A novel Bcl-2 inhibitor, BM-1197, Induces apoptosis in Malignant Lymphoma Cells through the Endogenous Apoptotic Pathway

Abstract Background Bcl-2 family members play an important role in the development of malignant lymphoma and can induce drug resistance in anticancer treatment. The development of small molecules targeting Bcl-2 family proteins may be a new strategy for the treatment of malignant lymphoma. In this study, we investigate the antitumor effect and cellular mechanism of a novel Bcl-2/Bcl-xL dual inhibitor, BM-1197, in DCBCL and Burkitt lymphoma cells. Methods The CCK-8 assay was used to detect cell viability. Apoptosis was determined by Hoechst 33258 staining and flow cytometry. The activity of caspase-3/caspase-9 was determined using a caspase-3/caspase-9 activity kit. Western blotting analysis was performed to evaluate the changes in protein expression. Functional analysis was performed via immunoprecipitation and siRNA interference. Human malignant lymphoma xenograft models in nude mice were established for in vivo efficacy detection. Results We find that BM-1197 exerts potent growth-inhibitory activity against lymphoma cells that harbor high expression of Bcl-2 and Bcl-xL in vitro and has a synergistic effect with chemotherapeutic drugs. Mechanistically, we see that the intrinsic apoptosis pathway is activated upon BM-1197 treatment. BM-1197 affects the protein interactions of Bak/Bcl-xl, Bim/Bcl-2, Bim/Bcl-xl, and PUMA/Bcl-2 and induces conformational changes in the Bax protein, which result in the activation of Bax and release of cytochrome c, activate caspase − 9, − 3, and − 7 and finally induce cell apoptosis. Furthermore, our data demonstrate that BM-1197 exhibits strong anti-tumor effects against established human malignant lymphoma xenograft models. Conclusions Our study demonstrated BM-1197 exerts potent antitumor effects both in vitro and in vivo and provides promising preclinical data for the further development of BM-1197 in malignant lymphoma.

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A novel Bcl-2 inhibitor, BM-1197, Induces apoptosis in Malignant Lymphoma Cells through the Endogenous Apoptotic Pathway

10.21203/rs.2.11086/v1 ◽

2019 ◽

Author(s):

Yue-Li Sun ◽

Wen-Qi Jiang ◽

Qiu-Yun Luo ◽

Da-Jun Yang ◽

Yu-Chen Cai ◽

...

Keyword(s):

Malignant Lymphoma ◽

Caspase 3 ◽

Caspase 9 ◽

Antitumor Effects ◽

Lymphoma Cells ◽

Dual Inhibitor ◽

Apoptosis Pathway ◽

Xenograft Models

Abstract Background: Bcl-2 family members play an important role in the development of malignant lymphoma and can induce drug resistance in anticancer treatment. The development of small molecules targeting Bcl-2 family protein can be new strategy for malignant lymphoma treatment. In this study, we investigate the antitumor effect and the cellular mechanism of a novel Bcl-2/Bcl-xL dual inhibitor BM-1197 in DCBCL and Burkitt lymphoma cells. Methods: CCK-8 assay was used to detect cell viability. Apoptosis was determined by Hoechst 33258 staining and flow cytometry. The activity of caspase-3/caspase-9 was determined using the caspase-3/ caspase-9 activity kit. Western blotting analysis was performed to evaluate the change of protein expression. The functional analysis was evalueated via immunoprecipitation and siRNA interference. Human malignant lymphoma xenograft models in nude mice were established for in vivo efficacy detection. Results: We find that BM-1197 exerts potent growth-inhibitory activity against lymphoma cells which harbor Bcl-2 and Bcl-xL high expression in vitro and has synergistic effect with chemotherapeutic drugs. Mechanistically, we see that the intrinsic apoptosis pathway is activated upon BM-1197 treatment. BM-1197 affects the protein interaction of Bak/Bcl-xl, Bim/Bcl-2, Bim/Bcl-xl, PUMA/Bcl-2 and induced conformational change in the Bax protein.which results in activation of Bax and release cytochrome c, and activated caspase -9, -3, -7 and finally induce cell apoptosis. Furthermore, our data demonstrates that BM-1197 exhibits strong anti-tumor effects against established human malignant lymphoma xenograft models. Conclusions: Our study demonstrated BM-1197 exerts potent antitumor effects both in vitro and in vivo, and provides promising preclinical data for further development of BM-1197 in malignant lymphoma.

