Abstract

Enzyme replacement therapy (ERT) is indispensable for patients of hemophilia or lysosome storage disease (LSD) lifetime. However, the efficacy of ERT is diminished or lost in some patients who acquired the immune responses by the repeated enzyme infusions. If reduced the immune responses such as neutralizing IgG antibody formation and/or anaphylaxis, one enzyme for ERT would be continuously administered without loss of efficacy and/or anaphylaxis risk. We here show that the pre-treatment with α-galactosylceramide (αGC), a representative ligand for invariant natural killer T (iNKT) cells prior to the ERT could remarkably suppress not only the enzyme-specific IgG antibody response but also total IgE formation in a mice model.

In our previous studies, liposomal, but not aqueous, αGC formulations were delivered to the splenic marginal zone (MZ) B cells which could enhance IL-10 production after the interaction with iNKT cells, and then the expansion of tolerogenic dendritic cells (DCs) and Foxp3-positive regulatory T cells (Tregs) (Ishii et al. Front Biosci 2008). The pretreatment with a liposomal αGC (lipo-αGC) formulation resulted in the remarkable suppression of both IgG and IgE antibody responses in mice immunized with alum-absorbed ovalbumin. In addition, administration of RGI-2001, a clinical study grade of lipo-αGC formulation, at the timing of bone marrow transplantation could induce the expansion of tolerogenic DCs and/or allo-specific Tregs in mouse models of either acute graft versus host disease (GvHD) or the mixed chimera formation (Duramad et al. Biol Blood Marrow Transplant 2011, Hirai et al. Am J Transplant 2014 and Hirai et al. Am J Transplant2016). Taken together, it was suggested that administration of lipo-αGC formulations prior to the ERT could work for the protection of the enzyme-specific antibody responses.

To test the suppressive capability of a lipo-αGC for ERT application, a mouse system to monitor a factor VIII-specific IgG antibody titer and IgE level in bloods was established. In BALB/c mice given the repeated intravenous injections of recombinant factor VIII, the factor VIII-specific IgG titer and total IgE concentration were substantially elevated (ERT-mice). The mice pretreated with the intravenous single injection of the lipo-αGC showed that not only the factor VIII-specific IgG but also total IgE was significantly suppressed compared to the untreated control mice. The antibody-suppressive efficacy by multiple injection of the lipo-αGC was comparable with that by single injection in the ERT-mice. However, cytokine profile of splenic iNKT cells was dramatically changed in BALB/c mice pretreated with either single or multiple lipo-αGC injections. The iNKT cells derived from mice pretreated with the single the lipo-αGC injection expressed high level of IFN-γ, IL-4 and IL-13 mRNAs but not IL-10 mRNA. Conversely, the iNKT cells pretreated with multiple the lipo-αGC injections expressed higher level of IL-10 mRNA more than IFN-γ mRNA, but remarkably diminished both IL-4 and IL-13 mRNAs. As known that αGC-activated iNKT cells could rapidly produce cytokines within a few hours, we then examined the last injection timing of multiple injections into the ERT-mice. The last injection at the timing of the ERT start did not show stronger efficacy than that at three day before the ERT start. Results collectively suggested that the suppression of IgG/IgE formations in the ERT-mice might not be caused by high IL-10 expression of iNKT cells by pretreatment with multiple the lipo-αGC injections, but other phenomena generated in three days after the interaction of activated iNKT cells with αGC-presenting MZ B cells.

We conclude that the lipo-αGC pretreatment is capable of protecting harmful formation of both the enzyme-specific neutralizing IgG and anaphylaxis-inducing IgE antibodies during ERT.

Disclosures

No relevant conflicts of interest to declare.

Author notes

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Asterisk with author names denotes non-ASH members.

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