Red Ginseng Improves D-galactose-Induced Premature Ovarian Failure in Mice Based on Network Pharmacology.

In this study, we evaluated the ameliorative effect and molecular mechanism of red ginseng (Panax ginseng C.A. Meyer) extract (RGE) on D-galactose (D-gal)-induced premature ovarian failure (POF) using network pharmacology analysis. Ginsenosides are important active ingredients in ginseng, which also contains some sugar and amino acid derivatives. We aimed to determine the key proteins through which RGE regulates POF. In this work, we retrieved and screened for active ingredients in ginseng and the corresponding POF disease targets in multiple databases. A PPI network of genes was constructed in the STRING database and core targets were screened using topological analysis. Gene ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses were conducted in R software. Finally, molecular docking was conducted to validate the results. Female ICR mice were used to establish a POF mouse model for in vivo experiments. Serum levels of relevant estrogens were determined using ELISA and expression levels of relevant proteins in ovarian tissues were detected using immunofluorescence and western blot analysis. Network pharmacology analysis predicted that PI3K, Akt, Bax, Bcl-2, p16, and other proteins were highly correlated with POF and RGE. The results clearly showed that RGE could increase estradiol (E2) and lower follicle-stimulating hormone (FSH) levels in D-gal-fed mice. RGE restored the expression levels of related proteins by reducing Nrf2-mediated oxidative stress, PI3K/Akt-mediated apoptosis, and senescence signaling pathways. Overall, RGE has the potential to prevent and treat POF and is likely to be a promising natural protector of the ovaries.

Specific immune biomarker monitoring in two children with severe IgA nephropathy and successful therapy with immunoadsorption in a rapidly progressive case.

BACKGROUND: Childhood IgA nephropathy (cIgAN) is one of the most common primary glomerulonephritides with the potential to evolve to kidney failure. IgAN is an autoimmune disease involving 3 key factors: galactose-deficient IgA1 (Gd-IgA1), anti-IgA1 autoantibodies, and soluble (s)CD89 IgA Fc receptor. These molecules and immune complexes have been described recently as potential biomarkers of disease progression in childhood IgAN but their evolution in time under immunosuppressive treatment remains unknown. METHODS: We performed a prospective study of two proliferative cIgAN patients by sequentially biomonitoring immune IgA complexes (sCD89-IgA, IgG-IgA), sCD89, and Gd-IgA1 and correlating them with clinical and histological outcome after treatment. RESULTS: After patient 1's treatment, a decrease in sCD89-IgA, IgG-IgA, and free sCD89 was linked to a decrease in proteinuria whereas eGFR (estimated glomerular filtration rate) and Gd-IgA1 levels remained stable. Patient 1 received tacrolimus and monthly intramuscular steroid injections of Kenacort for 10 months. At the end, a relapse induced an increase in proteinuria consistent with an increase of the 3 biomarkers. Patient 2 displayed rapidly progressive IgAN with crescents in more than 90% of glomeruli and received intense immunosuppression treatment associated with the immunoadsorption (IA) approach. During IA, proteinuria decreased rapidly, as well as levels of CD89-IgA, IgG-IgA, sCD89, and Gd-IgA1 biomarkers. After discontinuation of IA, proteinuria increased as well as IgG-IgA complexes whereas sCD89-IgA and sCD89 remained low. Further re-intensification of IA and addition of cyclophosphamide improved proteinuria again with reduced IgG-IgA. A second biopsy was performed showing a reduction of extracapillary proliferation to 6% of glomeruli and only 9% glomerulsoclerosis. CONCLUSIONS: In conclusion, sequential biomonitoring of Gd-IgA1, IgA-immune complexes, and sCD89 in cIgAN was found to be valuable, by correlating with clinical features and glomerular proliferative lesions in cIgAN. These biomarkers could represent useful tools to evaluate kidney injury without repeat kidney biopsies.

Consumption of Galacto-Oligosaccharides Increases Iron Absorption from Ferrous Fumarate: A Stable Iron Isotope Study in Iron-Depleted Young Women.

