A novel estrogen receptor 1: sphingomyelin phosphodiesterase acid-like 3B pathway mediates rituximab response in myositis patients.

OBJECTIVES: The B-cell depleting biologic, rituximab, is used to treat refractory autoimmune myositis. However, the beneficial effects of rituximab appear to outweigh the known contribution of B cells in myositis. We aimed to elucidate how myositis patients respond differently to rituximab and possible alternative mechanisms of action. METHODS: Here we have: (i) comprehensively investigated concurrent mRNA and microRNA expression in muscle biopsies taken at baseline and 16 weeks post treatment in 10 patients who were part of the rituximab in myositis (RIM) trial; and (ii) investigated the beneficial effect of rituximab on myositis muscle cells. RESULTS: Our analyses identified an increased number of changes in gene expression in biopsies from patients who had a clinical response to rituximab (n = 5) compared with non-responders (n = 5). The two groups had completely different changes in microRNA and mRNA expression following rituximab therapy, with the exception of one mRNA, BHMT2. Networks of mRNA and microRNA with opposite direction of expression changes highlighted ESR1 as upregulated in responders. We confirmed ESR1 upregulation upon rituximab treatment of immortalized myotubes and primary human dermatomyositis muscle cells in vitro, demonstrating a direct effect of rituximab on muscle cells. Notably, despite showing a response to rituximab, human dermatomyositis primary muscle cells did not express the rituximab target, CD20. However, these cells expressed a possible alternative target of rituximab, sphingomyelinase-like phosphodiesterase 3 b (SMPDL3B). CONCLUSION: In addition to B-cell depletion, rituximab may be beneficial in myositis due to increased ESR1 signalling mediated by rituximab binding to SMPDL3B on skeletal muscle cells.

The Effect of Immunomodulatory Treatments on Anti-Dystrophin Immune Response After AAV Gene Therapy in Dystrophin Deficient mdx Mice.

BACKGROUND: AAV-based gene therapy is an attractive approach to treat Duchenne muscular dystrophy (DMD) patients. Although the long-term consequences of a gene therapy approach for DMD are unknown, there is evidence in both DMD patients and animal models that dystrophin replacement by gene therapy leads to an anti-dystrophin immune response that is likely to limit the long-term use of these therapeutic strategies. OBJECTIVE: Our objective is to test whether the anti-dystrophin immune response is affected by immunomodulatory drugs in mdx mice after rAAV gene therapy. METHODS: mdx mice were treated with rAAV microdystrophin alone or in combination with immunomodulatory drugs. Dystrophin expression in skeletal muscle was assessed by mass spectrometry. Immune responses were assessed by immunophenotyping, western blot for anti-dystrophin antibodies and flow cytometry assays for antigen-specific T-cell cytokine expression. The impact on muscle was measured by grip strength assessment, in vivo torque, optical imaging for inflammation and H&E staining of sections to assess muscle damage. RESULTS: We found that AAV-9-microdystrophin gene therapy induced expression of microdystrophin, anti-dystrophin antibodies, and T-cell cytokine responses. Immunomodulatory treatments, rituximab and VBP6 completely abrogated the anti-dystrophin antibody response. Prednisolone, CTLA4-Ig, and eplerenone showed variable efficacy in blocking the anti-dystrophin immune response. In contrast, none of the drugs completely abrogated the antigen specific IFN-γ response. AAV-microdystrophin treatment significantly reduced inflammation in both forelimbs and hindlimbs, and the addition of prednisolone and VBP6 further reduced muscle inflammation. Treatment with immunomodulatory drugs, except eplerenone, enhanced the beneficial effects of AAV-microdystrophin therapy in terms of force generation. CONCLUSIONS: Our data suggest that AAV-microdystrophin treatment results in anti-dystrophin antibody and T-cell responses, and immunomodulatory treatments have variable efficacy on these responses.