Systems-Level Change to Alleviate Barriers to Cancer Clinical Trial Access for Adolescents and Young Adults in Australia.

Purpose: International data demonstrate association between clinical trial participation and reduced cancer mortality. Adolescents and young adults (AYA) have low clinical trial enrollment rates. We established a program to understand local barriers and develop targeted solutions that lead to greater AYA clinical trial participation. Methods: A steering committee (SC) with expertise in adult and pediatric oncology, research ethics, and consumer representation was formed. The SC mapped barriers related to AYA trial access and established working groups (WGs) around three themes. Results: The Regulatory Awareness WG identified a lack of understanding of processes that support protocol approval for clinical trials across the AYA age range. A guideline to raise awareness was developed. The Access WG identified challenges for young adults (18-25 years) to access a pediatric hospital to enroll in a pediatric trial. A procedure was developed to streamline applications for access. The first six applications using this procedure have been successful. The Availability WG identified lack of pediatric-adult oncology reciprocal relationships as a barrier to awareness of open trials, and future collaboration. An AYA Craft Group Framework was established to grow relationships within tumor streams across institutions; two craft groups are now operating locally. An additional achievement was a successful request to the Therapeutic Goods Administration for Australian adoption of the Food and Drug Administration Guidance on Considerations for the Inclusion of Adolescent Patients in Adult Oncology Clinical Trials. Conclusion: This multipronged approach to improving AYA clinical trial access has relevance for other health environments. Our knowledge products are available as an online toolkit.

Genomic risk scores for juvenile idiopathic arthritis and its subtypes.

OBJECTIVES: Juvenile idiopathic arthritis (JIA) is an autoimmune disease and a common cause of chronic disability in children. Diagnosis of JIA is based purely on clinical symptoms, which can be variable, leading to diagnosis and treatment delays. Despite JIA having substantial heritability, the construction of genomic risk scores (GRSs) to aid or expedite diagnosis has not been assessed. Here, we generate GRSs for JIA and its subtypes and evaluate their performance. METHODS: We examined three case/control cohorts (UK, US-based and Australia) with genome-wide single nucleotide polymorphism (SNP) genotypes. We trained GRSs for JIA and its subtypes using lasso-penalised linear models in cross-validation on the UK cohort, and externally tested it in the other cohorts. RESULTS: The JIA GRS alone achieved cross-validated area under the receiver operating characteristic curve (AUC)=0.670 in the UK cohort and externally-validated AUCs of 0.657 and 0.671 in the US-based and Australian cohorts, respectively. In logistic regression of case/control status, the corresponding odds ratios (ORs) per standard deviation (SD) of GRS were 1.831 (1.685 to 1.991) and 2.008 (1.731 to 2.345), and were unattenuated by adjustment for sex or the top 10 genetic principal components. Extending our analysis to JIA subtypes revealed that the enthesitis-related JIA had both the longest time-to-referral and the subtype GRS with the strongest predictive capacity overall across data sets: AUCs 0.82 in UK; 0.84 in Australian; and 0.70 in US-based. The particularly common oligoarthritis JIA also had a GRS that outperformed those for JIA overall, with AUCs of 0.72, 0.74 and 0.77, respectively. CONCLUSIONS: A GRS for JIA has potential to augment clinical JIA diagnosis protocols, prioritising higher-risk individuals for follow-up and treatment. Consistent with JIA heterogeneity, subtype-specific GRSs showed particularly high performance for enthesitis-related and oligoarthritis JIA.

Effect of MUC1 length polymorphisms on the NLRP3 inflammasome response of human macrophages.

Mucin 1 is a cell-membrane associated mucin, expressed on epithelial and immune cells that helps protect against pathogenic infections. In humans, MUC1 is highly polymorphic, predominantly due to the presence of a variable number tandem repeat (VNTR) region in the extracellular domain that results in MUC1 molecules of typically either short or long length. A genetic link is known between these MUC1 polymorphisms and inflammation-driven diseases, although the mechanism is not fully understood. We previously showed that MUC1 on murine macrophages specifically restricts activation of the NLRP3 inflammasome, thereby repressing inflammation. This study evaluated the effect of MUC1 VNTR polymorphisms on activity of the NLRP3 inflammasome in human macrophages, finding that long MUC1 alleles correlated with increased IL-1ß production following NLRP3 inflammasome activation. This indicates that the length of MUC1 can influence IL-1ß production, thus providing the first evidence of an immune-modulatory role of MUC1 VNTR polymorphisms in human macrophages.

