Molecular construction of HIV-gp120 discontinuous epitope mimics by assembly of cyclic peptides on an orthogonal alkyne functionalized TAC-scaffold

Mimics of discontinuous epitopes of for example bacterial or viral proteins may have considerable potential for the development of synthetic vaccines, especially if conserved epitopes can be mimicked. However, due to the structural complexity and size of discontinuous epitopes molecular construction of these mimics remains challeging. We present here a convergent route for the assembly of discontinuous epitope mimics by successive azide alkyne cycloaddition on an orthogonal alkyne functionalized scaffold. Here the synthesis of mimics of the HIV gp120 discontinuous epitope that interacts with the CD4 receptor is described. The resulting protein mimics are capable of inhibition of the gp120–CD4 interaction. The route is convergent, robust and should be applicable to other discontinuous epitopes

Variability of Serum Proteins in Chinese and Dutch Human Milk during Lactation

To better understand the variability of the type and level of serum proteins in human milk, the milk serum proteome of Chinese mothers during lactation was investigated using proteomic techniques and compared to the milk serum proteome of Dutch mothers. This showed that total milk serum protein concentrations in Chinese human milk decreased over a 20-week lactation period, although with variation between mothers in the rate of decrease. Variation was also found in the composition of serum proteins in both colostrum and mature milk, although immune-active proteins, enzymes, and transport proteins were the most abundant for all mothers. These three protein groups account for many of the 15 most abundant proteins, with these 15 proteins covering more than 95% of the total protein concentrations, in both the Chinese and Dutch milk serum proteome. The Dutch and Chinese milk serum proteome were also compared based on 166 common milk serum proteins, which showed that 22% of the 166 serum proteins differed in level. These differences were observed mainly in colostrum and concern several highly abundant proteins. This study also showed that protease inhibitors, which are highly correlated to immune-active proteins, are present in variable amounts in human milk and could be relevant during digestion.</p

Serum Protein N-Glycans in Colostrum and Mature Milk of Chinese Mothers

To study the Chinese human milk N-glycome over lactation, N-glycans were released and separated from serum proteins, purified by solid-phase extraction, and analyzed by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS). In total, 66 different putative N-glycans were found in the colostrum (week 1) and mature milk (week 4) of seven Chinese mothers. A clear difference was observed between milk of five secretor and two nonsecretor mothers, based on the type and relative amounts of the individual N-glycans. The relative levels of the total neutral nonfucosylated and the fucosylated N-glycans in milk of five secretor mothers increased and decreased over lactation, respectively. This pattern could not be observed for the milk from the two nonsecretor mothers. Overall, this was the first study that provided detailed information on individual N-glycans in milk among mothers and over time as well as that fucosylation of N-glycans in milk was associated with the mother's secretor status. </p

Human Milk Oligosaccharides in Colostrum and Mature Milk of Chinese Mothers: Lewis Positive Secretor Subgroups

To study the variability in human milk oligosaccharide (HMO) composition of Chinese human milk over a 20-wk lactation period, HMO profiles of 30 mothers were analyzed using CE-LIF. This study showed that total HMO concentrations in Chinese human milk decreased significantly over a 20-wk lactation period, independent of the mother’s SeLe status, although with individual variations. In addition, total acidic and neutral HMO concentrations in Chinese human milk decreased over lactation, and levels are driven by their mother’s SeLe status. Analysis showed that total neutral fucosylated HMO concentrations in Chinese human milk were higher in the two secretor groups as compared to the nonsecretor group. On the basis of the total neutral fucosylated HMO concentrations in Chinese human milk, HMO profiles within the Se⁺Le⁺ group can be divided into two subgroups. HMOs that differed in level between Se⁺Le⁺ subgroups were 2′FL, DF-L, LNFP I, and F-LNO. HMO profiles in Dutch human milk also showed Se⁺Le⁺ subgroup division, with 2′FL, LNT, and F-LNO as the driving force

Human Milk Oligosaccharides in Colostrum and Mature Milk of Chinese Mothers: Lewis Positive Secretor Subgroups

To study the variability in human milk oligosaccharide (HMO) composition of Chinese human milk over a 20-wk lactation period, HMO profiles of 30 mothers were analyzed using CE-LIF. This study showed that total HMO concentrations in Chinese human milk decreased significantly over a 20-wk lactation period, independent of the mother’s SeLe status, although with individual variations. In addition, total acidic and neutral HMO concentrations in Chinese human milk decreased over lactation, and levels are driven by their mother’s SeLe status. Analysis showed that total neutral fucosylated HMO concentrations in Chinese human milk were higher in the two secretor groups as compared to the nonsecretor group. On the basis of the total neutral fucosylated HMO concentrations in Chinese human milk, HMO profiles within the Se⁺Le⁺ group can be divided into two subgroups. HMOs that differed in level between Se⁺Le⁺ subgroups were 2′FL, DF-L, LNFP I, and F-LNO. HMO profiles in Dutch human milk also showed Se⁺Le⁺ subgroup division, with 2′FL, LNT, and F-LNO as the driving force

CHOP-ASO ameliorates glomerular and tubular damage on top of ACE inhibition in diabetic kidney disease.

Background: Maladaptive ER stress signaling in diabetic kidney disease (DKD) is linked to increased glomerular and tubular expression of the cell death-promoting transcription factor C/EBP homologous protein (CHOP). We determined whether therapy with locked nucleic acid (LNA)-modified antisense oligonucleotides (ASOs) targeting CHOP ameliorates experimental DKD. Methods: Following an in vivo dose-escalation study, we determined the efficacy of CHOPASO in the early and later stages of experimental DKD (8- or 16-week-old db/db mice, respectively) alone or in combination with an angiotensin-converting enzyme inhibitor (ACEi). Renal functional parameters and morphological analyses were used to determine the effects. Renal gene expression profiling was conducted to determine differentially regulated genes and pathways. Several human CHOP-ASOs were tested in hyperglycemia-exposed human kidney cells. Results: CHOP-ASOs efficiently reduced renal CHOP expression in diabetic mice and reduced markers of DKD at early and late stages. Early combined intervention (CHOP-ASO and ACEi) efficiently prevented interstitial damage. At the later timepoint, the combined treatment reduced indices of both glomerular and tubular damage more efficiently than either intervention alone. A significantly larger number of genes and disease pathways were affected by CHOP-ASO, including reduced Slc5a2 (sodium-glucose transport protein 2) and PROM1 (CD133). Human CHOP-ASOs efficiently reduced glucose-induced CHOP and prevented cell death of human kidney cells in vitro Conclusions: The ASO-based approach efficiently reduced renal CHOP expression in a diabetic mouse model, providing an additional benefit to an ACEi in particular at later timepoints. These studies demonstrate that ASO-based therapies efficiently reduce maladaptive CHOP expression and ameliorate experimental DKD