Novel therapeutics for multiple sclerosis: altered peptides and stem cell-based approaches
thesisposted on 28.02.2017, 23:57 by McDonald, Courtney Anne
Multiple Sclerosis (MS) is an autoimmune disease of the central nervous system (CNS). The aetiology of the disease remains unclear, however, the fundamental pathological features of the disease include the breakdown of the blood brain barrier (BBB) and the formation of demarcated plaques within the white matter of the CNS. These plaques are characterised by infiltrating immune cells, localised myelin destruction, loss of oligodendrocytes and axonal degeneration. Despite these common features, MS is well recognised as a heterogeneous disease, both clinically and pathologically. Current immunosuppressive and immunomodulatory treatments have only been effective in a relatively small proportion of patients. Furthermore, patients with progressive neurological deterioration, benefit little from current MS therapies. Thus, there is an urgent need for the development of novel and more specific treatments for this severe neurological disorder. The aim of this thesis was to investigate the effect of two different treatment strategies on the development of experimental automimmune encephalomyelitis (EAE), an animal model of MS. The two strategies utilised novel altered peptides and stem cell transplantation. Both were aimed at modulating the immune system: the altered peptides specifically targeting the autoreactive T-cells, while stem cells having the ability to home to sites of inflammation, can modulate the immune response in situ, as well as exerting possible neuroprotective effects. The first part of this thesis describes a series of experiments performed with novel altered peptides. These peptide analogues were based on the peptide sequence of myelin oligodendrocyte glycoprotein (MOG), one of the auto-antigens implicated in the development of MS. These altered MOG peptides were modified by substituting a β-amino acid into a putative TCR contact residue. Such modifications not only lead to increased peptide stability but also rendered such modified peptides unable to stimulate auto-reactive T-cells. These altered MOG peptides were unable to induce disease themselves, making them safe potential therapeutics. Vaccination with one such peptide, MOG44βF, significantly ameliorated EAE. The suppression of disease involved a significant reduction in peripheral immune responses, coupled with an increase in regulatory mechanisms in both the periphery and CNS of these animals. Additional studies using MOG44βF in an oral treatment regime, administered early after EAE induction, were also shown to attenuate EAE severity. The therapeutic efficacy of stem cells, specifically amnion epithelial cells (AEC), is described in the second part of this thesis. AECs offer a further potential immunosuppressive therapy that can modulate the autoimmune response. Moreover, recent studies from our and other laboratories have demonstrated the efficacy of transplanted mesenchymal stem cells (MSCs), cells with similar immunosuppressive properties to AECs, to ameliorate EAE. Thus, we hypothesised that AECs may also have the potential to act specifically at the site of inflammation and aid in neuropreotection and regeneration of damaged nerve tissue. This is currently not achieved by any treatments in the clinic. Extensive immunological and phenotypic characterisation was performed on AECs from multiple donors; specifically assessing the expression of cell surface immune markers and chemokine receptors, as well as their ability to suppress T-cell responses. Transplantation of AECs ameliorated rMOG-induced EAE, following systemic delivery in both chronic and relapsing-remitting models. This suppressive effect correlated with peripheral immune suppression, with reduced MOG-specific T-cell responses, reduced pro-inflammatory cytokine secretion and an increased proportion of Th2 anti-inflammatory T-cells in the peripheral lymphoid organs. Most importantly, administration of AECs also modulated the immune response within the CNS, with an increase in the proportion of Th2 cells, resulting in a significant shift in the immunological environment towards a protective anti-inflammatory setting. The third part of this thesis reports on the use of gene modified AECs as vehicles to deliver anti-inflammatory cytokines, as a means to possibly enhance their therapeutic efficacy in vivo. More specifically, we used lentiviral transduced AECs engineered to secrete IL-4, an anti-inflammatory cytokine known to promote Th2 differentiation and reduce EAE severity. Transplantation of AEC-IL4 just prior to disease onset was less effective at reducing the severity of EAE compared to AECs alone. Yet when AEC-IL4 were injected during the induction phase of disease, the onset and severity was significantly ameliorated. These studies suggest that the window of opportunity for treatment with stem cells over-expressing IL4 is likely to be during the initial inflammatory phase and is less effective once inflammation is established. Taken together, the data presented in this thesis demonstrates that the two therapeutic strategies investigated are capable of dampening inflammatory immune responses, resulting in amelioration of EAE. Altered MOG peptides have the ability to specifically target the autoimmune response that is important for disease propagation, without suppressing and compromising the wider immune system. Conversely, the ability of stem cells to traffic to inflamed sites within the CNS, allows them to act specifically at the site of inflammation and dampen the in situ immune response and promotes a Th2 protective environment. Thus, further understanding of the suppressive mechanisms afforded by these different strategies in vivo, will be critical for the development of such novel treatments for diseases like MS.