BACKGROUND: Animal studies suggest prebiotics can increase iron absorption, but results from human studies are equivocal. OBJECTIVES: In iron-depleted women, before (baseline) and after daily consumption of galacto-oligosaccharides (GOS) for 4 wk, we sought to assess fractional iron absorption (FIA) from an iron supplement given with and without single doses of GOS in test meals or water. METHODS: In all women (n = 34; median serum ferritin concentration = 16.4 µg/L), FIA from doses of 14 mg iron labeled with stable isotopes was measured in the following conditions at baseline: 1) FIA from ferrous fumarate (FeFum) in water given with and without 15 g GOS; 2) FIA from FeFum in a test meal given with and without 15 g GOS; 3) FIA from ferrous sulfate (FeSO4) in a test meal given without 15 g GOS. All subjects then consumed âˆ¼15 g GOS daily for 4 wk. Then the following conditions were tested: 4) FIA from FeFum in a test meal with and without 15 g GOS; and 5) FIA from FeSO4 in a test meal with 15 g GOS. FIA was measured as erythrocyte incorporation of stable isotopes. RESULTS: At baseline, GOS significantly increased FIA from FeFum when given with water (+61%; P < 0.001) and the meal (+28%; P = 0.002). After 4 wk of GOS consumption, GOS again significantly increased FIA from FeFum in the meal (+29%; P = 0.044). However, compared with baseline, consumption of GOS for 4 wk did not significantly enhance absorption from FeFum in the meal given without GOS. FIA from FeSO4 given with GOS in a meal after 4 wk of GOS consumption was not significantly greater than FIA from FeSO4 in a meal without GOS at baseline. CONCLUSIONS: In iron-depleted women, GOS given with FeFum increases FIA, but 4 wk of GOS consumption did not enhance this effect. The study was registered at clinicaltrials.gov as NCT03325270.

Galectin Targeted Therapy in Oncology: Current Knowledge and Perspectives.

The incidence and mortality of cancer have increased over the past decades. Significant progress has been made in understanding the underpinnings of this disease and developing therapies. Despite this, cancer still remains a major therapeutic challenge. Current therapeutic research has targeted several aspects of the disease such as cancer development, growth, angiogenesis and metastases. Many molecular and cellular mechanisms remain unknown and current therapies have so far failed to meet their intended potential. Recent studies show that glycans, especially oligosaccharide chains, may play a role in carcinogenesis as recognition patterns for galectins. Galectins are members of the lectin family, which show high affinity for ß-galactosides. The galectin-glycan conjugate plays a fundamental role in metastasis, angiogenesis, tumor immunity, proliferation and apoptosis. Galectins' action is mediated by a structure containing at least one carbohydrate recognition domain (CRD). The potential prognostic value of galectins has been described in several neoplasms and helps clinicians predict disease outcome and determine therapeutic interventions. Currently, new therapeutic strategies involve the use of inhibitors such as competitive carbohydrates, small non-carbohydrate binding molecules and antibodies. This review outlines our current knowledge regarding the mechanism of action and potential therapy implications of galectins in cancer.

Hyperinsulinemic hypoglycemia: clinical, molecular and therapeutical novelties.

Hyperinsulinemic hypoglycemia (HI) is the most common cause of hypoglycemia in children. Impairment of cellular pathways involved in insulin secretion from pancreatic ß-cells, broadly classified as channelopathies and metabolopathies, have been discovered in the past two decades. The increasing use of NGS target panels, combined with clinical, biochemical and imaging findings allows differentiating the diagnostic management of children with focal forms, surgically curable, from those with diffuse forms, more conservatively treated with pharmacological and nutritional interventions. Specific approaches according to the subtype of HI have been established and novel therapies are currently under investigation. Despite diagnostic and therapeutic advances, HI remains an important cause of morbidity in children, still accounting for 26-44% of permanent intellectual disabilities, especially in neonatal-onset patients. Initial insult from recurrent hypoglycemia in early life greatly contributes to the poor outcomes. Therefore, patients need to be rapidly identified and treated aggressively, and require at follow-up a complex and regular monitoring, managed by a multidisciplinary HI team. This review gives an overview on the more recent diagnostic and therapeutic tools, on the novel drug and nutritional therapies, and on the long-term neurological outcomes.