Higher Sun Exposure is Associated With Lower Risk of Pediatric Inflammatory Bowel Disease: A Matched Case-control Study.

OBJECTIVES: The incidence of pediatric inflammatory bowel disease (IBD) is increasing worldwide. Ecological studies show higher incidence in regions at higher latitude or lower ambient ultraviolet radiation; individual-level associations with sun exposure have not been assessed. METHODS: We recruited children (0-17 years) with IBD from 2 large hospitals in Melbourne, Australia. Control participants were recruited from the day surgery unit of one of the same hospitals. Questionnaires provided data on demographics, past sun exposure, the likelihood of sunburn (skin sensitivity) or tanning following sun exposure, use of sun protection, physical activity, and parental smoking and education. Grandparent ancestry was used to determine participant ethnicity. Cases and controls were matched on age and sex. We used conditional logistic regression to test the association between being an IBD case and past sun exposure at different ages, adjusted for a range of other factors. RESULTS: After matching, n = 99 cases and n = 396 controls were included in the analysis. In multivariable analysis, for each 10 min increment in leisure-time sun exposure in summer or winter there was a linear 6% reduction in the odds of having IBD (P = 0.002). Results were similar in sensitivity analyses including only the most recently diagnosed cases, only Caucasian cases and controls, only those with symptom onset within the year before study entry, or additionally adjusted for age or physical activity. CONCLUSIONS: Higher sun exposure in the previous summer or winter was associated with a reduced risk of having IBD. There are plausible pathways that could mediate this effect.

Extensive Ethnic Variation and Linkage Disequilibrium at the FCGR2/3 Locus: Different Genetic Associations Revealed in Kawasaki Disease.

The human Fc-gamma receptors (FcγRs) link adaptive and innate immunity by binding immunoglobulin G (IgG). All human low-affinity FcγRs are encoded by the FCGR2/3 locus containing functional single nucleotide polymorphisms (SNPs) and gene copy number variants. This locus is notoriously difficult to genotype and high-throughput methods commonly used focus on only a few SNPs. We performed multiplex ligation-dependent probe amplification for all relevant genetic variations at the FCGR2/3 locus in >4,000 individuals to define linkage disequilibrium (LD) and allele frequencies in different populations. Strong LD and extensive ethnic variation in allele frequencies was found across the locus. LD was strongest for the FCGR2C-ORF haplotype (rs759550223+rs76277413), which leads to expression of FcγRIIc. In Europeans, the FCGR2C-ORF haplotype showed strong LD with, among others, rs201218628 (FCGR2A-Q27W, r2 = 0.63). LD between these two variants was weaker (r2 = 0.17) in Africans, whereas the FCGR2C-ORF haplotype was nearly absent in Asians (minor allele frequency <0.005%). The FCGR2C-ORF haplotype and rs1801274 (FCGR2A-H131R) were in weak LD (r2 = 0.08) in Europeans. We evaluated the importance of ethnic variation and LD in Kawasaki Disease (KD), an acute vasculitis in children with increased incidence in Asians. An association of rs1801274 with KD was previously shown in ethnically diverse genome-wide association studies. Now, we show in 1,028 European KD patients that the FCGR2C-ORF haplotype, although nearly absent in Asians, was more strongly associated with susceptibility to KD than rs1801274 in Europeans. Our data illustrate the importance of interpreting findings of association studies concerning the FCGR2/3 locus with knowledge of LD and ethnic variation.

Genetic determinants of paediatric food allergy: A systematic review.

BACKGROUND: The genetic determinants of food allergy have not been systematically reviewed. We therefore systematically reviewed the literature on the genetic basis of food allergy, identifying areas for further investigation. METHODS: We searched three electronic databases (MEDLINE, EMBASE and PubMed) on 9 January 2018. Two authors screened retrieved articles for review according to inclusion criteria and extracted relevant information on study characteristics and measures of association. Eligible studies included those that reported an unaffected nonatopic control group, had genetic information and were carried out in children. RESULTS: Of the 2088 studies retrieved, 32 met our inclusion criteria. Five were genome-wide association studies, and the remaining were candidate gene studies. Twenty-two of the studies were carried out in a predominantly Caucasian population with the remaining 10 from Asian-specific populations or unspecified ethnicity. We found FLG, HLA, IL10, IL13, as well as some evidence for other variants (SPINK5, SERPINB and C11orf30) that are associated with food allergy. CONCLUSIONS: Little genetic research has been carried out in food allergy, with FLG, HLA and IL13 being the most reproducible genes for an association with food allergy. Despite promising results, existing genetic studies on food allergy are inundated with issues such as inadequate sample size and absence of multiple testing correction. Few included replication analyses or population stratification measures. Studies addressing these limitations along with functional studies are therefore needed to unravel the mechanisms of action of the identified genes.

