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Inhibition of the immunoproteasome ameliorates experimental autoimmune encephalomyelitis.

Basler M, Mundt S, Muchamuel T, Moll C, Jiang J, Groettrup M, Kirk CJ - EMBO Mol Med (2014)

Bottom Line: The immunoproteasome is a distinct class of proteasomes found predominantly in monocytes and lymphocytes.Additionally, ONX 0914 treatment prevented disease exacerbation in a relapsing-remitting model.These results implicate the immunoproteasome in the development of EAE and suggest that immunoproteasome inhibitors are promising drugs for the treatment of MS.

View Article: PubMed Central - PubMed

Affiliation: Biotechnology Institute Thurgau (BITg) at the University of Konstanz, Kreuzlingen, Switzerland.

ABSTRACT
Multiple sclerosis (MS) is a chronic demyelinating immune mediated disease of the central nervous system. The immunoproteasome is a distinct class of proteasomes found predominantly in monocytes and lymphocytes. Recently, we demonstrated a novel function of immunoproteasomes in cytokine production and T cell differentiation. In this study, we investigated the therapeutic efficacy of an inhibitor of the immunoproteasome (ONX 0914) in two different mouse models of MS. ONX 0914 attenuated disease progression after active and passive induction of experimental autoimmune encephalomyelitis (EAE), both in MOG₃₅-₅₅ and PLP₁₃₉₋₁₅₁-induced EAE. Isolation of lymphocytes from the brain or spinal cord revealed a strong reduction of cytokine-producing CD4(+) cells in ONX 0914 treated mice. Additionally, ONX 0914 treatment prevented disease exacerbation in a relapsing-remitting model. An analysis of draining lymph nodes after induction of EAE revealed that the differentiation to Th17 or Th1 cells was strongly impaired in ONX 0914 treated mice. These results implicate the immunoproteasome in the development of EAE and suggest that immunoproteasome inhibitors are promising drugs for the treatment of MS.

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ONX 0914 blocks GM-CSF production of T cells.Cytokine concentrations measured by ELISA are presented as the mean s.e.m. from triplicate wells. Data represent one out of three experiments.GM-CSF production of ONX 0914 (300 nM) treated splenocytes stimulated with plate bound anti-CD3/anti-CD28 Abs for 96 h as analyzed by ELISA.GM-CSF production of ONX 0914 (300 nM) treated splenocytes stimulated with plate bound anti-CD3/anti-CD28 Abs in the presence of neutralizing Abs to IFN-c and IL-12 for 96 h as analyzed by ELISA.Assessment of GM-CSF and IL-23 production of human PBMCs. PBMCs were incubated with indicated concentrations of ONX 0914 for 2 h and stimulated with plate bound anti-CD3/anti-CD28 Abs (GM-CSF) or LPS (IL-23) for 48 h.
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fig05: ONX 0914 blocks GM-CSF production of T cells.Cytokine concentrations measured by ELISA are presented as the mean s.e.m. from triplicate wells. Data represent one out of three experiments.GM-CSF production of ONX 0914 (300 nM) treated splenocytes stimulated with plate bound anti-CD3/anti-CD28 Abs for 96 h as analyzed by ELISA.GM-CSF production of ONX 0914 (300 nM) treated splenocytes stimulated with plate bound anti-CD3/anti-CD28 Abs in the presence of neutralizing Abs to IFN-c and IL-12 for 96 h as analyzed by ELISA.Assessment of GM-CSF and IL-23 production of human PBMCs. PBMCs were incubated with indicated concentrations of ONX 0914 for 2 h and stimulated with plate bound anti-CD3/anti-CD28 Abs (GM-CSF) or LPS (IL-23) for 48 h.

Mentions: Recently, we have demonstrated that LMP7 regulates secretion of IL-23 and IL-6 by activated monocytes and IFN-γ and IL-2 by T cells (Muchamuel et al, 2009). GM-CSF has been previously reported to play a pivotal role in the initiation of autoimmune neuroinflammation (Codarri et al, 2011; El-Behi et al, 2011). To investigate whether LMP7 inhibition is able to influence the secretion of this crucial cytokine in neuroinflammation, splenocytes were stimulated with plate bound CD3/CD28 antibodies in the presence or absence of ONX 0914 and GM-CSF secretion was measured 24 h later in the supernatant by ELISA (Fig 5A). LMP7-selective inhibition with ONX 0914 reduced the secretion of GM-CSF to background levels. A similar result was found when T cells were stimulated with plate bound CD3/CD28 antibodies under GM-CSF polarizing conditions (i.e. in the presence of neutralizing Abs to IFN-γ and IL-12) for 96 h as previously reported (Fig 5B) (Codarri et al, 2011). ONX 0914 treatment at 300 nM reduced cytokine secretion to background levels. To determine whether ONX 0914 can inhibit GM-CSF and IL-23 secretion of human cells, human PBMCs were incubated with varying concentrations of ONX 0914. Twenty-four hours post stimulation with plate bound CD3/CD28 antibodies, GM-CSF secretion into the supernatant was analyzed by ELISA whereas IL-23 production was assessed after LPS stimulation of PBMCs. ONX 0914 reduced IL-23 secretion to background levels at 100 nM (Fig 5C bottom panel) in accordance with our previous report (Muchamuel et al, 2009), while it diminished GM-CSF production by human PBMCs in a dose-dependent manner to approximately 50% at 300 nM.