Galactose conjugated platinum(II) complex targeting the Warburg effect for treatment of non-small cell lung cancer and colon cancer.

Malignant neoplasms exhibit a higher rate of glycolysis than normal cells; this is known as the Warburg effect. To target it, a galactose-conjugated (trans-R,R-cyclohexane-1,2-diamine)-2-chloromalonato-platinum(II) complex (Gal-Pt) was designed, synthesized, and evaluated in five human cancer cell lines and against two different xenograft tumour models. Gal-Pt exhibits much higher aqueous solubility (over 25 times) and improved cytotoxicity than oxaliplatin, especially in human colon (HT29) and lung (H460) cancer cell lines. The safety profile of Gal-Pt was investigated in vivo by exploring the maximum tolerated dose (MTD) and animal mortality rate. The ratios of the animal lethal dosage values to the cytotoxicity in HT29 (LD50/IC50) showed that Gal-Pt was associated with an increased therapeutic index by over 30-fold compared to cisplatin and oxaliplatin. We evaluated in vivo antitumor activity by single agent intravenous treatment comparison studies of Gal-Pt (50 mg/kg as 65% MTD) and cisplatin (3 mg/kg, as 80% MTD) in a H460 lung cancer xenograft model, and with oxaliplatin (7 mg/kg, as 90% MTD) in a HT29 colon cancer xenograft model. The results show that Gal-Pt was more efficacious against H460 than cisplatin, and had superior potency in HT29 cells compared to oxaliplatin under nontoxic dosage conditions. The dependency between cytotoxicity of Gal-Pt and glucose transporters (GLUTs) was investigated by using quercetin as an inhibitor of GLUTs in HT29 cells. The cytotoxic potency of Gal-Pt was highly reduced by the inhibitor, suggesting that the uptake of Gal-Pt was regulated by glucose transporters. The GLUT mediated transportability and cellular uptake of Gal-Pt was also demonstrated using a fluorescent glucose bioprobe in HT29 competition assay.

Synthesis of novel galactose functionalized gold nanoparticles and its radiosensitizing mechanism.

BACKGROUND: Biocompatible gold nanoparticles (GNPs) are potentially practical and efficient agents in cancer radiotherapy applications. In this study, we demonstrated that GNPs can significantly modulate irradiation response of hepatocellular carcinoma cells in vitro and investigated the underlying mechanisms. We co-grafted galactose (GAL) targeting hepatocyte specific asialoglycoprotein receptor and Polyethylene Glycol (PEG) onto GNPs surfaces to increase GNPs targeting specificity and stability. RESULTS: This novel GAL-PEG-GNPs and bare GNPs show similar appearance and cytotoxicity profiles, while more GAL-PEG-GNPs can be effectively uptaken and could enhance cancer cell killing. CONCLUSION: GAL-PEG-GNPs have better radiosensitization to HepG2. The sensitization mechanism of GAL-PEG-GNPs is related to the apoptotic gene process activated by generation of a large amount of free radicals induced by GNPs.

A CE-Marked Drug Used for Localized Adiposity Reduction: A 4-Year Experience.

