrous ion chelating (FIC), and β-carotene bleaching (BCB) activity) of crude extract and total saponin fraction of C. borivilianum tubers were performed. Crude extract was found to possess higher free radical scavenging activity (ascorbic acid equivalents 2578 ± 111 mg AA/100 g) and bleaching activity (IC50 = 0.7 mg mL−1), while total saponin fraction displayed higher ferrous ion chelating (EC50 = 1 mg mL−1). Cytotoxicity evaluation of crude extract and total saponin fraction against MCF-7, PC3, and HCT-116 cancer cell lines using 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide (MTT) cell viability assay indicated a higher cytotoxicity activity of the crude extract than the total saponin fraction on all cell lines, being most effective and selective on MCF-7 human breast cancer cell line.

The development of novel chemical entities, novel pharmaceuticals, and novel drug leads frequently involves the use of natural sources. One of the most well-known plants, Pandanus odoratissimus (Pandanaceae), is native to Malaysia and has long been used in Ayurvedic medicine to treat a variety of illnesses. The purpose of this study is to determine P odoratissimus's antioxidant and anticancer properties against MCF-7 human breast cancer cell lines. The 2, 2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging activity was used to assess antioxidant activity, and among the other extracts, core exhibited the highest antioxidant value. An MTT assay was used to assess the effects of plant components on MCF-7 cell lines (3-(4, 5-dimethylthiazolyl)-2, 5-diphenyl-tetrazolium bromide). The concentration of samples was shown to considerably boost the antioxidant activities in P. odoratissimus extracts. Our findings demonstrated that, after receiving treatment for 72 hours, sample treated cancer cells experienced a decline in cell viability. The IC50 value of 88.00 mg/ml indicates that the extracts have a poor cytotoxic impact and have no promise as anticancer agents against MCF-7, which is why it’s killing efficiency in cancer cells is regarded as lower.

Stem cell-based therapies hold significant potential for cancer treatment due to their unique properties, including migration toward tumor niche, secretion of bioactive molecules, and immunosuppression. Mesenchymal stem cells (MSCs) from adult tissues can inhibit tumor progression, angiogenesis, and apoptosis of cancer cells. We have previously reported the isolation and characterization of placenta-derived decidua basalis mesenchymal stem cells (DBMSCs), which demonstrated higher levels of pro-migratory and anti-apoptotic genes, indicating potential anti-cancer effects. In this study, we analyzed the anti-cancer effects of DBMSCs on human breast cancer cell lines MDA231 and MCF7, with MCF 10A used as control. We also investigated how these cancer cells lines affect the functional competence of DBMSCs. By co-culturing DBMSCs with cancer cells, we analyzed changes in functions of both cell types, as well as alterations in their genomic and proteomic profile. Our results showed that treatment with DBMSCs significantly reduced the functionality of MDA231 and MCF7 cells, while MCF 10A cells remained unaffected. DBMSC treatment decreased epithelial-to-mesenchymal transition (EMT)-related protein levels in MDA231 cells and modulated expression of other cancer-related genes in MDA231 and MCF7 cells. Although cancer cells reduced DBMSC proliferation, they increased their expression of anti-apoptotic genes. These findings suggest that DBMSCs can inhibit EMT-related proteins and reduce the invasive characteristics of MDA231 and MCF7 breast cancer cells, highlighting their potential as candidates for cell-based cancer therapies.

We report here that the antiestrogen tamoxifen (TAM) induces cell death in human breast cancer cell line MCF-7. We assessed the type of cell death induced by TAM in this breast cancer cell line on the basis of morphological and biochemical characteristics. Dying cells showed morphological characteristics of apoptosis, such as chromatin condensation and nuclear disintegration. DNA isolated from these cells revealed a pattern of distinctive DNA bands on agarose gel. The DNA fragmentation in MCF-7 cells induced by TAM could also be detected by terminal deoxynucleotidyl transferase-mediated dUTP-biotin end labeling. Northern blot hybridization revealed a substantial increase in the amounts of TRPM-2 and TGF-beta 1 mRNAs in MCF-7 cells after treatment with TAM. In contrast, the mRNA level of the estrogen-induced pS2 gene was strongly suppressed. The biological activity of TGF-beta was increased at least fourfold in the media from MCF-7 cells treated with TAM. The results presented in this study suggest that TAM induces apoptosis of MCF-7 cells and it may be mediated by the secretion of active TGF-beta.

