The clinical spectrum of SARS-CoV-2 infection in Gaucher disease: Effect of both a pandemic and a rare disease that disrupts the immune system.

INTRODUCTION: The impact of SARS-CoV-2 in rare disease populations has been underreported. Gaucher disease (GD) is a prototype rare disease that shares with SARS-CoV-2 a disruption of the lysosomal pathway. MATERIALS-METHODS: Retrospective analysis of 11 patients with Type 1 GD who developed COVID-19 between March 2020 and March 2021. RESULTS: Seven male and 4 female patients with Type 1 GD developed COVID-19. One was a pediatric patient (8 years old) while the remainder were adults, median age of 44 years old (range 21 to 64 years old). Two patients required hospitalization though none required intensive care or intubation. All 11 patients recovered from COVID-19 and there were no reported deaths. CONCLUSIONS: Our case series suggests that GD patients acquired COVID-19 at a similar frequency as the general population, though experienced a milder overall course despite harboring underlying immune system dysfunction and other known co-morbidities that confer high risk of adverse outcomes from SARS-CoV-2 infection.

C-X-C Motif Chemokine Ligand 9 and Its CXCR3 Receptor Are the Salt and Pepper for T Cells Trafficking in a Mouse Model of Gaucher Disease.

Gaucher disease is a lysosomal storage disease, which happens due to mutations in GBA1/Gba1 that encodes the enzyme termed as lysosomal acid ß-glucosidase. The major function of this enzyme is to catalyze glucosylceramide (GC) into glucose and ceramide. The deficiency of this enzyme and resultant abnormal accumulation of GC cause altered function of several of the innate and adaptive immune cells. For example, augmented infiltration of T cells contributes to the increased production of pro-inflammatory cytokines, (e.g., IFNγ, TNFα, IL6, IL12p40, IL12p70, IL23, and IL17A/F). This leads to tissue damage in a genetic mouse model (Gba19V/-) of Gaucher disease. The cellular mechanism(s) by which increased tissue infiltration of T cells occurs in this disease is not fully understood. Here, we delineate role of the CXCR3 receptor and its exogenous C-X-C motif chemokine ligand 9 (CXCL9) in induction of increased tissue recruitment of CD4+ T and CD8+ T cells in Gaucher disease. Intracellular FACS staining of macrophages (Mϕs) and dendritic cells (DCs) from Gba19V/- mice showed elevated production of CXCL9. Purified CD4+ T cells and the CD8+ T cells from Gba19V/- mice showed increased expression of CXCR3. Ex vivo and in vivo chemotaxis experiments showed CXCL9 involvement in the recruitment of Gba19V/- T cells. Furthermore, antibody blockade of the CXCL9 receptor (CXCR3) on T cells caused marked reduction in CXCL9- mediated chemotaxis of T cells in Gba19V/- mice. These data implicate abnormalities of the CXCL9-CXCR3 axis leading to enhanced tissue recruitment of T cells in Gaucher disease. Such results provide a rationale for blockade of the CXCL9/CXCR3 axis as potential new therapeutic targets for the treatment of inflammation in Gaucher disease.

Dendritic cells and monocyte subsets in children with Gaucher disease.

BACKGROUND: There are minimal data on the frequencies of monocyte subsets and dendritic cells (DCs) in children with Gaucher disease (GD), as nearly all previous studies have involved adult patients. Consequently, we aimed to describe the changes in these cell subpopulations in children with GD type 1 who were on regular enzyme replacement therapy (ERT). METHODS: This case-control study included 25 children with GD1 and 20 healthy controls. All participants underwent investigations such as complete blood count and flow cytometric assessment of DC and monocyte frequencies and phenotype. RESULTS: We found that GD1 children had significantly reduced percentages of both types of DCs, i.e., plasmacytoid DCs and myeloid DCs, compared to the control group. There was also a significant reduction in absolute monocyte numbers and percentage of classical monocyte. Moreover, the GD1 children had higher frequencies of non-classical and intermediate monocytes than the control group. CONCLUSIONS: Our results so far indicate that, when compared to the control group, the GD1 children had significantly reduced total and classical monocyte, with significantly decreased frequencies for both types of DCs. These changes can contribute to immunological abnormalities in pediatric patients with GD1. IMPACT: Children with Gaucher disease type 1 (GD1) have significantly reduced total and classical monocyte frequencies, with decreasing percentages for both types of dendritic cells. GD1 children had significantly reduced frequencies of myeloid and plasmacytoid dendritic cells as compared to the controls. The GD1 children also had significant changes in monocyte subsets when compared to the controls. Our results show that monocytes and dendritic cells' significant changes could contribute to immunological abnormalities in pediatric patients with GD1.

