Innate Immunity in Mucopolysaccharide Diseases.

Mucopolysaccharidoses are rare paediatric lysosomal storage disorders, characterised by accumulation of glycosaminoglycans within lysosomes. This is caused by deficiencies in lysosomal enzymes involved in degradation of these molecules. Dependent on disease, progressive build-up of sugars may lead to musculoskeletal abnormalities and multi-organ failure, and in others, to cognitive decline, which is still a challenge for current therapies. The worsening of neuropathology, observed in patients following recovery from flu-like infections, suggests that inflammation is highly implicated in disease progression. This review provides an overview of the pathological features associated with the mucopolysaccharidoses and summarises current knowledge regarding the inflammatory responses observed in the central nervous system and periphery. We propose a model whereby progressive accumulation of glycosaminoglycans elicits an innate immune response, initiated by the Toll-like receptor 4 pathway, but also precipitated by secondary storage components. Its activation induces cells of the immune system to release pro-inflammatory cytokines, such as TNF-α and IL-1, which induce progression through chronic neuroinflammation. While TNF-α is mostly associated with bone and joint disease in mucopolysaccharidoses, increasing evidence implicates IL-1 as a main effector of innate immunity in the central nervous system. The (NOD)-like receptor protein 3 inflammasome is therefore implicated in chronic neuroinflammation and should be investigated further to identify novel anti-inflammatory treatments.

Evading the AAV Immune Response in Mucopolysaccharidoses.

The humoral immune response elicited by adeno-associated virus (AAV)-mediated gene therapy for the treatment of mucopolysaccharidoses (MPS) poses a significant challenge to achieving therapeutic levels of transgene expression. Antibodies targeting the AAV capsid as well as the transgene product diminish the production of glycosaminoglycan (GAG)-degrading enzymes essential for the treatment of MPS. Patients who have antibodies against AAV capsid increase in number with age, serotype, and racial background and are excluded from the clinical trials at present. In addition, patients who have undergone AAV gene therapy are often excluded from the additional AAV gene therapy with the same serotype, since their acquired immune response (antibody) against AAV will limit further efficacy of treatment. Several methods are being developed to overcome this immune response, such as novel serotype design, antibody reduction by plasmapheresis and immunosuppression, and antibody evasion using empty capsids and enveloped AAV vectors. In this review, we examine the mechanisms of the anti-AAV humoral immune response and evaluate the strengths and weaknesses of current evasion strategies in order to provide an evidence-based recommendation on evading the immune response for future AAV-mediated gene therapies for MPS.

The role of innate immunity in mucopolysaccharide diseases.

Mucopolysaccharidoses are lysosomal storage disorders characterised by accumulation of abnormal pathological glycosaminoglycans, cellular dysfunction and widespread inflammation, resulting in progressive cognitive and motor decline. Lysosomes are important mediators of immune cell function, and therefore accumulation of glycosaminoglycans (GAGs) and other abnormal substrates could affect immune function and directly impact on disease pathogenesis. This review summarises current knowledge with regard to inflammation in mucopolysaccharidosis, with an emphasis on the brain and outlines a potential role for GAGs in induction of inflammation. We propose a model by which the accumulation of GAGs and other factors may impact on innate immune signalling with particular focus on the Toll-like receptor 4 pathway. Innate immunity appears to have a dominating role in mucopolysaccharidosis; however, furthering understanding of innate immune signalling would have significant impact on highlighting novel anti-inflammatory therapeutics for use in mucopolysaccharide diseases. This article is part of the Special Issue "Lysosomal Storage Disorders".

Hematopoietic cell transplantation for mucopolysaccharidosis patients is safe and effective: results after implementation of international guidelines.

