Immunological effects of interferon-beta and glatiramer acetate in multiple sclerosis

Abstract: Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system, which results in demyelination, axonal loss and neurological deficits. The aetiology is unknown, however, immunopathological events, which are believed to be of an autoimmune nature, are considered to be responsible for the development of the disease. Interferonbeta (IFN-beta) and glatiramer acetate (GA; Copaxone) are disease-modifying therapeutic drugs for MS, and their efficacy is attributed to their immune regulatory properties. Studying their immunological effects in NIS may lead to a better understanding of the pathogenesis of MS, and more importantly, may result in more efficient therapies. The aims of this thesis were to examine the possible involvement of blood dendritic cells (DCs) in MS, and to investigate in vitro responses of DCs to IFN-beta and GA separately, as well as in combination, and the possible synergistic effects between IFN-beta and GA (papers I-II). Another aim was to examine responses of MS patients' blood mononuclear cells (MNCs) during combined treatment with IFN-beta and GA, and their effects on the immunological markers under investigation in NIS (papers III-IV). Using flow cytometry and enzyme-linked immunosorbent assay (ELISA), we found that DCs from MS patients expressed higher levels of CD 1a and produced lower amounts of interleukin-10 (IL-10) than DCs from healthy controls (HCs). IFN-beta or GA upregulated HLA-DR and CD86, and reduced CD 1a expression on DCs from both MS patients and HCs. IFN-beta induced Th2-driving DCs, which produced increased amounts of IL-10 and reduced amounts of IL-12 as compared to control DCs. Accordingly, IFN-beta-treated DCs, when cocultured with MNCs from HCs, reduced the production of IFN-gamma, but enhanced the production of IL-10 by MNCs. GA reduced IL-12 production by DCs in both NIS patients and HCs. GA and IFN-beta synergistically suppressed CD 1a and enhanced CD86 expression on DCs. IFN-beta induced Th2 immune responses via the DCs pathway. This may represent one mechanism for the action of IFN-beta in MS. We assume that increased CD1a expression on DCs is relevant to autoaggressive immunity against myelin autoantigens in NIS and that CD86 promotes Th2 responses, and that suppression of CD1a and up-regulation of CD86 by DCs could be beneficial in MS. Cross-sectional comparisons revealed that the proportion of CD1a+ HLA-DR+ MNCs was higher and the production of IL-10 was lower in untreated MS patients as compared to patients treated with IFN-beta or IFN-beta and GA, or HCs. MNCs from untreated MS patients also had higher IFN-gamma and IL-12 p70 production as compared to HCs or patients treated with IFN-beta and GA, but not when compared to patients on monotherapy with IFN-beta. The proportion of CD 1a+ HLA-DR+MNCs and the production of IL-12 p70 and IFN-gamma were even lower in patients treated with IFN-gamma and GA in combinaton. Follow-up studies revealed that treatment with IFN-beta and GA in combination, but not with IFN-beta alone, resulted in normalization of CD 1a+ HLA-DR+ MNCs levels, and of IL, 12p70 and IFN-gamma production by MNCs. Using an automated computer-assisted enzyme-linked immunoadsorbent spot (ELISPOT) assay, we demonstrated that IL-12 and IFN-gamma secreting MNCs were higher, and IL-10 were fewer in untreated NIS patients than in patients treated with IFN-beta or GA or IFN-beta and GA, or in HCs. By contrast, MS patients treated with IFN-beta or GA or IFN-beta and GA in combination had similar numbers of IL-12, IFN-gamma and IL-10 secreting MNCs compared to HCs. NIS patients treated with IFN-beta and GA in combination had similar numbers of IL-12, IFN-gamma and IL-10 secreting MNCs compared to patients treated with IFN-beta or GA. No differences were observed between NIS patients treated with IFN-beta, or GA and MS patients treated with IFNbeta and GA, nor between any of these groups and HCs, for numbers of IL-4 secreting MNCs. The same trends for the IFN-gamma, IL-10 and IL-4 responses to in vitro exposure to GA were found in all groups of MS. Suppression of IL-12 and IFN-gamma, and promotion of IL-10 secretion by MNCs by combination therapy with IFN-beta and GA was similar as monotherapy with IFN-beta or GA. Treatment with IFNbeta and GA in combination may not alter cytokine profiles by MNCs in response to GA. In conclusion, both IFN-beta and GA act in MS through distinct immunomodulatory effects and the immunological effects of IFN-beta and GA in combination are complex. However, whether or not these immunological changes imply clinical benefits in MS, remains to be investigated in clinical trials.