A Phase 1B Trial in GBA1-Associated Parkinson's Disease of BIA-28-6156, a Glucocerebrosidase Activator.

BACKGROUND: Loss-of-function mutations in the GBA1 gene are one of the most common genetic risk factors for onset of Parkinson's disease and subsequent progression (GBA-PD). GBA1 encodes the lysosomal enzyme glucocerebrosidase (GCase), a promising target for a possible first disease-modifying therapy. LTI-291 is an allosteric activator of GCase, which increases the activity of normal and mutant forms of GCase. OBJECTIVES: This first-in-patient study evaluated the safety, tolerability, pharmacokinetics, and pharmacodynamics of 28 daily doses of LTI-291 in GBA-PD. METHODS: This was a randomized, double-blind, placebo-controlled trial in 40 GBA-PD participants. Twenty-eight consecutive daily doses of 10, 30, or 60 mg of LTI-291 or placebo were administered (n = 10 per treatment allocation). Glycosphingolipid (glucosylceramide and lactosylceramide) levels were measured in peripheral blood mononuclear cells (PBMCs), plasma, and cerebrospinal fluid (CSF), and a test battery of neurocognitive tasks, the Movement Disorder Society-Unified Parkinson's Disease Rating Scale and the Mini-Mental State Exam, were performed. RESULTS: LTI-291 was generally well tolerated, no deaths or treatment-related serious adverse events occurred, and no participants withdrew due to adverse events. Cmax , and AUC0-6 of LTI-291 increased in a dose-proportional manner, with free CSF concentrations equal to the free fraction in plasma. A treatment-related transient increase in intracellular glucosylceramide (GluCer) in PBMCs was measured. CONCLUSION: These first-in-patient studies demonstrated that LTI-291 was well tolerated when administered orally for 28 consecutive days to patients with GBA-PD. Plasma and CSF concentrations that are considered pharmacologically active were reached (ie, sufficient to at least double GCase activity). Intracellular GluCer elevations were detected. Clinical benefit will be assessed in a larger long-term trial in GBA-PD. © 2023 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

A 20-Year Longitudinal Study of Plasma Chitotriosidase Activity in Treated Gaucher Disease Type 1 and 3 Patients-A Qualitative and Quantitative Approach.

Chitotriosidase is an enzyme produced and secreted in large amounts by activated macrophages, especially macrophages loaded with phagocytozed glycosphingolipid in Gaucher disease. Macrophages phagocytose decayed blood cells that contain a lot of sphingolipid-rich cell membranes. In Gaucher disease, due to a deficit in beta-glucocerebrosidase activity, the phagocytozed substrate glucocerebroside cannot undergo further catabolism. In such a situation, macrophages secrete chitotriosidase in proportion to the degree of overload. Gaucher disease (GD) is a recessively inherited disorder resulting in storage of glucosylceramide (GlcCer) in lysosomes of tissue macrophages. It is directly caused by the deficiency of beta-glucocerebrosidase (GBA) activity. Chitotriosidase has been measured systematically each year in the same group of 49 patients with type 1 and 3 GD for over 20 years. Our analysis showed that chitotriosidase is very sensitive biomarker to enzyme replacement therapy (ERT). The response to treatment introduction is of an almost immediate nature, lowering pathologically high chitotriosidase levels by a factor of 2 in a time scale of 8 months, on average. Long term enzyme replacement therapy (ERT) brings chitotriosidase activity close to reference values. Finally, reducing the dose of ERT quickly boosts chitotriosidase activity, but restoring the initial dose of treatment brings chitotriosidase level of activity back onto the decreasing time trajectory.

Direct activation of microglia by ß-glucosylceramide causes phagocytosis of neurons that exacerbates Gaucher disease.