Allogeneic hematopoietic cell transplantation (HCT) is the only treatment able to prevent progressive neurodegenerative disease in a selected group of mucopolysaccharidosis (MPS) disorders. However, its use was historically limited by the high risk of graft failure and transplantation-related morbidity and mortality. Therefore, since 2005 new international HCT guidelines for MPS disorders were proposed. The survival and graft outcomes of MPS patients receiving HCT according to these guidelines in 2 European centers of expertise were evaluated. Two consecutive conditioning regimens were used, busulfan/cyclophosphamide or fludarabine/busulfan-based, both with exposure-targeted i.v. busulfan. A noncarrier matched sibling donor (MSD), matched unrelated cord blood (UCB), or matched unrelated donor (MUD) were considered to be preferred donors. If not available, a mismatched UCB donor was used. Participants were 62 MPS patients (56 MPS type I-Hurler, 2 MPS type II, 2 MPS type III, and 2 MPS type VI) receiving HCT at median age 13.5 months (range, 3 to 44). Forty-one patients received a UCB donor, 17 MSD, and 4 MUD. High overall survival (95.2%) and event-free survival (90.3%) were achieved with only low toxicity: 13.3% acute graft-versus-host disease aGVHD) grades II to IV and 14.8% chronic GVHD (1.9% extensive). A mismatched donor predicted for lower event-free survival (P = .04). A higher age at HCT was a predictor for both aGVHD (P = .001) and chronic GVHD (P = .01). The use of a mismatched donor was a predictor for aGVHD (P = .01). Higher rates of full-donor chimerism were achieved in successfully transplanted UCB recipients compared with MSD/MUD (P = .002). If complying with the international HCT guidelines, HCT in MPS patients results in high safety and efficacy. This allows extension of HCT to more attenuated MPS types. Because a younger age at HCT is associated with reduction of HCT-related toxicity, newborn screening may further increase safety.

Outcomes of donor lymphocyte infusion for treatment of mixed donor chimerism after a reduced-intensity preparative regimen for pediatric patients with nonmalignant diseases.

Mixed donor chimerism is increasingly common in the pediatric hematopoietic stem cell transplantation (HSCT) setting because of the increased use of reduced-intensity preparative regimens for nonmalignant diseases. Donor lymphocyte infusion (DLI) is potentially useful in the treatment of mixed donor chimerism, but little are data available on the use of DLI in this setting. We conducted a retrospective review of 27 pediatric patients who received DLI for mixed donor chimerism between January 2006 and December 2010 after receiving a preparative regimen of alemtuzumab, fludarabine, and melphalan. Twenty-one patients (78%) were alive at a median of 35 months post-transplant. Seven patients (26%) sustained full donor chimerism after DLI only at a median of 35 months post-HSCT. Nine patients (33%) continued with mixed donor chimerism (median, 38% [range, 18% to 70%]) at a median of 37 months after DLI only. Five patients underwent unconditioned stem cell boosts or second conditioned transplants after no improvement in donor chimerism was seen following DLI. Donor source appeared to contribute to outcomes after DLI; patients with mismatched unrelated donors had earlier first decline in chimerism and timing of first DLI, a higher response rate to DLI, and an increased rate of graft-versus-host disease (GVHD). There was no response to DLI in patients with matched sibling donors. Ten patients, all with improvement in chimerism after DLI, developed acute GVHD after DLI, with 3 having grade III GVHD. Three patients developed chronic GVHD after DLI. These data illustrate the potential efficacy of DLI in the treatment of mixed donor chimerism after a reduced-intensity preparative regimen.

Avaliação do comprometimento auditivo em pacientes com mucopolissacaridose / Evaluation of hearing loss in patients with mucopolysaccharides Psicol

Introdução: Mucopolissacaridose (MPS) é um conjunto de doenças raras causadas pela deficiência de enzimas lisossômicas levando ao acúmulo de glicosaminoglicanos (GAG) em órgãos e tecidos, responsáveis pelo quadro clínico multissistêmico, crônico e progressivo. O comprometimento auditivo é frequente. Objetivo: Avaliar manifestações auditivas de pacientes com MPS. Metodologia: Estudo descritivo, série de casos do comprometimento auditivo de pacientes com MPS. Foi realizada avaliação retrospectiva através de revisão de prontuário e avaliação prospectiva de dezembro de 2012 a outubro de 2014. Foram analisados a primeira e a última avaliação otorrinolaringológica (ORL) e audiológica realizada. Resultados: A principal queixa auditiva foi a hipoacusia. Aperdaauditiva estava presente em quase todos os pacientes, sendo que a perda auditiva condutiva foi a mais frequente, especialmente nos pacientes com MPS VI. Conclusão: A perda auditiva é muito frequente em pacientes com MPS, devendo o acompanhamento audiológico ser realizado precocemente.