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A novel Bcl-2 inhibitor, BM-1197, Induces apoptosis in Malignant Lymphoma Cells through the Endogenous Apoptotic Pathway

10.21203/rs.2.11086/v3 ◽

2019 ◽

Author(s):

Yue-Li Sun ◽

Wen-Qi Jiang ◽

Qiu-Yun Luo ◽

Da-Jun Yang ◽

Yu-Chen Cai ◽

...

Keyword(s):

Malignant Lymphoma ◽

Caspase 3 ◽

Caspase 9 ◽

Antitumor Effects ◽

Lymphoma Cells ◽

Dual Inhibitor ◽

Apoptosis Pathway ◽

Xenograft Models

Abstract Background: Bcl-2 family members play an important role in the development of malignant lymphoma and can induce drug resistance in anticancer treatment. The development of small molecules targeting Bcl-2 family protein can be new strategy for malignant lymphoma treatment. In this study, we investigate the antitumor effect and the cellular mechanism of a novel Bcl-2/Bcl-xL dual inhibitor BM-1197 in DCBCL and Burkitt lymphoma cells. Methods: CCK-8 assay was used to detect cell viability. Apoptosis was determined by Hoechst 33258 staining and flow cytometry. The activity of caspase-3/caspase-9 was determined using the caspase-3/ caspase-9 activity kit. Western blotting analysis was performed to evaluate the change of protein expression. The functional analysis was evalueated via immunoprecipitation and siRNA interference. Human malignant lymphoma xenograft models in nude mice were established for in vivo efficacy detection. Results: We find that BM-1197 exerts potent growth-inhibitory activity against lymphoma cells which harbor Bcl-2 and Bcl-xL high expression in vitro and has synergistic effect with chemotherapeutic drugs. Mechanistically, we see that the intrinsic apoptosis pathway is activated upon BM-1197 treatment. BM-1197 affects the protein interaction of Bak/Bcl-xl, Bim/Bcl-2, Bim/Bcl-xl, PUMA/Bcl-2 and induced conformational change in the Bax protein.which results in activation of Bax and release cytochrome c, and activated caspase -9, -3, -7 and finally induce cell apoptosis. Furthermore, our data demonstrates that BM-1197 exhibits strong anti-tumor effects against established human malignant lymphoma xenograft models. Conclusions: Our study demonstrated BM-1197 exerts potent antitumor effects both in vitro and in vivo, and provides promising preclinical data for further development of BM-1197 in malignant lymphoma.

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A novel Bcl-2 inhibitor, BM-1197, Induces apoptosis in Malignant Lymphoma Cells through the Endogenous Apoptotic Pathway

10.21203/rs.2.11086/v4 ◽

2019 ◽

Author(s):

Yue-Li Sun ◽

Wen-Qi Jiang ◽

Qiu-Yun Luo ◽

Da-Jun Yang ◽

Yu-Chen Cai ◽

...

Keyword(s):