Innate immune response in neuronopathic forms of Gaucher disease confers resistance against viral-induced encephalitis.

Both monogenic diseases and viral infections can manifest in a broad spectrum of clinical phenotypes that range from asymptomatic to lethal, suggesting that other factors modulate disease severity. Here, we examine the interplay between the genetic neuronopathic Gaucher's disease (nGD), and neuroinvasive Sindbis virus (SVNI) infection. Infection of nGD mice with SVNI had no influence on nGD severity. However, nGD mice were more resistant to SVNI infection. Significantly different inflammatory responses were seen in nGD brains when compared with SVNI brains: the inflammatory response in the nGD brains consisted of reactive astrocytes and microglia with no infiltrating macrophages, but the inflammatory response in the brains of SVNI-infected mice was characterized by infiltration of macrophages and altered activation of microglia and astrocytes. We suggest that the innate immune response activated in nGD confers resistance against viral infection of the CNS.

Downregulation of B regulatory cells and upregulation of T helper 1 cells in children with Gaucher disease undergoing enzyme replacement therapy.

Gaucher disease (GD) involves a broad spectrum of immunological cells, including T helper (Th) cells and regulatory B cells (Bregs), which function to resolve the immune response and inhibit excessive inflammation. This study aimed to explore T helper cells, B cells, and Bregs in GD children undergoing enzyme replacement therapy (ERT). Our study included 20 GD patients; six patients were categorized as type 1 and 14 as type 3 GD. All patients were on regular ERT. Twenty healthy children were enrolled as controls. All patients and controls were subjected to complete blood analysis, abdominal ultrasound, and flow cytometric detection of T helper cells, B cells, and Bregs. Despite undergoing ERT, CD4+ T helper lymphocytes and Bregs were still significantly lower in patients with GD compared with the controls. Th1 and B cells were more in the patients than in the healthy controls. Lower levels of Bregs were found in type 3, compared with type 1 patients. Increased platelet count was directly associated with increased levels of Bregs and lower levels of B cells. Elevated children's height was also accompanied by decreasing levels of Th1. Our results propose that ERT in GD is associated with partial improvement in immune status, and long-term ERT might be needed for the restoration of the desired immune response levels. Levels of Bregs and Th1 can be employed for monitoring improvement of immune status in GD patients undergoing ERT.

Effect of Substrate Reduction Therapy in Comparison to Enzyme Replacement Therapy on Immune Aspects and Bone Involvement in Gaucher Disease.

Gaucher disease (GD) is caused by mutations in the GBA gene, leading to deficient activity of the lysosomal enzyme glucocerebrosidase. Among all the symptoms across various organ systems, bone disease is a major concern as it causes high morbidity and reduces quality of life. Enzyme replacement therapy (ERT) is the most accepted treatment; however, there are still unmet needs. As an alternative, substrate reduction therapy (SRT) was developed using glucosylceramide synthase inhibitors. In the current study, the effects of ERT vs. SRT were compared, particularly the immunological and bone remodeling aspects. GD subjects were divided into three cohorts based on their treatment at initial visit: ERT, SRT, and untreated (UT). Immunophenotyping showed no significant immune cell alterations between the cohorts. Expression of RANK/RANKL/Osteoprotegerin pathway components on immune cells and the secreted markers of bone turnover were analyzed. In the ERT cohort, no significant changes were observed in RANK, RANKL or serum biomarkers. RANKL on T lymphocytes, Osteopontin and MIP-1ß decreased with SRT treatment indicating probable reduction in osteoclast activity. Other secreted factors, Osteocalcin and RANKL/Osteoprotegerin did not change with the treatment status. Insights from the study highlight personalized differences between subjects and possible use of RANK pathway components as markers for bone disease progression.

Hyperimmunoglobulinemia and IgG Subclass Abnormalities in Children With Gaucher Disease.