Gaucher disease (GD) is the most common lysosomal storage disease caused by recessive mutations in the degrading enzyme of ß-glucosylceramide (ß-GlcCer). However, it remains unclear how ß-GlcCer causes severe neuronopathic symptoms, which are not fully treated by current therapies. We herein found that ß-GlcCer accumulating in GD activated microglia through macrophage-inducible C-type lectin (Mincle) to induce phagocytosis of living neurons, which exacerbated Gaucher symptoms. This process was augmented by tumor necrosis factor (TNF) secreted from activated microglia that sensitized neurons for phagocytosis. This characteristic pathology was also observed in human neuronopathic GD. Blockade of these pathways in mice with a combination of FDA-approved drugs, minocycline (microglia activation inhibitor) and etanercept (TNF blocker), effectively protected neurons and ameliorated neuronopathic symptoms. In this study, we propose that limiting unrestrained microglia activation using drug repurposing provides a quickly applicable therapeutic option for fatal neuronopathic GD.

Venglustat combined with imiglucerase for neurological disease in adults with Gaucher disease type 3: the LEAP trial.

Gaucher disease type 3 is a chronic neuronopathic disorder with wide-ranging effects, including hepatosplenomegaly, anaemia, thrombocytopenia, skeletal disease and diverse neurological manifestations. Biallelic mutations in GBA1 reduce lysosomal acid ß-glucosidase activity, and its substrates, glucosylceramide and glucosylsphingosine, accumulate. Enzyme replacement therapy and substrate reduction therapy ameliorate systemic features of Gaucher disease, but no therapies are approved for neurological manifestations. Venglustat is an investigational, brain-penetrant, glucosylceramide synthase inhibitor with potential to improve the disease by rebalancing influx of glucosylceramide with impaired lysosomal recycling. The Phase 2, open-label LEAP trial (NCT02843035) evaluated orally administered venglustat 15 mg once-daily in combination with maintenance dose of imiglucerase enzyme replacement therapy during 1 year of treatment in 11 adults with Gaucher disease type 3. Primary endpoints were venglustat safety and tolerability and change in concentration of glucosylceramide and glucosylsphingosine in CSF from baseline to Weeks 26 and 52. Secondary endpoints included change in plasma concentrations of glucosylceramide and glucosylsphingosine, venglustat pharmacokinetics in plasma and CSF, neurologic function, infiltrative lung disease and systemic disease parameters. Exploratory endpoints included changes in brain volume assessed with volumetric MRI using tensor-based morphometry, and resting functional MRI analysis of regional brain activity and connectivity between resting state networks. Mean (SD) plasma venglustat AUC0-24 on Day 1 was 851 (282) ng•h/ml; Cmax of 58.1 (26.4) ng/ml was achieved at a median tmax 2.00 h. After once-daily venglustat, plasma concentrations (4 h post-dose) were higher compared with Day 1, indicating ∼2-fold accumulation. One participant (Patient 9) had low-to-undetectable venglustat exposure at Weeks 26 and 52. Based on mean plasma and CSF venglustat concentrations (excluding Patient 9), steady state appeared to be reached on or before Week 4. Mean (SD) venglustat concentration at Week 52 was 114 (65.8) ng/ml in plasma and 6.14 (3.44) ng/ml in CSF. After 1 year of treatment, median (inter-quartile range) glucosylceramide decreased 78% (72, 84) in plasma and 81% (77, 83) in CSF; median (inter-quartile range) glucosylsphingosine decreased 56% (41, 60) in plasma and 70% (46, 76) in CSF. Ataxia improved slightly in nine patients: mean (SD, range) total modified Scale for Assessment and Rating of Ataxia score decreased from 2.68 [1.54 (0.0 to 5.5)] at baseline to 1.55 [1.88 (0.0 to 5.0)] at Week 52 [mean change: -1.14 (95% CI: -2.06 to -0.21)]. Whole brain volume increased slightly in patients with venglustat exposure and biomarker reduction in CSF (306.7 ± 4253.3 mm3) and declined markedly in Patient 9 (-13894.8 mm3). Functional MRI indicated stronger connectivity at Weeks 26 and 52 relative to baseline between a broadly distributed set of brain regions in patients with venglustat exposure and biomarker reduction but not Patient 9, although neurocognition, assessed by Vineland II, deteriorated in all domains over time, which illustrates disease progression despite the intervention. There were no deaths, serious adverse events or discontinuations. In adults with Gaucher disease type 3 receiving imiglucerase, addition of once-daily venglustat showed acceptable safety and tolerability and preliminary evidence of clinical stability with intriguing but intrinsically inconsistent signals in selected biomarkers, which need to be validated and confirmed in future research.

