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Maritoclax and dinaciclib inhibit MCL-1 activity and induce apoptosis in both a MCL-1-dependent and -independent manner.

Varadarajan S, Poornima P, Milani M, Gowda K, Amin S, Wang HG, Cohen GM - Oncotarget (2015)

Bottom Line: Furthermore, maritoclax induced extensive mitochondrial fragmentation, and a Bax/Bak- but MCL-1-independent accumulation of mitochondrial reactive oxygen species (ROS), with an accompanying loss of complexes I and III of the electron transport chain.ROS scavengers, such as MitoQ, could not salvage maritoclax-mediated effects on mitochondrial structure and function.Although dinaciclib is clearly not a specific MCL-1 inhibitor, its ability to rapidly downregulate MCL-1 may be beneficial in many clinical settings, where it may reverse chemoresistance or sensitize to other chemotherapeutic agents.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Clinical Cancer Medicine, University of Liverpool, Liverpool, UK.

ABSTRACT
The anti-apoptotic BCL-2 family proteins are important targets for cancer chemotherapy. Specific and potent inhibitors of the BCL-2 family, such as ABT-263 (navitoclax) and ABT-199, are only effective against some members of the BCL-2 family but do not target MCL-1, which is commonly amplified in tumors and associated with chemoresistance. In this report, the selectivity and potency of two putative MCL-1 inhibitors, dinaciclib and maritoclax, were assessed. Although both compounds induced Bax/Bak- and caspase-9-dependent apoptosis, dinaciclib was more potent than maritoclax in downregulating MCL-1 and also in inducing apoptosis. However, the compounds induced apoptosis, even in cells lacking MCL-1, suggesting multiple mechanisms of cell death. Furthermore, maritoclax induced extensive mitochondrial fragmentation, and a Bax/Bak- but MCL-1-independent accumulation of mitochondrial reactive oxygen species (ROS), with an accompanying loss of complexes I and III of the electron transport chain. ROS scavengers, such as MitoQ, could not salvage maritoclax-mediated effects on mitochondrial structure and function. Taken together, our data demonstrate that neither dinaciclib nor maritoclax exclusively target MCL-1. Although dinaciclib is clearly not a specific MCL-1 inhibitor, its ability to rapidly downregulate MCL-1 may be beneficial in many clinical settings, where it may reverse chemoresistance or sensitize to other chemotherapeutic agents.

No MeSH data available.


Related in: MedlinePlus

Dinaciclib and maritoclax exert their effects upstream of cytochrome c release and loss in mitochondrial membrane potential(A) H460 cells, grown on coverslips, were exposed for 8 or 16 h to dinaciclib (30 nM) or maritoclax (3 μM), stained with the indicated primary and secondary antibodies and subjected to confocal microscopy. Scale bar – 10 μm. (B and C) H460 cells exposed to dinaciclib (30 nM) or maritoclax (3 μM) for the indicated times were stained with (B) TMRE to monitor changes in mitochondrial membrane potential (φm) or (C) Annexin V-FITC to assess cell death by measuring PS externalization. Error bars represent the Mean ± SEM from three independent experiments. (D) Whole cell lysates from H460 cells, exposed to dinaciclib (30 nM), maritoclax (3 μM) or dimethoxymaritoclax (3 μM) for the indicated times, were immunoblotted with the indicated antibodies. Recombinant BFL-1 protein was used as a positive control for the BFL-1 blot.
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Figure 3: Dinaciclib and maritoclax exert their effects upstream of cytochrome c release and loss in mitochondrial membrane potential(A) H460 cells, grown on coverslips, were exposed for 8 or 16 h to dinaciclib (30 nM) or maritoclax (3 μM), stained with the indicated primary and secondary antibodies and subjected to confocal microscopy. Scale bar – 10 μm. (B and C) H460 cells exposed to dinaciclib (30 nM) or maritoclax (3 μM) for the indicated times were stained with (B) TMRE to monitor changes in mitochondrial membrane potential (φm) or (C) Annexin V-FITC to assess cell death by measuring PS externalization. Error bars represent the Mean ± SEM from three independent experiments. (D) Whole cell lysates from H460 cells, exposed to dinaciclib (30 nM), maritoclax (3 μM) or dimethoxymaritoclax (3 μM) for the indicated times, were immunoblotted with the indicated antibodies. Recombinant BFL-1 protein was used as a positive control for the BFL-1 blot.

Mentions: Although our data in MEFs demonstrated that dinaciclib and maritoclax could induce apoptosis in a MCL-1-independent manner, these compounds exerted apoptosis in a Bax/Bak and caspase-9-dependent manner (Fig. 1B). Moreover, dinaciclib and maritoclax caused a time-dependent release of cytochrome c (Fig. 3A), with a concomitant loss in mitochondrial membrane potential (Fig. 3B) and an increase in PS externalization (Fig. 3C). As these observations placed the immediate effects of dinaciclib and maritoclax at the level of mitochondria, we wished to assess if the compounds modulated other anti-apoptotic members of the BCL-2 family, which may in turn result in apoptosis. In H460 cells, none of the anti-apoptotic members such as BCL-2, BCL-XL or BCL-w were downregulated following either dinaciclib or maritoclax (Fig. 3D). Furthermore, these cells did not express detectable levels of BFL-1 (Fig. 3D). Taken together, our data revealed that maritoclax and dinaciclib may inhibit MCL-1 and induce apoptosis in a cell-type specific manner.