BACKGROUND: Injectable fat-reducing therapies are not an alternative to liposuction. Rather, they may be best suited for patients who are unwilling or unable to undergo surgical reduction of small collections of fat, and for patients who desire touchups for liposuction-induced irregularities. OBJECTIVES: The authors report their 4-year experience with a novel injectable CE-marked drug, used in an off-label manner. METHODS: Between October 2009 and November 2013, 186 patients were treated by injection of an adipocitolytic solution in 1 of 4 private Italian aesthetic facilities, by 1 of 4 independent physicians. Treated areas included the neck, hips/saddlebags, abdomen/love handles, inner thighs, and buffalo hump. Complications and side effects were documented. RESULTS: All patients experienced mild to moderate swelling and reddening of the skin, which resolved 3 to 5 days after injection. No major complications or side effects occurred, such as necrosis. Rates of transient events were as follows: hematoma, 1.61%; paresthesia, 1.07%; and ecchymosis, 6.45%. Pruritus was reported by 21.5% of patients, which began 3 to 7 days following injection. Subcutaneous nodules were noted in 1.61% and resolved within 4 months of injection. A transitory "unusual sensation" was reported by 12.9% of patients, which lasted up to 2 months after final injection. CONCLUSIONS: Results demonstrate that this CE-marked agent appears to be effective and safe for medical treatment of fat reduction.

Efficacy of galactose and adalimumab in patients with resistant focal segmental glomerulosclerosis: report of the font clinical trial group.

BACKGROUND: Patients with resistant focal segmental glomerulosclerosis (FSGS) who are unresponsive to corticosteroids and other immunosuppressive agents are at very high risk of progression to end stage kidney disease. In the absence of curative treatment, current therapy centers on renoprotective interventions that reduce proteinuria and fibrosis. The FONT (Novel Therapies for Resistant FSGS) Phase II clinical trial (NCT00814255, Registration date December 22, 2008) was designed to assess the efficacy of adalimumab and galactose compared to standard medical therapy which was comprised of lisinopril, losartan, and atorvastatin. METHODS: Key eligibility criteria were biopsy confirmed primary FSGS or documentation of a causative genetic mutation, urine protein:creatinine ratio >1.0 g/g, and estimated glomerular filtration rate (eGFR) >40 ml/min/1.73 m(2). The experimental treatments - adalimumab, galactose, standard medical therapy-- were administered for 26 weeks. The primary endpoint was a 50 % reduction in proteinuria with stable eGFR. RESULTS: Thirty-two subjects were screened and 21 were assigned to one of the three study arms. While none of the adalimumab-treated subjects achieved the primary outcome, 2 subjects in the galactose and 2 in the standard medical therapy arm had a 50 % reduction in proteinuria without a decline in eGFR. The proteinuria response did not correlate with serial changes in the serum glomerular permeability activity measured by the Palb assay or soluble urokinase plasminogen activator receptor (suPAR). There were no serious adverse effects related to treatments in the study. CONCLUSIONS: Recruitment into this trial that addressed patients with resistant FSGS fell short of the enrollment goal. Our findings suggest that future studies of novel therapies for rare glomerular diseases such as FSGS may benefit from enrollment of patients earlier in the course of their disease. In addition, better identification of patients who are likely to respond to a new treatment based on biomarkers suggesting involvement of the disease pathway targeted by the experimental agent may reduce the required sample size and increase the likelihood of a favorable outcome.

Galactose therapy reduces proteinuria in patients with recurrent focal segmental glomerulosclerosis after kidney transplantation.

Primary focal segmental glomerulosclerosis is an important cause of end-stage kidney disease with a high rate of recurrent disease after kidney transplantation. Current therapy achieves remission in only half of patients. Recent interest has focused on the potential role of galactose in binding and inactivating the putative circulating permeability factor, supported by in vitro and clinical case report studies. Orally active and without major adverse effects, galactose has a favourable treatment profile compared with current immunosuppressive treatment options. We describe our experience using galactose therapy in two patients with recurrent focal segmental glomerulosclerosis after renal transplantation. Galactose was associated with symptomatic improvement and stabilization of graft function in one case; the other case was complicated by concurrent malignancy. In both cases, we observed a marked reduction in proteinuria with galactose treatment.

Inhibition of fucosylation reshapes inflammatory macrophages and suppresses type II collagen-induced arthritis.