Children with East Asian-Born Parents Have an Increased Risk of Allergy but May Not Have More Asthma in Early Childhood.

BACKGROUND: We previously reported that infants with Asian-born parents are 3 times more likely to have IgE-mediated food allergy than those with Australian-born parents. It is unknown whether this translates to the increased risk of other allergic diseases later in childhood and whether ancestry interacts with other risk factors for allergic disease development. OBJECTIVE: To compare prevalence and risk factors for allergic rhinitis, asthma, and aeroallergen sensitization at age 6 between children with East Asian-born and Caucasian-born parents. METHODS: A total of 5276 1-year-old infants were recruited into a population-based longitudinal study of allergy. A total of 4455 children participated in age 6 follow-up (84.4%), including 3015 with Caucasian-born parents and 415 with East Asian-born parents. Children underwent skin prick tests to aeroallergens and questionnaires captured data on asthma, eczema, and allergic rhinitis. RESULTS: Compared with children with Caucasian-born parents, children of East Asian-born parents had more allergic rhinitis (19.9% [95% confidence interval (CI) 14.9-26] vs 9.3% [95% CI 8-10.8], P < .001) and aeroallergen sensitization (64.3% [95% CI 57.5-70.5] vs 34.4% [95% CI 32.2-36.7], P < .001) at age 6. Asthma was similar in both groups (9.1% [95% CI 6.2-13.2] vs 11.7% [95% CI 10.4-13.1]), P = .21. Children with IgE-mediated food allergy and eczema in infancy were 3 times more likely to have asthma and 2 times more likely to have allergic rhinitis at age 6, irrespective of ancestry. CONCLUSIONS: Children of East Asian ancestry born in Australia have a higher burden of most allergic diseases in the first 6 years of life, whereas asthma may follow a different pattern. IgE-mediated food allergy and eczema at age 1 increase the risk of asthma and allergic rhinitis irrespective of ancestry.

Association of Increased Sun Exposure Over the Life-course with a Reduced Risk of Juvenile Idiopathic Arthritis.

Cutaneous sun exposure is an important determinant of circulating vitamin D. Both sun exposure and vitamin D have been inversely associated with risk of autoimmune disease. In juvenile idiopathic arthritis (JIA), low circulating vitamin D appears common, but disease-related behavioral changes may have influenced sun exposure. We therefore aimed to determine whether predisease sun exposure is associated with JIA. Using validated questionnaires, we retrospectively measured sun exposure for 202 Caucasian JIA case-control pairs born in Victoria Australia, matched for birth year and time of recruitment. Measures included maternal sun exposure at 12 weeks of pregnancy and child sun exposure across the life-course prediagnosis. We converted exposure to UVR dose and looked for case-control differences using logistic regression, adjusting for potential confounders. Higher cumulative prediagnosis UVR exposure was associated with reduced risk of JIA, with a clear dose-response relationship (trend P = 0.04). UVR exposure at 12 weeks of pregnancy was similarly inversely associated with JIA (trend P = 0.011). Associations were robust to sensitivity analyses for prediagnosis behavioral changes, disease duration and knowledge of the hypothesis. Our data indicate that lower UVR exposure may increase JIA risk. This may be through decreased circulating vitamin D, but prospective studies are required to confirm this.

Ximmer: a system for improving accuracy and consistency of CNV calling from exome data.

Background: While exome and targeted next-generation DNA sequencing are primarily used for detecting single nucleotide changes and small indels, detection of copy number variants (CNVs) can provide highly valuable additional information from the data. Although there are dozens of exome CNV detection methods available, these are often difficult to use, and accuracy varies unpredictably between and within datasets. Findings: We present Ximmer, a tool that supports an end-to-end process for evaluating, tuning, and running analysis methods for detection of CNVs in germline samples. Ximmer includes a simulation framework, implementations of several commonly used CNV detection methods, and a visualization and curation tool that together enable interactive exploration and quality control of CNV results. Using Ximmer, we comprehensively evaluate CNV detection on four datasets using five different detection methods. We show that application of Ximmer can improve accuracy and aid in quality control of CNV detection results. In addition, Ximmer can be used to run analyses and explore CNV results in exome data. Conclusions: Ximmer offers a comprehensive tool and method for applying and improving accuracy of CNV detection methods for exome data.