Malignant Lymphoma ◽

Caspase 3 ◽

Caspase 9 ◽

Antitumor Effects ◽

Lymphoma Cells ◽

Dual Inhibitor ◽

Apoptosis Pathway ◽

Xenograft Models

Abstract Background: Bcl-2 family members play an important role in the development of malignant lymphoma and can induce drug resistance in anticancer treatment. The development of small molecules targeting Bcl-2 family protein can be new strategy for malignant lymphoma treatment. In this study, we investigate the antitumor effect and the cellular mechanism of a novel Bcl-2/Bcl-xL dual inhibitor BM-1197 in DCBCL and Burkitt lymphoma cells. Methods: CCK-8 assay was used to detect cell viability. Apoptosis was determined by Hoechst 33258 staining and flow cytometry. The activity of caspase-3/caspase-9 was determined using the caspase-3/ caspase-9 activity kit. Western blotting analysis was performed to evaluate the change of protein expression. The functional analysis was evalueated via immunoprecipitation and siRNA interference. Human malignant lymphoma xenograft models in nude mice were established for in vivo efficacy detection. Results: We find that BM-1197 exerts potent growth-inhibitory activity against lymphoma cells which harbor Bcl-2 and Bcl-xL high expression in vitro and has synergistic effect with chemotherapeutic drugs. Mechanistically, we see that the intrinsic apoptosis pathway is activated upon BM-1197 treatment. BM-1197 affects the protein interaction of Bak/Bcl-xl, Bim/Bcl-2, Bim/Bcl-xl, PUMA/Bcl-2 and induced conformational change in the Bax protein.which results in activation of Bax and release cytochrome c, and activated caspase -9, -3, -7 and finally induce cell apoptosis. Furthermore, our data demonstrates that BM-1197 exhibits strong anti-tumor effects against established human malignant lymphoma xenograft models. Conclusions: Our study demonstrated BM-1197 exerts potent antitumor effects both in vitro and in vivo, and provides promising preclinical data for further development of BM-1197 in malignant lymphoma.

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Silica nanoparticles inducing the apoptosis of spermatocyte cell through microRNA-450b-3p targeting MTCH2-mediating mitochondrial signaling pathway

10.21203/rs.3.rs-24673/v1 ◽

2020 ◽

Author(s):

Guiqing Zhou ◽

Jianhui Liu ◽

Xiangyang Li ◽

Yujian Sang ◽

Yue Zhang ◽

...

Keyword(s):

Cytochrome C ◽

Silica Nanoparticles ◽

Caspase 3 ◽

Reproductive Toxicity ◽

Control Group ◽

Caspase 9 ◽

Protein Expressions ◽

Underlying Mechanisms

Abstract Background: Silica nanoparticles (SiNPs) are found in environmental particulate matter and are proven to have adverse effects on fertility. The relationship and underlying mechanisms between miRNAs and apoptosis induced by SiNPs during spermatogenesis is currently ambiguous. Experimental design: The present study was designed to investigate the role of miRNA-450b-3p in the reproductive toxicity caused by SiNPs. In vivo, 40 male mice were randomly divided into control and SiNPs groups, 20 per group. The mice in the SiNPs group were administrated 20 mg/kg SiNPs by tracheal perfusion once every 5 days, for 35 days, and the control group were given the equivalent of a normal luminal saline. In vitro, spermatocyte cells were divided into 0 and 5 μg/mL SiNPs groups, after passaged for 30 generations, the GC-2spd cells in 5 μg/mL SiNPs groups were transfected with miRNA-450b-3p and its mimic and inhibitor. Results: In vivo, the results showed that SiNPs damaged tissue structures of testis, decreased the quantity and quality of the sperm, reduced the expression of miR-450b-3p, and increased the protein expressions of the MTCH2, BID, BAX, Cytochrome C, Caspase-9, and Caspase-3 in the testis. In vitro, SiNPs obviously repressed the viability and increased the LDH level and apoptosis rate, decreased the levels of the miR-450b-3p, significantly enhanced the protein expressions of the MTCH2, BID, BAX, Cytochrome C, Caspase-9, Caspase-3; while the mimic of miR-450b-3p reversed the changes induced by SiNPs, but inhibitor further promoted the effects induced by SiNPs.Conclusion: The result suggested that SiNPs could induce the spermatocyte apoptosis by inhibiting the miR-450b-3p expression to target promoting the MTCH2 resulting in activating mitochondrial apoptotic signaling pathways in the spermatocyte cells.