Terfezia claveryi is a species that belongs to the genera of Terfeziaceae or desert truffles, which is a family of truffles. In the present study, silver nanoparticles were synthesized from aqueous extract of T. claveryi which are in the range of 25 to 60 nm. The synthesized nanoparticles were characterized by ultraviolet-visible (UV-Vis) spectroscopy, fourier transform infrared spectroscopy (FT-IR), X-ray diffraction (XRD), scanning electron microscopy (FESEM) and transmission electron microscopy (TEM). The effect of the silver nanoparticles on human breast cancer cell line has been tested. Peak absorption was recorded at 440 nm in UV-Vis spectra of silver nanoparticles. The XRD data reports that the silver nanoparticles are crystalline in nature and have face centered cubic geometry. FESEM showed the size range of synthesized silver nanoparticles as 25 to 50 nm. The TEM image represents that the majority of silver nanoparticles are in spherical shape with sizes ranging between 40 and 60 nm. The aim of the present study was to report for the first time fruit mediated synthesis of silver nanoparticles using the extract of T. claveryi and showed remarkable cytotoxicity activity against human breast MCF-7 cancer cell line. Key words: Silver nanoparticles, Terfezia claveryi, fourier transform infrared spectroscopy (FT-IR), X-ray diffraction (XRD), scanning electron microscopy (FESEM), transmission electron microscopy (TEM), MCF-7 cancer cell line.

Breast and cervical cancers are the most common gender-specific cancers affecting women worldwide. In this investigation, we highlighted the synthesis, VEGFR-2 and p38α MAPK inhibitory activity of new series of fluorinated coumarin-based derivatives featuring a variety of bioactive chemical moieties attached or fused to the coumarin nucleus at the 3 and/or 4 position. The bioactive inhibitors were further assessed for their anti-proliferative effect against human MCF-7 breast cancer and HeLa cervical cancer cell lines, respectively. Most of the tested compounds showed potent preferential inhibition effects against human VEGFR-2 and remarkable anticancer activities in the human breast cancer cell line MCF-7. Compounds 29, 24, and 2 displayed the highest inhibitory activity against VEGFR-2 (94% inhibition) and they were the most potent anticancer agents toward MCF-7 cancer cells with IC50 values of 7.90, 8.28, and 8.30 μg/mL, respectively. Compound 13 inhibited p38α MAPK phosphorylation with a significant reduction in % cell viability against HeLa cancer cells at 10 and 30 µM. Docking experiments carried out on VEGFR-2 and p38 MAPK crystallographic structures revealed that the active compounds bind to the active sites through H-bonds, arene-cation, and hydrophobic π-π interactions. QSAR analysis demonstrated considerable correlation coefficient (R2 = 0.76969) and root mean square error (RMSE = 0.10446) values. Also, the residual values between the experimental pIC50 and predicted pIC50 are very close, indicating the reliability of the established QSAR model.

