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Apoptosis

"One of the mechanisms by which CELL DEATH occurs (compare with NECROSIS and AUTOPHAGOCYTOSIS). Apoptosis is the mechanism responsible for the physiological deletion of cells and appears to be intrinsically programmed. It is characterized by distinctive morphologic changes in the nucleus and cytoplasm, chromatin cleavage at regularly spaced sites, and the endonucleolytic cleavage of genomic DNA; ( DNA FRAGMENTATION); at internucleosomal sites. This mode of cell death serves as a balance to mitosis in regulating the size of animal tissues and in mediating pathologic processes associated with tumor growth." (Source: MeSH)

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Latest Research Publications

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Latest Research Publications

This list of publications is regularly updated (Source: PubMed).

Jovanovic KK, Gligorijevic N, Gaur R, et al.
Anticancer activity of two ruthenium(II)-DMSO-chalcone complexes: Comparison of cytotoxic, pro-apoptotic and antimetastatic potential.
J BUON. 2016 Mar-Apr; 21(2):482-90 [PubMed] Related Publications
PURPOSE: Recently, we reported the synthesis and characterization of two complexes of general formula cis-[Ru(S-DMSO)3(R-CO-CH=CH-R')Cl] (R = 2-hydroxyphenyl for both, R' = thiophene (1), 3-methyl thiophene (2)) that showed remarkable topoisomerase II inhibition and strong binding with DNA. The aim of this study was the investigation of cytotoxic properties of these complexes against a panel of human tumor cell lines, with elucidation of their anticancer mechanisms in HeLa cells.
METHODS: Characterization of anticancer activity of the investigated ruthenium complexes 1 and 2 included analysis of cytotoxicity by MTT assay. Cell cycle phase disruption of HeLa cells treated with complexes 1 and 2 was analyzed by flow cytometry after propidium iodide (PI) staining. Annexin V-FITC/PI double staining and further flow cytometry analysis and acridine orange (AO)/ethidium bromide (EB) double staining and fluorescent microscopy were used to determine the apoptotic potential of the investigated ruthenium complexes. The inhibitory effect on gelatinases (MMP-2 and MMP-9) as an indication of possible antimetastatic potential was also analyzed using gelatine zymography.
RESULTS: The 50% cell growth inhibition (IC50) values of the investigated complexes ranged between 22.9 and 76.8 μM, with complex 2 being more cytotoxic. Both complexes induced G2 phase cell cycle arrest and apoptosis in HeLa cells. Inhibitory effect of complex 2 on MMP-2 activity was detected.
CONCLUSIONS: This work revealed the potential of the investigated Ru(II)-DMSO-chalcone complexes as anticancer agents with cytotoxic and pro-apoptotic activity and indicated complex 2 as leading compound for further chemical modifications and anticancer research.

Wang BZ, Yu ZG
The combination use of 1-O-acetylbritannilactone (ABL) and gemcitabine inhibits cell growth and induces cell apoptosis in lung adenocarcinoma cells.
Pharmazie. 2016; 71(4):213-7 [PubMed] Related Publications
1-O-acetylbritannilactone (ABL), a natural chemical component obtained from Chinese traditional medicine, Inula britannica, has been demonstrated to have anticancer activities. In the present study, we evaluated the anti-proliferative and the pro-apoptotic abilities of ABL alone or in combination with gemcitabine in human NSCLC cell line. A549 cells were treated, in vitro, with ABL, gemcitabine, and the combination of ABL and gemcitabine for 72 h. Our results showed ABL and gemcitabine inhibited cell growth and induced apoptosis of A549 cells. These effects after the combination of ABL and gemcitabine were superior to those of each alone. Furthermore, signal transduction analysis revealed NF-κB expression was significantly decreased by ABL and the combination treatment. IκBα and Bax levels were up regulated whereas Bcl-2 was substantially downregulated after all treatments. Our findings suggest that ABL combined with gemcitabine elicits a potent apoptosis of lung cancer cell and hence ABL has the potential to be developed as a chemotherapeutic agent.

Shintia C, Endang H, Diani K
Assessment of pathological response to neoadjuvant chemotherapy in locally advanced breast cancer using the Miller-Payne system and TUNEL.
Malays J Pathol. 2016; 38(1):25-32 [PubMed] Related Publications
BACKGROUND: Responses to neoadjuvant (before surgery) chemotherapy in locally advanced breast cancer (LABC) consist of clinical and pathological responses. Evaluating chemotherapy response is essential to predict survival rate and guide future chemotherapy. Until now, the evaluation of pathological response mainly involves quantitative assessment and is often inconsistent with clinical response. We explored the evaluation of pathological responses by both quantitative and qualitative methods, i.e. by evaluating the cellularity of tumour cells and the percentage of apoptosis.
MATERIALS AND METHOD: A cross-sectional analytical retrospective study was conducted on tissue of LABC diagnosed between 2010 and 2014 at the Department of Anatomical Pathology, Faculty of Medicine, Universitas Indonesia Cipto Mangunkusumo Hospital and Division of Surgical Oncology, Cipto Mangunkusumo Hospital. Biopsy and resection specimens were compared to evaluate reduction in cellularity, which were subsequently categorized into stages of Miller-Payne (MP) classification. The resection specimens were stained with TUNEL and the percentage of apoptosis was calculated. Reduction in cellularity between biopsy and mastectomy specimens with TUNEL staining is evaluated as a modification of the MP method.
RESULTS: We found no association between clinical responses with percentage of apoptosis, MP pathological responses and modified MP. There was a correlation between the dead cell evaluated by MP and by modified MP (p=0.000).
CONCLUSION: Modified MP increases the degree or grading of pathological responses, but it does not improve the correlation with clinical response.