OBJECTIVE: Fucosylation catalyzed by fucosyltransferases (FUTs) is an important posttranslational modification involved in a variety of biologic processes. This study was undertaken to determine the roles of fucosylation in rheumatoid arthritis (RA) and to assess the efficacy of reestablishing immune homeostasis with the use of 2-deoxy-d-galactose (2-d-gal), a fucosylation inhibitor. METHODS: Quantitative polymerase chain reaction was performed to determine the expression of FUT genes in synovial tissue from RA and osteoarthritis (OA) patients and in fluorescence-activated cell-sorted cells from RA synovial fluid. The in vivo inhibitory effect of 2-d-gal was evaluated in a murine collagen-induced arthritis (CIA) model. The in vitro effects of 2-d-gal on differentiation of inflammatory macrophages, production of cytokines, and antigen uptake, processing, and presentation functions were analyzed. RESULTS: FUTs that are involved in terminal or subterminal fucosylation, but not those involved in core fucosylation or O-fucosylation, were up-regulated in RA compared to OA synovial tissue. The expression of terminal FUTs was highly positively correlated with the expression of TNF (encoding for tumor necrosis factor α). Terminal FUTs were predominantly expressed in M1 macrophages. In vivo, 2-d-gal treatment of mice precluded the development of CIA by reducing inflammatory macrophages and Th17 cells in the draining lymph nodes and decreasing the levels of TNFα, interleukin-6 (IL-6), and antibodies to type II collagen in the serum. In vitro, treatment with 2-d-gal skewed the differentiation of M1 macrophages to IL-10-producing M2 macrophages. Furthermore, 2-d-gal significantly inhibited the antigen-presenting function of M1 macrophages. CONCLUSION: Terminal fucosylation is a novel hallmark of inflammatory macrophages. Inhibition of terminal FUTs reshapes the differentiation and functions of M1 macrophages, leading to resolution of inflammation in arthritis.

Multiple phenotypes in phosphoglucomutase 1 deficiency.

BACKGROUND: Congenital disorders of glycosylation are genetic syndromes that result in impaired glycoprotein production. We evaluated patients who had a novel recessive disorder of glycosylation, with a range of clinical manifestations that included hepatopathy, bifid uvula, malignant hyperthermia, hypogonadotropic hypogonadism, growth retardation, hypoglycemia, myopathy, dilated cardiomyopathy, and cardiac arrest. METHODS: Homozygosity mapping followed by whole-exome sequencing was used to identify a mutation in the gene for phosphoglucomutase 1 (PGM1) in two siblings. Sequencing identified additional mutations in 15 other families. Phosphoglucomutase 1 enzyme activity was assayed on cell extracts. Analyses of glycosylation efficiency and quantitative studies of sugar metabolites were performed. Galactose supplementation in fibroblast cultures and dietary supplementation in the patients were studied to determine the effect on glycosylation. RESULTS: Phosphoglucomutase 1 enzyme activity was markedly diminished in all patients. Mass spectrometry of transferrin showed a loss of complete N-glycans and the presence of truncated glycans lacking galactose. Fibroblasts supplemented with galactose showed restoration of protein glycosylation and no evidence of glycogen accumulation. Dietary supplementation with galactose in six patients resulted in changes suggestive of clinical improvement. A new screening test showed good discrimination between patients and controls. CONCLUSIONS: Phosphoglucomutase 1 deficiency, previously identified as a glycogenosis, is also a congenital disorder of glycosylation. Supplementation with galactose leads to biochemical improvement in indexes of glycosylation in cells and patients, and supplementation with complex carbohydrates stabilizes blood glucose. A new screening test has been developed but has not yet been validated. (Funded by the Netherlands Organization for Scientific Research and others.).

Childhood nephrotic syndrome--current and future therapies.