Candidate Gene Testing in Clinical Cohort Studies with Multiplexed Genotyping and Mass Spectrometry.

Complex diseases are often underpinned by multiple common genetic variants that contribute to disease susceptibility. Here, we describe a cost-effective tag single nucleotide polymorphism (SNP) approach using a multiplexed genotyping assay with mass spectrometry, to investigate gene pathway associations in clinical cohorts. We investigate the food allergy candidate locus Interleukin13 (IL13) as an example. This method efficiently maximizes the coverage by taking advantage of shared linkage disequilibrium (LD) within a region. Selected LD SNPs are then designed into a multiplexed assay, enabling up to 40 different SNPs to be analyzed simultaneously, boosting cost-effectiveness. Polymerase chain reaction (PCR) is used to amplify the target loci, followed by single nucleotide extension, and the amplicons are then measured using matrix-assisted laser desorption/ionization-time of flight(MALDI-TOF) mass spectrometry. The raw output is analyzed with the genotype calling software, using stringent quality control definitions and cut-offs, and high probability genotypes are determined and output for data analysis.

Higher parental occupational social contact is associated with a reduced risk of incident pediatric type 1 diabetes: Mediation through molecular enteroviral indices.

We aimed to examine the association between parental occupational social contact and hygiene factors on type 1 diabetes (T1D) risk and possible mediation of these effects through child enteroviral infection. We interviewed 333 incident T1D cases and 660 controls from 2008-2011 in Melbourne, Australia. Enteroviral indices (ribonucleic acid by reverse transcription polymerase chain reaction and Coxsackie B virus antibody levels) in peripheral blood were measured in nested case control samples. Parent occupational social contact was assessed by the number of well or sick children, adults or animals contacted daily through work. Higher parental occupational social contact was strongly associated with reduced T1D risk with evidence of dose response (contact with the well or sick score, Adjusted odds ratio (AOR) per category: 0.73 (95% Confidence Interval (CI): 0.66, 0.81); P<0.001 or AOR 0.63 (95% CI: 0.53, 0.75); P<0.001) respectively). Nine of the ten parental social contact indices, were significant mediated through one or more enteroviral indices. The strength of association between enterovirus presence and T1D onset increased with child age (1.2 fold increase per year; P = 0.05). Lower child hand hygiene enhanced the adverse effect of low parental occupational contact with the sick; Synergy Index 5.16 (95% CI: 3.61, 7.36). The interaction between hand washing and parental occupational contact is more consistent with protection against parental enteroviral shedding than the sharing of a protective infectious agent or microbiome.

The DNA methylation landscape of CD4+ T cells in oligoarticular juvenile idiopathic arthritis.

Juvenile idiopathic arthritis (JIA) is presumed to be driven by an adverse combination of genes and environment. Epigenetic processes, including DNA methylation, act as a conduit through which the environment can regulate gene activity. Altered DNA methylation has been associated with adult autoimmune rheumatic diseases such as rheumatoid arthritis, but studies are lacking for paediatric autoimmune rheumatic diseases including JIA. Here, we performed a genome-scale case-control analysis of CD4+ T cell DNA methylation from 56 oligoarticular JIA (oJIA) cases and 57 age and sex matched controls using Illumina HumanMethylation450 arrays. DNA methylation at each array probe was tested for association with oJIA using RUV (Remove Unwanted Variation) together with a moderated t-test. Further to this 'all-inclusive' analysis, we stratified by age at diagnosis (≤6yrs, >6yrs) and by sex as potential sources of heterogeneity. Following False Discovery Rate (FDR) adjustment, no probes were associated with oJIA in the all-inclusive, >6yrs-diagnosed, or sex-stratified analyses, and only one probe was associated with oJIA in the ≤6yrs-diagnosed analysis. We attempted technical validation and replication of 14 probes (punadj<0.01) at genes of known/potential relevance to disease. At VPS53, we demonstrated a regional shift towards higher methylation in oJIA (all-inclusive) compared to controls. At REEP3, where polymorphism has been previously associated with JIA, we demonstrated higher DNA methylation in male oJIA compared to male controls. This is the most comprehensive JIA case-control analysis of DNA methylation to date. While we have generated some evidence of altered methylation in oJIA, substantial differences are not apparent in CD4+ T cells. This may indicate a lesser relevance of DNA methylation levels in childhood, compared to adult, rheumatic disease.

