Predictors of the utility of clinical exome sequencing as a first-tier genetic test in patients with Mendelian phenotypes: results from a referral center study on 603 consecutive cases.

BACKGROUND: Clinical exome sequencing (CES) provides a comprehensive and effective analysis of relevant disease-associated genes in a cost-effective manner compared to whole exome sequencing. Although several studies have focused on the diagnostic yield of CES, no study has assessed predictors of CES utility among patients with various Mendelian phenotypes. We assessed the effectiveness of CES as a first-level genetic test for molecular diagnosis in patients with a Mendelian phenotype and explored independent predictors of the clinical utility of CES. RESULTS: Between January 2016 and December 2019, 603 patients (426 probands and 177 siblings) underwent CES at the Department of Molecular Medicine of the University Hospital of Nancy. The median age of the probands was 34 years (IQR, 12-48), and the proportion of males was 46.9% (200/426). Adults and children represented 64.8% (276/426) and 35.2% (150/426), respectively. The median test-to-report time was 5.6 months (IQR, 4.1-7.2). CES revealed 203 pathogenic or likely pathogenic variants in 160 patients, corresponding to a diagnostic yield of 37.6% (160/426). Independent predictors of CES utility were criteria strongly suggestive of an extreme phenotype, including pediatric presentation and patient phenotypes associated with an increased risk of a priori probability of a monogenic disorder, the inclusion of at least one family member in addition to the proband, and a CES prescription performed by an expert in the field of rare genetic disorders. CONCLUSIONS: Based on a large dataset of consecutive patients with various Mendelian phenotypes referred for CES as a first-tier genetic test, we report a diagnostic yield of ~ 40% and several independent predictors of CES utility that might improve CES diagnostic efficiency.

Gas-phase structure of polymer ions: Tying together theoretical approaches and ion mobility spectrometry.

An increasing number of studies take advantage of ion mobility spectrometry (IMS) coupled to mass spectrometry (IMS-MS) to investigate the spatial structure of gaseous ions. Synthetic polymers occupy a unique place in the field of IMS-MS. Indeed, due to their intrinsic dispersity, they offer a broad range of homologous ions with different lengths. To help rationalize experimental data, various theoretical approaches have been described. First, the study of trend lines is proposed to derive physicochemical and structural parameters. However, the evaluation of data fitting reflects the overall behavior of the ions without reflecting specific information on their conformation. Atomistic simulations constitute another approach that provide accurate information about the ion shape. The overall scope of this review is dedicated to the synergy between IMS-MS and theoretical approaches, including computational chemistry, demonstrating the essential role they play to fully understand/interpret IMS-MS data.

SARS-CoV-2 Variability in Patients and Wastewaters-Potential Immuno-Modulation during the Shift from Delta to Omicron.

The continuous emergence of SARS-CoV-2 variants favors potential co-infections and/or viral mutation events, leading to possible new biological properties. The aim of this work was to characterize SARS-CoV-2 genetic variability during the Delta-Omicron shift in patients and in a neighboring wastewater treatment plant (WWTP) in the same urban area. The surveillance of SARS-CoV-2 was performed by routine screening of positive samples by single nucleotide polymorphism analysis within the S gene. Moreover, additionally to national systematic whole genome sequencing (WGS) once a week in SARS-CoV-2-positive patients, WGS was also applied when mutational profiles were difficult to interpret by routine screening. Thus, WGS was performed on 414 respiratory samples and on four wastewater samples, northeastern France. This allowed us to report (i) the temporally concordant Delta to Omicron viral shift in patients and wastewaters; (ii) the characterization of 21J (Delta) and 21K (Omicron)/BA.1-21L (Omicron)/BA.2-BA.4 mixtures from humans or environmental samples; (iii) the mapping of composite mutations and the predicted impact on immune properties in the viral Spike protein.

Next-generation sequencing and genotype association studies reveal the association of HLA-DRB3*02:02 with delayed hypersensitivity to penicillins.