An observational study to investigate the relationship between plasma glucosylsphingosine (lyso-Gb1) concentration and treatment outcomes of patients with Gaucher disease in Japan.

BACKGROUND: Gaucher disease (GD) is an autosomal recessive disease caused by GBA1 mutations resulting in glucosylceramide accumulation in macrophages. GD is characterized by hepatosplenomegaly, anemia, thrombocytopenia, bone complications, and neurological complications. Glucosylsphingosine (lyso-Gb1), a deacylated form of glucosylceramide, has been identified as a promising biomarker for the diagnosis and treatment response in GD. The aim of this study was to examine the relationship between plasma lyso-Gb1 and therapeutic goals for GD (improvements in hepatomegaly, splenomegaly, anemia, thrombocytopenia, bone pain, and bone crisis), as well as disease type and GBA1 mutation type, in Japanese patients with GD receiving velaglucerase alfa, an enzyme replacement therapy (ERT). Furthermore, this study compared the plasma lyso-Gb1 concentration observed in Japanese patients included in this study with that observed in a previous non-Japanese clinical study. RESULTS: This non-interventional, open-label, multicenter observational cohort study (October 2020 to March 2021) included a total of 20 patients (of any age) with GD (type 1: n = 8; type 2: n = 9; type 3: n = 3) treated with velaglucerase alfa for ≥ 3 months. Median (minimum-maximum) duration of velaglucerase alfa treatment was 49.5 (3-107) months. A total of 14 (70.0%) patients achieved all therapeutic goals (i.e., 100% achievement; improvements in hepatomegaly, splenomegaly, anemia, thrombocytopenia, bone pain, and bone crisis). Overall, median (minimum-maximum) lyso-Gb1 concentration was 24.3 (2.1-150) ng/mL. Although not statistically significant, numerically lower plasma lyso-Gb1 concentrations were observed in patients with 100% achievement compared with those without; no statistically significant difference in plasma lyso-Gb1 concentration was observed between patients with different disease type or mutation type. Furthermore, lyso-Gb1 concentrations observed in Japanese patients were numerically lower than that observed in a previous study of non-Japanese patients with GD receiving ERT. CONCLUSIONS: In this study, high achievement rates of therapeutic goals with low lyso-Gb1 concentration were observed, demonstrating a correlation between therapeutic goals and lower plasma lyso-Gb1 concentration in Japanese patients with GD treated with velaglucerase alfa. This study further suggests that plasma lyso-Gb1 concentration may be a useful biomarker for treatment response in patients with GD.

Gaucher Disease: An Unusual Cause of Knee Pain.

INTRODUCTION: Gaucher disease (GD) is a genetic lysosomal disorder leading to storage of the glycolipid molecule glucocerebroside in macrophages, causing multiorgan dysfunction. Bone marrow involvement may result in painful bone crisis and hematologic disturbance. CASE REPORT: We present a case of a 13-year-old adolescent boy with right knee pain. Radiograph and magnetic resonance imaging of the distal femur indicated possible osteomyelitis or bone tumor. However, histologic examination of bone biopsy material suggested the diagnosis of GD, which was confirmed by detection of decreased ß-glucocerebrosidase activity and identification of the exact gene mutation. DISCUSSION: Many visceral and bone abnormalities of GD have been described. The diagnosis of GD is based on clinical and laboratory findings and is established by the measurement of ß-glucocerebrosidase dysfunction and the study of GBA gene mutations. Treatment is currently based on enzyme replacement and substrate reduction. CONCLUSION: This is a rare case of GD presenting initially with knee pain. Because early diagnosis is important for the treatment of this condition, orthopaedic surgeons should consider this uncommon cause in the differential diagnosis of joint pain.

Discovery of Brain-Penetrant Glucosylceramide Synthase Inhibitors with a Novel Pharmacophore.