Gaucher disease (GD) is the most common lysosomal storage disorder, the aim of the current study was to investigate hyperimmunoglobulinemia and abnormalities of serum immunoglobulin G (IgG) subclasses in children with GD and the relation of those findings to the GD phenotype and genotype, duration of enzyme replacement therapy (ERT), and infection frequency. The study included 20 Egyptian children with GD receiving ERT and 20 age-matched and sex-matched healthy children as controls. Serum Ig and serum IgG subclass levels were measured in the children with GD. Serum IgG subclass levels were measured in the control subjects. Hyperimmunoglobulinemia was present in 15 of the 20 (75%) children with GD. In addition, it is found significantly lower IgG2 levels and significantly higher IgG3 levels in the GD group than in the control group (P<0.001 and <0.006, respectively). Patients with 12 infections per year had significantly higher IgG3 levels compared with patients with 6 infections per year (P=0.022). In conclusion, hyperimmunoglobulinemia and IgG subclass abnormalities occur in children with GD who are on ERT and may be related to recurrent infections.

Gene expression profile in patients with Gaucher disease indicates activation of inflammatory processes.

Gaucher disease (GD) is a rare inherited metabolic disease caused by pathogenic variants in the GBA1 gene. So far, the pathomechanism of GD was investigated mainly in animal models. In order to delineate the molecular changes in GD cells we analysed gene expression profile in cultured skin fibroblasts from GD patients, control individuals and, additionally, patients with Niemann-Pick type C disease (NPC). We used expression microarrays with subsequent validation by qRT-PCR method. In the comparison GD patients vs. controls, the most pronounced relative fold change (rFC) in expression was observed for genes IL13RA2 and IFI6 (up-regulated) and ATOH8 and CRISPLD2 (down-regulated). Products of up-regulated and down-regulated genes were both enriched in genes associated with immune response. In addition, products of down-regulated genes were associated with cell-to-cell and cell-to-matrix interactions, matrix remodelling, PI3K-Akt signalling pathway and a neuronal survival pathway. Up-regulation of PLAU, IFIT1, TMEM158 and down-regulation of ATOH8 and ISLR distinguished GD patients from both NPC patients and healthy controls. Our results emphasize the inflammatory character of changes occurring in human GD cells indicating that further studies on novel therapeutics for GD should consider anti-inflammatory agents.

An unexpected player in Gaucher disease: The multiple roles of complement in disease development.

The complement system is well appreciated for its role as an important effector of innate immunity that is activated by the classical, lectin or alternative pathway. C5a is one important mediator of the system that is generated in response to canonical and non-canonical C5 cleavage by circulating or cell-derived proteases. In addition to its function as a chemoattractant for neutrophils and other myeloid effectors, C5a and its sister molecule C3a have concerted roles in cell homeostasis and surveillance. Through activation of their cognate G protein coupled receptors, C3a and C5a regulate multiple intracellular pathways within the mitochondria and the lysosomal compartments that harbor multiple enzymes critical for protein, carbohydrate and lipid metabolism. Genetic mutations of such lysosomal enzymes or their receptors can result in the compartmental accumulation of specific classes of substrates in this organelle summarized as lysosomal storage diseases (LSD). A frequent LSD is Gaucher disease (GD), caused by autosomal recessively inherited mutations in GBA1, resulting in functional defects of the encoded enzyme, acid ß-glucosidase (glucocerebrosidase, GCase). Such mutations promote excessive accumulation of ß-glucosylceramide (GC or GL1) in innate and adaptive immune cells frequently associated with chronic inflammation. Recently, we uncovered an unexpected link between the C5a and C5a receptor 1 (C5aR1) axis and the accumulation of GL1 in experimental and clinical GD. Here, we will review the pathways of complement activation in GD, its role as a mediator of the inflammatory response, and its impact on glucosphingolipid metabolism. Further, we will discuss the potential role of the C5a/C5aR1 axis in GL1-specific autoantibody formation and as a novel therapeutic target in GD.

Could enzyme replacement therapy promote immune tolerance in Gaucher disease type 1?

