A novel, highly sensitive and specific biomarker for Niemann-Pick type C1 disease

Background Lysosomal storage disorders (LSDs), are a heterogeneous group of rare disorders caused by defects in genes encoding for proteins involved in the lysosomal degradation of macromolecules. They occur at a frequency of about 1 in 5,000 live births, though recent neonatal screening suggests a higher incidence. New treatment options for LSDs demand a rapid, early diagnosis of LSDs if maximal clinical benefit is to be achieved. Methods Here, we describe a novel, highly specific and sensitive biomarker for Niemann-Pick Type C disease type 1 (NPC1), lyso-sphingomyelin-509. We cross-validate this biomarker with cholestane-3β,5α,6β-triol and relative lysosomal volume. The primary cohort for establishment of the biomarker contained 135 NPC1 patients, 66 NPC1 carriers, 241 patients with other LSDs and 46 healthy controls. Results With a sensitivity of 100.0% and specificity of 91.0% a cut-off of 1.4 ng/ml was established. Comparison with cholestane-3β,5α,6β-triol and relative acidic compartment volume measurements were carried out with a subset of 125 subjects. Both cholestane-3β,5α,6β-triol and lyso-Sphingomyelin-509 were sufficient in establishing the diagnosis of NPC1 and correlated with disease severity. Conclusion In summary, we have established a new biomarker for the diagnosis of NPC1, and further studies will be conducted to assess correlation to disease progress and monitoring treatment

Asymmetrische Totalsynthese von 16(S)-Iloprost und 3-Oxa-16(S)-Iloprost mittels der Azoen-Strategie

The present dissertation describes completely stereocontrolled asymmetric syntheses of 16S-iloprost and of 16S-3-oxa-iloprost by a azoene strategy. As Ilomedin and Ventavis registrated iloprost (1:1 mixture of diastereomers at C16) serves as a drug against ischemic heart disease, peripheral vascular disease and primary pulmonary hypertension. The 16S-diastereomer is 5 to 20 times more potent than the 16R-diastereomer in inhibiting collagen-induced platelet aggregation. 3-oxa-16S-iloprost should be more stable against beta-oxidation of the alpha side chain. A bicyclic azoene was prepared in 5 steps with an overall yield of 27% (95% ee). The omega side chain building block was obtained diastereomerically and enantiomerically pure in 9 steps with an overall yield of 22%-26%. Key steps are the diastereoselective alkylation of an acylated oxazolidinone, the synthesis of an Enantiomerically pure Weinreb-Amid and its coupling with a monolithiated stannane with formation of stannylated keton. The diastereoselective reduction of the ketone was performed by using catecholborane and an oxazaborolidine catalyst. One of the major key steps is the conjugate 1,4 addition of an C13-C20 organocopper building block to the bicyclic C6-C12 azoene with formation of the corresponding hydrazone in 84% yield. The E-Isomer of 16S-iloprost was obtained in 11 steps with an overall yield of 5%. 3-Oxa-16S-iloprost was successfully prepared by using an asymmetric HWE-reaction with a chiral phosphonate as a key step in an overall yield of 6% over 14 steps with >99% de and >99% ee. Furthermore a new, asymmetric synthesis of a central building block was developed. From an enantiomerically pure aldehyde the corresponding alkyne was synthesized by using a new method. This alkyne was successfully transformed into an alkenylstannane with a palladium catalysed hydrostannation. After coupling of the Alkenylstannane with an enantiomerically pure Weinreb-amid the resulting ketone was reduced using the oxazaborolidine catalysed CBS-reduction. The corresponding alcohol was obtained in high yield and high diastereoselectivity. Following this route 3-oxa-16S-iloprost was prepared in an overall yield of 14% over 12 steps with >99% de and >99% ee

Asymmetrische Totalsynthese von 16(S)-Iloprost und 3-Oxa-16(S)-Iloprost mittels der Azoen-Strategie

The present dissertation describes completely stereocontrolled asymmetric syntheses of 16S-iloprost and of 16S-3-oxa-iloprost by a azoene strategy. As Ilomedin and Ventavis registrated iloprost (1:1 mixture of diastereomers at C16) serves as a drug against ischemic heart disease, peripheral vascular disease and primary pulmonary hypertension. The 16S-diastereomer is 5 to 20 times more potent than the 16R-diastereomer in inhibiting collagen-induced platelet aggregation. 3-oxa-16S-iloprost should be more stable against beta-oxidation of the alpha side chain. A bicyclic azoene was prepared in 5 steps with an overall yield of 27% (95% ee). The omega side chain building block was obtained diastereomerically and enantiomerically pure in 9 steps with an overall yield of 22%-26%. Key steps are the diastereoselective alkylation of an acylated oxazolidinone, the synthesis of an Enantiomerically pure Weinreb-Amid and its coupling with a monolithiated stannane with formation of stannylated keton. The diastereoselective reduction of the ketone was performed by using catecholborane and an oxazaborolidine catalyst. One of the major key steps is the conjugate 1,4 addition of an C13-C20 organocopper building block to the bicyclic C6-C12 azoene with formation of the corresponding hydrazone in 84% yield. The E-Isomer of 16S-iloprost was obtained in 11 steps with an overall yield of 5%. 3-Oxa-16S-iloprost was successfully prepared by using an asymmetric HWE-reaction with a chiral phosphonate as a key step in an overall yield of 6% over 14 steps with >99% de and >99% ee. Furthermore a new, asymmetric synthesis of a central building block was developed. From an enantiomerically pure aldehyde the corresponding alkyne was synthesized by using a new method. This alkyne was successfully transformed into an alkenylstannane with a palladium catalysed hydrostannation. After coupling of the Alkenylstannane with an enantiomerically pure Weinreb-amid the resulting ketone was reduced using the oxazaborolidine catalysed CBS-reduction. The corresponding alcohol was obtained in high yield and high diastereoselectivity. Following this route 3-oxa-16S-iloprost was prepared in an overall yield of 14% over 12 steps with >99% de and >99% ee