The introduction of corticosteroids more than 50 years ago dramatically improved the prognosis of children with nephrotic syndrome. Corticosteroids remain the standard initial treatment for children with this disease, but a considerable proportion of patients do not respond and are therefore at risk of progressing to end-stage renal disease. Because of this risk, new therapeutic strategies are needed for steroid-resistant nephrotic syndrome. These strategies have historically focused on identifying effective alternative immunosuppressive agents, such as ciclosporin and tacrolimus, yet evidence now indicates that nephrotic syndrome results from podocyte dysfunction. Even conventional immunosuppressive agents, such as glucocorticoids and ciclosporin, directly affect podocyte structure and function, challenging the 'immune theory' of the pathogenesis of childhood nephrotic syndrome in which disease is caused by T cells. This Review summarizes the currently available treatments for childhood nephrotic syndrome, and discusses selected novel pathways in podocytes that could be targeted for the development of next-generation treatments for children with this syndrome.

Partial remission of resistant nephrotic syndrome after oral galactose therapy.

Focal segmental glomerulosclerosis is sometimes associated with a circulating permeability factor. It was proposed that this factor interacts with the sugars of the glycocalyx, and its high affinity for galactose was shown on the basis of chromatographic studies. Galactose inactivates it and seems to lead to its clearance from plasma. A toddler with a nephrotic syndrome resistant to corticosteroids was admitted. A renal biopsy revealed minimal change disease with deposition of immunoglobulin M. Immunosuppressive therapy with pulses of cyclophosphamide, low-dose combination immunosuppressive therapy, and later with mycophenolate mofetil failed to induce remission. A renal biopsy six years later showed transformation to FSGS. After unsuccessful treatment with monthly pulses of cyclophosphamide, we began therapy with tacrolimus, which showed no effect. After two months, we added oral galactose to tacrolimus for one month, after which proteinuria decreased by 50%. Seven months later, galactose was again added for six months, after which proteinuria remained below 2 g/24 h and the plasma albumin and cholesterol concentrations normalized. An adolescent girl with a nephrotic syndrome resistant to corticosteroids was admitted. A renal biopsy revealed mesangioproliferative glomerulonephritis with C1q nephropathy. Therapy with tacrolimus failed to induce remission. After six months, we added galactose for three months, which reduced proteinuria to 0.76 g/24 h. After the discontinuation of galactose therapy, proteinuria increased to 2.48 g/24 h, despite further treatment with tacrolimus. It seems that oral galactose at a dose of 0.2 g/kg twice a day could be a promising new and nontoxic therapy for the treatment of resistant nephrotic syndrome.

Oral administration of a galactooligosaccharide preparation inhibits development of atopic dermatitis-like skin lesions in NC/Nga mice.

Anti-allergic effects of galactooligosaccharide (GOS), which is found in breast milk and frequently added to food for promoting health, were evaluated in a human-like mouse model of atopic dermatitis (AD). NC/Nga mice were fed 5.5% GOS for 8 weeks, and we examined whether this treatment suppressed the development of AD-like skin lesions in these mice. Mice fed GOS exhibited significantly less symptoms of dermatitis, reduced scratching frequency, and lower levels of serum total immunoglobulin E compared to control. At the end of the 8-week-experimental period, spleens were removed, and the splenocytes were stimulated with phorbol 12-myristate 13-acetate and ionomycin, following which production of cytokines and a chemokine was analyzed. Elevated levels of Th1 cytokines such as interferon-gamma were observed in splenocytes from GOS-fed mice. However, the levels of Th2 cytokines such as interleukin (IL)-13 were unchanged. Furthermore, GOS inhibited the production of inflammatory cytokines such as IL-1beta, IL-6, IL-17, and tumor necrosis factor-alpha but enhanced production of immunomodulatory IL-10. The results indicate that GOS effectively blocked AD-like skin lesions in the mice by at least partly inducing production of IL-10 and suppressing the production of cytokines such as IL-17, which are involved in skin inflammation.

Inhalation with fucose and galactose for treatment of Pseudomonas aeruginosa in cystic fibrosis patients.

