Putative mechanisms of action of statins in multiple sclerosis – comparison to interferon-β and glatiramer acetate

Journal of the Neurological Sciences 233 (2005) 173 – 177 www.elsevier.com/locate/jns

Putative mechanisms of action of statins in multiple sclerosis – comparison to interferon-h and glatiramer acetate Oliver Neuhausa,*, Olaf Stu¨vea, Juan J. Archelosb, Hans-Peter Hartunga a

Department of Neurology, Heinrich Heine University, Moorenstraße 5, D-40225 Du¨sseldorf, Germany b Department of Neurology, Medical University, Graz, Austria Available online 20 April 2005

Abstract Statins are inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A reductase and are widely prescribed as cholesterol-lowering agents. They are promising candidates for future treatment in multiple sclerosis (MS) as they have been shown to exhibit immunomodulatory effects. Recent reports have demonstrated that statins are effective in preventing and reversing chronic and relapsing experimental autoimmune encephalomyelitis (EAE), an animal model of MS. Furthermore, in vitro experiments with human immune cells have documented an immunomodulatory mode of action of statins comparable to that of interferon (IFN)-h. An open label clinical trial assessing simvastatin in MS revealed a significant decrease in the number and volume of new MRI lesions and a favourable safety profile. This article reviews data thus far present on the putative mechanisms of action of statins in the immunopathogenesis of MS. Furthermore, the role of statins as potential pharmacotherapy for MS is discussed in the context of the mechanisms of approved immunotherapies in MS, namely IFN-h and glatiramer acetate (GA). D 2005 Elsevier B.V. All rights reserved. Keywords: Multiple sclerosis; Immunotherapy; Immunomodulation; Statins; Interferon-h; Glatiramer acetate

1. Multiple sclerosis Multiple sclerosis (MS) is the most common chronic central nervous system (CNS) disorder of younger adults and a major cause of lasting neurological disability [1]. Deviations of immune responses play a central role in its pathogenesis [2] by contributing to the formation and pertubation of MS lesion [3]. The initial inflammatory phase is characterized by selective demyelination, and eventually subsides to a neurodegenerative phase with axonal loss and gliosis [2,4]. With ever novel scientific information emerging, concepts regarding the pathogenesis of MS are in constant flux and shown in Fig. 1 [5]. In genetically susceptible individuals, an activation of antigenspecific, encephalitogenic T cells occurs in response to a yet unidentified trigger, probably in the periphery. Once activated, T lymphocytes are able to migrate through the blood – brain barrier and invade the CNS. Reactivated there, * Corresponding author. Tel.: +49 211 81 17880; fax: +49 211 81 18469. E-mail address: [email protected] (O. Neuhaus). 0022-510X/$ - see front matter D 2005 Elsevier B.V. All rights reserved. doi:10.1016/j.jns.2005.03.030

these T cells release pro-inflammatory cytokines and orchestrate destruction of the myelin sheath by various immune cell types following four different pathologic patterns [3]: (i) T cell and macrophage-mediated demyelination; (ii) antibody-mediated demyelination involving complement activation; (iii) distal oligodendrogliopathy and oligodendrocyte apoptosis; (iv) primary oligodendrocyte degeneration. Interestingly, the presence of pattern (iii), oligodendrocyte apoptosis, has recently been demonstrated in very early MS lesions [6]. In parallel to the autoaggressive inflammatory phase causing demyelination, recent evidence suggests that axonal loss responsible for irreversible disability occurs already early in the disease course and – as the disease evolves – predominates the underlying pathogenetic mechanisms [2,7,8].

2. Statins Statins, which are widely used as lipid-lowering agents, inhibit 3-hydroxy-3-methylglutaryl coenzyme A (HMG-

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O. Neuhaus et al. / Journal of the Neurological Sciences 233 (2005) 173 – 177

Antigen-presenting cell

Effector cell: macrophage

Periphery Activation Macrophage Antigenpresenting cell MHC-II Microbial Ag TCR

Central nervous system Migration Adhesion Penetration

Adhesion molecules, proteases

Reactivation

Inflammation

Microglia

Macrophage

MHC-II CNS Ag TCR

Neuron Oligodendrocyte TNF-α O2 NO

IFN-γ

TNF-α

Autoreactive T cell

Blood-brain barrier

Autoreactive T cell TH1

(iii)