BACKGROUND: Nonimmediate (delayed)-allergic reactions to penicillins are common and some of them can be life-threatening. The genetic factors influencing these reactions are unknown/poorly known/poorly understood. We assessed the genetic predictors of a delayed penicillin allergy that cover the HLA loci. METHODS: Using next-generation sequencing (NGS), we genotyped the MHC region in 24 patients with delayed hypersensitivity compared with 20 patients with documented immediate hypersensitivity to penicillins recruited in Italy. Subsequently, we analyzed in silico Illumina Immunochip genotyping data that covered the HLA loci in 98 Spanish patients with delayed hypersensitivity and 315 with immediate hypersensitivity compared to 1,308 controls. RESULTS: The two alleles DRB3*02:02:01:02 and DRB3*02:02:01:01 were reported in twenty cases with delayed reactions (83%) and ten cases with immediate reactions (50%), but not in the Allele Frequency Net Database. Bearing at least one of the two alleles increased the risk of delayed reactions compared to immediate reactions, with an OR of 8.88 (95% CI, 3.37-23.32; p < .0001). The haplotype (ACAA) from rs9268835, rs6923504, rs6903608, and rs9268838 genetic variants of the HLA-DRB3 genomic region was significantly associated with an increased risk of delayed hypersensitivity to penicillins (OR, 1.7; 95% CI: 1.06-1.92; p = .001), but not immediate hypersensitivity. CONCLUSION: We showed that the HLA-DRB3 locus is strongly associated with an increased risk of delayed penicillin hypersensitivity, at least in Southwestern Europe. The determination of HLA-DRB3*02:02 alleles in the risk management of severe delayed hypersensitivity to penicillins should be evaluated further in larger population samples of different origins.

Performance evaluation of an automated SARS-CoV-2 Ag test for the diagnosis of COVID-19 infection on nasopharyngeal swabs.

OBJECTIVES: The detection of SARS-CoV-2 in infected people is a key tool to help in controlling COVID-19 pandemic. Like rapid antigenic tests, automated antigen tests, that present the advantage of a higher throughput flow, may be of interest. The LIAISON® SARS-CoV-2 Ag test was evaluated for the quantification of SARS-CoV-2 nucleocapsid antigen in nasopharyngeal swabs by comparison to RT-PCR. METHODS: The study involved 378 nasopharyngeal samples (UTM® and FLOQSwab™, Copan Diagnostics), including 46 swabs positive for SARS-CoV-2 by RT-PCR. These samples came from asymptomatic (n=99, 26.2%) or symptomatic people (n=279, 73.8%), at different times from symptom onset. The samples were analyzed on LIAISON® XL. RESULTS: The overall specificity was 99.4% (CI95% [98.6-100]). The negative predictive value reached 100% in asymptomatic people. Among the 46 positive samples, the overall sensitivity was 84.8% (CI95% [74.4-95.2]), reached 91.9% (CI95% [83.1-100]) in the first fourth days after symptoms onset and was 100% for Cq values ≤25. Antigen was not detected in samples with Cq values >25. Similar results were observed on nasopharyngeal swabs coming from patients infected with the 20I/501Y.V1 variant or the 20H/501Y.V2 variant. CONCLUSIONS: According to technical performances, the LIAISON® SARS-CoV-2 Ag test may be a useful tool for COVID-19 diagnosis, especially during the first four days of symptoms.

Population and evolutionary genetics of the PAH locus to uncover overdominance and adaptive mechanisms in phenylketonuria: Results from a multiethnic study.

