O6-[(2”,3”-O-Isopropylidene-5”-O-tbutyldimethylsilyl)pentyl]- 5′-O-tbutyldiphenylsilyl-2′,3′-O-isopropylideneinosine

Cyclic adenosine diphosphate ribose (cADPR) is a cyclic nucleotide involved in the Ca2+ homeostasis. In its structure, the northern ribose, bonded to adenosine through an N1 glycosidic bond, is connected to the southern ribose through a pyrophosphate bridge. Due to the chemical instability at the N1 glycosidic bond, new bioactive cADPR derivatives have been synthesized. One of the most interesting analogues is the cyclic inosine diphosphate ribose (cIDPR), in which the hypoxanthine replaced adenosine. The efforts for synthesizing new linear and cyclic northern ribose modified cIDPR analogues led us to study in detail the inosine N1 alkylation reaction. In the last few years, we have produced new flexible cIDPR analogues, where the northern ribose has been replaced by alkyl chains. With the aim to obtain the closest flexible cIDPR analogue, we have attached to the inosine N1 position a 2”,3”-dihydroxypentyl chain, possessing the two OH groups in a ribose-like fashion. The inosine alkylation reaction afforded also the O6-alkylated regioisomer, which could be a useful intermediate for the construction of new kinds of cADPR mimics

Step-growth glycopolymers with a defined tacticity for selective carbohydrate–lectin recognition

Glycopolymers are potent candidates for biomedical applications by exploiting multivalent carbohydrate–lectin interactions. Owing to their specific recognition capabilities, glycosylated polymers can be utilized for targeted drug delivery to certain cell types bearing the corresponding lectin receptors. A fundamental challenge in glycopolymer research, however, is the specificity of recognition to receptors binding to the same sugar unit (e.g., mannose). Variation of polymer backbone chirality has emerged as an effective method to distinguish between lectins on a molecular level. Herein, we present a facile route toward producing glycopolymers with a defined tacticity based on a step-growth polymerization technique using click chemistry. A set of polymers have been fabricated and further functionalized with mannose moieties to enable lectin binding to receptors relevant to the immune system (mannose-binding lectin, dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin, and dendritic and thymic epithelial cell-205). Surface plasmon resonance spectrometry was employed to determine the kinetic parameters of the step-growth glycopolymers. The results highlight the importance of structural complexity in advancing glycopolymer synthesis, yet multivalency remains a main driving force in lectin recognition

Degradable thioester core-crosslinked star-shaped polymers

Degradable polymers are considered to present a promising solution to combat plastic pollution. However, many polymers are based on ester and amide bonds, which often require high temperatures and acidic/basic catalysis and hence do not easily degrade under typical environmental conditions. Thus, more readily degradable polymer structures that enable the use of milder conditions are highly sought after. Herein, degradable core-crosslinked star-shaped branched polymers have been synthesised via a two-step one-pot reversible addition–fragmentation chain transfer polymerisation. For the first time, a bifunctional thiomethacrylate crosslinker was used to prepare a range of star-shaped poly(methyl methacrylate) structures which were characterised by advanced viscometry gel permeation chromatography. The increased reactivity of thioesters over their oxoester analogues was exploited to degrade the resulting star-shaped polymers via amidation, as evidenced by the complete degradation of the crosslinked core upon heating in the presence of amines. The successful degradation of polymers containing thioesters shows the potential of thioester chemistry for producing more sustainable/degradable polymers with more complex and defined architectures

Low serum total testosterone level as a predictor of upstaging and upgrading in low-risk prostate cancer patients meeting the inclusion criteria for active surveillance

Active surveillance (AS) is currently a widely accepted treatment option for men with clinically localized prostate cancer (PCa). Several reports have highlighted the association of low serum testosterone levels with high-grade, high-stage PCa. However, the impact of serum testosterone as a predictor of progression in men with low-risk PCa has been little assessed.In this study, we evaluated the association of circulating testosterone concentrations with a staging/grading reclassification in a cohort of low-risk PCa patients meeting the inclusion criteria for the AS protocol but opting for radical prostatectomy.Radical prostatectomy (RP) was performed in 338 patients, eligible for AS according to the following criteria: clinical stage T2a or less, PSApT2) and upgrading (GS≥7; primary Gleason pattern 4) disease. Unfavorable disease was defined as the occurrence of pathological stage>pT2 and predominant Gleason score 4. Total testosterone was measured before surgery.Low serum testosterone levels (<300 ng/dL) were significantly associated with upgrading, upstaging, unfavorable disease and positive surgical margins. The addition of testosterone to a base model, including age, PSA, PSA density, clinical stage and positive cancer involvement in cores, showed a significant independent influence of this variable on upstaging, upgrading and unfavorable disease.In conclusion, our results support the idea that total testosterone should be a selection criterion for inclusion of low-risk PCa patients in AS programs and suggest that testosterone level less than 300 ng/dL should be considered a discouraging factor when a close AS program is considered as treatment option