Some new complexes of mefenamic acid with potentially interesting biological activity are described. The complexes of mefenamic acid [Mn(mef)2(H2O)2], 1, [Co(mef)2(H2O)2], 2, [Ni(mef)2(H2O)2], 3, [Cu(mef)2(H2O)]2, 4 and [Zn(mef)2], 5, were prepared by the reaction of mefenamic acid, a potent anti-inflammatory drug with metal salts. Optical and infrared spectral data of these new complexes are reported. Monomeric six-coordinated species were isolated in the solid state for Mn(II), Ni(II) and Co(II), dimeric five-coordinated for Cu(II) and monomeric four-coordinated for Zn(II). In DMF or CHCl3 solution the coordination number is retained and the coordinated molecules of water are replaced by solvent molecules. The anti-oxidant properties of the complexes were evaluated using the 1,1-diphenyl-2-picrylhydrazyl, DPPH, free radical scavenging assay. The scavenging activities of the complexes were measured and compared with those of the free drug and vitamin C. We have explored their ability to inhibit soybean lipoxygenase, β-glucuronidase and trypsin- induced proteolysis. The complex [Mn(mef)2(H2O)2] exhibits the highest antioxidant activity and the highest inhibitory effect against the soybean lipogygenase (LOX), properties that are not demonstrated by mefenamic acid. Their inhibitory effects on rat paw edema induced by Carrageenan was studied and compared with those of mefenamic acid. The complex [Zn(mef)2] exhibited a strong inhibitory effect at 0.1 mmol/Kg B.W. (81.5 ± 1.3% inhibition), superior to the inhibition induced by mefenamic acid at the same dose (61.5 ± 2.3% inhibition). Mefenamic acid and its metal complexes have been evaluated for antiproliferative activity in vitro against the cells of three human cancer cell lines: MCF-7 (human breast cancer cell line), T24 (bladder cancer cell line), A-549 (non-small cell lung carcinoma) and a mouse fibroblast L-929 cell line. The copper(II) complex displays against T24, MCF-7 and L-929 cancer cell lines, IC50 values in a μM range similar to that of the antitumor drug cis-platin and they are considered for further stages of screening in vitro and/or in vivo as agents with potential antitumor activity.

To evaluate the effect of schedule on the interaction of etoposide with paclitaxel in vitro against the A549 human lung cancer cell line and the MDA-231 and MCF-7 human breast cancer cell lines. Exposure schedules that were 24-h concurrent, 24-h sequential, and sequential 24-h with a 24-h intervening drug-free period were quantitatively evaluated by the use of the median-effect principle and the combination index. The clonogenic assay was used to assess cytotoxicity, and calculations were done with computer software. Concurrent exposures were less than additive in two of the three cell lines tested. Sequential 24-hour and sequential 24-h with an intervening 24-h drug-free period showed synergism at high effect levels in all three cell lines. Similar synergistic interactions were found when either agent was administered first. These results show a schedule-dependent cytotoxic interaction between etoposide and paclitaxel in the human lung and breast cancer cell lines evaluated, with optimal synergism occurring with sequential, but not with concurrent, treatment.

Objective To study the effects of tumor-associated calcium signal transducer-2 (TROP-2) gene small interfering RNA(siRNA) on adhesion and invasion of human breast cancer cell. Methods Real time PCR was used to evaluate the TROP-2 mRNA of seven human breast cancer cell lines Bcap-37 ,LCC1 ,MCF-7 ,MDA-MB-231,MDA-MB-435, MDA-MB-468 ,and ZR75-1. The cell line of TROP-2 highest expression was transfected with different dose of TROP-2 siRNA. The expression of TROP-2 mRNA and protein were determined by Real-time quantitative PCR and immumoflurescence method. The cell adhesion was evaluated by MTT assay,and invasion was exmined by hoyden chamber,respectively. Results Cell line MCF-7 showed the highest elevation of TROP-2 mRNA in seven breast cancer cell lines. The results from real-time quantitative PCR and immumoflurescence method showed that TROP-2 mRNA and protein reduced in time-and dose-dependent manners( P ＜ 0.01 ;P ＜ 0.01 ). The adhesive rate of siRNA groups(5 nM,10 nM,and 20 nM)was(52.9 +2.5)％ ,(25.6 ±2.3)％, ( 12.8 +2.2)％ (P ＜0.01 ) ,respectively.The transwell results showed that the invasion cells was(78 ± 17), (39 ± 15), ( 19 ± 16), ( 136 +25 ) and( 139 ±21 )in different groups(5,10,20 nM siRNA,and controls) ,respectively(P ＜0.01). Conclusion TROP-2 gene might play an important role in adhesion and invasion of human breast cancer cell. siRNA targeted TROP-2 could effectively inhibit adhesion and invasion of human breast cancer cell. Key words: Breast Neoplasms; Tumor-associated calcium signal transducer-2