BACKGROUND: Phenylketonuria (PKU) is the most common inborn error of amino acid metabolism in Europe. The reasons underlying the high prevalence of heterozygous carriers are not clearly understood. We aimed to look for pathogenic PAH variant enrichment according to geographical areas and patients' ethnicity using a multiethnic nationwide cohort of patients with PKU in France. We subsequently appraised the population differentiation, balancing selection and the molecular evolutionary history of the PAH locus. METHODS: The French nationwide PKU study included patients who have been referred at the national level to the University Hospital of Nancy, and for whom a molecular diagnosis of phenylketonuria was made by Sanger sequencing. We performed enrichment analyses by comparing alternative allele frequencies using Fisher's exact test with Bonferroni adjustment. We estimated the amount of genetic differentiation among populations using Wright's fixation index (Fst). To estimate the molecular evolutionary history of the PAH gene, we performed phylogenetic and evolutionary analyses using whole-genome and exome-sequencing data from healthy individuals and non-PKU patients, respectively. Finally, we used exome-wide association study to decipher potential genetic loci associated with population divergence on PAH. FINDINGS: The study included 696 patients and revealed 132 pathogenic PAH variants. Three geographical areas showed significant enrichment for a pathogenic PAH variant: North of France (p.Arg243Leu), North-West of France (p.Leu348Val), and Mediterranean coast (p.Ala403Val). One PAH variant (p.Glu280Gln) was significantly enriched among North-Africans (OR = 23·23; 95% CI: 9·75-55·38). PAH variants exhibiting a strong genetic differentiation were significantly enriched in the 'Biopterin_H' domain (OR = 6·45; 95% CI: 1·99-20·84), suggesting a balancing selection pressure on the biopterin function of PAH. Phylogenetic and timetree analyses were consistent with population differentiation events on European-, African-, and Asian-ancestry populations. The five PAH variants most strongly associated with a high selection pressure were phylogenetically close and were located within the biopterin domain coding region of PAH or in its vicinity. Among the non-PAH loci potentially associated with population divergence, two reached exome-wide significance: SSPO (SCO-spondin) and DBH (dopamine beta-hydroxylase), involved in neuroprotection and metabolic adaptation, respectively. INTERPRETATION: Our data provide evidence on the combination of evolutionary and adaptive events in populations with distinct ancestries, which may explain the overdominance of some genetic variants on PAH. FUNDING: French National Institute of Health and Medical Research (INSERM) UMR_S 1256.

Exome sequencing of cases with neural tube defects identifies candidate genes involved in one-carbon/vitamin B12 metabolisms and Sonic Hedgehog pathway.

Neural tube defects (NTD) result from complex mechanisms between genes, nutrition and environment. The identification of genetic predictors by genome exome sequencing and their influence on genome methylation need further consideration. Gene variants related to 1-CM metabolism (1-CM) could influence the methylation of genes involved in neural tube embryogenesis through impaired synthesis of S-adenosyl methionine. We performed exome sequencing of 6116 genes referenced in OMIM and NTD risk and genome-wide methylation in 23 NTD cases. We replicated the most significant associations in 81 other cases. The analysis of exome sequencing identified one gene of 1-CM, LRP2, and one gene of Sonic Hedgehog (SHH), GLI3, in the 23 NTD cases. The analysis restricted to genes of 1-CM and neural tube embryogenesis identified five gene predictors of 1-CM (LRP2, rs137983840; MMAA, rs148142853; TCN2, rs35838082; FPGS, rs41306702; BHMT, rs763726268) and two of SHH (GLI3, rs35364414; MKS1, rs151023718). We replicated the association of TCN2, BHMT and GLI3 with NTD risk in the 81 cases. We found a significant hemimethylation of CFAP46 that may influence SHH activation in one case, who carried risk alleles in BHMT, LRP2, MMAA and GLI3. In conclusion, we identified new candidate genes and rare variants that highlight an interacting influence of genes involved in SHH and 1-CM in the puzzle of genetic components of NTD risk.

Vitamin B-12 and liver activity and expression of methionine synthase are decreased in fetuses with neural tube defects.

