The HMG-Co-A reductase inhibitor, atorvastatin, promotes a Th2 bias and reverses paralysis in central nervous system autoimmune disease Youssef, et. al. Presented by Andrea Crowell November 1, 2004

Multiple Sclerosis Immune system attacks myelin Immune system attacks myelin Disruption of electrical conductivity results in fatigue and disturbances of vision, strength, balance, coordination, sensations, and bladder and bowel function. Disruption of electrical conductivity results in fatigue and disturbances of vision, strength, balance, coordination, sensations, and bladder and bowel function. Commonly treated with Interferon Beta (injectable) Commonly treated with Interferon Beta (injectable)

Geographical distribution of MS in the United States

Statins and Inflammation Given to heart transplant recipients who have high cholesterol levels and need to be immunosuppressed Given to heart transplant recipients who have high cholesterol levels and need to be immunosuppressed Lower cholesterol AND suppress immune system, esp. when taken with cyclosporin Lower cholesterol AND suppress immune system, esp. when taken with cyclosporin Findings from more later studies suggest statins might inhibit antigen presentation to pro-inflammatory Th1 cells Findings from more later studies suggest statins might inhibit antigen presentation to pro-inflammatory Th1 cells

Acetyl-CoA (citric acid cycle) and acetoacetyl-CoA to 3-hydroxy-3- methylglutaryl-CoA (HMG-CoA) Acetyl-CoA (citric acid cycle) and acetoacetyl-CoA to 3-hydroxy-3- methylglutaryl-CoA (HMG-CoA) HMG-CoA to mevalonate – by HMG-CoA reductase (target of statins) HMG-CoA to mevalonate – by HMG-CoA reductase (target of statins) Mevalonate to IPP by mevalonate kinase (mutation -> HIDS: Hyperimmunoglobinemia D – recurrent fevers) Mevalonate to IPP by mevalonate kinase (mutation -> HIDS: Hyperimmunoglobinemia D – recurrent fevers) IPP to geranyl-PP by farnesyl-PP synthetase IPP to geranyl-PP by farnesyl-PP synthetase Geranyl-PP to farnesyl-PP by farnesyl-PP synthetase (target of bisphosphonates – to treat osteoporosis) Geranyl-PP to farnesyl-PP by farnesyl-PP synthetase (target of bisphosphonates – to treat osteoporosis) HMG-CoA Reductase Pathway

EAE model Experimental Autoimmune Encephalomyelitis Experimental Autoimmune Encephalomyelitis 3 types, reflecting different disease phenotypes 3 types, reflecting different disease phenotypes –MOG p35-55 chronic –PLP p relapsing-remitting –MBP Ac1-11 specific TCR transgenic fulminant

Atorvastatin prevented or reversed chronic and relapsing paralysis in EAE mice a, b: MOG p35-55 mice c, d: PLP p mice e: TCR transgenic mice – atorvastatin administered before immunization f: PLP p mice – atorvastatin dose response curve 0.01–10 mg/kg (0.01mg/kg dose did not significantly reduce EAE)

Histological evidence of EAE reduced with atorvastatin treatment Atorvastatin treatment started before EAE induction prevented (10mg/kg) or decreased the incidence and severity of relapse in SJL/J mice Treatment started during acute EAE decreased the incidence and severity of relapse in SJL/J mice

MHC II expression in CNS white matter – assessed by histology Naïve CNS PBS 1mg/kg 10mg/kg SJL/J C57BL/6 High levels of MHC II expression were observed on microglia within or adjacent to EAE lesions Atorvastatin treatment downregulated MHC II expression on these cells.

MHC II expression in CNS white matter – assessed by FACS a)MHC II expression increased when microglia were treated with IFN-gamma, compared with untreated cells b)Treating the cells with IFN-gamma + atorvastatin, MHC II expression did not differ from the untreated cells c)With IFN-gamma + atorvastatin + L mevalonate treatment, MHC II expression increased

MHC II expression in CNS white matter – assessed by immunohistochemistry e) Untreated microglia (inset shows isotype-matched control IgG staining) f) Treatment with IFN-gamma g) Treatment with IFN-gamma + atorvastatin h) Treatment with IFN-gamma + atorvastatin + L-mevalonate

MHC II expression in CNS white matter – assessed by FACS Mean peak fluorescence intensity for MHC II expression Atorvastatin reverses the effect of IFN-gamma L-mevalonate reversed the effect of atorvastatin

CIITA MHC class II transactivator (CIITA) is the master regulator of MHC II expression in antigen presenting cells CIITA has multiple promoter regions, with pIV being predominant in epithelial cells and pI predominant in bone marrow derived cells. A previous study reported that statins suppressed IFN-gamma inducible CIITA transcription.

