Haematopoietic stem cell mobilization and re-arrangement of immune cell subsets following therapeutic alpha 4-integrin blockade in multiple sclerosis: functional relevance, clinical correlations and applications

Abstract

The anti α-4 integrin antibody natalizumab is broadly used in clinical practice for treatment of active multiple sclerosis (MS). Previous studies reported an increase of circulating haematopoietic stem cells in natalizumab-treated MS patients yet the biological significance remained unclear. This research wanted to ascertain the mobilization from the bone marrow (BM) and the functional relevance of the increased number of circulating hematopoietic stem and progenitor cells (HSPC) induced by natalizumab in patients with MS. CD45lowCD34+ HSPC frequency was evaluated by flow-cytometry in the peripheral blood of 45 natalizumab-treated patients (12 of which were prospectively followed during the first year of treatment as part of a pilot cohort and 16 were prospectively followed for validation), 10 untreated MS patients and 24 healthy donors. In the natalizumab-treated group the cell cycle status of sorted HSPC was also evaluated, together with T- and B lymphocyte (n=29) and myeloid-derived suppressor cells subpopulation frequencies and HSPC differentiation potential (n=10). Moreover, an in vitro model was designed for studying the immune-modulatory effect of increasing concentrations of HSCP on the distribution of lymphocytes subsets in HSPC/mononuclear cells (MNC) co-cultures started from Cord Blood samples (n=5). Natalizumab-induced circulating HSPC were predominantly quiescent, suggesting recent mobilization from the BM, and were capable of differentiation ex vivo. Circulating HSPC numbers were significantly increased during natalizumab although heterogeneously, allowing the stratification of “Mobilizer” and “Non-Mobilizer” sub-groups. "Non-Mobilizer" status was associated with persistence of disease activity during treatment. The frequency of B cells, Transitional B regulatory cells and CD103+ CD8+ regulatory T cells persistently increased, more significantly in “Mobilizer” patients who also showed a specific naïve/memory B cell profile. Also the proportion of myeloid-derived suppressor cells (MDSC) was found to increase during natalizumab treatment, correlating with HSPC counts. In vitro, B regulatory cells proportions increased significantly upon addition of increasing concentrations of HSPC in co-cultures, while IL-6 concentration consistently decreased in co-cultures supernatant and level of IL-10 relatively increase. The data suggest that the natalizumab-induced circulating HSPC increase is the result of true mobilization from the BM and has clinical and immunological relevance. HSPC mobilization, associated with clinical remission and with increased proportion of circulating B and regulatory T cells and MDSC, may contribute to the treatment’s mode of action. Thus, HSPC blood counts could represent an early biomarker of responsiveness to natalizumab.