Introduction: Mucopolysaccharidosis (MPS) is a set of rare diseases caused by deficiency of lysosomal enzymes leading to accumulation of glicosaminoglicanos (GAG) in tissues and organs responsible for the multisystemic clinical, chronic and progressive symptons. Objective: Todescribe the profile of otorhinolaryngological clinical examination and audiology tests of patients with MPS disease. Methods:Study of case series. The evaluation was performed, at the beginning, in 31 patients with MPS I, II, IIIA, IV and VI. Results: The most common hearing complaint was hearing loss and it was confirmed by audiology tests in almost 100% of patients, mostly with condutive hearing loss. Conclusions: It is important to evaluate the complaints, physical examination and audiology tests in MPS disease. Otorhinolaryngologist should be part of professional group that follow these patients in order to better monitor their hearing and provide early hearing rehabilitation.

Mucopolysaccharide diseases: a complex interplay between neuroinflammation, microglial activation and adaptive immunity.

Mucopolysaccharide (MPS) diseases are lysosomal storage disorders (LSDs) caused by deficiencies in enzymes required for glycosaminoglycan (GAG) catabolism. Mucopolysaccharidosis I (MPS I), MPS IIIA, MPS IIIB and MPS VII are deficient in the enzymes α-L-Iduronidase, Heparan-N-Sulphatase, N-Acetylglucosaminidase and Beta-Glucuronidase, respectively. Enzyme deficiency leads to the progressive multi-systemic build-up of heparan sulphate (HS) and dermatan sulphate (DS) within cellular lysosomes, followed by cell, tissue and organ damage and in particular neurodegeneration. Clinical manifestations of MPS are well established; however as lysosomes represent vital components of immune cells, it follows that lysosomal accumulation of GAGs could affect diverse immune functions and therefore influence disease pathogenesis. Theoretically, MPS neurodegeneration and GAGs could be substantiating a threat of danger and damage to alert the immune system for cellular clearance, which due to the progressive nature of MPS storage would propagate disease pathogenesis. Innate immunity appears to have a key role in MPS; however the extent of adaptive immune involvement remains to be elucidated. The current literature suggests a complex interplay between neuroinflammation, microglial activation and adaptive immunity in MPS disease.

Osteoimmunology in mucopolysaccharidoses type I, II, VI and VII. Immunological regulation of the osteoarticular system in the course of metabolic inflammation.

BACKGROUND: Mucopolysaccharidoses (MPSs) are rare genetic diseases caused by a deficient activity of one of the lysosomal enzymes involved in the glycosaminoglycan (GAG) breakdown pathway. These metabolic blocks lead to the accumulation of GAGs in various organs and tissues, resulting in a multisystemic clinical picture. The pathological GAG accumulation begins a cascade of interrelated responses: metabolic, inflammatory and immunological with systemic effects. Metabolic inflammation, secondary to GAG storage, is a significant cause of osteoarticular symptoms in MPS disorders. OBJECTIVE AND METHOD: The aim of this review is to present recent progress in the understanding of the role of inflammatory and immune processes in the pathophysiology of osteoarticular symptoms in MPS disorders and potential therapeutic interventions based on published reports in MPS patients and studies in animal models. RESULTS AND CONCLUSIONS: The immune and skeletal systems have a number of shared regulatory molecules and many relationships between bone disorders and aberrant immune responses in MPS can be explained by osteoimmunology. The treatment options currently available are not sufficiently effective in the prevention, inhibition and treatment of osteoarticular symptoms in MPS disease. A lot can be learnt from interactions between skeletal and immune systems in autoimmune diseases such as rheumatoid arthritis (RA) and similarities between RA and MPS point to the possibility of using the experience with RA in the treatment of MPS in the future. The use of different anti-inflammatory drugs requires further study, but it seems to be an important direction for new therapeutic options for MPS patients.

Involvement of the Toll-like receptor 4 pathway and use of TNF-alpha antagonists for treatment of the mucopolysaccharidoses.