Inhibition of the immunoproteasome ameliorates experimental autoimmune encephalomyelitis.

Basler M, Mundt S, Muchamuel T, Moll C, Jiang J, Groettrup M, Kirk CJ - EMBO Mol Med (2014)

ONX 0914 blocks GM-CSF production of T cells.Cytokine concentrations measured by ELISA are presented as the mean s.e.m. from triplicate wells. Data represent one out of three experiments.GM-CSF production of ONX 0914 (300 nM) treated splenocytes stimulated with plate bound anti-CD3/anti-CD28 Abs for 96 h as analyzed by ELISA.GM-CSF production of ONX 0914 (300 nM) treated splenocytes stimulated with plate bound anti-CD3/anti-CD28 Abs in the presence of neutralizing Abs to IFN-c and IL-12 for 96 h as analyzed by ELISA.Assessment of GM-CSF and IL-23 production of human PBMCs. PBMCs were incubated with indicated concentrations of ONX 0914 for 2 h and stimulated with plate bound anti-CD3/anti-CD28 Abs (GM-CSF) or LPS (IL-23) for 48 h.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3927957&req=5

fig05: ONX 0914 blocks GM-CSF production of T cells.Cytokine concentrations measured by ELISA are presented as the mean s.e.m. from triplicate wells. Data represent one out of three experiments.GM-CSF production of ONX 0914 (300 nM) treated splenocytes stimulated with plate bound anti-CD3/anti-CD28 Abs for 96 h as analyzed by ELISA.GM-CSF production of ONX 0914 (300 nM) treated splenocytes stimulated with plate bound anti-CD3/anti-CD28 Abs in the presence of neutralizing Abs to IFN-c and IL-12 for 96 h as analyzed by ELISA.Assessment of GM-CSF and IL-23 production of human PBMCs. PBMCs were incubated with indicated concentrations of ONX 0914 for 2 h and stimulated with plate bound anti-CD3/anti-CD28 Abs (GM-CSF) or LPS (IL-23) for 48 h.
Mentions: Recently, we have demonstrated that LMP7 regulates secretion of IL-23 and IL-6 by activated monocytes and IFN-γ and IL-2 by T cells (Muchamuel et al, 2009). GM-CSF has been previously reported to play a pivotal role in the initiation of autoimmune neuroinflammation (Codarri et al, 2011; El-Behi et al, 2011). To investigate whether LMP7 inhibition is able to influence the secretion of this crucial cytokine in neuroinflammation, splenocytes were stimulated with plate bound CD3/CD28 antibodies in the presence or absence of ONX 0914 and GM-CSF secretion was measured 24 h later in the supernatant by ELISA (Fig 5A). LMP7-selective inhibition with ONX 0914 reduced the secretion of GM-CSF to background levels. A similar result was found when T cells were stimulated with plate bound CD3/CD28 antibodies under GM-CSF polarizing conditions (i.e. in the presence of neutralizing Abs to IFN-γ and IL-12) for 96 h as previously reported (Fig 5B) (Codarri et al, 2011). ONX 0914 treatment at 300 nM reduced cytokine secretion to background levels. To determine whether ONX 0914 can inhibit GM-CSF and IL-23 secretion of human cells, human PBMCs were incubated with varying concentrations of ONX 0914. Twenty-four hours post stimulation with plate bound CD3/CD28 antibodies, GM-CSF secretion into the supernatant was analyzed by ELISA whereas IL-23 production was assessed after LPS stimulation of PBMCs. ONX 0914 reduced IL-23 secretion to background levels at 100 nM (Fig 5C bottom panel) in accordance with our previous report (Muchamuel et al, 2009), while it diminished GM-CSF production by human PBMCs in a dose-dependent manner to approximately 50% at 300 nM.

Bottom Line: The immunoproteasome is a distinct class of proteasomes found predominantly in monocytes and lymphocytes.Additionally, ONX 0914 treatment prevented disease exacerbation in a relapsing-remitting model.These results implicate the immunoproteasome in the development of EAE and suggest that immunoproteasome inhibitors are promising drugs for the treatment of MS.

View Article: PubMed Central - PubMed

Affiliation: Biotechnology Institute Thurgau (BITg) at the University of Konstanz, Kreuzlingen, Switzerland.

ABSTRACT
Multiple sclerosis (MS) is a chronic demyelinating immune mediated disease of the central nervous system. The immunoproteasome is a distinct class of proteasomes found predominantly in monocytes and lymphocytes. Recently, we demonstrated a novel function of immunoproteasomes in cytokine production and T cell differentiation. In this study, we investigated the therapeutic efficacy of an inhibitor of the immunoproteasome (ONX 0914) in two different mouse models of MS. ONX 0914 attenuated disease progression after active and passive induction of experimental autoimmune encephalomyelitis (EAE), both in MOG₃₅-₅₅ and PLP₁₃₉₋₁₅₁-induced EAE. Isolation of lymphocytes from the brain or spinal cord revealed a strong reduction of cytokine-producing CD4(+) cells in ONX 0914 treated mice. Additionally, ONX 0914 treatment prevented disease exacerbation in a relapsing-remitting model. An analysis of draining lymph nodes after induction of EAE revealed that the differentiation to Th17 or Th1 cells was strongly impaired in ONX 0914 treated mice. These results implicate the immunoproteasome in the development of EAE and suggest that immunoproteasome inhibitors are promising drugs for the treatment of MS.

Show MeSH
Related in: MedlinePlus