scholarly journals The application of BH3 mimetics in myeloid leukemias

2021 ◽  
Vol 12 (2) ◽  
Author(s):  
Narissa Parry ◽  
Helen Wheadon ◽  
Mhairi Copland
Keyword(s):  
Specific Inhibitor ◽  
Standard Of Care ◽  
Mitochondrial Outer Membrane ◽  
Great Success ◽  
Intermembrane Space ◽  
Cytochrome C Release ◽  
Apoptotic Pathway ◽  
Bh3 Mimetics ◽  
Myeloid Leukemias ◽  
Need To Evaluate

AbstractExecution of the intrinsic apoptotic pathway is controlled by the BCL-2 proteins at the level of the mitochondrial outer membrane (MOM). This family of proteins consists of prosurvival (e.g., BCL-2, MCL-1) and proapoptotic (e.g., BIM, BAD, HRK) members, the functional balance of which dictates the activation of BAX and BAK. Once activated, BAX/BAK form pores in the MOM, resulting in cytochrome c release from the mitochondrial intermembrane space, leading to apoptosome formation, caspase activation, and cleavage of intracellular targets. This pathway is induced by cellular stress including DNA damage, cytokine and growth factor withdrawal, and chemotherapy/drug treatment. A well-documented defense of leukemia cells is to shift the balance of the BCL-2 family in favor of the prosurvival proteins to protect against such intra- and extracellular stimuli. Small molecule inhibitors targeting the prosurvival proteins, named ‘BH3 mimetics’, have come to the fore in recent years to treat hematological malignancies, both as single agents and in combination with standard-of-care therapies. The most significant example of these is the BCL-2-specific inhibitor venetoclax, given in combination with standard-of-care therapies with great success in AML in clinical trials. As the number and variety of available BH3 mimetics increases, and investigations into applying these novel inhibitors to treat myeloid leukemias continue apace the need to evaluate where we currently stand in this rapidly expanding field is clear.

Mitochondria in cell death

2010 ◽  
Vol 47 ◽  
pp. 99-114 ◽  
Author(s):  
Melissa J. Parsons ◽  
Douglas R. Green
Keyword(s):  
Cell Death ◽  
Neurological Diseases ◽  
Mitochondrial Outer Membrane ◽  
Intermembrane Space ◽  
Apoptotic Pathway ◽  
Life And Death ◽  
Mitochondrial Outer Membrane Permeabilization ◽  
Mitochondrial Functions ◽  
A Cell ◽  
Signalling Cascade

Apoptosis can be thought of as a signalling cascade that results in the death of the cell. Properly executed apoptosis is critically important for both development and homoeostasis of most animals. Accordingly, defects in apoptosis can contribute to the development of autoimmune disorders, neurological diseases and cancer. Broadly speaking, there are two main pathways by which a cell can engage apoptosis: the extrinsic apoptotic pathway and the intrinsic apoptotic pathway. At the centre of the intrinsic apoptotic signalling pathway lies the mitochondrion, which, in addition to its role as the bioenergetic centre of the cell, is also the cell’s reservoir of pro-death factors which reside in the mitochondrial IMS (intermembrane space). During intrinsic apoptosis, pores are formed in the OMM (outer mitochondrial membrane) of the mitochondria in a process termed MOMP (mitochondrial outer membrane permeabilization). This allows for the release of IMS proteins; once released during MOMP, some IMS proteins, notably cytochrome c and Smac/DIABLO (Second mitochondria-derived activator of caspase/direct inhibitor of apoptosis-binding protein with low pI), promote caspase activation and subsequent cleavage of structural and regulatory proteins in the cytoplasm and the nucleus, leading to the demise of the cell. MOMP is achieved through the co-ordinated actions of pro-apoptotic members and inhibited by anti-apoptotic members of the Bcl-2 family of proteins. Other aspects of mitochondrial physiology, such as mitochondrial bioenergetics and dynamics, are also involved in processes of cell death that proceed through the mitochondria. Proper regulation of these mitochondrial functions is vitally important for the life and death of the cell and for the organism as a whole.