Inhibition of glucosylceramide synthase (GCS) is a major therapeutic strategy for Gaucher's disease and has been suggested as a potential target for treating Parkinson's disease. Herein, we report the discovery of novel brain-penetrant GCS inhibitors. Assessment of the structure-activity relationship revealed a unique pharmacophore in this series. The lipophilic ortho-substituent of aromatic ring A and the appropriate directionality of aromatic ring B were key for potency. Optimization of the absorption, distribution, metabolism, elimination, toxicity (ADMETox) profile resulted in the discovery of T-036, a potent GCS inhibitor in vivo. Pharmacophore-based scaffold hopping was performed to mitigate safety concerns associated with T-036. The ring opening of T-036 resulted in another potent GCS inhibitor with a lower toxicological risk, T-690, which reduced glucosylceramide in a dose-dependent manner in the plasma and cortex of mice. Finally, we discuss the structural aspects of the compounds that impart a unique inhibition mode and lower the cardiovascular risk.

Effect of Torula Yeast (Candida utilis)-Derived Glucosylceramide on Skin Dryness and Other Skin Conditions in Winter.

Glucosylceramide (GlcCer) is present in foods such as barley, corn, and wheat flour. GlcCer derived from different foods has differences in its physiological effects, depending on the sphingoid backbone and constituent fatty acids. In this study, we investigated the moisturizing and skin conditioning effects of GlcCer derived from torula yeast (Candida utilis) in healthy human subjects. The participants were randomly distributed in a crossover, double-blind comparative manner. Seventeen volunteers were orally administered both 1.8 mg/d of GlcCer derived from torula yeast and a placebo for 4 wk. Before and after oral administration, transepidermal water loss (TEWL) was measured and the objective skin condition observation and a questionnaire on skin condition were conducted. The primary endpoint was TEWL; secondary endpoints included the objective and subjective skin conditions. The change in TEWL over the study period on the forearm was -0.97±0.48 and -1.26±0.46 g/m2•h in the placebo and GlcCer groups, respectively, with significantly lower (p=0.01) TEWL observed in the GlcCer group. Brown spots increased in the placebo group but significantly decreased in the GlcCer group (p=0.04). Although chapped skin worsened in the placebo group, it significantly improved in the GlcCer group (p=0.04). The use of torula yeast-derived GlcCer as a functional cosmeceutical food is a viable option to ameliorate skin conditions, including improvement in skin barrier function, reduction of brown spots, and fixation of chapped skin.

Combined beta-glucosylceramide and ambroxol hydrochloride in patients with Gaucher related Parkinson disease: From clinical observations to drug development.

Both patients with non-neuronopathic Gaucher disease (GD) and heterozygous GBA mutation carrier are at increased risk for Parkinson disease (PD). The risk for PD in these groups does not linearly increase with glucosylceramide (GC) accumulation or with acid ß-glucocerebrosidase (GCase) activity. This observation, together with other clinical systemic observations raises the possibility that extra-cellular GC actually has beneficial, anti-inflammatory, properties. Based on this hypothesis, we suggest here that the administration of supplementary oral GC to GBA carriers at risk for PD may slow inflammatory-driven secondary neuronal death. Such a treatment may act synergistically in GBA carriers once given in combination with an agent that prevent the primary pathologic process that leads to cell death. Ambroxol hydrochloride, a pharmacological chaperone, which reduces endoplasmic reticulum (ER) stress induced by accumulation of mutant misfolded GCase could serve as such an agent. The efficacy of this combined therapy, derived from clinical observations, in vivo and in vitro studies, should be evaluated in clinical trials.

Glucosylceramide attenuates the inflammatory mediator expression in lipopolysaccharide-stimulated RAW264.7 cells.

The positive effect of glucosylceramide (GlcCer) on skin conditions is well known. Recently, there has been increasing interest in the potential antiinflammatory effects of GlcCer due to its efficacy in relieving atopic skin symptoms. However, the role of GlcCer in inflammation has not been investigated completely. Thus, we hypothesized that GlcCer might exhibit the antiinflammatory effects through the inhibition of nuclear factor κB (NF-κB) and mitogen-activated protein kinase (MAPK) signaling. To test this hypothesis, the antiinflammatory effects and signaling mechanisms of GlcCer were investigated in lipopolysaccharide (LPS)-induced RAW 264.7 cells. We report that GlcCer inhibited messenger RNA and protein expression of tissue necrosis factor α and interleukin 1ß without cytotoxicity. However, it did not affect interleukin 6 production in LPS-stimulated RAW 264.7 macrophages. Glucosylceramide also suppressed prostaglandin E2 but not nitric oxide production, consistent with its inhibition of cyclooxygenase 2 but not of inducible nitric oxide synthase expression. The molecular mechanism of GlcCer-mediated inhibition of LPS-induced inflammation in RAW 264.7 cells is closely related to suppression of NF-κB p65 subunit nuclear translocation as well as to phosphorylation of extracellular signal-regulated kinase and, in particular, p38 MAPK. In addition, GlcCer did not affect c-Jun N-terminal kinase phosphorylation. In conclusion, GlcCer inhibits LPS-induced inflammation by blocking the nuclear translocation of NF-κB and inhibiting the phosphorylation of extracellular signal-regulated kinase/p38 MAPK pathways in macrophages, suggesting that it might be a promising potential drug candidate for various inflammatory diseases.