Enzyme replacement therapy is currently available for three of the mucopolysaccharidoses (MPSs) but has limited effects on the skeletal lesions. We investigated the involvement of the Toll-like receptor 4 (TLR4) signaling pathway in the pathogenesis of MPS bone and joint disease, and the use of the anti-TNF-alpha drug, Remicade (Centocor, Inc.), for treatment. TLR4 KO (TLR4(lps-/-)) mice were interbred with MPS VII mice to produce double-KO (DKO) animals. The DKO mice had longer and thinner faces and longer femora as revealed by micro-computed tomography analysis compared with MPS VII mice. Histological analyses also revealed more organized and thinner growth plates. The serum levels of TNF-alpha were normalized in the DKO animals, and the levels of phosphorylated STAT1 and STAT3 in articular chondrocytes were corrected. These findings led us to evaluate the effects of Remicade in MPS VI rats. When initiated at 1 month of age, i.v. treatment prevented the elevation of TNF-alpha, receptor activator of NF-kappaB, and other inflammatory molecules not only in the blood but in articular chondrocytes and fibroblast-like synoviocytes (FLSs). Treatment of 6-month-old animals also reduced the levels of these molecules to normal. The number of apoptotic articular chondrocytes in MPS VI rats was similarly reduced, with less infiltration of synovial tissue into the underlying bone. These studies revealed the important role of TLR4 signaling in MPS bone and joint disease and suggest that targeting TNF-alpha may have positive therapeutic effects.

Reconstitution of lymphocyte subpopulations in children with inherited metabolic storage diseases after haematopoietic cell transplantation.

We prospectively evaluated the reconstitution of lymphocyte subpopulations in nine children with lysosomal diseases who underwent 11 allogeneic haematopoietic cell transplants (HCTs) following CD34(+) immunomagnetic enrichment, limited T-cell addback and in vivo B-cell depletion. Absolute lymphocyte count recovery was slow to cross the 5th percentile, occurring at a median of 10 months after HCT in patients with full chimaerism. Natural killer cells represented up to 90% of the total lymphoid population during the first 3 months. CD4(+) lymphocyte recovery occurred 9-18 months after HCT. In most patients, CD8(+) lymphocyte recovery was slow and comparable with that of CD4(+) lymphocytes. The CD4(+)/CD8(+) ratio normalised by 3-7 months after HCT in 50% of the patients. CD8(+) lymphocyte recovery was enhanced in patients with viral reactivation. Reconstitution of B-lymphocytes was particularly delayed in patients treated with rituximab. Declining chimaerism, rejection and viral reactivation were the most common problems in our series. Because of the unique graft manipulation, the pace of lymphocyte reconstitution was particularly slow, suggesting that these patients are at a significantly increased risk of infections for up to 2 years after HCT.

Immunelectronmicroscopic characterization of T4 and T8 lymphocytes and natural killer cells in neuronal ceroid-lipofuscinosis.

CD4+, CD8+, and CD56+ cells were isolated with the immunomagnetic separation technique from peripheral blood mononuclear cells (PBMC) of 3 patients with neuronal ceroid-lipofuscinosis: one patient each with infantile (INCL), late infantile (LINCL), and juvenile (JNCL) neuronal ceroid-lipofuscinoses, all studied by light (LM) and electron (EM) microscopy. To compare the pathology of these cells with affected cells in other types of lysosomal diseases, the separation was also performed with PBMC of 1 patient with mucolipidosis (ML) type II, 2 patients with mucopolysaccharidosis (MPS) type I, and 4 patients with MPS type III. Disease-specific lysosomal inclusions were identified in CD4+, CD8+, and CD56+ cells. Subclass-specific storage products could not be found. T4 and T8 lymphocytes showed nearly the same ratio of affected to nonaffected cells. These results suggest that lysosomal storage can be found in most of the lymphocyte subclasses in these forms of neuronal ceroid-lipofuscinoses and other lysosomal diseases.

Characterization of T-cell subclasses and NK-cells in lysosomal disorders by immuno-electron microscopy.