Li HG, Chen JX, Xiong JH, Zhu JW
Myricetin exhibits anti-glioma potential by inducing mitochondrial-mediated apoptosis, cell cycle arrest, inhibition of cell migration and ROS generation.
J BUON. 2016 Jan-Feb; 21(1):182-90 [PubMed] Related Publications
PURPOSE: To study the antiproliferative effects of myricetin in human glioma U251 cells together with assessing its effects on cell cycle, apoptosis, apoptosis-related proteins, reactive oxygen species (ROS) generation and cell migration.
METHODS: Cell viability of human glioma cells after myricetin treatment was assessed by MTT assay. Phase-contrast and confocal fluorescence microscopies were used to assess the morphological changes that occured in these cells following myricetin treatment. Flow cytometry using propidium iodide (PI) and Annexin-V FITC as probes was employed to evaluate the effects on cell cycle arrest and apoptosis induction, respectively. The effect of myricetin on intracellular ROS production was measured by flow cytometry with a fluorescent probe CM-DCFH2-DA.
RESULTS: Myricetin induced a dose-dependent as well as time-dependent growth inhibitory effect in U251 human glioma cells. Myricetin treatment resulted in U251 cells detachment from adjacent cells making clusters of cells floating in the medium. Detached cells had irregular shape and incapable to maintain their membranes intact. Apoptotic cell death was induced by myricetin treatment as witnessed by fluorescence microscopy. The percentage of early and late apoptotic cells increased from 0.41% and 8.2% to 23.1% and 10.2%, 25.2% and 19.4%, to finally 36.2% and 28.4% after treatment with 15 μM, 60 μM and 120 μM of myricetin, respectively. We also observed a dose-dependent increase in Bax and Bad levels and a dose-dependent decrease in Bcl-2 and Bcl-xl expression levels following myricetin treatment. Cell cycle arrest in G2/M phase of the cell cycle was also induced by the drug treatment. A concentration-dependent ROS generation was also witnessed and a 3-fold increase of ROS production was seen after 60 μM myricetin treatment.
CONCLUSION: Myricetin exerts anticancer effects in U251 human glioma cells by inducing mitochondrial-mediated apoptosis, G2/M phase cell cycle arrest, ROS generation and inhibition of cell migration.

Li QC, Liang Y, Tian Y, Hu GR
Arctigenin induces apoptosis in colon cancer cells through ROS/p38MAPK pathway.
J BUON. 2016 Jan-Feb; 21(1):87-94 [PubMed] Related Publications
PURPOSE: In the current study the antiproliferative effect of arctigenin, plant lignin, was evaluated on human colon cancer cell line HT-29. Furthermore, attempts were made to explore the signaling mechanism which may be responsible for its effect.
METHODS: Cell growth inhibition was assessed by MTT and LDH assays. Flow cytometric analysis was performed to determine cell arrest in the cell cycle phase and apoptosis. Furthermore, to confirm the apoptotic activity of arctigenin, caspase-9 and -3 activities analysis was performed. The levels of reactive oxygen species (ROS) and p38 mitogen activated protein kinase (MAPK) were investigated to determine their role in inducing apoptosis in arctigenin-treated HT-29 colon cancer cell line.
RESULTS: MTT and LDH results demonstrated significant cell growth inhibitory effect of arctigenin on HT-29 cells in a dose-dependent manner. Furthermore, increase in cell number arrested at G2/M phase was observed in flow cytometric analysis upon arctigenin treatment. In addition, arctigenin increased the apoptotic ratio in a dose-dependent manner. The involvement of intrinsic apoptotic pathway was indicated by the activation of caspase-9 and -3. Moreover, increased ROS production, activation of p38 MAPK and changes in mitochondrial membrane potential (ΔΨm) also revealed the role of intrinsic apoptotic signaling pathway in cell growth inhibition after arctigenin exposure.
CONCLUSION: Arctigenin induces apoptosis in HT-29 colon cancer cells by regulating ROS and p38 MAPK pathways.

Zhu YQ, Wang BY, Wu F, et al.
Influence of Tanshinone IIA on the Apoptosis of Human Esophageal Ec-109 Cells.
Nat Prod Commun. 2016; 11(1):17-9 [PubMed] Related Publications
The induced-apoptosis effect and mechanism of human esophageal cancer Ec-109 cells via tanshinone IIA was investigated. The Ec-109 cells were cultured in vitro with different concentrations of tanshinone IIA (2 µg/mL, 4 µg/mL, or 8 µg/mL) for 12, 24, 36, and 48 hours. MTT assay was used to evaluate the proliferative inhibition rate of tanshinone IIA on esophageal Ec-109 cells. After 24 hours of culturing in vitro, a control group was assigned. The apoptosis rate was detected by the AO/EB and annexin V-FITC/propidium iodide assay, and the protein levels of Caspase-4 and CHOP were determined by the Western blot technique. MTT data showed that tanshinone IIA could significantly inhibit the proliferation of Ec-109 cells with a dose- and time-dependent mode. Compared with the control group, tanshinone IIA could apparently induce apoptosis of Ec-109 cells, and the level of Caspase-4 and CHOP (p < 0.01) obviously increased. Tanshinone IIA can significantly induce the apoptosis of Ec-109 cells, which may take effect by the stress pathway of the endoplasmic reticulum.

Chen YJ, Jiang HT, Cao JY
Influence of Photodynamic Therapy on Apoptosis and Invasion of Human Cholangiocarcinoma QBC939 Cell Line.
Chin Med Sci J. 2015; 30(4):252-9 [PubMed] Related Publications
OBJECTIVE: To investigate the effect of photodynamic therapy (PDT) mediated by hematoporphyrin derivative (HPD) on apoptosis and invasion of cholangiocarcinoma QBC939 cell lines.
METHODS: In vitro cultured cholangiocarcinoma QBC939 cell line was exposed to 2, 4, 6, 8, 10, 12, and 14 μg/ml HPD with 5, 10, and 15 J/cm2 light intensity, respectively. The optical density at 450 nm of the QBC939 cells was measured by CCK8 assay and its growth inhibition ratio was calculated. Flow cytometry and transwell migration assay were applied to detect cell apoptosis and invasion respectively. RT-PCR and immunocytochemistry analyses were used to detect expressions of vascular endothelial growth factor-C (VEGF-C), cyclooxygenase-2 (COX-2), and proliferating cell nuclear antigen (PCNA). Enzyme-linked immunosorbent assay (ELISA) was carried out to examine the secretion of VEGF-C and COX-2 in QBC939 cells.
RESULTS: Exposure to HPD-PDT can significantly suppress the growth of QBC939 cells (all P<0.05). HPD-PDT can promote apoptosis of QBC939 cells at the early stage. When the concentration of HPD was 2 μg/ml and light irradiation was 5 J/cm2, HPD-PDT had no obvious inhibitory effect on QBC939 cell growth, but can obviously inhibit cell invasion, and significant difference was observed between the HPD-PDT and control groups (P<0.01). The HPD-PDT can reduce the mRNA and protein expressions of VEGF-C, COX-2, and PCNA, and decrease the secretion of VEGF-C and COX-2 in QBC939 cells.
CONCLUSION: PDT could promote apoptosis and inhibit growth and invasion of cholangiocarcinoma cells QBC939 in vitro.