BACKGROUND: The risk of neural tube defects (NTDs) is influenced by nutritional factors and genetic determinants of one-carbon metabolism. A key pathway of this metabolism is the vitamin B-12- and folate-dependent remethylation of homocysteine, which depends on methionine synthase (MS, encoded by MTR), methionine synthase reductase, and methylenetetrahydrofolate reductase. Methionine, the product of this pathway, is the direct precursor of S-adenosylmethionine (SAM), the universal methyl donor needed for epigenetic mechanisms. OBJECTIVES: This study aimed to evaluate whether the availability of vitamin B-12 and folate and the expression or activity of the target enzymes of the remethylation pathway are involved in NTD risk. METHODS: We studied folate and vitamin B-12 concentrations and activity, expression, and gene variants of the 3 enzymes in liver from 14 NTD and 16 non-NTD fetuses. We replicated the main findings in cord blood from pregnancies of 41 NTD fetuses compared with 21 fetuses with polymalformations (metabolic and genetic findings) and 375 control pregnancies (genetic findings). RESULTS: The tissue concentration of vitamin B-12 (P = 0.003), but not folate, and the activity (P = 0.001), transcriptional level (P = 0.016), and protein expression (P = 0.003) of MS were decreased and the truncated inactive isoforms of MS were increased in NTD livers. SAM was significantly correlated with MS activity and vitamin B-12. A gene variant in exon 1 of GIF (Gastric Intrinsic Factor gene) was associated with a dramatic decrease of liver vitamin B-12 in 2 cases. We confirmed the decreased vitamin B-12 in cord blood from NTD pregnancies. A gene variant of GIF exon 3 was associated with NTD risk. CONCLUSIONS: The decreased vitamin B-12 in liver and cord blood and decreased expression and activity of MS in liver point out the impaired remethylation pathway as hallmarks associated with NTD risk. We suggest evaluating vitamin B-12 in the nutritional recommendations for prevention of NTD risk beside folate fortification or supplementation.

Plasma mSEPT9: A Novel Circulating Cell-free DNA-Based Epigenetic Biomarker to Diagnose Hepatocellular Carcinoma.

BACKGROUND: Patients with cirrhosis are at high risk of hepatocellular carcinoma (HCC). The SEPT9 gene is a key regulator of cell division and tumor suppressor whose hypermethylation is associated with liver carcinogenesis. The primary aim of this study was to evaluate the diagnostic accuracy of a PCR-based assay for the analysis of SEPT9 promoter methylation in circulating cell-free DNA (mSEPT9) for diagnosing HCC among cirrhotic patients. METHODS: We report two phase II biomarker studies that included cirrhotic patients with or without HCC from France (initial study) and Germany (replication study). All patients received clinical and biological evaluations, and liver imaging according to current recommendations. The primary outcome was defined as the presence of HCC according to guidelines from the American Association for the Study of Liver Diseases. The diagnosis of HCC was confirmed by abdominal contrast-enhanced computed tomography scan and systematically discussed in a multidisciplinary consultation meeting. HCC-free cirrhotic patients were recruited if the screening abdominal ultrasound showed no evidence of HCC at the time of blood sampling for the mSEPT9 test and on the next visit six months later. The adjudicating physicians were blinded to patient results associated with the mSEPT9 test. FINDINGS: We included 289 patients with cirrhosis (initial: 186; replication: 103), among whom 98 had HCC (initial: 51; replication: 47). The mSEPT9 test exhibited high diagnostic accuracy for HCC diagnosis, with an area under the receiver operating characteristic curve (AUROC) of 0.944 (0.900-0.970, p<0.0001) in the initial study (replication: 0.930 [0.862-0.971, p<0.0001]; meta-analysis: AUROC=0.940 [0.910-0.970, p<0.0001], no heterogeneity: I2=0%, p=0.67; and no publication bias). In multivariate logistic regression analysis, the number of positive mSEPT9 triplicates was the only independent variable significantly associated with HCC diagnosis (initial: OR=6.30, for each mSEPT9 positive triplicate [2.92-13.61, p<0.0001]; replication: OR=6.07 [3.25-11.35, p<0.0001]; meta-analysis: OR=6.15 [2.93-9.38, p<0.0001], no heterogeneity: I2=0%, p=0.95; no publication bias). AUROC associated with the discrimination of the logistic regression models in initial and validation studies were 0.969 (0.930-0.989) and 0.942 (0.878-0.978), respectively, with a pooled AUROC of 0.962 ([0.937-0.987, p<0.0001], no heterogeneity: I2=0%, p=0.36; and no publication bias). INTERPRETATION: Among patients with cirrhosis, the mSEPT9 test constitutes a promising circulating epigenetic biomarker for HCC diagnosis at the individual patient level. Future prospective studies should assess the mSEPT9 test in the screening algorithm for cirrhotic patients to improve risk prediction and personalized therapeutic management of HCC.