Maritoclax and dinaciclib inhibit MCL-1 activity and induce apoptosis in both a MCL-1-dependent and -independent manner.

Varadarajan S, Poornima P, Milani M, Gowda K, Amin S, Wang HG, Cohen GM - Oncotarget (2015)

Dinaciclib and maritoclax exert their effects upstream of cytochrome c release and loss in mitochondrial membrane potential(A) H460 cells, grown on coverslips, were exposed for 8 or 16 h to dinaciclib (30 nM) or maritoclax (3 μM), stained with the indicated primary and secondary antibodies and subjected to confocal microscopy. Scale bar – 10 μm. (B and C) H460 cells exposed to dinaciclib (30 nM) or maritoclax (3 μM) for the indicated times were stained with (B) TMRE to monitor changes in mitochondrial membrane potential (φm) or (C) Annexin V-FITC to assess cell death by measuring PS externalization. Error bars represent the Mean ± SEM from three independent experiments. (D) Whole cell lysates from H460 cells, exposed to dinaciclib (30 nM), maritoclax (3 μM) or dimethoxymaritoclax (3 μM) for the indicated times, were immunoblotted with the indicated antibodies. Recombinant BFL-1 protein was used as a positive control for the BFL-1 blot.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4494965&req=5

Figure 3: Dinaciclib and maritoclax exert their effects upstream of cytochrome c release and loss in mitochondrial membrane potential(A) H460 cells, grown on coverslips, were exposed for 8 or 16 h to dinaciclib (30 nM) or maritoclax (3 μM), stained with the indicated primary and secondary antibodies and subjected to confocal microscopy. Scale bar – 10 μm. (B and C) H460 cells exposed to dinaciclib (30 nM) or maritoclax (3 μM) for the indicated times were stained with (B) TMRE to monitor changes in mitochondrial membrane potential (φm) or (C) Annexin V-FITC to assess cell death by measuring PS externalization. Error bars represent the Mean ± SEM from three independent experiments. (D) Whole cell lysates from H460 cells, exposed to dinaciclib (30 nM), maritoclax (3 μM) or dimethoxymaritoclax (3 μM) for the indicated times, were immunoblotted with the indicated antibodies. Recombinant BFL-1 protein was used as a positive control for the BFL-1 blot.
Mentions: Although our data in MEFs demonstrated that dinaciclib and maritoclax could induce apoptosis in a MCL-1-independent manner, these compounds exerted apoptosis in a Bax/Bak and caspase-9-dependent manner (Fig. 1B). Moreover, dinaciclib and maritoclax caused a time-dependent release of cytochrome c (Fig. 3A), with a concomitant loss in mitochondrial membrane potential (Fig. 3B) and an increase in PS externalization (Fig. 3C). As these observations placed the immediate effects of dinaciclib and maritoclax at the level of mitochondria, we wished to assess if the compounds modulated other anti-apoptotic members of the BCL-2 family, which may in turn result in apoptosis. In H460 cells, none of the anti-apoptotic members such as BCL-2, BCL-XL or BCL-w were downregulated following either dinaciclib or maritoclax (Fig. 3D). Furthermore, these cells did not express detectable levels of BFL-1 (Fig. 3D). Taken together, our data revealed that maritoclax and dinaciclib may inhibit MCL-1 and induce apoptosis in a cell-type specific manner.

Bottom Line: Furthermore, maritoclax induced extensive mitochondrial fragmentation, and a Bax/Bak- but MCL-1-independent accumulation of mitochondrial reactive oxygen species (ROS), with an accompanying loss of complexes I and III of the electron transport chain.ROS scavengers, such as MitoQ, could not salvage maritoclax-mediated effects on mitochondrial structure and function.Although dinaciclib is clearly not a specific MCL-1 inhibitor, its ability to rapidly downregulate MCL-1 may be beneficial in many clinical settings, where it may reverse chemoresistance or sensitize to other chemotherapeutic agents.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Clinical Cancer Medicine, University of Liverpool, Liverpool, UK.

ABSTRACT
The anti-apoptotic BCL-2 family proteins are important targets for cancer chemotherapy. Specific and potent inhibitors of the BCL-2 family, such as ABT-263 (navitoclax) and ABT-199, are only effective against some members of the BCL-2 family but do not target MCL-1, which is commonly amplified in tumors and associated with chemoresistance. In this report, the selectivity and potency of two putative MCL-1 inhibitors, dinaciclib and maritoclax, were assessed. Although both compounds induced Bax/Bak- and caspase-9-dependent apoptosis, dinaciclib was more potent than maritoclax in downregulating MCL-1 and also in inducing apoptosis. However, the compounds induced apoptosis, even in cells lacking MCL-1, suggesting multiple mechanisms of cell death. Furthermore, maritoclax induced extensive mitochondrial fragmentation, and a Bax/Bak- but MCL-1-independent accumulation of mitochondrial reactive oxygen species (ROS), with an accompanying loss of complexes I and III of the electron transport chain. ROS scavengers, such as MitoQ, could not salvage maritoclax-mediated effects on mitochondrial structure and function. Taken together, our data demonstrate that neither dinaciclib nor maritoclax exclusively target MCL-1. Although dinaciclib is clearly not a specific MCL-1 inhibitor, its ability to rapidly downregulate MCL-1 may be beneficial in many clinical settings, where it may reverse chemoresistance or sensitize to other chemotherapeutic agents.

No MeSH data available.


Related in: MedlinePlus