Ma Q, Chang Z, Wang W, Wang B
Rapamycin-Mediated mTOR Inhibition Reverses Drug Resistance to Adriamycin in Colon Cancer Cells.
Hepatogastroenterology. 2015; 62(140):880-6 [PubMed] Related Publications
BACKGROUND/AIMS: To detect the cellular sensitivity to adriamycin (ADR) by assessing autophagy, apoptosis, and multidrug resistance gene 1 (mdr1) expression in LoVo/Adr cells.
METHODOLOGY: LoVo/Adr cells were designated accordingly into ADR group, Rapamycin (RAPA) group, ADR plus RAPA, and control group in main observations. Autophage, cell death and mdr1 were examined.
RESULTS: IC50 value of ADR in LoVo/Adr was significantly decreased in response to RAPA (P < 0.05). Autophagy rate of LoVo/Adr cells was higher in the ADR or RAPA-alone group than in control (p < 0.05), while ADR/RAPA combination has significantly increased autophagy rate compared to ADR or RAPA alone (p < 0.05). Compared with controls, apoptosis rate in the RAPA group had no difference (p > 0.05); whereas there was significant difference in ADR group (p < 0.05). Furthermore, apoptosis rate was significantly different in combined RAPA/ADR compared to ADR (p < 0.05). Expression of mRNA and protein P-gp level of mdr 1 gene in LoVo/Adr cells were significantly decreased under RAPA-treated groups at 25 µmol/L and 50 µmol/L (p < 0.05).
CONCLUSION: This study has indicated that the inhibition of the mTOR pathway reverses multidrug resistance in colorectal cancer cells, which is associated with increased autophagy, apoptosis and reduced mdr1 gene expression in drug-resistant cells treated with Adriamycin.

Ge Y, Yan D, Deng H, et al.
Novel Molecular Regulators of Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL)-Induced Apoptosis in NSCLC Cells.
Clin Lab. 2015; 61(12):1855-63 [PubMed] Related Publications
BACKGROUND: Although tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) can selectively induce apoptosis in some non-small cell lung cancer (NSCLC) cells, some NSCLC cells exhibit TRAIL-resistance. The underlying mechanisms that regulate TRAIL sensitivity in NSCLC cells are not well understood. The objective of this study was to investigate molecular regulators of the TRAIL pathway in NSCLC cells.
METHODS: The TRAIL-sensitive NSCLC cell line NCI-H358 and a TRAIL-resistant cell line A549 were treated with rmhTRAIL for 24 hours. Then cell viability were measured by MTT assay, meanwhile cell cycle and apoptosis were measured by flow cytometry. Furthermore, mass spectrometry (LC-MS/MS) was used to identify the difference in the protein expression profiles. Finally, real-time PCR was performed to detect the mRNA expression of TRAIL receptors and apoptotic related proteins.
RESULTS: These results confirmed that NCI-H358 cells were sensitive to TRAIL, whereas A549 cells were resistant. Both mRNA and protein levels of voltage-dependent anion-selective channel proteinl (VDAC1), caspase9 (CASP9), and cytochrome c1 (CYC1) were upregulated in H358 cells but downregulated in A549 cells, whereas antiapoptotic protein BAG-2 was downregulated. In addition, TRAIL also causes DR5 low expression in A549 cells.
CONCLUSIONS: These results indicate that rmhTRAIL had different anti-tumor activity in different NSCLC cell lines. Downregulation of VDAC1, CYC1, CASP9, and upregulation of BAG-2 might be associated with underlying TRAIL-resistance mechanisms. These findings motivated further studies to explore new therapeutic strategy overcoming TRAIL-resistance of NSCLC cells through modulating dysregulation of the proteins above.

Zhu X, Yi F, Chen P, et al.
5-Aza-2'-Deoxycytidine and CDDP Synergistically Induce Apoptosis in Renal Carcinoma Cells via Enhancing the APAF-1 Activity.
Clin Lab. 2015; 61(12):1821-30 [PubMed] Related Publications
BACKGROUND: It has been reported that the hypermethylation of APAF-1, DAPK-1 and other tumor suppressive genes (TSGs) correlates with progression of renal cell carcinoma and exerts prognostic and diagnostic relevance in renal cell carcinoma. A recent study has confirmed that demethylation regulates the TSGs expression and proliferation of various types of cancer cells. The present study was to recognize a potential anti-tumor effect of 5-aza-2'-deoxycytidine (DAC), a demethylation agent.
METHODS: We evaluated the DNA demethylation by DAC in human renal carcinoma cells and determined the synergism of the demethylation with the toxicity of Cisplatin (CDDP), which is a commonly utilized anti-tumor agent for renal carcinoma.
RESULTS: It was demonstrated that DAC promoted a significant global genomic demethylation and improved APAF-1 expression at both mRNA and protein levels. The DAC treatment deteriorated the CDDP-induced viability decreasing Caki or ACHN cells and synergized the apoptosis induction of CDDP in ACHN cells. The treatment with both DAC and CDDP promoted a significantly higher level of renal carcinoma cell apoptosis than singular DAC or CDDP treatment. The APAF-1 knockdown significantly inhibited the synergism of DAC with the CDDP-induced apoptosis in ACHN cells.
CONCLUSIONS: The present study confirmed that DAC demethylated the CpGs, particularly APAF-1 in renal carcinoma cells, and that the demethylation synergized the cytotoxity of CDDP in renal carcinoma cells via enhancing the CDDP-induced apoptosis.