Publisher Correction: A PRDX1 mutant allele causes a MMACHC secondary epimutation in cblC patients.

The original version of this Article contained an error in the title, which was incorrectly given as 'APRDX1 mutant allele causes a MMACHC secondary epimutation in cblC patients'. This has now been corrected in both the PDF and HTML versions of the Article to read 'A PRDX1 mutant allele causes a MMACHC secondary epimutation in cblC patients'.

APRDX1 mutant allele causes a MMACHC secondary epimutation in cblC patients.

To date, epimutations reported in man have been somatic and erased in germlines. Here, we identify a cause of the autosomal recessive cblC class of inborn errors of vitamin B12 metabolism that we name "epi-cblC". The subjects are compound heterozygotes for a genetic mutation and for a promoter epimutation, detected in blood, fibroblasts, and sperm, at the MMACHC locus; 5-azacytidine restores the expression of MMACHC in fibroblasts. MMACHC is flanked by CCDC163P and PRDX1, which are in the opposite orientation. The epimutation is present in three generations and results from PRDX1 mutations that force antisense transcription of MMACHC thereby possibly generating a H3K36me3 mark. The silencing of PRDX1 transcription leads to partial hypomethylation of the epiallele and restores the expression of MMACHC. This example of epi-cblC demonstrates the need to search for compound epigenetic-genetic heterozygosity in patients with typical disease manifestation and genetic heterozygosity in disease-causing genes located in other gene trios.

BRIP1 coding variants are associated with a high risk of hepatocellular carcinoma occurrence in patients with HCV- or HBV-related liver disease.

The molecular mechanisms of hepatocellular carcinoma (HCC) carcinogenesis are still not fully understood. DNA repair defects may influence HCC risk. The aim of the study was to look for potential genetic variants of DNA repair genes associated with HCC risk among patients with alcohol- or viral-induced liver disease. We performed four case-control studies on 2,006 European- (Derivation#1 and #2 studies) and African-ancestry (Validation#1 and #2 studies) patients originating from several cohorts in order to assess the association between genetic variants on DNA repair genes and HCC risk using a custom array encompassing 94 genes. In the Derivation#1 study, the BRIP1 locus reached array-wide significance (Chi-squared SV-Perm, P=5.00×10-4) among the 253 haplotype blocks tested for their association with HCC risk, in patients with viral cirrhosis but not among those with alcoholic cirrhosis. The BRIP1 haplotype block included three exonic variants (rs4986763, rs4986764, rs4986765). The BRIP1 'AAA' haplotype was significantly associated with an increased HCC risk [odds ratio (OR), 2.01 (1.19-3.39); false discovery rate (FDR)-P=1.31×10-2]. In the Derivation#2 study, results were confirmed for the BRIP1 'GGG' haplotype [OR, 0.53 (0.36-0.79); FDR-P=3.90×10-3]. In both Validation#1 and #2 studies, BRIP1 'AAA' haplotype was significantly associated with an increased risk of HCC [OR, 1.71 (1.09-2.68); FDR-P=7.30×10-2; and OR, 6.45 (4.17-9.99); FDR-P=2.33×10-19, respectively]. Association between the BRIP1 locus and HCC risk suggests that impaired DNA mismatch repair might play a role in liver carcinogenesis, among patients with HCV- or HBV-related liver disease.

Polymers for Traveling Wave Ion Mobility Spectrometry Calibration.

