Targeted inhibition of the deubiquitinating enzymes, USP14 and UCHL5, induces proteotoxic stress and apoptosis in Waldenström macroglobulinaemia tumour cells

Kasyapa Chitta, Aneel Paulus, Sharoon Akhtar, Maja Kristin K Blake, Thomas Caulfield, Anne J Novak, Stephen Maxted Ansell, Pooja Advani, Sikander Ailawadhi, Taimur Sher, Stig Linder, Asher A Chanan Khan

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28 Citations (Scopus)

Abstract

Deubiquitinase enzymes (DUBs) of the proteasomal 19S regulatory particle are emerging as important therapeutic targets in several malignancies. Here we demonstrate that inhibition of two proteasome-associated DUBs (USP14 and UCHL5) with the small molecule DUB inhibitor b-AP15, results in apoptosis of human Waldenström macroglobulinaemia (WM) cell lines and primary patient-derived WM tumour cells. Importantly, b-AP15 produced proteotoxic stress and apoptosis in WM cells that have acquired resistance to the proteasome inhibitor bortezomib. In silico modelling identified protein residues that were critical for the binding of b-AP15 with USP14 or UCHL5 and proteasome enzyme activity assays confirmed that b-AP15 does not affect the proteolytic capabilities of the 20S proteasome β-subunits. In vitro toxicity from b-AP15 appeared to result from a build-up of ubiquitinated proteins and activation of the endoplasmic reticulum stress response in WM cells, an effect that also disrupted the mitochondria. Focused transcriptome profiling of b-AP15-treated WM cells revealed modulation of several genes regulating cell stress and NF-κB signalling, the latter whose protein translocation and downstream target activation was reduced by b-AP15 in vitro. This is the first report to define the effects and underlying mechanisms associated with inhibition of USP14 and UCHL5 DUB.

Original languageEnglish (US)
Pages (from-to)377-390
Number of pages14
JournalBritish Journal of Haematology
Volume169
Issue number3
DOIs
StatePublished - May 1 2015

Fingerprint

Waldenstrom Macroglobulinemia
Apoptosis
Proteasome Endopeptidase Complex
Neoplasms
Ubiquitinated Proteins
Proteasome Inhibitors
Endoplasmic Reticulum Stress
Enzyme Assays
Gene Expression Profiling
Protein Transport
Computer Simulation
Mitochondria
Deubiquitinating Enzymes
Cell Line
Enzymes
Genes
Proteins

Keywords

  • Deubiquitinase enzymes
  • Preclinical
  • Proteasome
  • Transcriptome
  • Waldenströms macroglobulinaemia

ASJC Scopus subject areas

  • Hematology

Cite this

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title = "Targeted inhibition of the deubiquitinating enzymes, USP14 and UCHL5, induces proteotoxic stress and apoptosis in Waldenstr{\"o}m macroglobulinaemia tumour cells",
abstract = "Deubiquitinase enzymes (DUBs) of the proteasomal 19S regulatory particle are emerging as important therapeutic targets in several malignancies. Here we demonstrate that inhibition of two proteasome-associated DUBs (USP14 and UCHL5) with the small molecule DUB inhibitor b-AP15, results in apoptosis of human Waldenstr{\"o}m macroglobulinaemia (WM) cell lines and primary patient-derived WM tumour cells. Importantly, b-AP15 produced proteotoxic stress and apoptosis in WM cells that have acquired resistance to the proteasome inhibitor bortezomib. In silico modelling identified protein residues that were critical for the binding of b-AP15 with USP14 or UCHL5 and proteasome enzyme activity assays confirmed that b-AP15 does not affect the proteolytic capabilities of the 20S proteasome β-subunits. In vitro toxicity from b-AP15 appeared to result from a build-up of ubiquitinated proteins and activation of the endoplasmic reticulum stress response in WM cells, an effect that also disrupted the mitochondria. Focused transcriptome profiling of b-AP15-treated WM cells revealed modulation of several genes regulating cell stress and NF-κB signalling, the latter whose protein translocation and downstream target activation was reduced by b-AP15 in vitro. This is the first report to define the effects and underlying mechanisms associated with inhibition of USP14 and UCHL5 DUB.",
keywords = "Deubiquitinase enzymes, Preclinical, Proteasome, Transcriptome, Waldenstr{\"o}ms macroglobulinaemia",
author = "Kasyapa Chitta and Aneel Paulus and Sharoon Akhtar and Blake, {Maja Kristin K} and Thomas Caulfield and Novak, {Anne J} and Ansell, {Stephen Maxted} and Pooja Advani and Sikander Ailawadhi and Taimur Sher and Stig Linder and {Chanan Khan}, {Asher A}",
year = "2015",
month = "5",
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language = "English (US)",
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T1 - Targeted inhibition of the deubiquitinating enzymes, USP14 and UCHL5, induces proteotoxic stress and apoptosis in Waldenström macroglobulinaemia tumour cells