KIAA1840 mutations cause ARCMT2

Charcot–Marie–Tooth disease is a group of hereditary peripheral neuropathies that share clinical characteristics of progressive distal muscle weakness and atrophy, foot deformities, distal sensory loss, as well as diminished tendon reflexes. Hundreds of causative DNA changes have been found, but much of the genetic basis of the disease is still unexplained. Mutations in the ALS5/SPG11/ KIAA1840 gene are a frequent cause of autosomal recessive hereditary spastic paraplegia with thin corpus callosum and peripheral axonal neuropathy, and account for ∼40% of autosomal recessive juvenile amyotrophic lateral sclerosis. The overlap of axonal Charcot–Marie–Tooth disease with both diseases, as well as the common autosomal recessive inheritance pattern of thin corpus callosum and axonal Charcot–Marie–Tooth disease in three related patients, prompted us to analyse the ALS5/SPG11/ KIAA1840 gene in affected individuals with autosomal recessive axonal Charcot–Marie–Tooth disease. We investigated 28 unrelated families with autosomal recessive axonal Charcot–Marie–Tooth disease defined by clinical, electrophysiological, as well as pathological evaluation. Besides, we screened for all the known genes related to axonal autosomal recessive Charcot–Marie-Tooth disease (CMT2A2/HMSN2A2/ MFN2 , CMT2B1/ LMNA , CMT2B2/ MED25 , CMT2B5/ NEFL , ARCMT2F/dHMN2B/ HSPB1 , CMT2K/ GDAP1 , CMT2P/ LRSAM1 , CMT2R/ TRIM2 , CMT2S/ IGHMBP2 , CMT2T/ HSJ1 , CMTRID/ COX6A1 , ARAN-NM/ HINT and GAN/ GAN ), for the genes related to autosomal recessive hereditary spastic paraplegia with thin corpus callosum and axonal peripheral neuropathy (SPG7/ PGN , SPG15/ ZFYVE26, SPG21/ ACP33 , SPG35/ FA2H , SPG46/ GBA2 , SPG55/ C12orf65 and SPG56/ CYP2U1 ), as well as for the causative gene of peripheral neuropathy with or without agenesis of the corpus callosum ( SLC12A6 ) . Mitochondrial disorders related to Charcot–Marie–Tooth disease type 2 were also excluded by sequencing POLG and TYMP genes. An additional locus for autosomal recessive Charcot–Marie–Tooth disease type 2H on chromosome 8q13-21.1 was excluded by linkage analysis. Pedigrees originated in Italy, Brazil, Canada, England, Iran, and Japan. Interestingly, we identified 15 ALS5/SPG11/ KIAA1840 mutations in 12 families (two sequence variants were never reported before, p.Gln198* and p.Pro2212fs*5). No large deletions/duplications were detected in these patients. The novel mutations seemed to be pathogenic since they co-segregated with the disease in all pedigrees and were absent in 300 unrelated controls. Furthermore, in silico analysis predicted their pathogenic effect. Our results indicate that ALS5/SPG11/ KIAA1840 is the causative gene of a wide spectrum of clinical features, including autosomal recessive axonal Charcot–Marie–Tooth disease

Aspetti metodologici del monitoraggio di inquinanti aerodiffusi persistenti mediante “MOSS-BAGS” di nuova generazione