Among the lysosomal storage disorders, Gaucher disease (GD) features some of the most striking alterations in the immune system, including increased levels of cytokines and chemokines. Although studies have demonstrated the efficacy of enzyme replacement therapy (ERT) for GD, the ideal dosage remains controversial. In this study, we report differences in levels of cytokines (IL-6, TNF-a, and IFN-y) and chemokines (IL-8, IP-10, and MCP-1) in patients with GD type 1 treated with different ERT dosages and treatment durations. Patients were recruited from two ERT centers in Brazil and divided into two groups according to treatment facility. Comparison between groups showed that patients in group 1 had received ERT for longer (p=0.0078) and at higher doses (p=0.0002) than those in group 2. Patients in group 1 exhibited decreased levels of IL-6 (p=0.0006), TNF-α (p<0.0001), IFN-γ (p<0.0001), IL-8 (p=0.0083), IP-10 (p<0.0001), and MCP-1 (p<0.0001) when compared to patients in group 2. Otherwise, patients in both groups were clinically similar, with no differences in hemoglobin, platelet, or leukocyte counts. Our data suggest that in GD type 1 the dosage and duration of therapy may be associated with establishment of peripheral tolerance and, consequently, decreased serum levels of inflammatory cytokines and chemokines.

Prevalence of autoantibodies in the course of Gaucher disease type 1: A multicenter study comparing Gaucher disease patients to healthy subjects.

OBJECTIVES: Type 1 Gaucher disease may be related to the presence of autoantibodies. Their clinical significance is questioned. Primary endpoint was to compare the prevalence of autoantibodies in type 1 Gaucher disease patients with healthy subjects, seeking correlations with autoimmune characteristics. Secondary endpoints were to determine whether patients with autoantibodies reported autoimmunity-related symptoms and if genotype, splenectomy or treatment influenced autoantibodies presence. METHODS: Type 1 Gaucher disease patients and healthy volunteers were included in this national multicenter exploratory study. Autoantibodies presence was compared in both groups and assessed regarding to genotype, splenectomy, Gaucher disease treatment and autoimmunity-related symptoms. RESULTS: Twenty healthy subjects and 40 type 1 Gaucher disease patients were included. Of the studied group: 15 patients undergone splenectomy, 37 were treated either with enzyme replacement therapy (34) or with substrate reduction therapy (3), 25 were homozygous/heterozygous for the N370S mutation. In type 1 Gaucher disease group (studied group), 52% had positive autoantibodies versus 26% in control group. Antiphospholipid antibodies were more frequent in the studied group (30% vs. 5%), but without correlation to thrombosis, osteonecrosis or bone infarcts. In the studied group, antinuclear antibodies were more frequent (25% vs. 16%). None of the patients with autoantibodies had clinical manifestations of autoimmune diseases. Autoantibodies were not correlated with treatment, genotype, or splenectomy, except for anticardiolipid, more frequent in splenectomized patients. CONCLUSIONS: In type 1 Gaucher disease, autoantibodies were more frequent compared to a healthy population. However, they were not associated with an increased prevalence of clinical active autoimmune diseases.

Enzyme replacement therapy reverses B lymphocyte and dendritic cell dysregulations in patients with Gaucher Disease.

Gaucher disease (GD) is caused by mutations in the GBA gene encoding lysosomal enzyme, ß-glucocerebrosidase (GCase). GCase deficiency results in accumulation of its substrates in cells of macrophage lineage, affecting multiple organ systems. Enzyme replacement therapy (ERT) with recombinant human GCase is the standard of care to treat GD. In GD, it is well established that there are immune alterations, clinically presenting as lymphadenopathy, gammopathies, and predisposition to hematological cancers. We examined the effect of ERT on immune dysregulations in treatment-naïve GD patients longitudinally after the initiation of ERT. Immunophenotyping was performed in peripheral blood samples obtained before and after ERT. T and B lymphocyte subsets, NK, NKT and dendritic cells were evaluated. In all treatment naïve patients at baseline, transitional B cells, characterized by CD21low expression were markedly elevated. After establishment of stable-dose therapy, CD21low cells were significantly reduced and subsequent increase in CD21Hi B lymphocytes indicated improved B cell maturation. Class-switching and memory B cell defects which were noted prior to treatment were found to be normalized. An increase in dendritic cells also resulted after the treatment. Our data shows that GD affects across various immune cell types and ERT or its effects directly improve affected immunological parameters.

