Metachromatic leukodystrophy (MLD) is an inherited lysosomal storage disease caused by arylsulfatase A (ARSA) deficiency. Patients with MLD exhibit progressive motor and cognitive impairment and die within a few years of symptom onset. We used a lentiviral vector to transfer a functional ARSA gene into hematopoietic stem cells (HSCs) from three presymptomatic patients who showed genetic, biochemical, and neurophysiological evidence of late infantile MLD. After reinfusion of the gene-corrected HSCs, the patients showed extensive and stable ARSA gene replacement, which led to high enzyme expression throughout hematopoietic lineages and in cerebrospinal fluid. Analyses of vector integrations revealed no evidence of aberrant clonal behavior. The disease did not manifest or progress in the three patients 7 to 21 months beyond the predicted age of symptom onset. These findings indicate that extensive genetic engineering of human hematopoiesis can be achieved with lentiviral vectors and that this approach may offer therapeutic benefit for MLD patients.

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Metachromatic leukodystrophy is a neurodegenerative disorder characterized by progressive demyelination. The disease is caused by variants in the ARSA gene, which codes for the lysosomal enzyme arylsulfatase A, or, more rarely, in the PSAP gene, which codes for the activator protein saposin B. In this Mutation Update, an extensive review of all the ARSA- and PSAP-causative variants published in the literature to date, accounting for a total of 200 ARSA and 10 PSAP allele types, is presented. The detailed ARSA and PSAP variant lists are freely available on the Leiden Online Variation Database (LOVD) platform at http://www.LOVD.nl/ARSA and http://www.LOVD.nl/PSAP, respectively.

Metachromatic leukodystrophy (MLD) is a rare lysosomal storage disorder resulting from the inherited deficiency of the arylsulfatase A (ARSA) enzyme. Currently, no valid therapeutic options are available for affected patients. A thorough knowledge of disease progression in its diverse clinical variants, together with the identification of reliable prognostic factors, could be instrumental in accurate patient selection for new upcoming therapeutic opportunities, such as enzyme replacement and gene therapy. The described correlation between genotype and clinical presentation proved helpful in predicting patient's prognosis, only in the minority of MLD patients harboring common mutations. Molecular characterization of a cohort of 26 MLD patients allowed us to identify 18 mutations, excluding the common 0 and R alleles, 10 of which are rare and 8 are novel. By categorizing the rare mutations, we were able to confirm a correlation between ARSA gene mutations, age at onset and patterns of disease progression, not only in those patients bearing common mutations, but also in those carrying rare mutant alleles. Moreover, in the case of absent or delayed molecular diagnosis, or of newly identified mutations, the involvement of peripheral nervous system from disease onset proved to be a sensitive prognostic marker predicting a severe progression.

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Summary Background Effective treatment for metachromatic leukodystrophy (MLD) remains a substantial unmet medical need. In this study we investigated the safety and efficacy of atidarsagene autotemcel (arsa-cel) in patients with MLD. Methods This study is an integrated analysis of results from a prospective, non-randomised, phase 1/2 clinical study and expanded-access frameworks. 29 paediatric patients with pre-symptomatic or early-symptomatic early-onset MLD with biochemical and molecular confirmation of diagnosis were treated with arsa-cel, a gene therapy containing an autologous haematopoietic stem and progenitor cell (HSPC) population transduced ex vivo with a lentiviral vector encoding human arylsulfatase A (ARSA) cDNA, and compared with an untreated natural history (NHx) cohort of 31 patients with early-onset MLD, matched by age and disease subtype. Patients were treated and followed up at Ospedale San Raffaele, Milan, Italy. The coprimary efficacy endpoints were an improvement of more than 10% in total gross motor function measure score at 2 years after treatment in treated patients compared with controls, and change from baseline of total peripheral blood mononuclear cell (PBMC) ARSA activity at 2 years after treatment compared with values before treatment. This phase 1/2 study is registered with ClinicalTrials.gov , NCT01560182 . Findings At the time of analyses, 26 patients treated with arsa-cel were alive with median follow-up of 3·16 years (range 0·64–7·51). Two patients died due to disease progression and one due to a sudden event deemed unlikely to be related to treatment. After busulfan conditioning, all arsa-cel treated patients showed sustained multilineage engraftment of genetically modified HSPCs. ARSA activity in PBMCs was significantly increased above baseline 2 years after treatment by a mean 18·7-fold (95% CI 8·3–42·2; p<0·0001) in patients with the late-infantile variant and 5·7-fold (2·6–12·4; p<0·0001) in patients with the early-juvenile variant. Mean differences in total scores for gross motor function measure between treated patients and age-matched and disease subtype-matched NHx patients 2 years after treatment were significant for both patients with late-infantile MLD (66% [95% CI 48·9–82·3]) and early-juvenile MLD (42% [12·3–71·8]). Most treated patients progressively acquired motor skills within the predicted range of healthy children or had stabilised motor performance (maintaining the ability to walk). Further, most displayed normal cognitive development and prevention or delay of central and peripheral demyelination and brain atrophy throughout follow-up; treatment benefits were particularly apparent in patients treated before symptom onset. The infusion was well tolerated and there was no evidence of abnormal clonal proliferation or replication-competent lentivirus. All patients had at least one grade 3 or higher adverse event; most were...

Lysosomal storage diseases (LSDs) are rare inherited metabolic disorders characterized by a dysfunction in lysosomes, leading to waste material accumulation and severe organ damage. Enzyme replacement therapy (ERT) and haematopoietic stem cell transplant (HSCT) have been exploited as potential treatments for LSDs but pre-clinical and clinical studies have shown in some cases limited efficacy. Intravenous ERT is able to control the damage of visceral organs but cannot prevent nervous impairment. Depending on the disease type, HSCT has important limitations when performed for early variants, unless treatment occurs before disease onset. In the attempt to overcome these issues, gene therapy has been proposed as a valuable therapeutic option, either ex vivo, with target cells genetically modified in vitro, or in vivo, by inserting the genetic material with systemic or intra-parenchymal, in situ administration. In particular, the use of autologous haematopoietic stem cells (HSC) transduced with a viral vector containing a healthy copy of the mutated gene would allow supra-normal production of the defective enzyme and cross correction of target cells in multiple tissues, including the central nervous system. This review will provide an overview of the most recent scientific advances in HSC-based gene therapy approaches for the treatment of LSDs with particular focus on metachromatic leukodystrophy (MLD) and mucopolysaccharidosis type I (MPS-I).

Adenosine Deaminase (ADA) deficiency is an autosomal recessive variant of severe combined immunodeficiency (SCID) caused by systemic accumulation of ADA substrates. Neurological and behavioral abnormalities observed in ADA-SCID patients surviving after stem cell transplantation or gene therapy represent an unresolved enigma in the field. We found significant neurological and cognitive alterations in untreated ADA-SCID patients as well as in two groups of patients after short- and long-term enzyme replacement therapy with PEG-ADA. These included motor dysfunction, EEG alterations, sensorineural hypoacusia, white matter and ventricular alterations in MRI as well as a low mental development index or IQ. Ada-deficient mice were significantly less active and showed anxiety-like behavior. Molecular and metabolic analyses showed that this phenotype coincides with metabolic alterations and aberrant adenosine receptor signaling. PEG-ADA treatment corrected metabolic adenosine-based alterations, but not cellular and signaling defects, indicating an intrinsic nature of the neurological and behavioral phenotype in ADA deficiency.