One of the main issues when using traveling wave ion mobility spectrometry (TWIMS) for the determination of collisional cross-section (CCS) concerns the need for a robust calibration procedure built from referent ions of known CCS. Here, we implement synthetic polymer ions as CCS calibrants in positive ion mode. Based on their intrinsic polydispersities, polymers offer in a single sample the opportunity to generate, upon electrospray ionization, numerous ions covering a broad mass range and a large CCS window for different charge states at a time. In addition, the key advantage of polymer ions as CCS calibrants lies in the robustness of their gas-phase structure with respect to the instrumental conditions, making them less prone to collisional-induced unfolding (CIU) than protein ions. In this paper, we present a CCS calibration procedure using sodium cationized polylactide and polyethylene glycol, PLA and PEG, as calibrants with reference CCS determined on a home-made drift tube. Our calibration procedure is further validated by testing the polymer calibration to determine CCS of numerous different ions for which CCS are reported in the literature. Graphical Abstract ᅟ.

Synthesis of Quercetin-imprinted Polymer Spherical Particles with Improved Ability to Capture Quercetin Analogues.

INTRODUCTION: Molecularly imprinted polymers (MIPs) are composed of specific cavities able to selectively recognise a template molecule. Used as chromatographic sorbents, MIPs may not trap related structures due to the high rigidity of their cross-linking. OBJECTIVE: To improve the capture of quercetin analogues by modulating the synthesis strategy for a quercetin-imprinted polymer (Qu MIP). METHODOLOGY: An additional comonomer bearing a short oligoethylene glycol (OEG) unit was used to prepare a Qu MIP that was compared to a traditional one formulated in a similar fashion, but without the OEG-comonomer. The Qu MIPs were prepared in bead form through fluorocarbon suspension polymerisation. After solid phase extraction (SPE) assessment of their imprinted cavities, the MIPs were evaluated by HPLC for their recognition properties towards quercetin and other polyphenols, including flavonoids, phenolic acids and curcumin. The Qu MIPs were finally SPE-tested on a white onion extract. RESULTS: The incorporation of OEG units modulated the selectivity of the Qu MIP by improving the recognition of quercetin related structures (12-61% increase in the imprinting effect for distant analogues). It also allowed limiting or suppressing non-specific hydrophobic interactions (decrease of about 10% in the rate of quercetin retention on the non-imprinted polymer). The SPE application of the MIP to a white onion extract indicates its interest for the selective extraction of quercetin and its analogues. CONCLUSION: The OEG-modified Qu MIP appears to be an attractive tool to discover new drug candidates from natural sources by extracting, amongst interfering compounds, structural analogues of quercetin. Copyright © 2017 John Wiley & Sons, Ltd.

Cyclic Polymers by Ring-Closure Strategies.

The preparation of cyclic macromolecules has always represented a challenging task for polymer science, mainly because of difficulties in connecting chain extremities together. Initiated by the pioneering studies of Jacobson and Stockmayer, preparative pathways to cyclic polymers have been considerably improved within the last two decades thanks to the advent of both controlled polymerizations and efficient coupling reactions in organic chemistry. This Review aims to provide a critical up-to-date overview and illustrate the considerable efforts that have been made in the past few years to improve the availability of macrocycles for industrial and academic investigations through the use of the ring-closure approach. Particular attention is paid to methods for the preparation of monocycles over more complex architectures, since the latter are usually derived from the former.

Cystathionine ß-synthase genetic variant rs2124459 is associated with a reduced risk of cleft palate in French and Belgian populations.