scholarly journals Regorafenib Alteration of the BCL-xL/MCL-1 Ratio Provides a Therapeutic Opportunity for BH3-Mimetics in Hepatocellular Carcinoma Models

Cancers ◽  
2020 ◽  
Vol 12 (2) ◽  
pp. 332 ◽  
Author(s):  
Blanca Cucarull ◽  
Anna Tutusaus ◽  
Miguel Subías ◽  
Milica Stefanovic ◽  
Tania Hernáez-Alsina ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Death ◽  
Hepatoma Cell ◽  
Mitochondrial Outer Membrane ◽  
Cytochrome C Release ◽  
Multikinase Inhibitor ◽  
Bh3 Mimetics ◽  
Mitochondrial Outer Membrane Permeabilization

Background: The multikinase inhibitor regorafenib, approved as second-line treatment for hepatocellular carcinoma (HCC) after sorafenib failure, may induce mitochondrial damage. BH3-mimetics, inhibitors of specific BCL-2 proteins, are valuable drugs in cancer therapy to amplify mitochondrial-dependent cell death. Methods: In in vitro and in vivo HCC models, we tested regorafenib’s effect on the BCL-2 network and the efficacy of BH3-mimetics on HCC treatment. Results: In hepatoma cell lines and Hep3B liver spheroids, regorafenib cytotoxicity was potentiated by BCL-xL siRNA transfection or pharmacological inhibition (A-1331852), while BCL-2 antagonism had no effect. Mitochondrial outer membrane permeabilization, cytochrome c release, and caspase-3 activation mediated A-1331852/regorafenib-induced cell death. In a patient-derived xenograft (PDX) HCC model, BCL-xL inhibition stimulated regorafenib activity, drastically decreasing tumor growth. Moreover, regorafenib-resistant HepG2 cells displayed increased BCL-xL and reduced MCL-1 expression, while A-1331852 reinstated regorafenib efficacy in vitro and in a xenograft mouse model. Interestingly, BCL-xL levels, associated with poor prognosis in liver and colorectal cancer, and the BCL-xL/MCL-1 ratio were detected as being increased in HCC patients. Conclusion: Regorafenib primes tumor cells to BH3-mimetic-induced cell death, allowing BCL-xL inhibition with A-1331852 or other strategies based on BCL-xL degradation to enhance regorafenib efficacy, offering a novel approach for HCC treatment, particularly for tumors with an elevated BCL-xL/MCL-1 ratio.

scholarly journals The Mitochondrial TOM Complex Is Required for tBid/Bax-induced Cytochrome c Release

2007 ◽  
Vol 282 (38) ◽  
pp. 27633-27639 ◽  
Author(s):  
Martin Ott ◽  
Erik Norberg ◽  
Katharina M. Walter ◽  
Patrick Schreiner ◽  
Christian Kemper ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cytochrome C ◽  
Outer Membrane ◽  
Recombinant Proteins ◽  
Mitochondrial Membrane ◽  
Mitochondrial Outer Membrane ◽  
Intermembrane Space ◽  
Outer Mitochondrial Membrane ◽  
Cytochrome C Release ◽  
Unknown Mechanism

Cytochrome c release from mitochondria is a key event in apoptosis signaling that is regulated by Bcl-2 family proteins. Cleavage of the BH3-only protein Bid by multiple proteases leads to the formation of truncated Bid (tBid), which, in turn, promotes the oligomerization/insertion of Bax into the mitochondrial outer membrane and the resultant release of proteins residing in the intermembrane space. Bax, a monomeric protein in the cytosol, is targeted by a yet unknown mechanism to the mitochondria. Several hypotheses have been put forward to explain this targeting specificity. Using mitochondria isolated from different mutants of the yeast Saccharomyces cerevisiae and recombinant proteins, we have now investigated components of the mitochondrial outer membrane that might be required for tBid/Bax-induced cytochrome c release. Here, we show that the protein translocase of the outer mitochondrial membrane is required for Bax insertion and cytochrome c release.