Li M, Long C, Yang G, et al.
MiR-26b inhibits melanoma cell proliferation and enhances apoptosis by suppressing TRAF5-mediated MAPK activation.
Biochem Biophys Res Commun. 2016; 471(3):361-7 [PubMed] Related Publications
Alterations in microRNA-26b (miR-26b) expression have been shown to participate in various malignant tumor developments. However, the possible function of miR-26b in human melanoma cells remains unclarified. In this study, quantitative polymerase chain reaction was used to explore the expression profiles of miR-26b in melanoma cells. The effect of miR-26b on cell viability was determined by using MTT assays and colony formation assay. The apoptosis levels were evaluated by using Annexin V/fluorescein isothiocyanate (FITC) apoptosis detection kit and the apoptosis cells were confirmed by Transmission Electron Microscopy (TEM). Luciferase reporter plasmids were constructed to confirm direct targeting. Our study found that the expression of miR-26b was downregulated in human melanoma specimens. Overexpression of miR-26b significantly increased the anti-proliferative effects and apoptosis in A375 and B16F10 melanoma cells. In addition, luciferase gene reporter assays confirmed that TRAF5 was a direct target gene of miR-26b and the anti-tumor effect of miR-26b in melanoma cells was significantly counteracted by treatment with TRAF5 overexpression. Furthermore, the molecular mechanisms underlying the tumor suppressor of miR-26b in malignant melanomas may be due to the dephosphorylation of MAPK pathway caused by the decrease in TRAF5 expression when miR-26b is up-regulated in melanoma cells. These findings indicate that miR-26b might influence TRAF5-MAPK signaling pathways to facilitate the malignant progression of melanoma cells.

Ham J, Costa C, Sano R, et al.
Exploitation of the Apoptosis-Primed State of MYCN-Amplified Neuroblastoma to Develop a Potent and Specific Targeted Therapy Combination.
Cancer Cell. 2016; 29(2):159-72 [PubMed] Free Access to Full Article Related Publications
Fewer than half of children with high-risk neuroblastoma survive. Many of these tumors harbor high-level amplification of MYCN, which correlates with poor disease outcome. Using data from our large drug screen we predicted, and subsequently demonstrated, that MYCN-amplified neuroblastomas are sensitive to the BCL-2 inhibitor ABT-199. This sensitivity occurs in part through low anti-apoptotic BCL-xL expression, high pro-apoptotic NOXA expression, and paradoxical, MYCN-driven upregulation of NOXA. Screening for enhancers of ABT-199 sensitivity in MYCN-amplified neuroblastomas, we demonstrate that the Aurora Kinase A inhibitor MLN8237 combines with ABT-199 to induce widespread apoptosis. In diverse models of MYCN-amplified neuroblastoma, including a patient-derived xenograft model, this combination uniformly induced tumor shrinkage, and in multiple instances led to complete tumor regression.

Li WL, Lee MR, Cho MY
The small molecule survivin inhibitor YM155 may be an effective treatment modality for colon cancer through increasing apoptosis.
Biochem Biophys Res Commun. 2016; 471(2):309-14 [PubMed] Related Publications
Survivin has a known beneficial role in the survival of both cancer cells and normal cells. Therapies targeting survivin have been proposed as an alternative treatment modality for various tumors; however, finding the proper indication for this toxic therapy is critical for reducing unavoidable side effects. We recently observed that high survivin expression in CD133(+) cells is related to chemoresistance in Caco-2 colon cancer cells. However, the effect of survivin-targeted therapy on CD133(+) colon cancer is unknown. In this study, we investigated the roles of CD133 and survivin expression in colon cancer biology in vitro and comparatively analyzed the anticancer effects of survivin inhibitor on CD133(+) cells (ctrl-siRNA group) and small interfering RNA (siRNA)-induced CD133(-) cells (CD133-siRNA group) obtained from a single colon cancer cell line. CD133 knockdown via siRNA transfection did not change the tumorigenicity of cells, although in vitro survivin expression levels in CD133(+) cells were higher than those in siRNA-induced CD133(-) cells. The transfection procedure seemed to induce survivin expression. Notably, a significant number of CD133(-) cells (33.8%) was found in the cell colonies of the CD133-siRNA group. In the cell proliferation assay after treatment, YM155 and a combination of YM155 and 5-fluorouracil (5-FU) proved to be far more effective than 5-FU alone. A significantly increased level of apoptosis was observed with increasing doses of YM155 in all groups. However, significant differences in therapeutic effect and apoptosis among the mock, ctrl-siRNA, and CD133-siRNA groups were not detected. Survivin inhibitor is an effective treatment modality for colon cancer; however, the role of CD133 and the use of survivin expression as a biomarker for this targeted therapy must be verified.

Zhang CG, Huang JC, Liu T, Li XY
Anticancer effects of bishydroxycoumarin are mediated through apoptosis induction, cell migration inhibition and cell cycle arrest in human glioma cells.
J BUON. 2015 Nov-Dec; 20(6):1592-600 [PubMed] Related Publications
PURPOSE: To evaluate the cytotoxic and apoptotic effects of bishydroxycoumarin (BHC) against human glioma cells and to study their mode of action.
METHODS: Three cells were used in the experiments. MTT and LDH assays were used to assess the cytotoxic effects of BHC while an in vitro wound healing assay was used to study the effect of BHC on cell migration. Fluorescence microscopy and annexin V-FITC assay were used to study the cellular morphology and apoptotic effects while flow cytometry in combination with propidium iodide (PI) were used to study cell cycle arrest induced by BHC.
RESULTS: BHC induced substantial and dose-dependent cytotoxic effects against all three cell lines with U87MG cell line being most susceptible. BHC also inhibited cell migration and induced characteristic morphological changes including chromatin condensation, nuclear shrinkage which increased with increasing dose of BHC. The apoptotic cell population (both early and late apoptotic cells) increased with increasing dose of BHC which also induced substantial G0/G1 cell cycle growth arrest in U87MG cells.
CONCLUSION: This study provides help to elucidate the translational potential of the in vitro results to be used for further in vivo studies on human glioma using animal or human subjects. The mechanistic pathway studied in this report could be helpful in explaining the mode of action of these classes of natural products.