Familial Analysis of Epistatic and Sex-Dependent Association of Genes of the Renin-Angiotensin-Aldosterone System and Blood Pressure.

BACKGROUND: Renin-angiotensin-aldosterone system genes have been inconsistently associated with blood pressure, possibly because of unrecognized influences of sex-dependent genetic effects or gene-gene interactions (epistasis). METHODS AND RESULTS: We tested association of systolic blood pressure with single-nucleotide polymorphisms (SNPs) at renin (REN), angiotensinogen (AGT), angiotensin-converting enzyme (ACE), angiotensin II type 1 receptor (AGTR1), and aldosterone synthase (CYP11B2), including sex-SNP or SNP-SNP interactions. Eighty-eight tagSNPs were tested in 2872 white individuals in 809 pedigrees from the Victorian Family Heart Study using variance components models. Three SNPs (rs8075924 and rs4277404 at ACE and rs12721297 at AGTR1) were individually associated with lower systolic blood pressure with significant (P<0.00076) effect sizes ≈1.7 to 2.5 mm Hg. Sex-specific associations were seen for 3 SNPs in men (rs2468523 and rs2478544 at AGT and rs11658531 at ACE) and 1 SNP in women (rs12451328 at ACE). SNP-SNP interaction was suggested (P<0.005) for 14 SNP pairs, none of which had shown individual association with systolic blood pressure. Four SNP pairs were at the same gene (2 for REN, 1 for AGT, and 1 for AGTR1). The SNP rs3097 at CYP11B2 was represented in 5 separate pairs. CONCLUSIONS: SNPs at key renin-angiotensin-aldosterone system genes associate with systolic blood pressure individually in both sexes, individually in one sex only and only when combined with another SNP. Analyses that incorporate sex-dependent and epistatic effects could reconcile past inconsistencies and account for some of the missing heritability of blood pressure and are generally relevant to SNP association studies for any phenotype.

Conjunctival Ultraviolet Autofluorescence as a Measure of Past Sun Exposure in Children.

Background: Conjunctival ultraviolet autofluorescence (CUVAF) area detected from UVAF photographs is a recently developed potential marker for past sun exposure, but its relationship with sun-related factors has not been fully investigated.Methods: The study included 339 healthy children ages 5 to 15 years in Melbourne, Australia. Data were collected by questionnaire and examination at school. CUVAF area was measured using a computer program and analyzed as a continuous and dichotomous outcome (any/none).Results: Fifty-three children (15.6%) had detectable CUVAF, and the youngest age at which a child showed sun damage was 8 years. Compared with silicone skin cast score, there was good inter-grader agreement on CUVAF grading, with Cohen kappa 0.85 [95% confidence interval (CI), 0.65-1.00] for total CUVAF area using both eye photographs. Perfect intra-grader agreement was achieved. Fairer pigmentation, including medium/fair skin color [adjusted odds ratio (AOR), 3.42; 95% CI, 1.02-11.48 vs. dark/olive] and blue/gray eye color (AOR, 4.07; 95% CI, 1.73-9.55 vs. brown) was associated with increased odds of CUVAF. Increasing lifetime sunburn number (e.g., AOR, 2.89; 95% CI, 1.14-7.35 and 4.29; 1.04-17.76 for sunburns 2 to 4 and ≥ 5 times, respectively, vs. no sunburns, trend P = 0.004) and freckling by the end of last summer were associated with increased odds of CUVAF.Conclusions: CUVAF area can be an a priori objective measure of past sun exposure in pediatric populations for future research.Impact: To our knowledge, this is the first pediatric study that evaluated associations of sun-related risk factors with CUVAF. Cancer Epidemiol Biomarkers Prev; 26(7); 1146-53. ©2017 AACR.

A survey of national and multi-national registries and cohort studies in juvenile idiopathic arthritis: challenges and opportunities.

