Isolation and1H-NMR spectroscopic identification of the glucose-containing lipid-linked precursor oligosaccharide ofN-linked carbohydrate chains

The lipid-linked precursor ofN-type glycoprotein oligosaccharides was isolated from porcine thyroid microsomes after in cubation with UDP[³H] Glucose. The carbohydrate was released from dolichol pyrophosphate by mild acid hydrolysis, purified by gel filtration and characterized by 500-MHz¹H-NMR spectroscopy in combination with enzymatic degradation. The parent oligosaccharide was found to be Glc₃Man₉Glc-NAc₂. The three glucose residues are present in the linear sequence Glcα1-2Glα1-3 Glc, the latter being α(1-3)-linked to one of the mannose residues. In order to establish the branch location of the triglucosyl unit, the parent compound was digested with jack-bean α-mannosidase. The oligosaccharide product was purified by gel filtration, and identified by¹H-NMR as Glc₃Man₅GlcNAc₂ lacking the mannose residues A, D₂, B and D₃. Therefore, the structure of the precursor oligosaccharide is as follows: $$\begin{gathered} c b a D_1 C 4 \hfill \\ Glc\alpha 1 - 2Glc\alpha 1 - 3Glc\alpha 1 - 3Man\alpha 1 - 2Man\alpha 1 - 2Man\alpha 1 \hfill \\ 3 \swarrow 3 2 1 \hfill \\ Man\alpha 1 - 2Man\alpha 1 Man\beta 1 - 4GlcNAc\beta 1 - 4GlcNAc \hfill \\ D_{2 } A 3 6 \hfill \\ Man\alpha 1 \hfill \\ 6 \hfill \\ Man\alpha 1 - 2Man\alpha 1 \nwarrow 4 \hfill \\ D_3 B \hfill \\ \end{gathered} $$      

Clinical and pathological significance of programmed cell death 1 (PD-1)/programmed cell death ligand 1 (PD-L1) expression in high grade serous ovarian cancer

We investigated programmed cell death 1 (PD-1) / programmed cell death ligand 1 (PD-L1) expression in high grade serous ovarian cancer (HGSOC) and its relationship to tumor infiltrating lymphocytes (TIL) and prognosis. Formalin fixed paraffin embedded (FFPE) samples of 94 HGSOC cases were included in the study. Immunohistochemical analysis (CD3, CD4, CD8, PD-1 and PD-L1) was performed. Samples were analyzed for expression of immune proteins in the peritumoral stromal and intratumoral areas, scored, and expression was correlated with overall survival, stage, and age. PD-L1 staining ratio with a score greater than 0 was found to have lower survival. There were two positive staining patterns, patchy/diffuse and patchy/focal patterns, in 24 (25.5%) cases. Considering the threshold value ≥5%, we demonstrated that the PD-L1 positive cancer cell membrane immunoreactivity rate and patchy/diffuse PD-L1 expression were 9.6% (n = 9). There was statistically significant relationship between high PD-1 scores and PD-L1 cases of ≥ 5%. A statistically significant difference was found between PD-L1 staining and survival in patients with a threshold ≥ 5%. However an appropriate rate for treatment was determined in 9.6% cases. There was a statistically significant correlation between PD-1 positive TIL score and intratumoral CD3, peritumoral stromal CD3, intratumoral CD4 and intratumoral CD8 positive cells. Survival was lower in cases with higher PD-L1 positive stromal TIL score.     

The impact of binding site waters on the activity/selectivity trade-off of Janus kinase 2 (JAK2) inhibitors

Structure based optimization of B39, an indazole-based low micromolar JAK2 virtual screening hit is reported. Analysing the effect of certain modifications on the activity and selectivity of the analogues suggested that these parameters are influenced by water molecules available in the binding site. Simulation of water networks in combination with docking enabled us to identify the key waters and to optimize our primary hit into a low nanomolar JAK2 lead with promising selectivity over JAK1.     

Solid-phase synthesis of an N-(phenylalkyl)cinnamide library via Horner-Wadsworth-Emmons reaction

A readily automated solid-phase approach to the synthesis of diverse N-(phenylalkyl)cinnamides, analogues of the NR2B antagonist 2, is described. The procedure utilizes polymer supported N-(phenylalkyl)amines, (diethylphosphono)acetic acid and a wide range of commercially available hydroxybenzaldehydes. The key step, a Horner-Wadsworth-Emmons reaction is achieved under mild conditions and was found to be general for a large number of benzaldehydes. A 225-member focused library was synthesized using a Tecan Combitec synthesizer.     

The gene for juvenile hyaline fibromatosis maps to chromosome 4q21

Juvenile hyaline fibromatosis (JHF) is an autosomal recessive condition characterized by multiple subcutaneous nodular tumors, gingival fibromatosis, flexion contractures of the joints, and an accumulation of hyaline in the dermis. We performed a genomewide linkage search in two families with JHF from the same region of the Indian state of Gujarat and identified a region of homozygosity on chromosome 4q21. Dense microsatellite analyses within this interval in five families with JHF who were from diverse origins demonstrate that all are compatible with linkage to chromosome 4q21 (multipoint LOD score 5.5). Meiotic recombinants place the gene for JHF within a 7-cM interval bounded by D4S2393 and D4S395.     

Presumptive monosomy 21 with neuronal migration disorder re-diagnosed as de novo unbalanced translocation t(18p;21q) by fluorescence in situ hybridisation

We present clinical and cytogenetic data of a one year old boy with partial monosomy for both 21q and 18p, resulting from a de novo unbalanced translocation. The initial diagnosis of a seemingly full monosomy 21 was revised after fluorescence in situ hybridisation (FISH) with whole chromosome painting probes and a locus-specific chromosome 21 probe. The karyotype was reinterpreted as 45,XY,der(18)t(18;21)(p11.2;q22.1),-21. This karyotype, to our knowledge, has not been previously described. The boy presented with a spectrum of clinical features previously described for (partial) monosomy 18p only, for monosomy 21q only, or for both of these aneusomies. The radiological finding of a neuronal migration disorder with localised polymicrogyria (cortical dysplasia) has not been described for either monosomy before.     

Carbamoyloximes as novel non-competitive mGlu5 receptor antagonists

Hit-to-lead optimization of a HTS hit led to new carbamoyloxime derivatives. After identification of an advanced hit (8d) the CYP enzyme inhibitory activity of this class of compounds was successfully eliminated. Systematic exploration of different parts of the advanced hit led us to some promising lead compounds with mGluR5 affinities comparable to that of MPEP.     

Hit-to-lead optimization of disubstituted oxadiazoles and tetrazoles as mGluR5 NAMs

Here we report the discovery and early SAR of a series of mGluR5 negative allosteric modulators (NAMs). Starting from a moderately active HTS hit we synthesized 3,5-disubstituted-oxadiazoles and tetrazoles as mGluR5 NAMs. Based on the analysis of ligand efficiency and lipophilic efficiency metrics we identified a promising lead candidate as a starting point for further optimization.