Enzymes as Immunotherapeutics.

Enzymes are attractive as immunotherapeutics because they can catalyze shifts in the local availability of immunostimulatory and immunosuppressive signals. Clinical success of enzyme immunotherapeutics frequently hinges upon achieving sustained biocatalysis over relevant time scales. The time scale and location of biocatalysis are often dictated by the location of the substrate. For example, therapeutic enzymes that convert substrates distributed systemically are typically designed to have a long half-life in circulation, whereas enzymes that convert substrates localized to a specific tissue or cell population can be more effective when designed to accumulate at the target site. This Topical Review surveys approaches to improve enzyme immunotherapeutic efficacy via chemical modification, encapsulation, and immobilization that increases enzyme accumulation at target sites or extends enzyme half-life in circulation. Examples provided illustrate "replacement therapies" to restore deficient enzyme function, as well as "enhancement therapies" that augment native enzyme function via supraphysiologic doses. Existing FDA-approved enzyme immunotherapies are highlighted, followed by discussion of emerging experimental strategies such as those designed to enhance antitumor immunity or resolve inflammation.

Complement drives glucosylceramide accumulation and tissue inflammation in Gaucher disease.

Gaucher disease is caused by mutations in GBA1, which encodes the lysosomal enzyme glucocerebrosidase (GCase). GBA1 mutations drive extensive accumulation of glucosylceramide (GC) in multiple innate and adaptive immune cells in the spleen, liver, lung and bone marrow, often leading to chronic inflammation. The mechanisms that connect excess GC to tissue inflammation remain unknown. Here we show that activation of complement C5a and C5a receptor 1 (C5aR1) controls GC accumulation and the inflammatory response in experimental and clinical Gaucher disease. Marked local and systemic complement activation occurred in GCase-deficient mice or after pharmacological inhibition of GCase and was associated with GC storage, tissue inflammation and proinflammatory cytokine production. Whereas all GCase-inhibited mice died within 4-5 weeks, mice deficient in both GCase and C5aR1, and wild-type mice in which GCase and C5aR were pharmacologically inhibited, were protected from these adverse effects and consequently survived. In mice and humans, GCase deficiency was associated with strong formation of complement-activating GC-specific IgG autoantibodies, leading to complement activation and C5a generation. Subsequent C5aR1 activation controlled UDP-glucose ceramide glucosyltransferase production, thereby tipping the balance between GC formation and degradation. Thus, extensive GC storage induces complement-activating IgG autoantibodies that drive a pathway of C5a generation and C5aR1 activation that fuels a cycle of cellular GC accumulation, innate and adaptive immune cell recruitment and activation in Gaucher disease. As enzyme replacement and substrate reduction therapies are expensive and still associated with inflammation, increased risk of cancer and Parkinson disease, targeting C5aR1 may serve as a treatment option for patients with Gaucher disease and, possibly, other lysosomal storage diseases.

Efferocytosis is impaired in Gaucher macrophages.

Gaucher disease, the inherited deficiency of lysosomal glucocerebrosidase, is characterized by the presence of glucosylceramide-laden macrophages resulting from impaired digestion of aged erythrocytes or apoptotic leukocytes. Studies of macrophages from patients with type 1 Gaucher disease with genotypes N370S/N370S, N370S/L444P or N370S/c.84dupG revealed that Gaucher macrophages have impaired efferocytosis resulting from reduced levels of p67phox and Rab7. The decreased Rab7 expression leads to impaired fusion of phagosomes with lysosomes. Moreover, there is defective translocation of p67phox to phagosomes, resulting in reduced intracellular production of reactive oxygen species. These factors contribute to defective deposition and clearance of apoptotic cells in phagolysosomes, which may have an impact on the inflammatory response and contribute to the organomegaly and inflammation seen in patients with Gaucher disease.

Activated and Memory T Lymphocytes in Children with Gaucher Disease.