I muschi, grazie alla loro elevata efficienza nell’accumulo di particolato atmosferico, sono considerati degli ottimi biomonitor. Una tecnica oramai diffusa prevede l’esposizione del materiale all’interno di reticelle, formando le cosiddette “moss-bags”, che possono essere distribuite sul territorio secondo schemi espositivi ottimali. Tra i principali limiti di questa tecnica si ricorda che il muschio viene raccolto in natura, e ciò ha un certo impatto ambientale, mentre l’operazione di preparazione delle bags presenta aspetti “artigianali” che rappresentano un serio handicap per la sua diffusione. Per ovviare a questi problemi un team di ricercatori europei ha sviluppato il progetto FP7 MOSSCLONE, che ha tra gli obiettivi anche quelli di (1) individuare e caratterizzare un muschio particolarmente performante per clonarlo, coltivarlo in condizioni controllate, e quindi usarlo come materiale di uso standard, dopo un processo di devitalizzazione; (2) ottimizzare le moss-bags per forma, taglia, ampiezza delle maglie della rete di contenimento e quantità di muschio, proponendo un prototipo di nuova concezione da sottoporre a brevetto internazionale; (3) standardizzare le condizioni di esposizione, in termini di altezza e durata di esposizione; (4) confrontare i dati ottenuti con le moss-bags di nuova generazione con i dati ottenuti con i metodi chimicofisici tradizionali. In questa sede si presentano i risultati del primo anno e mezzo di attività, in particolare quelli relativi ai punti (2) e (3), basati sull’esposizione di più di 250 bags di vecchia e nuova generazione per periodi di diversa durata, da 3 a un massimo di 12 settimane, in sette siti campani con diverso uso del suolo; ciò in attesa di avere a disposizione i dati relativi ad altre esposizioni condotte in due regioni europee climaticamente molto diverse, la Galizia e l’Austria orientale. Viene inoltre illustrato in prima nazionale il nuovo prototipo

The In Vitro, Ex Vivo, and In Vivo Effect of Polymer Hydrophobicity on Charge-Reversible Vectors for Self-Amplifying RNA

RNA technology has the potential to revolutionize vaccination. However, the lack of clear structure-property relationships in relevant biological models mean there is no clear consensus on the chemical motifs necessary to improve RNA delivery. In this work, we describe the synthesis of a series of copolymers based on the self-hydrolyzing charge-reversible polycation poly(dimethylaminoethyl acrylate) (pDMAEA), varying the lipophilicity of the additional co-monomers. All copolymers formed stable polyplexes, showing efficient complexation with model nucleic acids from nitrogen/phosphate (N/P) ratios of N/P = 5, with more hydrophobic complexes exhibiting slower charge reversal and disassembly compared to hydrophilic analogues. The more hydrophobic copolymers outperformed hydrophilic versions, homopolymer controls and the reference standard polymer (polyethylenimine), in transfection assays on 2D cell monolayers, albeit with significantly higher toxicities. Similarly, hydrophobic derivatives displayed up to a 4-fold higher efficacy in terms of the numbers of cells expressing green fluorescent protein (GFP+) cells in ex vivo human skin (10%) compared to free RNA (2%), attributed to transfection enrichment in epithelial cells. In contrast, in a mouse model, we observed the reverse trend in terms of RNA transfection, with no observable protein production in more hydrophobic analogues, whereas hydrophilic copolymers induced the highest transfection in vivo. Overall, our results suggest an important relationship between the vector lipophilicity and RNA transfection in vaccine settings, with polymer biocompatibility potentially a key parameter in effective in vivo protein production

The global impact of the DRACMA guidelines cow’s milk allergy clinical practice

Background: The 2010 Diagnosis and Rationale for Action against Cow’s Milk Allergy (DRACMA) guidelines are the only Grading of Recommendations Assessment, Development and Evaluation (GRADE) guidelines for cow’s milk allergy (CMA). They indicate oral food challenge (OFC) as the reference test for diagnosis, and suggest the choice of specific alternative formula in different clinical conditions. Their recommendations are flexible, both in diagnosis and in treatment. Objectives & methods: Using the Scopus citation records, we evaluated the influence of the DRACMA guidelines on milk allergy literature. We also reviewed their impact on successive food allergy and CMA guidelines at national and international level. We describe some economic consequences of their application. Results: DRACMA are the most cited CMA guidelines, and the second cited guidelines on food allergy. Many subsequent guidelines took stock of DRACMA’s metanalyses adapting recommendations to the local context. Some of these chose not to consider OFC as an absolute requirement for the diagnosis of CMA. Studies on their implementation show that in this case, the treatment costs may increase and there is a risk of overdiagnosis. Interestingly, we observed a reduction in the cost of alternative formulas following the publication of the DRACMA guidelines. Conclusions: DRACMA reconciled international differences in the diagnosis and management of CMA. They promoted a cultural debate, improved clinician’s knowledge of CMA, improved the quality of diagnosis and care, reduced inappropriate practices, fostered the efficient use of resources, empowered patients, and influenced some public policies. The accruing evidence on diagnosis and treatment of CMA necessitates their update in the near futur