Yu EY, Zhao RY, Wang DS
Inhibitory effect of Aphidicolin - a tetracyclic diterpene - on the proliferation and apoptotic induction in human cervical cancer (HeLa) cells.
J BUON. 2015 Nov-Dec; 20(6):1480-6 [PubMed] Related Publications
PURPOSE: The objective of the present work was to investigate the antitumor and apoptotic effect of aphidicolin against human cervical cancer (HeLa) cells.
METHODS: Flow cytometry was performed to detect alterations in the mitochondrial membrane potential loss (ΔΨm) after aphidicolin treatment. Cell viability was assessed by the MTT assay while the apoptotic effects of the compound were examined by fluorescence microscopy and flow cytometry.
RESULTS: Aphidicolin induced dose-dependent as well as time-dependent cytotoxic effects in HeLa cells. Chromatin condensation and formation of apoptotic bodies were observed by Hoechst 33258 staining after drug treatment. Aphidicolin induced both early and late apoptosis in HeLa cells which were correlated with strong concentration of the compound. ΔΨm was also observed following aphidicolin treatment at varying doses.
CONCLUSION: Aphidicolin is a potent antitumor and apoptotic agent against human cervical carcinoma and its effects are mediated via chromatin condensation and mitochondrial membrane potential loss.

Sakatani A, Fujiya M, Ueno N, et al.
Polyphosphate Derived from Lactobacillus brevis Inhibits Colon Cancer Progression Through Induction of Cell Apoptosis.
Anticancer Res. 2016; 36(2):591-8 [PubMed] Related Publications
Although probiotics are known to have antitumor activity, few bacteria-derived antitumor molecules have been identified. The present study explored an antitumor molecule derived from Lactobacillus brevis SBL8803 (L. brevis 8803) and the mechanisms that underlie its effects. Cell viability and apoptosis were assessed by a sulforhodamine B assay and terminal deoxynucleotidyl transferase dUTP staining, respectively. Phosphorylated extracellular signal-regulated kinase (ERK) and cleaved poly ADP-ribose polymerase (PARP) expression were detected by western blotting. The conditioned medium of L. brevis 8803 inhibited SW620 cells viability and the effect was reduced by the degradation of polyphosphate (poly P) in the conditioned medium. A xenograft model showed that poly P inhibited the growth of SW620 cells. Poly P induced the apoptosis of SW620 cells through activation of the ERK pathway. In contrast, in primary cultured cells derived from normal mouse, poly P did not affect cell viability. Probiotic-derived poly P is expected to be an antitumor drug with fewer adverse effects than conventional drugs.

Jiang X, Kim KJ, Ha T, Lee SH
Potential Dual Role of Activating Transcription Factor 3 in Colorectal Cancer.
Anticancer Res. 2016; 36(2):509-16 [PubMed] Related Publications
BACKGROUND/AIM: Activating transcription factor 3 (ATF3) is a member of the ATF/CREB transcription factor family and has been proposed as a molecular target for cancer therapy. The present study was undertaken in order to investigate whether ATF3 influences cancer-related phenotypes in colorectal cancer.
MATERIALS AND METHODS: ATF3 was overexpressed in human colorectal cancer cells and the effects of ATF3 on apoptosis, cell cycle, cell migration and epithelial mesenchymal transition (EMT) were investigated. B-cell lymphoma-2 (Bcl-2) promoter was cloned and used for luciferase assay in cells transfected with control or ATF3 expression vector.
RESULTS: ATF3 down-regulated the expression of Bcl-2 and promoter activity of the Bcl-2 gene. ATF3 increased collective cell migration and expression of cluster of differentiation 44 (CD44) and decreased retinoblastoma (Rb) expression. In addition, ATF3 down-regulated EMT-inducing transcription factors and β-catenin.
CONCLUSION: ATF3 may play a dichotomous role in apoptosis and metastasis in human colorectal cancer cells.

Sun H, Luo L, Lal B, et al.
A monoclonal antibody against KCNK9 K(+) channel extracellular domain inhibits tumour growth and metastasis.
Nat Commun. 2016; 7:10339 [PubMed] Free Access to Full Article Related Publications
Two-pore domain potassium (K2P) channels act to maintain cell resting membrane potential--a prerequisite for many biological processes. KCNK9, a member of K2P family, is implicated in cancer, owing to its overexpression in human tumours and its ability to promote neoplastic cell survival and growth. However, KCNK9's underlying contributions to malignancy remain elusive due to the absence of specific modulators. Here we describe the development of monoclonal antibodies against the KCNK9 extracellular domain and their functional effects. We show that one antibody (Y4) with the highest affinity binding induces channel internalization. The addition of Y4 to KCNK9-expressing carcinoma cells reduces cell viability and increases cell death. Systemic administration of Y4 effectively inhibits growth of human lung cancer xenografts and murine breast cancer metastasis in mice. Evidence for Y4-mediated carcinoma cell autonomous and immune-dependent cytotoxicity is presented. Our study reveals that antibody-based KCNK9 targeting is a promising therapeutic strategy in KCNK9-expressing malignancies.

Makarević J, Tsaur I, Juengel E, et al.
Amygdalin delays cell cycle progression and blocks growth of prostate cancer cells in vitro.
Life Sci. 2016; 147:137-42 [PubMed] Related Publications
AIMS: Despite impressive survival benefits from new agents to treat metastasized prostate cancer (PCa), progressive drug resistance hinders long-term response and restricts the efficacy of subsequent therapy. Due to reported antitumor activity of amygdalin and growing popularity for complementary and alternative medicine the potential of this natural, widely used substance to exert antineoplastic effects on prostate cancer cells has been assessed.
MAIN METHODS: LNCaP (castration-sensitive), DU-145 and PC3 cells (castration-resistant) were exposed to different concentrations of amygdalin for 24h or 2weeks. Cell growth was measured by the MTT test, clonal formation by the clonogenic assay. Flow cytometry served to investigate apoptosis and cell cycle phases. Cell cycle regulating proteins and the mTOR-akt signaling axis were analyzed by western blotting.
KEY FINDINGS: Amygdalin dose-dependently diminished tumor cell growth with maximum effects at 10mg/ml. Apoptosis of PC3 and LNCaP but not of DU-145 cells was reduced, whereas colony formation was suppressed in all cell lines. A decrease in the number of G2/M- and S-phase cells along with an elevated number of G0/G1-phase cells was recorded. The cell cycle proteins cdk 1, cdk 2 and cdk 4 as well as cyclin A, cyclin B and cyclin D3 were modulated by amygdalin after both 24h and 2weeks. Distinct effects on p19 and p27 expression and on Akt, Rictor and Raptor activation became evident only after 2weeks.
SIGNIFICANCE: Amygdalin exhibits significant antitumor activity in both castration-sensitive and castration-resistant PCa cell lines and merits further evaluation for therapeutic purposes.