scholarly journals BCL2 inhibitors and MCL1 inhibitors for hematological malignancies

Blood ◽  
2021 ◽  
Author(s):  
Andrew W. Roberts ◽  
Andrew H Wei ◽  
David Ching Siang Huang
Keyword(s):  
Single Agent ◽  
Specific Inhibitor ◽  
Lymphocytic Leukemia ◽  
Tolerability Profile ◽  
Cell Of Origin ◽  
Apoptotic Pathway ◽  
Secondary Resistance ◽  
Bh3 Mimetics ◽  
Major Activity ◽  
Hematological Cancers

BCL2 and MCL1 are commonly expressed pro-survival (anti-apoptotic) proteins in hematological cancers and play important roles in their biology either through dysregulation or by virtue of intrinsic importance to the cell-of-origin of the malignancy. A new class of small molecule anti-cancer drugs, BH3-mimetics, now enable specific targeting of these proteins in patients. BH3-mimetics act by inhibiting the pro-survival BCL2 proteins to enable the activation of BAX and BAK, apoptosis effectors which permeabilize the outer mitochondrial membrane, triggering apoptosis directly in many cells and sensitizing others to cell death when combined with other anti-neoplastic drugs. Venetoclax, a specific inhibitor of BCL2, is the first approved in class, demonstrating striking single agent activity in chronic lymphocytic leukemia (CLL) and in other lymphoid neoplasms, as well as activity against acute myeloid leukemia (AML), especially when used in combination. Key insights from the venetoclax experience include that responses occur rapidly, with major activity as monotherapy proving to be the best indicator for success in combination regimens. This emphasizes the importance of adequate single agent studies for drugs in this class. Furthermore, secondary resistance is common with long-term exposure and often mediated by genetic or adaptive changes in the apoptotic pathway, suggesting that BH3-mimetics are better suited to limited-duration, rather than continuous, therapy. The success of venetoclax has inspired development of BH3-mimetics targeting MCL1. Despite promising preclinical activity against MYC-driven lymphomas, myeloma and AML, their success may particularly depend on their tolerability profile given physiological roles for MCL1 in several non-hematological tissues.

scholarly journals Chemical, Physical and Biological Triggers of Evolutionary Conserved Bcl-xL-Mediated Apoptosis

Cancers ◽  
2020 ◽  
Vol 12 (6) ◽  
pp. 1694 ◽  
Author(s):  
Aleksandr Ianevski ◽  
Evgeny Kulesskiy ◽  
Klara Krpina ◽  
Guofeng Lou ◽  
Yahyah Aman ◽  
...  
Keyword(s):  
Specific Inhibitor ◽  
Chemical Compounds ◽  
Mitochondrial Outer Membrane ◽  
Physical Factors ◽  
Apoptotic Pathway ◽  
C Elegans ◽  
Mitochondrial Outer Membrane Permeabilization ◽  
Evolutionarily Conserved ◽  
Novel Drug ◽  
Specific Inhibitors

Background: The evidence that pan-Bcl-2 or Bcl-xL-specific inhibitors prematurely kill virus-infected or RNA/DNA-transfected cells provides rationale for investigating these apoptotic inducers further. We hypothesized that not only invasive RNA or DNA (biological factors) but also DNA/RNA-damaging chemical or physical factors could trigger apoptosis that have been sensitized with pan-Bcl-2 or Bcl-xL-specific agents; Methods: We tested chemical and physical factors plus Bcl-xL-specific inhibitor A-1155463 in cells of various origins and the small roundworms (C. elegans); Results: We show that combination of a A-1155463 along with a DNA-damaging agent, 4-nitroquinoline-1-oxide (4NQO), prematurely kills cells of various origins as well as C. elegans. The synergistic effect is p53-dependent and associated with the release of Bad and Bax from Bcl-xL, which trigger mitochondrial outer membrane permeabilization. Furthermore, we found that combining Bcl-xL-specific inhibitors with various chemical compounds or physical insults also induced cell death; Conclusions: Thus, we were able to identify several biological, chemical and physical triggers of the evolutionarily conserved Bcl-xL-mediated apoptotic pathway, shedding light on strategies and targets for novel drug development.