Inhibition of invasion and metastasis of human liver cancer HCCLM3 cells by portulacerebroside A.

CONTEXT: Portulacerebroside A (PCA) is a novel cerebroside compound isolated from Portulaca oleracea L. (Portulacaceae), an edible and medicinal plant distributed in the temperate and tropical zones worldwide. OBJECTIVE: This study investigates the effects of PCA in human liver cancer HCCLM3 cells on metastasis and invasion. MATERIALS AND METHODS: After the cells were treated with PCA (2.5, 5, and 10 µg/ml) for 6, 12, 24, or 48 h, adhesion, transwell invasion, and scratch tests were conducted and cell functions were evaluated. Western blot and FQ-RT-PCR assays explored the mechanism of PCA-inhibited invasion and metastasis in the cells. RESULTS: The adhesion rate of the cells was suppressed at 0.5 h (79.4 ± 1.0, 68.7 ± 1.3, and 58.1 ± 1.3%, versus 100 ± 1.5% in the control), 1 h (78.2 ± 1.2, 70.9 ± 1.6, and 55.4 ± 1.9%, versus 100 ± 1.2% in the control), and 1.5 h (71.6 ± 1.1, 62.3 ± 0.9, and 50.4 ± 0.9%, versus 100 ± 1.1% in the control). The 24 h invasion ability was decreased (356.6 ± 11.2, 204.0 ± 17.6, and 113.0 ± 9.5%, versus 443.6 ± 15.4% in the control). The migration capability was also restrained by PCA for 24 h (324.8 ± 25.4, 250.4 ± 21.0, and 126.3 ± 10.1, versus 381.6 ± 30.6 in the control) and 48 h (470.3 ± 34.3, 404.0 ± 19.7, and 201.0 ± 15.4, versus 752.0 ± 63.6 in the control). There was an increase in the mRNA and protein expression levels of TIMP-2 and nm23-H1, inhibition in the mRNA expression of MTA1, MMP-2, and MMP-9, and suppression in the protein expression of MTA1, RhoA, Rac1/Cdc42, MMP-2, but not RhoC and MMP-9. CONCLUSION: PCA suppresses the invasion and metastasis of HCCLM3 cells possibly by modulation of the mRNA and protein expression of related parameters. This is the first study to reveal a new potential therapeutic application of PCA in antimetastatic therapy for liver cancer.

Orally administered glucosylceramide improves the skin barrier function by upregulating genes associated with the tight junction and cornified envelope formation.

Dietary glucosylceramide improves the skin barrier function. We used a microarray system to analyze the mRNA expression in SDS-treated dorsal skin of the hairless mouse to elucidate the molecular mechanisms involved. The transepidermal water loss of mouse skin was increased by the SDS treatment, this increase being significantly reduced by a prior oral administration of glucosylceramides. The microarray-evaluated mRNA expression ratio showed a statistically significant increase in the expression of genes related to the cornified envelope and tight junction formation when compared with all genes in the glucosylceramide-fed/SDS-treated mouse skin. We then examined the contribution of glucosylceramide metabolites to the tight junction formation of cultured keratinocytes. The SDS treatment of cultured keratinocytes significantly decreased the transepidermal electrical resistance, this decrease being significantly ameliorated in the presence of sphingosine or phytosphingosine, the major metabolites of glucosylceramide. These results suggest that an oral administration of glucosylceramide improved the skin barrier function by up-regulating genes associated with both the cornified envelope and tight junction formation.