Yan H, Guo BY, Zhang S
Cancer-associated fibroblasts attenuate Cisplatin-induced apoptosis in ovarian cancer cells by promoting STAT3 signaling.
Biochem Biophys Res Commun. 2016; 470(4):947-54 [PubMed] Related Publications
One of the main reasons for treatment failure in ovarian cancer is acquired drug resistance. Cancer associated fibroblasts (CAFs) are known to enhance chemoresistance in many human tumors. However, its contributions to chemoresistance acquisition in ovarian cancer are not well understood. Here, we provide the first evidence that the conditioned medium of CAFs (CAFs-CM) could attenuate the sensitivity to Cisplatin in A2780 and ES2 ovarian cancer cells and protect them from Cisplatin-induced apoptosis. We found the expression level of two anti-apoptotic proteins, Bcl-2 and Survivin, as well as their upstream controller p-STAT3 were significantly increased when ovarian cancer cells were exposed to CAFs-CM. Furthermore, inhibition of STAT3 signaling with Cryptotanshinone could down-regulate the expression of Bcl-2 and Survivin, thus weaken the post-target resistance to Cispaltin mediating by CAFs-CM in ovarian cancer cells. In conclusion, our data suggested that CAFs could activate the anti-apoptotic STAT3 signaling, thereby decrease the Cisplatin-induced apoptosis and promote chemoresistance in ovarian cancer.

Sophonnithiprasert T, Mahabusarakam W, Nakamura Y, Watanapokasin R
Antiproliferation and Apoptosis Induction in Colorectal Cancer Cells by Goniothalamin.
J Med Assoc Thai. 2015; 98 Suppl 9:S146-51 [PubMed] Related Publications
OBJECTIVE: To investigate the effect of goniothalamin on antiproliferation and apoptosis induction in three types of colorectal cancer cells.
BACKGROUND: Colorectal cancer is the third of the twentieth most commonly diagnosed cancer. Different types of colorectal cancer cells differ in genotype and characteristics leading to different responses to anticancer drugs. Therefore, finding new anticancer compound for the colorectal cancer cells is necessary.
MATERIAL AND METHOD: Antiproliferative response of goniothalamin on three colorectal cancer cell lines including Colo 205, SW480, and LoVo were determined by MTT assay. The antiproliferative response at different time and dose was also observed. Apoptosis induction by goniothalamin was observed in all three cell-lines via morphological changes and nuclear condensation by Hoechst33342 staining.
RESULTS: Goniothalamin showed different antiproliferative response on Colo 205, SW480, and Lo Vo cells at the IC50 value is 9.86 ± 0.38 µM, 22.00 ± 4.40 µM, and 65.25 ± 1.85 µM respectively. In addition, the antiproliferative response of goniothalamin was a time- and dose-dependent manner Apoptosis morphological changes and nuclear condensation were clearly observed in Colo 205, SW480 and LoVo cells treated with 10 µM, 25 µM, and 50 µM goniothalamin, respectively.
CONCLUSION: Goniothalamin showed antiproliferation and apoptosis induction in colorectal cancer cells with different sensitivity depending on cell type. Investigation of mechanisms underlying apoptosis and its potential use for colorectal cancer treatment should be further studied.

Amoury M, Kolberg K, Pham AT, et al.
Granzyme B-based cytolytic fusion protein targeting EpCAM specifically kills triple negative breast cancer cells in vitro and inhibits tumor growth in a subcutaneous mouse tumor model.
Cancer Lett. 2016; 372(2):201-9 [PubMed] Related Publications
Triple-negative breast cancer (TNBC) is associated with poor prognosis and high prevalence among young premenopausal women. Unlike in other breast cancer subtypes, no targeted therapy is currently available. Overexpression of epithelial cell adhesion molecule (EpCAM) in 60% of TNBC tumors correlates with poorer prognosis and is associated with cancer stem cell phenotype. Thus, selective elimination of EpCAM(+) TNBC tumor cells is of clinical importance. Therefore, we constructed a fully human targeted cytolytic fusion protein, designated GbR201K-αEpCAM(scFv), in which an EpCAM-selective single-chain antibody fragment (scFv) is genetically fused to a granzyme B (Gb) mutant with reduced sensitivity to its natural inhibitor serpin B9. In vitro studies confirmed its specific binding, internalization and cytotoxicity toward a panel of EpCAM-expressing TNBC cells. Biodistribution kinetics and tumor-targeting efficacy using MDA-MB-468 cells in a human TNBC xenograft model in mice revealed selective accumulation of GbR201K-αEpCAM(scFv) in the tumors after i.v. injection. Moreover, treatment of tumor-bearing mice demonstrated a prominent inhibition of tumor growth of up to 50 % in this proof-of-concept study. Taken together, our results indicate that GbR201K-αEpCAM(scFv) is a promising novel targeted therapeutic for the treatment of TNBC.