scholarly journals Bax-induced cytochrome c release from mitochondria depends on alpha-helices-5 and -6

2004 ◽  
Vol 378 (1) ◽  
pp. 247-255 ◽  
Author(s):  
Gerd HEIMLICH ◽  
Alastair D. McKINNON ◽  
Katussevani BERNARDO ◽  
Dieter BRDICZKA ◽  
John C. REED ◽  
...  
Keyword(s):  
Cytochrome C ◽  
Transmembrane Domain ◽  
Mitochondrial Outer Membrane ◽  
Deletion Mutants ◽  
Intermembrane Space ◽  
Mitochondrial Membranes ◽  
Cytochrome C Release ◽  
Alpha Helices ◽  
Bh3 Domain ◽  
Apoptotic Protein

The pro-apoptotic protein Bax plays a key role in the mitochondrial signalling pathway. Upon induction of apoptosis, Bax undergoes a conformational change and translocates to mitochondrial membranes, where it inserts and mediates the release of cytochrome c from the intermembrane space into the cytosol. However, the domains of Bax that are essential for the induction of cytochrome c release are still elusive. Therefore various Bax deletion mutants were generated and expressed in Escherichia coli. The proteins were then purified in order to delineate the function of the transmembrane domain, the BH3 (Bcl-2 homology 3) domain and the putative pore-forming α-helices-5 and -6. These proteins were used to analyse the mechanism of Bax-induced cytochrome c release from mitochondria. None of the Bax proteins caused cytochrome c release merely through physical perturbation of the mitochondrial outer membrane. The α-helices-5 and -6 of Bax were shown to mediate the insertion of the protein into mitochondrial membranes and to be essential for the cytochrome c-releasing activity of Bax. In contrast, neither the transmembrane domain nor a functional BH3 domain is required for the Bax-mediated release of cytochrome c from mitochondria.

scholarly journals The Pro-Apoptotic Proteins, Bid and Bax, Cause a Limited Permeabilization of the Mitochondrial Outer Membrane That Is Enhanced by Cytosol

1999 ◽  
Vol 147 (4) ◽  
pp. 809-822 ◽  
Author(s):  
Ruth M. Kluck ◽  
Mauro Degli Esposti ◽  
Guy Perkins ◽  
Christian Renken ◽  
Tomomi Kuwana ◽  
...  
Keyword(s):  
Cytochrome C ◽  
Membrane Permeability ◽  
Outer Membrane ◽  
Mitochondrial Outer Membrane ◽  
Intermembrane Space ◽  
Cytochrome C Release ◽  
Free System ◽  
Xenopus Egg ◽  
Apoptotic Proteins ◽  
Outer Membrane Permeability

During apoptosis, an important pathway leading to caspase activation involves the release of cytochrome c from the intermembrane space of mitochondria. Using a cell-free system based on Xenopus egg extracts, we examined changes in the outer mitochondrial membrane accompanying cytochrome c efflux. The pro-apoptotic proteins, Bid and Bax, as well as factors present in Xenopus egg cytosol, each induced cytochrome c release when incubated with isolated mitochondria. These factors caused a permeabilization of the outer membrane that allowed the corelease of multiple intermembrane space proteins: cytochrome c, adenylate kinase and sulfite oxidase. The efflux process is thus nonspecific. None of the cytochrome c-releasing factors caused detectable mitochondrial swelling, arguing that matrix swelling is not required for outer membrane permeability in this system. Bid and Bax caused complete release of cytochrome c but only a limited permeabilization of the outer membrane, as measured by the accessibility of inner membrane-associated respiratory complexes III and IV to exogenously added cytochrome c. However, outer membrane permeability was strikingly increased by a macromolecular cytosolic factor, termed PEF (permeability enhancing factor). We hypothesize that PEF activity could help determine whether cells can recover from mitochondrial cytochrome c release.