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Intracellular NAD+ Depletion Enhances Bortezomib-Induced Myeloma Cytotoxicity

Blood ◽

10.1182/blood.v120.21.330.330 ◽

2012 ◽

Vol 120 (21) ◽

pp. 330-330

Author(s):

Antonia Cagnetta ◽

Michele Cea ◽

Chirag Acharya ◽

Teresa Calimeri ◽

Yu-Tzu Tai ◽

...

Keyword(s):

Cell Death ◽

Synergistic Effect ◽

Cell Lines ◽

Caspase 3 ◽

Statistical Significance ◽

Caspase 8 ◽

Caspase 9 ◽

Nicotinamide Phosphoribosyltransferase

Abstract Abstract 330 Background: Our previous study demonstrated that inhibition of nicotinamide phosphoribosyltransferase (Nampt) acts by severely depleting intracellular NAD+ content and thus eliciting mitochondrial dysfunction and autophagic MM cell death. The proteasome inhibitor Bortezomib induces anti-MM activity by affecting a variety of signaling pathways. However, as with other agents, dose-limiting toxicities and the development of resistance limit its long-term utility. Here, we demonstrate that combining Nampt inhibitor and bortezomb induces synergistic anti-MM cell death both in vitro using MM cell lines or patient CD138+ MM cells and in vivo in a human plasmacytoma xenograft mouse model. Material and Methods: We utilized MM.1S, MM.1R, RPMI-8226, and U266 human MM cell lines, as well as purified tumor cells from patients relapsing after prior therapies. Cell viability and apoptosis assays were performed using Annexin V/PI staining. Intracellular NAD+ level and proteasome activity were quantified after 12, 24, and 48h exposure to single/combination drugs by specific assays. In vitro angiogenesis was assessed by Matrigel capillary-like tube structure formation assay. Immunoblot analysis was performed using antibodies to caspase-8, caspase-9, caspase-3, PARP, Bcl-2, and tubulin. CB-17 SCID male mice (n = 28; 7 mice/EA group) were subcutaneously inoculated with 5.0 × 106 MM.1S cells in 100 microliters of serum free RPMI-1640 medium. When tumors were measurable (3 weeks after MM cell injection), mice were treated for three weeks with vehicle alone, FK866 (30mg/kg 4 days weekly), Bortezomib (0.5 mg/kg twice weekly), or FK866 (30 mg/kg) plus Bortezomib (0.5 mg/kg). Statistical significance of differences observed in FK866, Bortezomib or combination-treated mice was determined using a Student t test. Isobologram analysis was performed using “CalcuSyn” software program. A combination index < 1.0 indicates synergism. Results/Discussion: Combining FK866 and Bortezomib induces synergistic anti-MM activity in vitro against MM cell lines (P<0.005, CI < 1) or patient CD138-positive MM cells (P< 0.004). FK866 plus Bortezomib-induced synergistic effect is associated with: 1)activation of caspase-8, caspase-9, caspase-3, and PARP; 2) improved intracellular NAD+ dissipation; 3) suppression of chymotrypsin-like, caspase-like, and trypsin-like proteolytic activities; 4) inhibition of NF-kappa B signaling; and 5) inhibition of angiogenesis. Importantly, the ectopic overexpression of Nampt rescues this observed synergistic effect; conversely, Nampt knockdown by RNAi significantly enhances the anti-MM effect of bortezomib. In the murine xenograft MM model, low dose combination FK866 (30 mg/kg) and Bortezomib (0.5 mg/kg) is well tolerated, significantly inhibits tumor growth (P < 0.001), and prolongs host survival (2–2.5 months in mice receiving combined drugs, P = 0.001). These findings demonstrate that intracellular NAD+ levels represent a major determinant in the ability of bortezomib to induce apoptosis of MM cells, providing the rationale for clinical protocols evaluating FK866 together with Bortezomib to improve patient outcome in MM. Disclosures: Munshi: Celgene: Consultancy; Millenium: Consultancy; Merck: Consultancy; Onyx: Consultancy.