Activated T cell (i) Complement

IFN-γ IL-2

Activated B cell Plasma cell

Demyelination patterns (i) - (iv)

(iv)

Axonal damage

(i) T-cell and Macrophage attack (ii) Antibody attack (iii) OG apoptosis (iv) Primary OG degeneration

Axonal swelling Lobulation Disconnection

(ii) Effector cells

Demyelinating Target cells antibodies

Effector cell: B cell

Fig. 1. Putative action sites of statins in the pathogenesis of MS. Via their T cell receptor (TCR), pro-inflammatory T cells are activated in the periphery by foreign or self-antigens (Ag) presented on major histocompatibility complex class II (MHC-II) by antigen-presenting cells (APC), including dendritic cells and B cells. Activated T cells migrate to, adhere at and penetrate through the blood – brain barrier, a step mediated by adhesion molecules, proteases and chemokines. Inside the central nervous system (CNS), T cells are reactivated in the context of MHC-II on resident APC, predominantly microglia cells. These reactivated T cells secrete pro-inflammatory cytokines, such as interferon (IFN)-g or interleukin (IL)-2 and induce CNS inflammation by subsequent activation of macrophages, other T cells and B cells as effector cells. Macrophages and T cells attack the oligodendrocytic myelin sheath by cytotoxic mediators, mainly tumor necrosis factor (TNF)-a, oxygen (O2) radicals and nitric oxide (NO). B cells differentiate to plasma cells that secrete demyelinating antibodies. They can guide and activate macrophages or ignite the complement cascade with assembly of the membrane attack complex which causes pore formation in myelin membranes. Statins have been shown to exert a variety of inhibitory effects on autoreactive T cells, B cells, macrophages and other antigen-presenting cells. Modified from [5], with permission from Elsevier.

CoA) reductase, the enzyme that catalyzes the rate-limiting step of the cholesterol biosynthesis pathway. Furthermore, statins decrease cardiovascular-related morbidity and mortality [9]. The safety profile of statins that has been in clinical use for almost two decades is favourable [10]. Infrequent side effects include hepatotoxicity and myopathy with a very low risk of rhabdomyolysis [11]. For many years, statins have been postulated to be potent immunomodulators [12 –16] but only recently, a series of publications brought statins into the focus of interest as a potential MS therapy [17 – 22]. Statins ameliorate or even prevent disease expression in several experimental autoimmune encephalomyelitis (EAE) animal models. Immunologically, a shift from pro-inflammatory towards antiinflammatory conditions was consistently observed [18,19]. Furthermore, in vitro studies with human peripheral blood lymphocytes revealed that simvastatin, lovastatin and mevastatin exerted strong anti-inflammatory properties comparable to those of IFN-h1b, although partial proinflammatory effects by heightened secretion of IFN-g and interleukin 12 were also observed [22]. More recently and in contrast to IFN-h-1b, simvastatin has been demonstrated to augment the proteolytic activity of matrix metalloproteinase 2, further suggesting partial pro-inflammatory effects of statins [23].

Sena et al. reported a small open-label study with lovastatin in MS [24]. Seven patients with relapsingremitting MS and at least 2 relapses during the previous 2 years received 20 mg lovastatin for 12 months. There were no serious adverse events. Although no clinical changes were observed, MRI revealed partial treatment effects in this small cohort. Vollmer et al. published the results of an open-label phase II study on simvastatin in MS [25]. Twenty-eight patients with relapsing-remitting MS who had at least one gadolinium enhancing lesion in one of three monthly MRI scans were treated for 6 months with 80 mg of simvastatin daily. Serial MRI scans were obtained at month 4, 5 and 6 during treatment. A 44% reduction in the mean number of new gadoliniumenhancing lesions ( p <0.0001) and a 41% reduction in the mean volume of new gadolinium-enhancing lesions ( p = 0.0018) were observed. No serious adverse events and no clinical changes were noted in this short study. Clearly, for further evaluation of statins in MS, prospective placebo-controlled studies are required as well as trials to test statins in combination with approved MS treatments. Those trials should answer the question if statins – either combined with an established disease modifying drug or alone – represent a viable therapeutic option in MS.

3. Statins—clinical data

4. Putative mechanisms of action of statins in MS

Two smaller studies examining treatment effects of statins on patients with MS have been presented recently.