The warburg effect and tumour cell survival in human GBMs

2007 ◽  
Vol 30 (4) ◽  
pp. 97 ◽  
Author(s):  
A Wolf ◽  
J Mukherjee ◽  
A Guha
Keyword(s):  
Cytochrome C ◽  
Cell Survival ◽  
Expression Profile ◽  
Tumour Cell ◽  
Warburg Effect ◽  
Glycolytic Enzymes ◽  
Cytochrome C Release ◽  
Apoptotic Pathway ◽  
The Warburg Effect

Introduction: GBMs are resistant to apoptosis induced by the hypoxic microenvironment and standard therapies including radiation and chemotherapy. We postulate that the Warburg effect, a preferential glycolytic phenotype of tumor cells even under aerobic conditions, plays a role in these aberrant pro-survival signals. In this study we quantitatively examined the expression profile of hypoxia-related glycolytic genes within pathologically- and MRI-defined “centre” and “periphery” of GBMs. We hypothesize that expression of hypoxia-induced glycolytic genes, particularly hexokinase 2 (HK2), favours cell survival and modulates resistance to tumour cell apoptosis by inhibiting the intrinsic mitochondrial apoptotic pathway. Methods: GBM patients underwent conventional T1-weighted contrast-enhanced MRI and MR spectroscopy studies on a 3.0T GE scanner, prior to stereotactic sampling (formalin and frozen) from regions which were T1-Gad enhancing (“centre”) and T2-positive, T1-Gad negative (“periphery”). Real-time qRT-PCR was performed to quantify regional gene expression of glycolytic genes including HK2. In vitro functional studies were performed in U87 and U373 GBM cell lines grown in normoxic (21% pO2) and hypoxic (< 1%pO2) conditions, transfected with HK2 siRNA followed by measurement of cell proliferation (BrdU), apoptosis (activated caspase 3/7, TUNEL, cytochrome c release) and viability (MTS assay). Results: There exists a differential expression profile of glycolytic enzymes between the hypoxic center and relatively normoxic periphery of GBMs. Under hypoxic conditions, there is increased expression of HK2 at the mitochondrial membrane in GBM cells. In vitro HK2 knockdown led to decreased cell survival and increased apoptosis via the intrinsic mitochondrial pathway, as seen by increased mitochondrial release of cytochrome-C. Conclusions: Increased expression of HK2 in the centre of GBMs promotes cell survival and confers resistance to apoptosis, as confirmed by in vitro studies. In vivo intracranial xenograft studies with injection of HK2-shRNA are currently being performed. HK2 and possibly other glycolytic enzymes may provide a target for enhanced therapeutic responsiveness thereby improving prognosis of patients with GBMs.

scholarly journals Posttranslational Modification of Bcl-2 Facilitates Its Proteasome-Dependent Degradation: Molecular Characterization of the Involved Signaling Pathway

2000 ◽  
Vol 20 (5) ◽  
pp. 1886-1896 ◽  
Author(s):  
Kristin Breitschopf ◽  
Judith Haendeler ◽  
Philipp Malchow ◽  
Andreas M. Zeiher ◽  
Stefanie Dimmeler
Keyword(s):  
Endothelial Cells ◽  
Protein Kinase ◽  
Map Kinase ◽  
Specific Inhibitor ◽  
26S Proteasome ◽  
Apoptotic Pathway ◽  
Critical Determinant ◽  
Intact Cells ◽  
Antiapoptotic Protein ◽  
Tnf Α

