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

Issue title: Special Issue Dedicated to Professor Terence Partridge

Guest editors: Jyoti Jaiswal and Kanneboyina Nagaraju

Article type: Research Article

Authors: Li, Ning^{a; 1} | Parkes, Joanna E.^{a; 1} | Spathis, Rita^a | Morales, Melissa^a | Mcdonald, John^a | Kendra, Ryan M.^a | Ott, Emily M.^b | Brown, Kristy J.^c | Lawlor, Michael W.^b | Nagaraju, Kanneboyina^{a; *}

Affiliations: [a] School of Pharmacy and Pharmaceutical Sciences, Binghamton University, Binghamton, NY, USA | [b] Department of Pathology and Neuroscience Research Center, Medical College of Wisconsin, Milwaukee, WI, USA | [c] Solid Biosciences Inc, Cambridge, MA, USA

Correspondence: [*] Correspondence to: Kanneboyina Nagaraju, DVM, PhD, Professor and Founding Chair, Department of Pharmaceutical sciences, School of Pharmacy and Pharmaceutical Sciences, Binghamton University, PO Box 6000, Binghamton, NY 13902-6000, USA. E-mail: Nagaraju@binghamton.edu.

Note: [1] Equal contribution.

Abstract: 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.

Keywords: Gene therapy, microdystrophin, immune response, adeno associated virus, anti-dystrophin antibodies, immunomodulatory treatments

DOI: 10.3233/JND-210706

Journal: Journal of Neuromuscular Diseases, vol. 8, no. s2, pp. S325-S340, 2021