Laquinimod interferes with migratory capacity of T cells and reduces IL-17 levels, inflammatory demyelination and acute axonal damage in mice with experimental autoimmune encephalomyelitis
Introduction
Multiple sclerosis (MS) is the most common chronic neurological disease leading to disability in early to middle adulthood. The majority of patients experience a relapsing–remitting course in the early phase of the disease. Over time, most patients develop secondary progression of disability, but the course of MS can vary considerably between patients. Pathologically, MS is characterized by inflammatory demyelinating lesions in the central nervous system (CNS). However, the progressive phase of the disease is characterized by chronic demyelinated lesions as well as axonal loss which is the major correlate of chronic disability in MS (Bjartmar et al., 2000, Ganter et al., 1999). Recent pathological studies indicate that axonal injury begins early at the disease onset, occurring within each focal inflammatory lesion (Trapp et al., 1998, Ferguson et al., 1997, Kuhlmann et al., 2002). Drugs with myelin and axon preservation effects might limit the tissue damage accumulating in the acute inflammatory process. Current approved treatments are all injectable drugs mainly targeted at the inflammatory aspects of the disease, which predominate in the relapsing–remitting phase.
Experimental autoimmune encephalomyelitis (EAE) represents the principle animal model of MS which has proven useful in the development of treatments for MS (Steinman and Zamvil, 2005). This inflammatory autoimmune disease can be induced by active immunization with myelin antigens or the adoptive transfer of encephalitogenic T cells (Gold et al., 2006). EAE shares clinical and histopathological characteristics with MS. Acute and chronic EAE models can imitate different phases of the human disease. These models are widely used as initial steps to study potential new treatments for MS. Interferon-β (IFN-β) (Yasuda et al., 1999) and copolymer-1 (i.e. glatiramer acetate) (Teitelbaum et al., 1996) have been shown to inhibit disease in EAE. Inhibitory activity in EAE has also been demonstrated for the oral immunmodulator roquinimex (Karussis et al., 1993a, Karussis et al., 1993b). In MS, roquinimex significantly reduced MRI and clinical activity in phase II and III studies (Wolinsky et al., 2000), but its application in MS was stopped due to undesirable side effects (Noseworthy et al., 2000).
Laquinimod is a new immunomodulatory substance that is structurally related to roquinimex, but pharmacologically and chemically distinct, which results in increased potency and improved safety profile. Phase II studies indicate that laquinimod is safe and well tolerated, reduces the number of relapses and significantly reduces the formation of MRI-active lesions in relapsing–remitting MS (Comi et al., 2008, Polman et al., 2005). Two phase III studies are underway to determine the effects on clinical parameters and MRI measures related to permanent axonal damage.
Laquinimod has been shown to inhibit EAE in mice (Brunmark et al., 2002) and Lewis rats (Yang et al., 2004). In addition, laquinimod also suppressed the development of various experimental autoimmune inflammatory-mediated animal models, including experimental autoimmune neuritis in Lewis rats (Zou et al., 2002). The analysis of cytokine profiles demonstrated that laquinimod redirected the cytokine production in favour of the TH2/TH3 cytokines IL-4, IL-10 and TGF-β (Yang et al., 2004). The effect of laquinimod has been reported to be independent of endogenous IFN-β (Runstrom et al., 2006), which indicates that laquinimod might also be a potential future treatment in MS patients who do not respond to INF-β. Previous experimental studies have mainly demonstrated that laquinimod inhibits relapses in rodent EAE models and have shown histological data after preventive laquinimod treatment (Brunmark et al., 2002, Yang et al., 2004, Runstrom et al., 2006).
In this study we assessed the effects of laquinimod on inflammation, cytokine production, migratory capacities of lymphocytes as well as demyelination and axonal damage in C57BL/6 mice with MOG35–55-induced EAE. Preventive and therapeutic laquinimod treatment reduced clinical symptoms, macrophage as well as T cell infiltration, and demyelination. In addition, we found 22% less acute axonal damage within spinal cord lesions in mice with therapeutic treatment compared to control animals. Marked decreases of IL-17 levels and down-regulation of the pro-inflammatory cytokines IL-13, IFN-γ and TNF-α were found in splenocytes from treated mice. Furthermore, lymphocytes from treated animals displayed reduced VLA-4-mediated adhesiveness. Our findings thus indicate that laquinimod might protect myelin and axons by decreasing pro-inflammatory cytokines and impairing the migratory capacity of lymphocytes in a model of inflammatory CNS demyelination.
Section snippets
Test compounds and formulations
Laquinimod (originally ABR-215062) (RLB#054 M0004) was synthesized at TEVA Pharmaceutical Industries, Ltd. The compound was dissolved in purified water and administered orally by gavage in a volume of 0.2 ml, beginning on the day of immunization for preventive treatment (days 0–28 post immunization (p.i.) for treatment with 5 mg/kg or 25 mg/kg laquinimod) or after disease onset (days 13–28 p.i. for treatment with 25 mg/kg laquinimod) for therapeutic treatment until the end of the experiment. The
Suppression of clinical disease, inflammation and demyelination after preventive treatment with laquinimod
All vehicle-treated mice immunized with MOG35–55 developed clinical signs of acute EAE. In both preventive treatment groups laquinimod was given from the first day after immunization until the last day (days 0–28). After preventive treatment with 5 mg/kg most animals (13/15) developed clinical signs of disease, but the cumulative disease severity score (1.5 ± 1.2) was reduced in comparison to control mice (2.7 ± 0.6) (Fig. 1). Preventive treatment with 25 mg/kg laquinimod led to a significantly
Discussion
The orally active immunomodulator laquinimod has been demonstrated to inhibit relapses in rodent EAE models when given before (Brunmark et al., 2002, Yang et al., 2004) and after disease onset (Runstrom et al., 2006). Here we extend these studies by providing evidence for reduced acute axonal damage, down-regulation of pro-inflammatory cytokines and impaired migratory capacity of lymphocytes following laquinimod treatment in an EAE model. We demonstrated less acute axonal damage within lesions
Acknowledgement
Christiane Wegner was supported by research funding of the University Medical Center Göttingen (Heidenreich von Siebold research grant), Germany. We would like to acknowledge Jasmin Held for excellent technical assistance. We also thank Uwe Hanisch for valuable advice on the manuscript.
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2017, Journal of NeuroimmunologyCitation Excerpt :Failure of laquinimod to reach secondary study outcomes may be the result of treatment dosing, frequency, and duration, as well as relatively small treatment groups. However, much lower cumulative doses of laquinimod showed detectable biological effects in the EAE model of MS (Wegner et al., 2010; Thone et al., 2012; Brunmark et al., 2002; Yang et al., 2004; Moore et al., 2013; Berg et al., 2016). To study the interaction between Aβ and tau and their effect on synaptic function, Oddo et al. generated a triple-transgenic (Tg) animal model (3xTg-AD mouse) (Oddo et al., 2003).