Jing Z, Sui X, Yao J, et al.
SKF-96365 activates cytoprotective autophagy to delay apoptosis in colorectal cancer cells through inhibition of the calcium/CaMKIIγ/AKT-mediated pathway.
Cancer Lett. 2016; 372(2):226-38 [PubMed] Related Publications
Store-operated Ca(2+) entry (SOCE) inhibitors are emerging as an attractive new generation of anti-cancer drugs. Here, we report that SKF-96365, an SOCE inhibitor, exhibits potent anti-neoplastic activity by inducing cell-cycle arrest and apoptosis in colorectal cancer cells. In the meantime, SKF-96365 also induces cytoprotective autophagy to delay apoptosis by preventing the release of cytochrome c (cyt c) from the mitochondria into the cytoplasm. Mechanistically, SKF-96365 treatment inhibited the calcium/calmodulin-dependent protein kinase IIγ (CaMKIIγ)/AKT signaling cascade in vitro and in vivo. Overexpression of CaMKIIγ or AKT abolished the effects of SKF-96365 on cancer cells, suggesting a critical role of the CaMKIIγ/AKT signaling pathway in SFK-96365-induced biological effects. Moreover, Hydroxychloroquine (HCQ), an FDA-approved drug used to inhibit autophagy, could significantly augment the anti-cancer effect of SFK-96365 in a mouse xenograft model. To our best knowledge, this is the first report to demonstrate that calcium/CaMKIIγ/AKT signaling can regulate apoptosis and autophagy simultaneously in cancer cells, and the combination of the SOCE inhibitor SKF-96365 with autophagy inhibitors represents a promising strategy for treating patients with colorectal cancer.

de Oliveira MR, Nabavi SF, Manayi A, et al.
Resveratrol and the mitochondria: From triggering the intrinsic apoptotic pathway to inducing mitochondrial biogenesis, a mechanistic view.
Biochim Biophys Acta. 2016; 1860(4):727-45 [PubMed] Related Publications
BACKGROUND: Mitochondria, the power plants of the cell, are known as a cross-road of different cellular signaling pathways. These cytoplasmic double-membraned organelles play a pivotal role in energy metabolism and regulate calcium flux in the cells. It is well known that mitochondrial dysfunction is associated with different diseases such as neurodegeneration and cancer. A growing body of literature has shown that polyphenolic compounds exert direct effects on mitochondrial ultra-structure and function. Resveratrol is known as one of the most common bioactive constituents of red wine, which improves mitochondrial functions under in vitro and in vivo conditions.
SCOPE OF REVIEW: This paper aims to review the molecular pathways underlying the beneficial effects of resveratrol on mitochondrial structure and functions. In addition, we discuss the chemistry and main sources of resveratrol.
MAJOR CONCLUSIONS: Resveratrol represents the promising effects on mitochondria in different experimental models. However, there are several reports on the detrimental effects elicited by resveratrol on mitochondria.
GENERAL SIGNIFICANCE: An understanding of the chemistry and source of resveratrol, its bioavailability and the promising effects on mitochondria brings a new hope to therapy of mitochondrial dysfunction-related diseases.

Alshatwi AA, Periasamy VS, Athinarayanan J, Elango R
Synergistic anticancer activity of dietary tea polyphenols and bleomycin hydrochloride in human cervical cancer cell: Caspase-dependent and independent apoptotic pathways.
Chem Biol Interact. 2016; 247:1-10 [PubMed] Related Publications
Bleomycin is a chemotherapeutic agent that is frequently used in the treatment of various cancers. Bleomycin causes serious adverse effects via antioxidant defense abnormalities against reactive oxygen species (ROS). However, the current cervical cancer monodrug therapy strategy has failed to produce the expected outcomes; hence, combinational therapies are gaining great interest. Tea polyphenols are also effective antioxidative and chemo-preventive agents. However, the combined effect of tea polyphenol (TPP) and bleomycin (BLM) against cervical cancer remains unknown. In this study, we focused on the potential of TPP on BLM anticancer activity against cervical cancer cells. Cervical cancer cells (SiHa) were treated with various concentrations of TPP, BLM and TPP combined with BLM (TPP-BLM), and their effects on cell growth, intracellular reactive oxygen species, poly-caspase activity, early apoptosis and the expression of caspase-3, caspase-8 and caspase-9, Bcl-2 and p53 were assessed. The MTT assay revealed that the SiHa cells were less sensitive to growth inhibition by TPP treatment compared with both BLM and the combination therapy. Nuclear staining indicated that exposure to TPP-BLM increased the percentage of apoptotic nuclei compared with a mono-agent treatment. Caspase activation assay demonstrated that proportion of early and late apoptotic/secondary necrotic cells was higher in the cells treated with the combination therapy than in those treated with either TPP or BLM alone. The TPP-BLM treatment synergistically induced apoptosis through caspase-3, caspase-8 and caspase-9 activation, Bcl-2 upregulation and p53 overexpression. This study suggests that TPP-BLM may be used as an efficient antioxidant-based combination therapy for cervical cancer.

Adamopoulos PG, Kontos CK, Tsiakanikas P, Scorilas A
Identification of novel alternative splice variants of the BCL2L12 gene in human cancer cells using next-generation sequencing methodology.
Cancer Lett. 2016; 373(1):119-29 [PubMed] Related Publications
The next-generation sequencing (NGS) technology has enabled genome-wide studies, providing massively parallel DNA sequencing. NGS applications constitute a revolution in molecular biology and genetics and have already paved new ways in cancer research. BCL2L12 is an apoptosis-related gene, previously cloned from members of our research group. Like most members of the BCL2 gene family, it is highly implicated in various types of cancer and hematological malignancies. In the present study, we used NGS to discover novel alternatively spliced variants of the apoptosis-related BCL2L12 gene in many human cancer cell lines, after 3'-RACE nested PCR. Extensive computational analysis uncovered new alternative splicing events and patterns, resulting in novel alternative transcripts of the BCL2L12 gene. PCR was then performed to validate NGS data and identify the derived novel transcripts of the BCL2L12 gene. Therefore, 50 novel BCL2L12 splice variants were discovered. Since BCL2L12 is involved in the apoptotic machinery, the quantification of distinct BCL2L12 transcripts in human samples may have clinical applications in different types of cancer.