The exact mechanisms of action of statins are still elusive. HMG-CoA reductase-dependent effects [26] and a

O. Neuhaus et al. / Journal of the Neurological Sciences 233 (2005) 173 – 177

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Nearly all immunomodulatory effects of statins, including those mediated by inhibition of isoprenylation of small GTP-binding proteins, can be reversed by treatment with L-mevalonate, the product of HMG-CoA reductase, indicating that statin-induced immunomodulation is mediated through inhibition of the mevanolate pathway. However, there is at least one exception. Statins have been observed to directly inhibit leukocyte function antigen (LFA)-1 [28], a cellular adhesion molecule that has key roles in both T cell activation and migration into the brain [29]. A number of immunological effects of statins have been described thus far. For example, statins block IFNg-inducible [30] and constitutive [18] transcription of the major histocompatibility complex (MHC) class II transactivator (CIITA), which regulates nearly all MHC class II gene expression and consequently inhibits activation of

direct impact on immune receptors [15] are conceivable. By inhibition of the product of HMG-CoA, mevalonate, the synthesis of other important intermediates of the cholesterol biosynthetic pathway is also impaired. Farnesylpyrophosphate and geranylgeranylpyrophosphate are important lipid attachments for the posttranslational modification of several proteins including the small GTPbinding proteins Ras, Rac and Rho [26]. Only this lipid attachment, termed isoprenylation, permits subsequent activation and membrane translocation of these proteins, which is crucial for a variety of cellular functions, such as cell shape, motility, secretion, differentiation and proliferation. Thus, prevention of isoprenylation of Rho by statins induces the accumulation of inactive Rho molecules in the cytosol and inhibits cellular functions vital for the activation of various cell types, including that of encephalitogenic leukocytes [27] (Fig. 2).

2 x Acetyl-CoA (C2) Cellular structure secretion migration differentiation proliferation

Statins

Acetoacetyl-CoA (C4) HMG-CoA (C6)

HMG-CoA reductase

Rho kinase

GTP

Mevalonate (C6) Mevalonate-P (C6) Mevalonate-PP (C6) Rho kinase

Mevalonate-PPP (C6) Dimethylallyl-PP (C5)

Isopentenyl-PP (C5) H2 N

Geranyl-PP (C10) Farnesylated proteins, e.g. Ras

GTP

GDP

COOCH 2 S

Active Rho with membrane anchor

Farnesyl-PP (C15)

Activation and membrane translocation

Statins Squalene (C30) Lanosterol (C30)

Geranylgeranyl-PP (C20) PP

H2 N

COOH Cys

Zymosterol (C27)

Inactive Rho

Accumulation of inactive Rho molecules, inhibition of cellular functions

Chlesterol (C27) Fig. 2. Cholesterol biosynthesis and isoprenylation of Rho. Farnesylpyrophosphate (-PP) and its derivative, geranylgeranyl-PP are lipid attachments for the posttranslational modification of immunologically important proteins (e.g. Ras, Rac, Rho). This ‘‘isoprenylation’’ permits subsequent activation and membrane translocation of this type of proteins that is crucial for a variety of cellular functions. Isoprenylated Rho binds to GDP and acts as a molecular switch between a dormant GDP-binding and an active GTP-binding state that exerts its biological function by binding to Rho kinase. Prevention of isoprenylation of Rho by statins induces the accumulation of inactive Rho molecules in the cytosol and inhibits cellular functions vital for leukocyte activation. All HMG-CoA reductase dependent effects of statins can be antagonized by L-mevalonate. The numbers in parentheses represent the numbers of carbon atoms. HMG-CoA, 3-hydroxy3-methylglutaryl coenzyme A; P, phosphate residue; GDP, guanosinediphosphate; GTP, guanosinetriphosphate. Modified from [41], with permission from Elsevier.

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CD4+ T lymphocytes. Possible sites of action of statins in the immunopathogenesis of MS are shown in Fig. 1.