BACKGROUND: Colonisation of cystic fibrosis (CF) lungs with Pseudomonas aeruginosa is facilitated by two lectins, which bind to the sugar coat of the surface lining epithelia and stop the cilia beating. OBJECTIVES: We hypothesized that P. aeruginosa lung infection should be cleared by inhalation of fucose and galactose, which compete for the sugar binding site of the two lectins and thus inhibit the binding of P. aeruginosa. METHODS: 11 adult CF patients with chronic infection with P. aeruginosa were treated twice daily with inhalation of a fucose/galactose solution for 21 days (4 patients only received inhalation, 7 patients received inhalation and intravenous antibiotics). Microbial counts of P. aeruginosa, lung function measurements, and inflammatory markers were determined before and after treatment. RESULTS: The sugar inhalation was well tolerated and no adverse side effects were observed. Inhalation alone as well as combined therapy (inhalation and antibiotics) significantly decreased P. aeruginosa in sputum (P < 0.05). Both therapies also significantly reduced TNFalpha expression in sputum and peripheral blood cells (P < 0.05). No change in lung function measurements was observed. CONCLUSIONS: Inhalation of simple sugars is a safe and effective measure to reduce the P. aeruginosa counts in CF patients. This may provide an alternative therapeutical approach to treat infection with P. aeruginosa.

A glycosylated nitric oxide donor, beta-Gal-NONOate, and its site-specific antitumor activity.

So far, nitric oxide (NO) donors have been applied to various aspects of antitumor therapy. To selectively sensitize tumor cells and avoid unwanted side effects, we recently synthesized a beta-galactosidase-activatable NO-releasing compound, beta-galactosyl-pyrrolidinyl diazeniumdiolate (beta-Gal-NONOate). In this study, we first verified its superiority over its parent diazeniumdiolate (NONOate) in terms of targeted intracellular NO-releasing and antitumor activity with 9L/LacZ cells (rat glioma cell line 9L with transformed LacZ gene) in vitro. beta-Gal-NONOate only released NO when hydrolyzed by induced beta-galactosidase in 9L/LacZ cells, which led to its more powerful cytotoxicity than that of NONOate. The results showed that beta-Gal-NONOate produced higher NO levels than NONOate in 9L/LacZ cells at equal concentration, and hence induced optimal NO levels for antitumor activity. However, in 9L cells, beta-Gal-NONOate showed less toxicity than NONOate. Therefore, it is demonstrated that beta-Gal-NONOate is a site-specific prodrug for targeting NO intracellularly as a beta-galactosidase-sensitive NO donor, and it is also expected to be a promising probe in numerous experimental settings and a potential therapeutic drug for antitumor treatment.

The heart in Anderson Fabry disease.

Anderson Fabry disease is a life threatening, X-linked inborn metabolic defect of the lysosomal enzyme áalpha-galactosidase A. The deficiency of alpha-galactosidase A leads to a progressive accumulation of globotriaosylceramide (Gb(3)), the major glycosphingolipid substrate of the enzyme, within vulnerable cells, tissues, and organs, including the cardiovascular system. Cardiac involvement is frequent and patients with cardiac affection develop progressive hypertrophic infiltrative cardiomyopathy, valvular abnormalities, arrhythmias, and conduction abnormalities and may develop coronary heart disease. Hemizygous male patients have no detectable alpha-galactosidase A activity, while affected heterozygous females may have normal level of alpha-galactosidase A activity. Death occurs in male patients at 45 to 50 years, about 15 to 20 years earlier than in female patients due to a vicious circle from chronic renal insufficiency, arterial hypertension, atherosclerotic lesions and cerebrovascular hemorrhage or insults, and cardiomyopathy. Cardiac involvement in hetero- and hemizygotes will be discussed as well as the influence of enzyme replacement of alpha-galactosidase A.

Enzyme-based therapy for autism spectrum disorders -- is it worth another look?