ABSTRACT The ratio of proapoptotic versus antiapoptotic Bcl-2 members is a critical determinant that plays a significant role in altering susceptibility to apoptosis. Therefore, a reduction of antiapoptotic protein levels in response to proximal signal transduction events may switch on the apoptotic pathway. In endothelial cells, tumor necrosis factor alpha (TNF-α) induces dephosphorylation and subsequent ubiquitin-dependent degradation of the antiapoptotic protein Bcl-2. Here, we investigate the role of different putative phosphorylation sites to facilitate Bcl-2 degradation. Mutation of the consensus protein kinase B/Akt site or of potential protein kinase C or cyclic AMP-dependent protein kinase sites does not affect Bcl-2 stability. In contrast, inactivation of the three consensus mitogen-activated protein (MAP) kinase sites leads to a Bcl-2 protein that is ubiquitinated and subsequently degraded by the 26S proteasome. Inactivation of these sites within Bcl-2 revealed that dephosphorylation of Ser87 appears to play a major role. A Ser-to-Ala substitution at this position results in 50% degradation, whereas replacement of Thr74 with Ala leads to 25% degradation, as assessed by pulse-chase studies. We further demonstrated that incubation with TNF-α induces dephosphorylation of Ser87 of Bcl-2 in intact cells. Furthermore, MAP kinase triggers phosphorylation of Bcl-2, whereas a reduction in Bcl-2 phosphorylation was observed in the presence of MAP kinase-specific phosphatases or the MAP kinase-specific inhibitor PD98059. Moreover, we show that oxidative stress mediates TNF-α-stimulated proteolytic degradation of Bcl-2 by reducing MAP kinase activity. Taken together, these results demonstrate a direct protective role for Bcl-2 phosphorylation by MAP kinase against apoptotic challenges to endothelial cells and other cells.

Sab concentrations indicate chemotherapeutic susceptibility in ovarian cancer cell lines

2018 ◽  
Vol 475 (21) ◽  
pp. 3471-3492 ◽  
Author(s):  
Iru Paudel ◽  
Sean M. Hernandez ◽  
Gilda M. Portalatin ◽  
Tara P. Chambers ◽  
Jeremy W. Chambers
Keyword(s):  
Ovarian Cancer ◽  
Cell Lines ◽  
Ovarian Cancer Cell ◽  
Prognostic Biomarker ◽  
Chemotherapeutic Agents ◽  
Mitochondrial Outer Membrane ◽  
Expression Data ◽  
Apoptotic Pathway ◽  
Gynecological Malignancy ◽  
Ovarian Cancer Cell Lines

The occurrence of chemotherapy-resistant tumors makes ovarian cancer (OC) the most lethal gynecological malignancy. While many factors may contribute to chemoresistance, the mechanisms responsible for regulating tumor vulnerability are under investigation. Our analysis of gene expression data revealed that Sab, a mitochondrial outer membrane (MOM) scaffold protein, was down-regulated in OC patients. Sab-mediated signaling induces cell death, suggesting that this apoptotic pathway is diminished in OC. We examined Sab expression in a panel of OC cell lines and found that the magnitude of Sab expression correlated to chemo-responsiveness; wherein, OC cells with low Sab levels were chemoresistant. The Sab levels were reflected by a corresponding amount of stress-induced c-Jun N-terminal kinase (JNK) on the MOM. BH3 profiling and examination of Bcl-2 and BH3-only protein concentrations revealed that cells with high Sab concentrations were primed for apoptosis, as determined by the decrease in pro-survival Bcl-2 proteins and an increase in pro-apoptotic BH3-only proteins on mitochondria. Furthermore, overexpression of Sab in chemoresistant cells enhanced apoptotic priming and restored cellular vulnerability to a combination treatment of cisplatin and paclitaxel. Contrariwise, inhibiting Sab-mediated signaling or silencing Sab expression in a chemosensitive cell line resulted in decreased apoptotic priming and increased resistance. The effects of silencing on Sab on the resistance to chemotherapeutic agents were emulated by the silencing or inhibition of JNK, which could be attributed to changes in Bcl-2 protein concentrations induced by sub-chronic JNK inhibition. We propose that Sab may be a prognostic biomarker to discern personalized treatments for OC patients.

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