Oral glucosylceramide reduces 2,4-dinitrofluorobenzene induced inflammatory response in mice by reducing TNF-alpha levels and leukocyte infiltration.

Sphingolipids are constituents of cellular membranes and play important roles as second messengers mediating cell functions. As significant components in foods, sphingolipids have been proven to be critical for human health. Moreover, diverse metabolic intermediates of sphingolipids are known to play key roles both in proinflammatory and in anti-inflammatory effects. However, the effect of dietary sphingolipids on inflammation is a complicated field that needs to be further assessed. Our study evaluated the effects of orally administered maize glucosylceramide (GluCer), one of the most conventional dietary sphingolipids, on inflammation using the 2,4-dinitro-1-fluorobenzene-treated BALB/c murine model. Oral administration of GluCer inhibited ear swelling and leukocyte infiltration to the inflammatory site, suggesting that dietary GluCer has anti-inflammatory properties. ELISA analyses revealed that oral administration of GluCer for 6 days had not modified the Th1/Th2 balance, but significantly down-regulated the activation of TNF-α at the inflammatory site. Based on these results, the down-regulation of TNF-α by dietary GluCer may suppress vascular permeability and reduce the migration of inflammatory cells. Our findings increase understanding of the actions of dietary sphingolipids on the balance of the immune response.

Velaglucerase alfa, a human recombinant glucocerebrosidase enzyme replacement therapy for type 1 Gaucher disease.

Gaucher disease (GD) is caused by a deficiency of the lysosomal enzyme glucocerebrosidase, which results in the accumulation of its substrate, glucocerebroside, in macrophages. This excess in lipid storage within macrophages (subsequently recognized as Gaucher cells) leads to the development of disease, which presents clinical features including anemia, thrombocytopenia and hepatosplenomegaly, and can also lead to the development of neurological problems or bone disease. Velaglucerase alfa is a gene-activated human recombinant glucocerebrosidase being developed by Shire Human Genetic Therapies Inc as an enzyme replacement therapy for type 1 GD. In vitro, velaglucerase alfa was internalized by human macrophages more rapidly than imiglucerase, which has been the sole standard of care for GD for over 15 years. Clinical trials in patients with GD demonstrated that the safety and efficacy of velaglucerase alfa appeared to be comparable with historical imiglucerase data, although head-to-head data were unavailable. Recent problems with the production of imiglucerase led to the unanticipated introduction of velaglucerase alfa to patients with GD through a pre-approval expanded access protocol. Whether this will prove beneficial, in terms of uptake and prescribing of the enzyme, remains to be seen in a market dominated by imiglucerase.

Beta-glucosylceramide ameliorates liver inflammation in murine autoimmune cholangitis.

We have demonstrated spontaneous development of autoimmune cholangitis, similar to human primary biliary cirrhosis, in mice expressing a dominant negative form of the transforming growth factor-beta receptor (dnTGF-betaRII) restricted to T cells. The autoimmune cholangitis appears to be mediated by autoreactive CD8(+) T lymphocytes that home to the portal tracts and biliary system. Because the liver pathology is primarily secondary to CD8(+) T cells, we have determined herein whether administration of beta-glucosylceramide (GC), a naturally occurring plant glycosphingolipid, alters the natural history of disease in this model. We chose GC because previous work has demonstrated its ability to alter CD8(+) T cell responses and to down-regulate tissue inflammation. Accordingly, dnTGF-betaRII mice were treated with either GC or control for a period of 18 weeks beginning at 6 weeks of age. Importantly, in mice that received GC, there was a significant decrease in the frequency and absolute number of autoreactive liver-infiltrating CD8(+) T cells, accompanied by a significant decrease in activated CD44(high) CD8(+) T cell populations. Further, there was a significant reduction in portal inflammation in GC-treated mice. Interestingly, there were no changes in anti-mitochondrial antibodies, CD4(+) T cells, CD19(+) B cells or natural killer (NK) T cell populations, indicating further that the beneficial effects of GC on liver inflammation were targeted specifically to liver-infiltrating CD8(+) T cells. These data suggest that further work on GC in models of CD8(+) T-mediated inflammation are needed and point to a new therapeutic venue for potentially treating and/or modulating autoimmune disease.