BACKGROUND: Orofacial cleft (OFC) is the most prevalent craniofacial birth defect. Genes involved in one-carbon, folate and vitamin B12 metabolisms have been associated with OFC but no study performed a concomitant assessment on genes involved in these three pathways. OBJECTIVE: We looked for potential genetic variants associated with OFC using an exhaustive gene panel of one-carbon metabolism. METHODS: We performed a case-control discovery study on children with OFC (236 cases, 145 controls) and their related mothers (186 cases, 127 controls). We performed a replication study on the top significant genetic variant in an independent group from Belgium (248 cases, 225 controls). RESULTS: In the discovery study on 'mothers', the CBS locus reached array-wide significance (p=9.13×10-6; Bonferroni p=4.77×10-3; OR 0.47 (0.33 to 0.66)) among the 519 haplotypes tested for their association with OFC risk. Within the CBS haplotype block (rs2124459, rs6586282, rs4920037, rs234705, rs234709), the rs2124459 was the most significantly associated with a reduced risk of OFC (p=1.77×10-4; Bonferroni p=2.00×10-2; OR 0.53 (0.38 to 0.74), minor allele). The rs2124459 was associated with a reduced risk of cleft palate (CP) (p=6.78×10-5; Bonferroni p=7.80×10-3; OR 0.40 (0.25 to 0.63)). In the 'children' group, the rs2124459 was associated with a reduced risk of CP (p=0.02; OR 0.61 (0.40 to 0.93), minor allele). The association between rs2124459 and reduced risk of CP was replicated in an independent children population from Belgium (p=0.02; OR 0.64 (0.44 to 0.93), minor allele). CONCLUSIONS: The CBS rs2124459 was associated with a reduced risk of CP in both French and Belgian populations. These results highlight the prominent involvement of the vitamin B6-dependent transsulfuration pathway of homocysteine in OFC risk and the interest for evaluating vitamin B6 status in further population studies.

Synthesis and Characterization of Double Crystalline Cyclic Diblock Copolymers of Poly(ε-caprolactone) and Poly(l(d)-lactide) (c(PCL-b- PL(D)LA)).

The synthesis of symmetric cyclo poly(ε-caprolactone)-block-poly(l(d)-lactide) (c(PCL-b-PL(D)LA)) by combining ring-opening polymerization of ε-caprolactone and lactides and subsequent click chemistry reaction of the linear precursors containing antagonist functionalities is presented. The two blocks can sequentially crystallize and self-assemble into double crystalline spherulitic superstructures. The cyclic chain topology significantly affects both the nucleation and the crystallization of each constituent, as gathered from a comparison of the behavior of linear precursors and cyclic block copolymers. The stereochemistry of the PLA block does not have a significant effect on the nonisothermal crystallization of both linear and cyclo PCL-b-PDLA and PCL-b-PLLA copolymers.

Binary Mixed Homopolymer Brushes Tethered to Cellulose Nanocrystals: A Step Towards Compatibilized Polyester Blends.

This article reports on the successful preparation and characterization of cellulose nanocrystals (CNCs) surface-modified with polylactide (PLA) and poly(butylene succinate) (PBS) binary mixed homopolymer brushes. Their synthesis was designed as a three-step procedure combining polyester synthesis and surface-modification of CNCs with simultaneous polyester grafting via a heterogeneous copper(I)-catalyzed azide-alkyne cycloaddition reaction. For comparison, single homopolymer brushes tethered to CNCs (PLLA-g-CNC and PBSBDEMPAM-g-CNC) were obtained applying the same procedure. The hairy nanoparticles were characterized in terms of chemical composition and thermal properties. Spectroscopic analyses suggested "rippled" microphase separation of both immiscible homopolyesters in the mixed brushes, while others showed impeded homopolyester crystallization after surface-grafting. A synergistic relationship between the polyesters and CNCs was also suggested, i.e., the polyester grafting increases the CNC thermal resistance, while CNC presence imparts char formation. The as-obtained binary homopolymer brushes tethered to nanoparticles makes these surface-modified cellulosic nanomaterials attractive as compatibilization/reinforcement agents for PLA/PBS blends.

Exome-Wide Association Study Identifies New Low-Frequency and Rare UGT1A1 Coding Variants and UGT1A6 Coding Variants Influencing Serum Bilirubin in Elderly Subjects: A Strobe Compliant Article.

