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TRAIL resistance in human neuroblastoma SK-N-AS cells is dependent on protein kinase C and involves inhibition of caspase-3 proteolytic processing
Stockholm University, Faculty of Science, Department of Neurochemistry.
Stockholm University, Faculty of Science, Department of Neurochemistry.ORCID iD: 0000-0002-1007-747X
Stockholm University, Faculty of Science, Department of Neurochemistry.ORCID iD: 0000-0002-0308-1964
2012 (English)In: Journal of Neuro-Oncology, ISSN 0167-594X, E-ISSN 1573-7373, Vol. 109, no 3, 503-512 p.Article in journal (Refereed) Published
Abstract [en]

Neuroblastoma is the most common solid extracranial cancer form in childhood with an etiology that is mostly unknown. Although tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has been proposed as a promising future anticancer drug candidate, highly malignant neuroblastoma has been reported to acquire TRAIL resistance by mechanisms that are poorly understood. Here, we show by western blot analysis, and live cell imaging using anchored FRET sensors, that the resistance to TRAIL-induced apoptosis in human neuroblastoma SK-N-AS cells depends on an incomplete processing of procaspase-3, generating an immature and catalytically inactive 21 kDa fragment. We have previously shown that the naturally occurring compound curcumin can sensitize SK-N-AS cells to TRAIL. In the present study, we show that curcumin also has a similar effect on human neuroblastoma SHEP1 cells. Furthermore, we show that curcumin and TRAIL co-treatment induces complete maturation and activation of caspase-3 in both cell lines. The mechanisms behind this effect seem to be dependent on protein kinase C (PKC), since inhibition of PKC using bisindolylmaleimide XI, could also sensitize these cells to TRAIL through a similar effect on caspase-3 activation. Moreover, TRAIL co-treatment with bisindolylmaleimide XI or curcumin resulted in down-regulation of X-linked inhibitor of apoptosis protein. In conclusion, our study shows that PKC can be involved in TRAIL resistance in human neuroblastoma cells by preventing caspase-3 maturation.

Place, publisher, year, edition, pages
2012. Vol. 109, no 3, 503-512 p.
Keyword [en]
AFS, Caspases, Curcumin, Neuroblastoma, PKC, TRAIL
National Category
Natural Sciences
Research subject
Neurochemistry with Molecular Neurobiology
Identifiers
URN: urn:nbn:se:su:diva-79146DOI: 10.1007/s11060-012-0932-2ISI: 000308441700007PubMedID: 22798207OAI: oai:DiVA.org:su-79146DiVA: diva2:547696
Funder
Swedish Research Council
Available from: 2012-08-28 Created: 2012-08-28 Last updated: 2017-12-07Bibliographically approved
In thesis
1. Regulation of apoptotic processes in neurodegeneration and cancer: In vitro studies on human neuroblastoma cells
Open this publication in new window or tab >>Regulation of apoptotic processes in neurodegeneration and cancer: In vitro studies on human neuroblastoma cells
2012 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Apoptosis is a highly controlled process of cell death, which is vital for maintenance of all multicellular organisms. Aberrant regulation of apoptosis can give rise to pathological conditions such as neurodegenerative diseases and cancer. Here, we used different human neuroblastoma cell lines to study mechanisms that may be involved in either neurodegeneration or resistance to cancer treatment. First, we have designed and developed tau-anchored FRET sensors (tAFSs) for live cell imaging of local caspase activation. Using these sensors we showed that the Alzheimer’s disease related neurotoxic peptide, amyloid-β, induced a global activation of caspase-3 and -6, but not -9, in neuronally differentiated SH-SY5Y cells. We also investigated the possible role of NF-κB in the resistance to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) in different human neuroblastoma cell lines. While N-type SH-SY5Y and IMR32 cells were unaffected, S-type SK-N-AS cells were clearly sensitized to TRAIL by different NF-κB inhibitory agents. However, no correlation between NF-κB inhibition and sensitization to TRAIL could be observed. Instead, induction of reactive oxygen species (ROS) seemed to play a more important role. Furthermore, using tAFSs we also showed that TRAIL resistance in SK-N-AS cells is mainly due to incomplete activation of caspase-3, and could be reversed by different PKC inhibitors.

Place, publisher, year, edition, pages
Stockholm: Department of Neurochemistry, Stockholm University, 2012. 71 p.
National Category
Natural Sciences
Research subject
Neurochemistry with Molecular Neurobiology
Identifiers
urn:nbn:se:su:diva-79157 (URN)978-91-7447-553-1 (ISBN)
Public defence
2012-10-19, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 13:00 (English)
Opponent
Supervisors
Note

At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 2: Manuscript.

Available from: 2012-09-27 Created: 2012-08-28 Last updated: 2012-09-05Bibliographically approved
2. Caspase activation in human neuroblastoma cells: mechanisms and spatiotemporal aspects
Open this publication in new window or tab >>Caspase activation in human neuroblastoma cells: mechanisms and spatiotemporal aspects
2015 (English)Licentiate thesis, comprehensive summary (Other academic)
Abstract [en]

Apoptosis is one of the modes of programmed cell death, in which several members of the caspase family of proteases play the central role. However, activation of apoptotic caspases does not necessarily lead to cell death. Instead, these caspases may mediate, for instance, differentiation or synaptic plasticity, if their activity is restricted in space and time. Such localized caspase activation has been also implicated in the initial stages of neurodegeneration. In order to assess this kind of events at a subcellular level, our research group has previously constructed tau-anchored FRET-based caspase sensors (tAFSs). Here, we demonstrate that localization of tAFSs to the cytoskeleton results in enrichment of the sensors in neuritic processes and enables increased spatiotemporal resolution for live cell imaging of caspase activation, as compared to soluble FRET sensors. This feature is particularly beneficial for investigation of neurodegeneration-related processes.

tAFSs were further employed for investigation of caspase activation in neuroblastoma, an extracranial solid pediatric tumor. Tumor necrosis factor-related apoptosis inducing factor (TRAIL) is a promising candidate for cancer treatment due to its ability to selectively trigger apoptosis malignant cells. However, many cancer cells, including neuroblastoma, acquire resistance to TRAIL. Here, we show that in S-type neuroblastoma cell lines, TRAIL resistance is dependent on incomplete activation of apoptotic caspase-3. Sensitization to TRAIL was achieved with protein kinase C (PKC)-inhibiting compounds, suggesting a role for this kinase in blocking the apoptotic response to TRAIL. This effect of PKC could possibly involve stabilization of XIAP, an endogenous caspase inhibitor, as PKC inhibition, in combination with TRAIL treatment, led to downregulation of XIAP.

Place, publisher, year, edition, pages
Stockholm: Department of Neurochemistry, 2015
Keyword
apoptosis, caspase, local caspase activation, neurodegeneration, tau-anchored FRET sensors, tAFS, cancer, neuroblastoma, TNF-related apoptosis inducing ligand, TRAIL, protein kinase C, PKC, X-linked inhibitor of apoptosis protein, XIAP
National Category
Chemical Sciences Biochemistry and Molecular Biology Cell Biology
Research subject
Neurochemistry with Molecular Neurobiology
Identifiers
urn:nbn:se:su:diva-122651 (URN)978-91-7649-303-8 (ISBN)
Presentation
2015-11-27, Heilbronnsalen, C458, Svante Arrhenius väg 16B, Stockholm, 14:00 (English)
Opponent
Supervisors
Available from: 2015-11-12 Created: 2015-11-05 Last updated: 2015-11-12Bibliographically approved

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