Autism is a developmental disease usually manifesting within the first three years of life. To date, no causative agent has been found. Similarly, treatment options have been limited. Of the treatment options available, a number of them have been nutritionally based in an attempt to address one or more of the theories regarding the etiology of the disease. An example would be enzyme therapy for the digestion of purported offending neuroactive peptides collectively known as exorphins. This paper discusses the exorphin theory of autism and subsequent treatment with dietary enzyme therapy. Novel data are presented in support of the theory that enzymes play a critical role in autism. Forty-six patients between the ages of 5 and 31 were selected for inclusion in the study based on a diagnosis placing them in the category of the autism spectrum disorders (ASD). The diets were supplemented with a novel dietary enzyme formulation, ENZYMAID, for a period of 12 weeks. Progress was tracked according to the Symptom Outcome Survey (SOS) (1) form method of symptom charting and presented in a table for further analysis. The novel enzyme formula, ENZYMAID, beneficially and safely affected all 13 of the parameters measured. Improvements ranged from 50-90%, depending on the parameter measured. Enzyme therapy to treat ASD may indeed a viable option in treatment protocols. These results indicate that further controlled studies are warranted.

Depletion of anti-gal antibodies in baboons by intravenous therapy with bovine serum albumin conjugated to gal oligosaccharides.

BACKGROUND: Anti-Galalpha 1-3Gal (Gal) antibodies (Ab) play a key role in the rejection of pig cells or organs transplanted into primates. A course of extracorporeal immunoadsorption (EIA) of anti-Gal Ab using an immunoaffinity column of a Gal type 6 oligosaccharide depletes Ab successfully, but Ab returns during the next few days. Although therapy with an anti-CD154 monoclonal antibody (mAb) prevents an induced Ab response to Gal or non-Gal epitopes, T cell-independent natural anti-Gal IgM and IgG return to baseline (pretransplant) levels. We have investigated the capacity of continuous i.v. infusion of bovine serum albumin conjugated to Gal type 6 oligosaccharide (BSA-Gal) to deplete or maintain depletion of circulating anti-Gal Ab. METHODS: Porcine peripheral blood mobilized progenitor cells (PBPC) obtained by leukapheresis from MHC-inbred miniature swine (n=6) were transplanted into baboons. Group 1 baboons (n=4) underwent whole body (300 cGy) and thymic (700 cGy) irradiation, T cell depletion with antithymocyte globulin, complement depletion with cobra venom factor, short courses of anti-CD154 mAb therapy (20 mg/kg i.v. on alternate days), cyclosporine (CyA) (in two baboons only), mycophenolate mofetil, and porcine hematopoietic growth factors. Anti-Gal Ab depletion by EIA was carried out before transplantation of high doses (2-4x 1010 cells/kg) of PBPC. Group 2 baboons (n=3) received the group 1 regimen (including CyA) plus a continuous i.v. infusion of BSA-Gal. To prevent sensitization to BSA, anti-CD154 mAb therapy was continued until BSA-Gal administration was discontinued. RESULTS: In group 1, Gal-reactive Ab returned to pre-PBPC transplant levels within 15-21 days, but no induced Ab to Gal or non-Gal determinants developed while anti-CD154 mAb therapy was being administered. In group 2, anti-Gal Ab was either not measurable or minimally measurable while BSA-Gal was being administered. After discontinuation of BSA-Gal, Ab did not return to pre-PBPC transplant level for more than 40-60 days, and no sensitization developed even when all therapy was discontinued. In one baboon, however, Ab to Gal type 2, but not type 6, returned during BSA-Gal therapy. CONCLUSIONS: Prevention of the induced humoral response to Gal and non-Gal epitopes by anti-CD154 mAb therapy has been reported previously by our group, but our studies are the first to demonstrate a therapy that resulted in an absence of natural anti-Gal Ab for a prolonged period. The combination of BSA-Gal and T cell costimulatory blockade may facilitate survival of pig cells and organs transplanted into primates. The return in one baboon of Ab reactive with the Gal type 2 oligosaccharide, but not type 6, indicates some polymorphism of anti-Gal Ab and suggests that, to be effective in all cases, the infusion of a combination of type 6 and type 2 BSA-Gal may be required.