5. Statins, interferon-B and glatiramer acetate Both IFN-h and glatiramer acetate (GA) are approved for treatment of MS and a variety of immunomodulatory effects have been described [31 –33]. IFN-h exerts its immunological effects antigen-independently through reducing the secretion of proteolytic matrix metalloproteinases that mediate the migration of T cells across biological barriers [34,35], as well as possibly downregulating MHC class II on various antigen presenting cells (APC) [36,37]. GA acts on ‘‘signal one’’ of T cell activation by binding to MHC class II molecules irrespective of haplotype [38], and possibly crossreacts with several CNS antigens. It also serves as an altered peptide ligand (APL) [39] and induces antiinflammatory regulatory GA-reactive T cells capable of a ‘‘bystander suppression’’ once reactivated inside the CNS [32]. An overview of immunological effects of statins, IFN-h and GA is given in Table 1. As both approved immunomo-

dulatory agents and statins share common mechanisms of action in the inflammatory cascade of MS, yet differ in others, combinations of these agents could be reasonable. Consistent with this notion, in vitro experiments exhibited additive effects of IFN-h and statins [22]. In EAE, the combination of suboptimal doses of GA and atorvastatin ameliorated the clinical course of the disease which was not observed with GA or atorvastatin alone [40]. Furthermore, combination trials in MS are planned.

6. Conclusions In recent years, there have been major advances in understanding how cellular and humoral immune responses contribute to the pathogenesis of MS. Targeting of key elements of the immunological cascade that culminates in neural and glial tissue damage through statins could potentially offer a number of advantages over currently available drugs. They are administered orally, may exert additive or synergistic effects with established disease modifying agents and have a well-established safety profile.

Table 1 Immunological effects of statins, IFN-h, and glatiramer acetate [32,41] Immune mechanism

Statins

IFN-h

Glatiramer acetate

Proliferation of T cells in vitro B cell activation Regulation of MHC expression MHC binding

Antigen-independent suppression of T cell proliferation , Activation of B lymphocytes , MHC class II expression

, Proliferation of MBP-specific T cells No known effect No known effect

No known effect

Antigen-independent suppression of T cell proliferation No known effect , IFN-g-induced upregulation of MHC class II expression No known effect

T cell migration

, T cell migration

, T cell migration

Adhesion molecule expression

Binding to LFA-1, , binding to ICAM-1

MMP expression

, Expression and secretion of MMP9, j proteolytic activity of MMP-2 , Secretion of chemokines, , expression of chemokine receptors Cytokine shift from TH1 to TH2, j secretion of IFN-g and IL-12

j Soluble adhesion molecules (sICAM-1, sVCAM-1), , surface-expressed adhesion molecules (VLA-4) , Expression of MMP-9 , Secretion of chemokines, , expression of chemokine receptors Induction of TH2 cytokines and reduction of TH1 cytokines in PBL

APL effect on CNS-specific T cells Effects on APC

No known effect

No known effect

, Proliferation of macrophages

, IFN-g induced FcgRI expression in monocytes

Neuroprotection

Mediation of neuroprotection in an ischemia – reperfusion model

Positive clinical and MRI effects on disease progression

Chemokine and chemokine receptor expression Cytokine shift in PBL

Direct and promiscuous binding to HLA-DR molecules , Migration of PBL from GA-treated patients (unknown mechanism) No known effect

No known effect No known effect j IL-10 in serum and of mRNA for TGF-h and IL-4; , mRNA for TNF-a in PBL; shift of GA-reactive T cells from TH1 towards TH2 during GA treatment Induction of anergy in MBP-specific T cell clones , TNF-a and cathepsin-B production in a monocytic cell line, , monocyte and dendritic cell functions Secretion of BDNF by GA-reactive T cells; neuroprotection in EAE

j, increase; ,, inhibitory effect; APC, antigen presenting cells; APL, altered peptide ligand; BDNF, brain-derived neurotrophic factor; CNS, central nervous system; EAE, experimental autoimmune encephalomyelitis; GA, glatiramer acetate; HLA, human leukocyte antigen; ICAM, intercellular adhesion molecule; IFN, interferon; LFA, lymphocyte function-associated antigen; IL, interleukin; MBP, myelin-basic protein; MHC, major histocompatibility complex; MRI, magnetic resonance imaging; MMP, matrix metalloproteinase; PBL, peripheral blood lymphocytes; sICAM, soluble intercellular cell adhesion molecule; sVCAM, soluble vascular cell adhesion molecule; TGF, transforming growth factor; TH, T helper function; TNF, tumor necrosis factor; VLA, very late antigen.

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