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Tetrastigma hemsleyanum flavones exert anti-hepatic carcinoma property both in vitro and in vivo

Food Quality and Safety ◽

10.1093/fqsafe/fyab025 ◽

2021 ◽

Author(s):

Yangyang Liu ◽

Yonglu Li ◽

Wen Chen ◽

Xiang Ye ◽

Ruoyi Jia ◽

...

Keyword(s):

Hepg2 Cells ◽

Caspase 3 ◽

Expression Level ◽

Caspase 9 ◽

Tetrastigma Hemsleyanum ◽

Tumor Bearing ◽

And Migration ◽

Proliferation And Migration

Abstract: Tetrastigma hemsleyanum has been regarded as an anticancer food in China. However, its corresponding mechanisms remains unclear. Thus, in this study, the antitumor activity of flavones-rich fraction of root of Tetrastigma hemsleyanum (FRTH) was investigated in vitro and in vivo. The results indicated that FRTH could inhibit the proliferation and migration of HepG2 cells in vitro by PI3K/AKT pathway. FRTH could increase the ROS level and change the mitochondrial membrane potential (MMP) in HepG2 cells. In addition, FRTH treatment (300, 600 mg/kg BW) significantly suppressed tumor growth on HepG2 tumor-bearing nude mice. Besides, immunohistochemistry assays and western blotting revealed that FRTH enhanced the expression level of Bax/Bcl-2, cytochrome C, Caspase-3, caspase-9, Cleaved-caspase-3, and downregulated the expression level of CD31, ki67 and VEGF in HepG2 tumor-bearing mice. Our study suggests Tetrastigma hemsleyanum as a promising candidate medicine for liver cancer treatment.

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Ethanolic Extract of Senna velutina Roots: Chemical Composition, In Vitro and In Vivo Antitumor Effects, and B16F10-Nex2 Melanoma Cell Death Mechanisms

Oxidative Medicine and Cellular Longevity ◽

10.1155/2019/5719483 ◽

2019 ◽

Vol 2019 ◽

pp. 1-14 ◽

Cited By ~ 3

Author(s):

David Tsuyoshi Hiramatsu Castro ◽

Jaqueline Ferreira Campos ◽

Marcio José Damião ◽

Heron Fernandes Vieira Torquato ◽

Edgar Julian Paredes-Gamero ◽

...

Keyword(s):

Cell Cycle ◽

Chemical Composition ◽

Caspase 3 ◽

Tumor Volume ◽

Apoptotic Cell ◽

Ethanolic Extract ◽

Antitumor Effects ◽

Cycle Arrest

Cutaneous melanoma is among the most aggressive types of cancer, and its rate of occurrence increases every year. Current pharmacological treatments for melanoma are not completely effective, requiring the identification of new drugs. As an alternative, plant-derived natural compounds are described as promising sources of new anticancer drugs. In this context, the objectives of this study were to identify the chemical composition of the ethanolic extract of Senna velutina roots (ESVR), to assess its in vitro and in vivo antitumor effects on melanoma cells, and to characterize its mechanisms of action. For these purposes, the chemical constituents were identified by liquid chromatography coupled to high-resolution mass spectrometry. The in vitro activity of the extract was assessed in the B16F10-Nex2 melanoma cell line using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and based on the apoptotic cell count; DNA fragmentation; necrostatin-1 inhibition; intracellular calcium, pan-caspase, and caspase-3 activation; reactive oxygen species (ROS) levels; and cell cycle arrest. The in vivo activity of the extract was assessed in models of tumor volume progression and pulmonary nodule formation in C57Bl/6 mice. The chemical composition results showed that ESVR contains flavonoid derivatives of the catechin, anthraquinone, and piceatannol groups. The extract reduced B16F10-Nex2 cell viability and promoted apoptotic cell death as well as caspase-3 activation, with increased intracellular calcium and ROS levels as well as cell cycle arrest at the sub-G0/G1 phase. In vivo, the tumor volume progression and pulmonary metastasis of ESVR-treated mice decreased over 50%. Combined, these results show that ESVR had in vitro and in vivo antitumor effects, predominantly by apoptosis, thus demonstrating its potential as a therapeutic agent in the treatment of melanoma and other types of cancer.