CIITA transcription EOC microglia B10.PL microglia IFN-gamma increases CIITA transcription Atorvastatin reverses effect of IFN-gamma L-mevalonate reverses effect of atorvastatin Results suggest that statins inhibit CIITA transcription in general, rather than by selective activity at a particular promoter region.

Effect of atorvastatin on T cell activation and Th regulation Splenocyte proliferation (measured by 3H incorporation) was suppressed by 1mg/kg and 10mg/kg atorvastatin. - C57BL/6 mice were MOG p35-55 immunized - SJL/J mice were PLP p immunized Atorvastatin suppresses recall response to EAE-activating antigen in all 3 mouse models.

Effect of atorvastatin on T cell activation and Th regulation Th1-associated cytokine secretion suppressed by atorvastatin.

Effect of atorvastatin on T cell activation and Th regulation Th2-associated cytokine secretion suppressed by atorvastatin.

Results of atorvastatin treatment of naïve Th0 cells from TCR transgenic mouse: a: Suppressed proliferation of splenocytes in vitro b-e: Th1-associated cytokine secretion was suppressed f-i: Th2- associated cytokine secretion was enhanced Addition of L-mevalonate reversed these effects, indicating that atorvastatin acts via inhibition of the mevalonate pathway.

In vivo atorvastatin treatment promotes Th2 bias via STAT4 inhibition and STAT6 induction STAT 4 phosphorylation suppressed STAT4 is required for Th1 lineage commitment STAT6 phosphorylation is promoted STAT6 is required for Th2 lineage committment STAT: Signal Transducer and Activator of Transcription

Atorvastatin has immunomodulatory effects on both APC cells and T cells. In vivo atorvastatin treatment (A) or in vitro treatment (B) of only APC cells or only T cells suppressed proliferation.

Atorvastatin has immunomodulatory effects on both APC cells and T cells. Inhibition of proliferation by atorvastatin treatment of either APCs or T cells also produced a Th2 pattern of cytokine secretion. Th1 Th2

Atorvastatin has immunomodulatory effects on both APC cells and T cells. The expression levels of MHC II on primary macrophages, as well as the expression of the co-stimulatory molecules CD40, CD80, and CD86 were measured. IFN-gamma induced expression. Atorvastatin inhibited IFN-gamma-induced expression. L-Mevalonate reversed atorvastatin inhibition.

Adoptive transfer of atorvastatin-treated T cells prevents induction of EAE. Irradiated mice that received T cells from mice TCR transgenic mice treated with 10mg/kg of atorvastatin were protected against EAE induction following immunization. Irradiated mice that received T cells from mice TCR transgenic mice treated with 10mg/kg of atorvastatin were protected against EAE induction following immunization. Atorvastatin-treated T cells were divided into CD4+ and CD8+ cells. The CD4+ cells protected against EAE while the CD8+ cells did not. Atorvastatin-treated T cells were divided into CD4+ and CD8+ cells. The CD4+ cells protected against EAE while the CD8+ cells did not.

Conclusions Oral atorvastatin treatment prevented or reversed chronic and relapsing EAE paralysis. Oral atorvastatin treatment prevented or reversed chronic and relapsing EAE paralysis. Atorvastatin promoted differentiation of Th0 cells to Th2 cells. Atorvastatin promoted differentiation of Th0 cells to Th2 cells. Atorvastatin treatment induced a population of regulatory T cells that suppressed EAE paralysis. Atorvastatin treatment induced a population of regulatory T cells that suppressed EAE paralysis. Statins may be a good treatment for Multiple Sclerosis and other Th1-mediated autoimmune diseases due to its pleiotropic immunomodulatory effects. Statins may be a good treatment for Multiple Sclerosis and other Th1-mediated autoimmune diseases due to its pleiotropic immunomodulatory effects.