Cell-Mediated Immunity to NAGLU Transgene Following Intracerebral Gene Therapy in Children With Mucopolysaccharidosis Type IIIB Syndrome

Mucopolysaccharidosis type IIIB syndrome (Sanfilippo disease) is a rare autosomic recessif disorder caused by mutations in the α-N-acetylglucosaminidase (NAGLU) gene coding for a lysosomal enzyme, leading to neurodegeneration and progressive deterioration of cognitive abilities in affected children. To supply the missing enzyme, several recent human gene therapy trials relied on the deposit of adeno-associated virus (AAV) vectors directly into the brain. We reported safety and efficacy of an intracerebral therapy in a phase 1/2 clinical trial (https://clinicaltrials.gov/ct2/show/NCT03300453), with a recombinant AAV serotype 2/5 (rAAV2/5) coding human NAGLU in four children with MPS IIIB syndrome receiving immunosuppression. It was reported that AAV-mediated gene therapies might elicit a strong host immune response resulting in decreased transgene expression. To address this issue, we performed a comprehensive analysis of cellular immunity and cytokine patterns generated against the therapeutic enzyme in the four treated children over 5.5 years of follow-up. We report the emergence of memory and polyfunctional CD4+ and CD8+ T lymphocytes sensitized to the transgene soon after the start of therapy, and appearing in peripheral blood in waves throughout the follow-up. However, this response had no apparent impact on CNS transgene expression, which remained stable 66 months after surgery, possibly a consequence of the long-term immunosuppressive treatment. We also report that gene therapy did not trigger neuroinflammation, evaluated through the expression of cytokines and chemokines in patients’ CSF. Milder disease progression in the youngest patient was found associated with low level and less differentiated circulating NAGLU-specific T cells, together with the lack of proinflammatory cytokines in the CSF. Findings in this study support a systematic and comprehensive immunomonitoring approach for understanding the impact immune reactions might have on treatment safety and efficacy of gene therapies.

Genotype to Phenotype: Identification of Mucopolysaccharidosis Type IIIB (Sanfilippo's B) Case Using Whole Exome Sequencing

Mucopolysaccharidosis type IIIB (Sanfilippo's B; OMIM no.: 252920) is a lysosomal storage disorder caused by defective degradation of heparan sulfate. The enzyme that has decreased function in this disease is α-N acetylglucosaminidase. This enzyme is encoded by the NAGLU gene. A 9-year-old male patient was referred to us with speech disability, developmental delay, hepatomegaly, mild learning disability, and otitis media with effusion complaints. Whole exome sequencing (WES) was performed because of consanguinity between the parents of the patient and the lack of specific prediagnosis. As a result of the patient's WES analysis, a homozygous mutation was detected in the NAGLU gene. The leukocyte enzyme activity was then evaluated to confirm the diagnosis. Alpha-N acetylglucosaminidase deficiency was found. Alpha-N acetylglucosaminidase activity was 0.2 nmol/mLh. WES is a successful diagnostic method in the diagnosis of the mild clinical diseases with recessive inheritance. In addition, our case is a good example of genotype to phenotype diagnosis. Because in storage diseases, the diagnosis is made by leukocyte enzyme analysis first, and then the result is confirmed by gene analysis. The opposite situation occurred in our case.