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Small-molecule XIAP antagonist restores caspase-9–mediated apoptosis in XIAP-positive diffuse large B-cell lymphoma cells

Blood ◽

10.1182/blood-2007-04-085480 ◽

2008 ◽

Vol 111 (1) ◽

pp. 369-375 ◽

Cited By ~ 26

Author(s):

Saskia A. G. M. Cillessen ◽

John C. Reed ◽

Kate Welsh ◽

Clemencia Pinilla ◽

Richard Houghten ◽

...

Keyword(s):

B Cell ◽

Small Molecule ◽

Caspase 3 ◽

B Cell Lymphoma ◽

Caspase 9 ◽

Lymphoma Cells ◽

Apoptosis Pathway ◽

Expression Levels ◽

Apoptosis Protein ◽

Large B Cell

Clinical outcome in patients with primary nodal diffuse large B-cell lymphomas (DLBCLs) is correlated with expression of inhibitors of the intrinsic apoptosis pathway, including X-linked inhibitor of apoptosis protein (XIAP). XIAP suppresses apoptosis through inhibiting active caspase-3, caspase-7, and caspase-9. In this study, we investigated to see if the small-molecule XIAP antagonist 1396-12 induces cell death in cultured lymphoma cells of patients with DLBCL. Treatment with this XIAP antagonist resulted in relief of caspase-3 inhibition and in induction of apoptosis in 16 of 20 tested DLBCL samples. Sensitivity to the XIAP antagonist was observed in both chemotherapy-refractory and -responsive DLBCL, but did not affect peripheral blood mononuclear cells and tonsil germinal-center B cells from healthy donors. XIAP antagonist-sensitive samples were characterized by high expression levels of XIAP, relatively low expression levels of Bcl-2, and by constitutive caspase-9 activation. These data indicate that the small-molecule XIAP antagonist can induce apoptosis in cultured DLBCL cells and therefore should be considered for possible development as a therapy for these patients. In vitro sensitivity to the XIAP antagonist can be predicted based on biological markers, suggesting the possibility of predefining patients most likely to benefit from XIAP antagonist therapy.

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Indole-3-Carbinol Inhibits Nasopharyngeal Carcinoma

International Journal of Toxicology ◽

10.1177/1091581809356481 ◽

2010 ◽

Vol 29 (2) ◽

pp. 185-192 ◽

Cited By ~ 3

Author(s):

Wei Zhu ◽

Wenxue Li ◽

Guangyu Yang ◽

Quanxin Zhang ◽

Ming Li ◽

...

Keyword(s):

Superoxide Dismutase ◽

Nasopharyngeal Carcinoma ◽

Glutathione Peroxidase ◽

Tumor Growth ◽

Caspase 3 ◽

Caspase 9 ◽

Human Nasopharyngeal Carcinoma ◽

Induced Apoptosis

This study explored the effects of indole-3-carbinol on the proliferation of human nasopharyngeal carcinoma, both in vitro and in vivo, and the underlying mechanisms in inducing apoptosis of CNE1 cells. Proliferation, apoptosis, malondialdehyde, superoxide dismutase, glutathione peroxidase, expressions of caspase-9, and caspase-3 in human nasopharyngeal carcinoma cells CNE1 were examined. Indole-3-carbinol suppressed proliferation, induced apoptosis, decreased malondialdehyde level, increased the activity of superoxide dismutase and glutathione peroxidase, and up-regulated the expression of active fragments of caspase-9 and caspase-3 both in vitro and in vivo. It was concluded that indole-3-carbinol could inhibit proliferation and induce apoptosis of CNE1 cells and inhibit tumor growth in mice. Increased activity of superoxide dismutase and glutathione peroxidase and activated expression of caspase-9 and caspase-3 were also observed in indole-3-carbinol–treated tumors or tumor cells, suggesting that stress- and apoptosis-related molecules are involved in the indole-3-carbinol–induced apoptosis and inhibition of tumor growth.

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