Previous studies have shown that B and T lymphocytes are affected in lysosomal disorders. The aim of this study was to investigate the involvement of subclasses of T lymphocytes and natural killer cells in lysosomal diseases. CD4+, CD8+, and CD56+ cells were immunomagnetically separated from peripheral blood mononuclear cells in 10 patients with various lysosomal diseases--including one patient each with infantile, late infantile, and juvenile neuronal ceroid-lipfuscinoses, two patients with mucopolysaccharidosis (MPS) type I and four patients with MPS type III, and one patient with mucolipidosis type II; all lymphocytes were studied by light and electron microscopy. Respective vacuolar or non-vacuolar lysosomal inclusions were identified in CD4+, CD8+, and CD56+ cells. Subclass-specific abnormal lysosomes could not be found. Both subclasses of T lymphocytes investigated showed nearly the same ratio of affected to non-affected cells. These results were obtained by studying a small number of different lysosomal disorders. Nevertheless, they suggest that lysosomal storage in these subclasses of immunocompetent cells might also be found in other forms of lysosomal disease.

Review: the immunochemical analysis of enzyme from mucopolysaccharidoses patients.

The immunochemical analysis of enzyme from mucopolysaccharidoses (MPS) patients is aimed at defining the level and nature of the enzymically deficient protein produced by specific gene mutations. Immunochemical techniques allow purification of enzyme, characterization of the composite molecular species, measurement of cellular protein content, investigation of protein biosynthesis, determination of subcellular distribution, as well as information on protein structure and folding. This review focuses on the application of immunochemical techniques to the study of the aberrant protein produced in skin fibroblast cells derived from MPS patients. The analysis of enzyme protein has been applied to phenotype expression within single enzyme deficiency disorders. It is proposed that reliable prediction of MPS patient phenotype may require a combined approach utilizing immunochemical, biochemical, cell biological and gene analysis. However, this review will address the structure and nature of the protein produced in cells from MPS patients, the biological activity of this protein, and the incorporation of the protein into, and location within, subcellular fractions.

Treatment of mucopolysaccharidosis: clinical and biochemical aspects of leucocyte transfusion as compared with plasma infusion in patients with Hurler's and Scheie's syndromes.

The therapeutic effectiveness of leucocyte transfusion (LT) was compared with that of plasma infusion (PI) clinically by range of motion (ROM) of joints and biochemically from the standpoint of alpha-L-iduronidase activity and urinary excretion of acid mucopolysaccharides (AMPS) in 2 patients with Hurler's and Scheie's syndromes. Both syndromes are considered to be due to the lack of alpha-L-iduronidase activity, a congenital metabolic disorder. As a result, leukocyte transfusion surpasses plasma infusion with respect to enzyme content, the grade and duration of clinical improvement in the stiffness of joints. Clinical improvement in the stiffness of joints was correlated with the degradation of AMPS when the ratio of urinary AMPS fragments to the total large molecule AMPS has become 50% or more after the leucocyte transfusion and plasma infusion.

Human beta-glucuronidase deficiency mucopolysaccharidosis: identification of cross-reactive antigen in cultured fibroblasts of deficient patients by enzyme immunoassay.

An enzyme immunoassay for human beta-glucuronidase was developed to determine the presence or absence of antigenically cross-reactive material (CRM) in patients with beta-glucuronidase deficiency mucopolysaccharidosis. This assay provided a sensitive means of measuring the primary interaction between the enzyme molecule and antibody but required neither pure antigen nor monospecific antiserum, an important consideration, since neither of these was available. Goat antiserum to partially purified human placenta beta-glucuronidase did not recognize differences in normal enzyme from human placenta, liver, fibroblasts, or blood platelets. CRM was identified in fibroblast extracts from all four of the unrelated beta-glucuronidase-deficient patients studied, but titration patterns indicated genetic heterogeneity among these four mutant proteins. Fibroblast enzymes from two obligate heterozygotes were distinguishable immunologically from normal enzyme. The enzyme immunoassay was also used to compare human enzyme with liver enzyme from other mammalian species. CRM was present in liver extracts of all species tested, but the liver enzymes, except for the rabbit, were weakly cross-reactive. We conclude that despite certain limitations, the enzyme immunoassay for human beta-glucuronidase is useful and that all four beta-glucuronidase-deficient patients studied possess CRM.