Gaucher disease (GD) is the most prevalent lysosomal storage disorder. Gaucher disease is associated with remarkable alterations in the immune system, and GD patients are more susceptible to infections and are at a higher risk of developing autoimmune disorders and malignancies. In a case-control study, we used three-color flow cytometric immunophenotyping for determination of the frequency of lymphocyte subpopulations and activated T lymphocytes among 18 children with GD1 under enzyme replacement therapy managed in Assiut University Hospitals. We found significant increases in the frequencies of total lymphocytes, CD19+, CD3+, CD4+, and CD8+ in children with GD1 when compared to healthy control. The frequencies of activated T lymphocytes (CD3+HLA-DR+), activated T-helper cells (CD4+HLA-DR+), and activated T-suppressor/cytotoxic cells (CD8+HLA-DR+) were significantly higher in GD1 as compared to healthy children. Our data show that the increased proportion of activated T lymphocytes in children with GD1 raises the issue of their possible involvement in the pathogenesis of the immune dysfunction seen in these patients. Our data suggested that the activated T lymphocytes could play a role in the clinical course of GD1. The relationship of these cells to immune disorders in GD1 children remains to be determined.

Time of Initiating Enzyme Replacement Therapy Affects Immune Abnormalities and Disease Severity in Patients with Gaucher Disease.

Gaucher disease (GD) patients often present with abnormalities in immune response that may be the result of alterations in cellular and/or humoral immunity. However, how the treatment and clinical features of patients impact the perturbation of their immunological status remains unclear. To address this, we assessed the immune profile of 26 GD patients who were part of an enzyme replacement therapy (ERT) study. Patients were evaluated clinically for onset of GD symptoms, duration of therapy and validated outcome measures for ERT. According to DS3 disease severity scoring system criteria, they were assigned to have mild, moderate or severe GD. Flow cytometry based immunophenotyping was performed to analyze subsets of T, B, NK, NKT and dendritic cells. GD patients showed multiple types of immune abnormalities associated to T and B lymphocytes with respect to their subpopulations as well as memory and activation markers. Skewing of CD4 and CD8 T cell numbers resulting in lower CD4/CD8 ratio and an increase in overall T cell activation were observed. A decrease in the overall B cells and an increase in NK and NKT cells were noted in the GD patients compared to controls. These immune alterations do not correlate with GD clinical type or level of biomarkers. However, subjects with persistent immune alterations, especially in B cells and DCs correlate with longer delay in initiation of ERT (ΔTX). Thus, while ERT may reverse some of these immune abnormalities, the immune cell alterations become persistent if therapy is further delayed. These findings have important implications in understanding the immune disruptions before and after treatment of GD patients.

Clonal Immunoglobulin against Lysolipids in the Origin of Myeloma.

Antigen-driven selection has been implicated in the pathogenesis of monoclonal gammopathies. Patients with Gaucher's disease have an increased risk of monoclonal gammopathies. Here we show that the clonal immunoglobulin in patients with Gaucher's disease and in mouse models of Gaucher's disease-associated gammopathy is reactive against lyso-glucosylceramide (LGL1), which is markedly elevated in these patients and mice. Clonal immunoglobulin in 33% of sporadic human monoclonal gammopathies is also specific for the lysolipids LGL1 and lysophosphatidylcholine (LPC). Substrate reduction ameliorates Gaucher's disease-associated gammopathy in mice. Thus, long-term immune activation by lysolipids may underlie both Gaucher's disease-associated gammopathies and some sporadic monoclonal gammopathies.

CD4+CD25 high Foxp3+ Treg deficiency in a Brazilian patient with Gaucher disease and lupus nephritis.

Gaucher Disease (GD) is a rare autosomal recessive disorder caused by the deficient activity of beta-glucocerebrosidase. GD is one of the lysosomal storage diseases with the most remarkable alterations in the immune system, and that may manifest clinically as autoimmune disorders and malignancy. We reported the immunological evaluation of a patient with GD and lupus nephritis. Decreased absolute values of T, and NK, and an inversion of CD4(+)/CD8(+) ratio, low levels of IgM and normal B cells were found when compared to reference values in a Brazilian population. Absence ofCD4(+)CD25(high)Foxp3(+) Treg and high levels of total NKT, iNKT cells and CD8(+) iNKT subsets were also observed when compared to the healthy control and GD patient without lupus nephritis. Treg subset and CD8(+) iNKT abnormalities might be involved in severe lupus nephritis in a GD patient. We conclude by emphasizing the importance of the immunological evaluation on early diagnosis of autoimmunity contributing to reduce mortality and morbidity of these patients.