Das D, Persaud L, Dejoie J, et al.
Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) activates caspases in human prostate cancer cells through sigma 1 receptor.
Biochem Biophys Res Commun. 2016; 470(2):319-23 [PubMed] Related Publications
Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-based therapy is currently evaluated in clinical studies as a tumor cell-selective pro-apoptotic approach. Unfortunately, many clinical studies have shown that cancer cells acquire TRAIL resistance and finally avoid TRAIL-induced apoptosis. Therefore, defining the mechanisms that permit TRAIL to activate apoptosis is critical for the development of strategies that maximize the potential effectiveness of TRAIL in clinical applications. This study aims at understanding the molecular mechanisms underlying TRAIL-induced apoptosis and unraveling signaling pathways that could revert sensitivity to apoptosis stimuli. Our current study demonstrates for the first time that Sigma 1 Receptor (Sig1R), a ligand-regulated protein chaperone, contributes to TRAIL induction of apoptosis. We show that Sig1R agonist (+)-SKF10047 action or increasing Sig1R expression, significantly reduced apoptosis by TRAIL in prostate cell lines, indicating the importance of Sig1R and signifying that higher levels of Sig1R in prostate cancer cells make them more resistant to TRAIL treatment. Here we show that Sig1R is critically involved in TRAIL-induced caspase activation. Furthermore, we show that Sig1R protein is degraded upon TRAIL treatment. Knockdown of Sig1R, by siRNA transfection increased the sensitivity of breast cancer cells to TRAIL. These results indicate that Sig1R could represent a promising molecule to sensitize human breast cancers to TRAIL. Collectively, these studies define Sig1R as a key mediator of TRAIL induction of cancer-specific killing.

Matsumoto T, Jimi S, Migita K, et al.
Inhibition of glucose transporter 1 induces apoptosis and sensitizes multiple myeloma cells to conventional chemotherapeutic agents.
Leuk Res. 2016; 41:103-10 [PubMed] Related Publications
Despite the recent development of anti-myeloma drugs, the prognosis of high-risk multiple myeloma remains poor. Therefore, new effective treatment strategies for this disease are needed. It has been reported that high intensity of 18-fluorodeoxyglucose positron emission tomography is high-risk factor in myeloma, suggesting that glucose uptake can be therapeutic target in high-risk myeloma. In this study, we addressed the utility of glucose transporter 1 (GLUT1) as a therapeutic target for myeloma with increased glucose uptake. We found myeloma cell lines with elevated glucose uptake activity via GLUT1 up-regulation. STF-31, a selective GLUT1 inhibitor, completely suppressed the glucose uptake activity and induced apoptosis in GLUT1 expressing myeloma cells. On the other hand, this agent little shows the cytotoxicity in normal peripheral blood mononuclear cells. Moreover, STF-31 synergistically enhanced the cell death induced by melphalan, doxorubicin, and bortezomib. GLUT1 may be promising therapeutic target in myeloma with elevated glucose uptake.

Carneiro MC, Henriques CM, Nabais J, et al.
Short Telomeres in Key Tissues Initiate Local and Systemic Aging in Zebrafish.
PLoS Genet. 2016; 12(1):e1005798 [PubMed] Free Access to Full Article Related Publications
Telomeres shorten with each cell division and telomere dysfunction is a recognized hallmark of aging. Tissue proliferation is expected to dictate the rate at which telomeres shorten. We set out to test whether proliferative tissues age faster than non-proliferative due to telomere shortening during zebrafish aging. We performed a prospective study linking telomere length to tissue pathology and disease. Contrary to expectations, we show that telomeres shorten to critical lengths only in specific tissues and independently of their proliferation rate. Short telomeres accumulate in the gut but not in other highly proliferative tissues such as the blood and gonads. Notably, the muscle, a low proliferative tissue, accumulates short telomeres and DNA damage at the same rate as the gut. Together, our work shows that telomere shortening and DNA damage in key tissues triggers not only local dysfunction but also anticipates the onset of age-associated diseases in other tissues, including cancer.

Rajagopalan P, Alahmari KA, Elbessoumy AA, et al.
Biological evaluation of 2-arylidene-4, 7-dimethyl indan-1-one (FXY-1): a novel Akt inhibitor with potent activity in lung cancer.
Cancer Chemother Pharmacol. 2016; 77(2):393-404 [PubMed] Related Publications
PURPOSE: Human lung cancer is contributed to be a major mortality factor in the cancer-related deaths. Arylidene indan-ones constitute a new class of potential anti-tumor compounds. Herein, we report the biological efficacy of the 2-arylidene-4, 7-dimethyl indan-1-one (FXY-1), a potential lead molecule of arylidene indan-ones in lung cancer models. We previously described anticancer activity of FXY-1 against human breast adenocarcinoma.
METHODS: FXY-1 efficacy was assessed by standard anticancer screening in NCI-H 460, A549, NCI-H 1975 and NCI-H 2170 cells. Initial molecular docking analysis was performed to check the interaction of compound to Akt enzyme. Anti-proliferation, anti-metastatic and transendothelial cell migration were performed to check efficacy of the drug. Western blot analysis was performed to understand the regulation of pro-apoptotic and anti-apoptotic proteins by the compound. The effect of FXY-1 on caspase induction and Akt phosphorylation were checked using Western blot analysis. Flow cytometry was used to reveal the cell cycle changes and apoptosis-inducing properties of FXY-1 in the lung cancer cells. In-vitro Akt inhibition property of the compound was studied using a fluorescence-based, coupled-enzyme reaction. The in-vivo efficacy of the compound was determined using a mouse xenograft model.
RESULTS: Our molecular docking analysis reveals higher interaction of FXY-1 with Akt. FXY-1 depicted anti-proliferative and pro-apoptotic activity with higher therapeutic window in-vitro in NCI-H 460 and A549 cells. The compound treatment to lung cancer cells resulted in induction of DNA fragmentation, inhibition of transendothelial migration, caspase activation and poly (ADP-ribose) polymerase (PARP) cleavage. FXY-1 treatment resulted in G 0/G 1 arrest in both cell lines at lower concentrations, but led to apoptosis at higher doses. Western blot analysis revealed dephosphorylation of Akt (Ser 473) with activation of p53, Bax, Bak, Bid and reduction in Bcl-2 and Bcl-xL levels. Further mechanistic investigation showed that FXY-1 activity was facilitated through an allosteric inhibition of Akt enzyme. FXY-1 treatment significantly reduced the tumor growth and pAkt levels in mouse xenograft exhibiting the in-vivo efficacy in the compound.
CONCLUSION: Collectively, our results suggest DNA damage-mediated activation by FXY-1 in lung cancer cells leading to extensive apoptosis through the mitochondrial pathway.

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