AU - Chitta, Kasyapa

AU - Paulus, Aneel

AU - Akhtar, Sharoon

AU - Blake, Maja Kristin K

AU - Caulfield, Thomas

AU - Novak, Anne J

AU - Ansell, Stephen Maxted

AU - Advani, Pooja

AU - Ailawadhi, Sikander

AU - Sher, Taimur

AU - Linder, Stig

AU - Chanan Khan, Asher A

PY - 2015/5/1

Y1 - 2015/5/1

N2 - Deubiquitinase enzymes (DUBs) of the proteasomal 19S regulatory particle are emerging as important therapeutic targets in several malignancies. Here we demonstrate that inhibition of two proteasome-associated DUBs (USP14 and UCHL5) with the small molecule DUB inhibitor b-AP15, results in apoptosis of human Waldenström macroglobulinaemia (WM) cell lines and primary patient-derived WM tumour cells. Importantly, b-AP15 produced proteotoxic stress and apoptosis in WM cells that have acquired resistance to the proteasome inhibitor bortezomib. In silico modelling identified protein residues that were critical for the binding of b-AP15 with USP14 or UCHL5 and proteasome enzyme activity assays confirmed that b-AP15 does not affect the proteolytic capabilities of the 20S proteasome β-subunits. In vitro toxicity from b-AP15 appeared to result from a build-up of ubiquitinated proteins and activation of the endoplasmic reticulum stress response in WM cells, an effect that also disrupted the mitochondria. Focused transcriptome profiling of b-AP15-treated WM cells revealed modulation of several genes regulating cell stress and NF-κB signalling, the latter whose protein translocation and downstream target activation was reduced by b-AP15 in vitro. This is the first report to define the effects and underlying mechanisms associated with inhibition of USP14 and UCHL5 DUB.

AB - Deubiquitinase enzymes (DUBs) of the proteasomal 19S regulatory particle are emerging as important therapeutic targets in several malignancies. Here we demonstrate that inhibition of two proteasome-associated DUBs (USP14 and UCHL5) with the small molecule DUB inhibitor b-AP15, results in apoptosis of human Waldenström macroglobulinaemia (WM) cell lines and primary patient-derived WM tumour cells. Importantly, b-AP15 produced proteotoxic stress and apoptosis in WM cells that have acquired resistance to the proteasome inhibitor bortezomib. In silico modelling identified protein residues that were critical for the binding of b-AP15 with USP14 or UCHL5 and proteasome enzyme activity assays confirmed that b-AP15 does not affect the proteolytic capabilities of the 20S proteasome β-subunits. In vitro toxicity from b-AP15 appeared to result from a build-up of ubiquitinated proteins and activation of the endoplasmic reticulum stress response in WM cells, an effect that also disrupted the mitochondria. Focused transcriptome profiling of b-AP15-treated WM cells revealed modulation of several genes regulating cell stress and NF-κB signalling, the latter whose protein translocation and downstream target activation was reduced by b-AP15 in vitro. This is the first report to define the effects and underlying mechanisms associated with inhibition of USP14 and UCHL5 DUB.

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KW - Transcriptome

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