За кадры. 1948. № 25 (390)

Лучших товарищей на руководящую комсомольскую работуКомсорги групп / К. ЧадоваСочетать академическую учебу с общественной работой / А. ИльенковРазъяснение о новом порядке зачисления на стипендиюВ спортивном клубе института / И. ПавловНовые программы по иностранным языкам / В. М. ГладковаЯвка студентов к началу занятий / А. АстафуровНачало занятий в вечернем университете марксизма-ленинизмаЗа правильную организацию работы / В. Т. ЮринскийСистематически работать над лекционным материалом / С. П. КузнецовУпорно и настойчиво изучать основы марксизма-ленинизма / РабиновичКак добиться успеха в изучении графических дисциплин / Л. С. СкриповПрактические и лабораторные занятия по общей химии / Г. Н. ХодалевичКак я планирую свой рабочий день / Кузмиче

Diagnosis of Morquio Syndrome in Dried Blood Spots Based on a New MRM-MS Assay

<div><p>Background</p><p>Mucopolysaccharidosis IVA (MPS IVA; Morquio A disease) is an autosomal recessive disease caused and characterized by a decreased activity of N-acetylgalactosamine-6-sulfate sulfatase (GALNS), resulting in accumulation of keratan sulfate and chondroitin-6-sulfate in tissues and secondary organ damage. Recently approved enzyme replacement therapy renders the easy and early identification of MPS IVA of out-most importance.</p><p>Methodology</p><p>We propose a completely new assay for the stable and reproducible detection of GALNS deficiency in dry blood spots (DBS). For the validation blood samples were taken from 59 healthy individuals and 24 randomly selected genetically confirmed MPS IVA patients. The material extracted from DBS was incubated with a 4-methylumbelliferyl-β-D-galactopyranoside-6-sulfate as a specific substrate. Final enzymatic product, 4-methylumbelliferone, obtained after adding exogenous beta-galactosidase, was quantified by LC/MRM-MS (liquid-chromatography/multiple-reaction-monitoring mass-spectrometry). 4-propyl-5-hydroxy-7-methyl-2h-chromen-2-one was used as internal standard, a compound with a similar molecular structure and fragmentation pattern in negative ion mode as 4-methylumbelliferone.</p><p>Findings</p><p>The enzymatic assay yielded a positive and negative predictive value of 1.0 for genetically confirmed MPS IVA patients (GALNS activity of 0.35 ± 0.21 μmol/L/h) and for controls with normal GALNS activity (23.1 ± 5.3 μmol/L /h). With present enzymatic conditions, the reaction yield in dried blood spots is at least 20 fold higher than any previously reported data with other assays.</p><p>Interpretation</p><p>The present LC/MRM-MS based assay for MPS IVA diagnosis provides an easy, highly-standardized, accurate and innovative quantification of the enzymatic product in vitro and distinguishes perfectly between MPS IVA affected patients and normal controls. This technique will significantly simplify the early detection of MPS IVA patients.</p></div

Glucosylsphingosine Causes Hematological and Visceral Changes in Mice—Evidence for a Pathophysiological Role in Gaucher Disease

Glucosylceramide and glucosylsphingosine are the two major storage products in Gaucher disease (GD), an inherited metabolic disorder caused by a deficiency of the lysosomal enzyme glucocerebrosidase. The build-up of glucosylceramide in the endoplasmic reticulum and prominent accumulation in cell lysosomes of tissue macrophages results in decreased blood cell and platelet counts, and skeletal abnormalities. The pathological role of the deacylated form of glucosylceramide, glucosylsphingosine (lyso-Gb1), a recently identified sensitive and specific biomarker for GD, is not well investigated. We established a long-term infusion model in C57BL/6JRj mice to examine the effect of lyso-Gb1 on representative hallmark parameters of GD. Mice received lyso-Gb1 at a dosage of 10 mg·kg−1 per day as a continuous subcutaneous administration, and were routinely checked for blood lyso-Gb1 levels using liquid chromatography-multiple reaction monitoring mass spectrometry (LC/MRM-MS) measurements at four-weekly intervals throughout treatment. The C57BL/6JRj mice showed a stable increase of lyso-Gb1 up to-&gt;500-fold greater than the normal reflecting concentrations seen in moderately to severely affected patients. Furthermore, lyso-Gb1 accumulated in peripheral tissues. The mice developed hematological symptoms such as reduced hemoglobin and hematocrit, increased spleen weights and a slight inflammatory tissue response after eight weeks of treatment. The above findings indicate a measurable visceral and hematological response in treated mice that suggests a role for lyso-Gb1 in the development of peripheral signs of GD