Potential role of NKT regulatory cell ligands for the treatment of immune mediated colitis.

Natural killer T lymphocytes (NKT) have been implicated in the regulation of autoimmune processes in both mice and humans. In response to stimuli, this subset of cells rapidly produces large amounts of cytokines thereby provoking immune responses, including protection against autoimmune diseases. NKT cells are present in all lymphoid compartments, but are most abundant in the liver and bone marrow. They are activated by interaction of their T-cell receptor with glycolipids presented by CD1d, a nonpolymorphic, major histocompatibility complex class I-like molecule expressed by antigen presenting cells. Several possible ligands for NKT cells have recently been suggested. beta-glucosylceramide, a naturally occurring glycolipid, is a metabolic intermediate in the anabolic and catabolic pathways of complex glycosphingolipids. Like other beta-glycolipids, beta-glucosylceramide has an immunomodulatory effect in several immune mediated disorders, including immune mediated colitis. Due to the broad impact that NKT cells have on the immune system, there is intense interest in understanding how NKT cells are stimulated and the extent to which NKT cell responses can be controlled. These novel ligands are currently being evaluated in animal models of colitis. Here, we discuss strategies to alter NKT lymphocyte function in various settings and the potential clinical applications of natural glycolipids.

Glucocerebroside ameliorates the metabolic syndrome in OB/OB mice.

Glucocerebroside (GC) is a naturally occurring glycolipid that may alter natural killer T (NKT) cell function. To determine the effect of GC on the metabolic derangements and immune profile in leptin-deficient mice, Ob/Ob mice were treated by daily injections of GC for 8 weeks and followed for various metabolic and immunological parameters. Marked amelioration of the metabolic alterations characteristic of leptin-deficient mice was observed in GC-treated animals compared with controls. A significant decrease in liver size and hepatic fat content were observed in GC-treated mice. Near-normalization of glucose tolerance and decreased serum triglyceride levels were observed. Fluorescence-activated cell sorting analysis of peripheral and intrahepatic lymphocytes revealed a 1.6-fold increase of the peripheral/intrahepatic NKT lymphocyte ratio. A 33% decrease of serum interferon-gamma level and a 2.6-fold increase of serum interleukin 10 level were noted in GC-treated mice. Immune modulation by GC may have a role in the treatment of nonalcoholic steatohepatitis and other immune-mediated disorders.

Ceramide-beta-D-glucuronide: synthesis, digestion, and suppression of early markers of colon carcinogenesis.

Dietary sphingolipids inhibit chemically induced colon cancer in mice. The most likely mediators of this effect are the metabolites ceramide (Cer) and sphingosine, which induce growth arrest and apoptosis in transformed cells. Sphingolipids are digested in both the upper and the lower intestine; therefore, a more colon-specific method of delivery of sphingolipids might be useful. A Cer analogue with a D-glucuronic acid attached at the primary hydroxyl of N-palmitoyl-D-sphingosine (Cer-beta-glucuronide) was synthesized and evaluated as a substrate for Escherichia coli beta-glucuronidase and colonic digestion, as well as for suppression of early events in colon carcinogenesis in CFI mice treated with 1,2-dimethylhydrazine. Purified beta-glucuronidase (EC 3.2.1.31) and colonic segments (as a source of colonic enzymes and microflora) hydrolyzed Cer-beta-glucuronide to release Cer, as analyzed by tandem mass spectrometry. More than 75% of the Cer-beta-glucuronide was cleaved in an 8-h incubation with the colonic segments. When Cer-beta-glucuronide was administered for 4 weeks as 0.025% and 0.1% of the diet (AIN 76A) to 1,2-dimethylhydrazine-treated mice, there were significant reductions in colonic cell proliferation, as determined by in vivo BrdUrd incorporation, and in the appearance of aberrant crypt foci. The effect of dietary Cer-beta-glucuronide on aberrant crypt foci correlated significantly with the length of the colon, which suggests that Cer-beta-glucuronide was most effective when there was a larger compartment for digestion. Thus, synthetic sphingolipids that target the colon for the release of the bioactive backbones offer a promising approach to colon cancer prevention.