Genome-wide association studies (GWASs) have identified loci contributing to total serum bilirubin level. However, no exome-wide approaches have been performed to address this question. Using exome-wide approach, we assessed the influence of protein-coding variants on unconjugated, conjugated, and total serum bilirubin levels in a well-characterized cohort of 773 ambulatory elderly subjects from Italy. Coding variants were replicated in 227 elderly subjects from the same area. We identified 4 missense rare (minor allele frequency, MAF < 0.5%) and low-frequency (MAF, 0.5%-5%) coding variants located in the first exon of the UGT1A1 gene, which encodes for the substrate-binding domain (rs4148323 [MAF = 0.06%; p.Gly71Arg], rs144398951 [MAF = 0.06%; p.Ile215Val], rs35003977 [MAF = 0.78%; p.Val225Gly], and rs57307513 [MAF = 0.06%; p.Ser250Pro]). These variants were in strong linkage disequilibrium with 3 intronic UGT1A1 variants (rs887829, rs4148325, rs6742078), which were significantly associated with total bilirubin level (P = 2.34 × 10(-34), P = 7.02 × 10(-34), and P = 8.27 × 10(-34)), as well as unconjugated, and conjugated bilirubin levels. We also identified UGT1A6 variants in association with total (rs6759892, p.Ser7Ala, P = 1.98 × 10(-26); rs2070959, p.Thr181Ala, P = 2.87 × 10(-27); and rs1105879, p.Arg184Ser, P = 3.27 × 10(-29)), unconjugated, and conjugated bilirubin levels. All UGT1A1 intronic variants (rs887829, rs6742078, and rs4148325) and UGT1A6 coding variants (rs6759892, rs2070959, and rs1105879) were significantly associated with gallstone-related cholecystectomy risk. The UGT1A6 variant rs2070959 (p.Thr181Ala) was associated with the highest risk of gallstone-related cholecystectomy (OR, 4.58; 95% CI, 1.58-13.28; P = 3.21 × 10(-3)). Using an exome-wide approach we identified coding variants on UGT1A1 and UGT1A6 genes in association with serum bilirubin level and hyperbilirubinemia risk in elderly subjects. UGT1A1 intronic single-nucleotide polymorphisms (SNPs) (rs6742078, rs887829, rs4148324) serve as proxy markers for the low-frequency and rare UGT1A1 variants, thereby providing mechanistic explanation to the relationship between UGT1A1 intronic SNPs and the UGT1A1 enzyme activity. UGT1A1 and UGT1A6 variants might be potentially associated with gallstone-related cholecystectomy risk.

HLA-DRA variants predict penicillin allergy in genome-wide fine-mapping genotyping.

BACKGROUND: Immediate reactions to ß-lactams are the most common causes of anaphylactic reactions and can be life-threatening. The few known genetic factors influencing these reactions suggest a link with atopy and inflammation. OBJECTIVE: We performed a fine-mapping genome-wide association study of the genetic predictors of ß-lactam allergy to better understand the underlying mechanisms. METHODS: We studied 387 patients with immediate allergic reactions to ß-lactams and 1124 paired control subjects from Spain. We replicated the results in 299 patients and 362 paired control subjects from Italy. RESULTS: We found significant associations with the single nucleotide polymorphisms rs4958427 of ZNF300 (c.64-471G>A, P = 9.9 × 10(-9)), rs17612 of C5 (c.4311A>C [p.Glu1437Asp], P = 7.5 × 10(-7)), rs7754768 and rs9268832 of the HLA-DRA | HLA-DRB5 interregion (P = 1.6 × 10(-6) and 4.9 × 10(-6)), and rs7192 of HLA-DRA (c.724T>G [p.Leu242Val], P = 7.4 × 10(-6)) in an allelic model, with similar results in an additive model. Single nucleotide polymorphisms of HLA-DRA and ZNF300 predicted skin test positivity to amoxicillin and other penicillins but not to cephalosporins. A haplotype block in HLA-DRA and the HLA-DRA | HLA-DRB5 interregion encompassed a motif involved in balanced expression of the α- and ß-chains of MHC class II, whereas rs7192 was predicted to influence α-chain conformation. HLA-DRA rs7192 and rs8084 were significantly associated with allergy to penicillins and amoxicillin (P = 6.0 × 10(-4) and P = 4.0 × 10(-4), respectively) but not to cephalosporins in the replication study. CONCLUSIONS: Gene variants of HLA-DRA and the HLA-DRA | HLA-DRB5 interregion were significant predictors of allergy to penicillins but not to cephalosporins. These data suggest complex gene-environment interactions in which genetic susceptibility of HLA type 2 antigen presentation plays a central role.