BACKGROUND: To characterize the existing national and multi-national registries and cohort studies in juvenile idiopathic arthritis (JIA) and identify differences as well as areas of potential future collaboration. METHODS: We surveyed investigators from North America, Europe, and Australia about existing JIA cohort studies and registries. We excluded cross-sectional studies. We captured information about study design, duration, location, inclusion criteria, data elements and collection methods. RESULTS: We received survey results from 18 studies, including 11 national and 7 multi-national studies representing 37 countries in total. Study designs included inception cohorts, prevalent disease cohorts, and new treatment cohorts (several of which contribute to pharmacosurveillance activities). Despite numerous differences, the data elements collected across the studies was quite similar, with most studies collecting at least 5 of the 6 American College of Rheumatology core set variables and the data needed to calculate the 3-variable clinical juvenile disease activity score. Most studies were collecting medication initiation and discontinuation dates and were attempting to capture serious adverse events. CONCLUSION: There is a wide-range of large, ongoing JIA registries and cohort studies around the world. Our survey results indicate significant potential for future collaborative work using data from different studies and both combined and comparative analyses.

Environmental and genetic determinants of two vitamin D metabolites in healthy Australian children.

BACKGROUND: Vitamin D deficiency has been associated with adverse health outcomes. We examined genetic and environmental determinants of serum 25(OH)D3 and 1,25(OH)2D3 in childhood. METHODS: The study sample consisted of 322 healthy Australian children (predominantly Caucasians) who provided a venous blood sample. A parental interview was conducted and skin phototype and anthropometry measures were assessed. Concentrations of 25(OH)D3 and 1,25(OH)2D3 were measured by selective solid-phase extraction-capillary liquid chromatography-tandem mass spectrometry. These concentrations were deseasonalised where relevant to remove the effect of month of sampling. RESULTS: Deseasonalised log 25(OH)D3 and 1,25(OH)2D3 concentrations were only moderately correlated (r=0.42, p<0.001). The following predicted both 25(OH)D3 and 1,25(OH)2D3: UVR 6 weeks before the interview, natural skin and eye colour, height and vitamin D allelic metabolism score. The following predicted 25(OH)D3 only: lifetime sunburns and vitamin D allelic synthesis score. Overall, 43.5% and 25.6% of variation in 25(OH)D3 and 1,25(OH)2D3 could be explained. After accounting for 25(OH)D3 concentrations, higher UVR 6 weeks before the interview and vitamin D allelic metabolism score further predicted 1,25(OH)2D3 concentrations. CONCLUSIONS: Environmental factors and genetic factors contributed to both vitamin D metabolite concentrations. The intriguing finding that the higher ambient UVR contributed to higher 1,25(OH)2D3 after accounting for 25(OH)D3 concentrations requires further evaluation.

Determinants of Neonatal Vitamin D Levels as Measured on Neonatal Dried Blood Spot Samples.

BACKGROUND: Vitamin D deficiency is linked to adverse childhood health outcomes, yet data on the distribution and quantifiable determinants of neonatal 25-hydroxyvitamin D3 (25OHD) concentration, a vitamin D biomarker, are limited. OBJECTIVE: Our aim was to identify determinants of neonatal 25OHD concentration, measured using neonatal dried blood spots (DBS). METHODS: A total of 259 ethnically diverse children aged 0-16 years born in Victoria, Australia, were recruited. Data included maternal sun exposure, skin type, 25OHD concentration on stored neonatal DBS, and genotypes at the target genes. Associations were investigated using multiple linear regression models. RESULTS: The median 25OHD concentration was 29.2 nmol/l (IQR 18.0-47.4). Measured 25OHD was <50 nmol/l in almost half of the neonatal sample. Ambient ultraviolet radiation (UVR) 6 weeks before birth was the strongest predictor of neonatal 25OHD, accounting for 23% of its variation. A further 10% was explained by infant genetic variants at GC (rs2282679), the gene encoding the vitamin D binding protein, and DHCR7 (rs12785878), a gene required for synthesis of 7-dehydrocholesterol, a precursor to 25OHD. DBS age explained 7%, and patterns of maternal sun exposure and clothing choices accounted for 4%. A child's skin colour was strongly associated with GC gene variants and not independent of these variants in predicting 25OHD. The final model explained 43% of the total variance in neonatal 25OHD concentration. CONCLUSION: Maternal lifestyle factors and infant genetic variants predict neonatal 25OHD levels; the importance of maternal UVR exposure in late pregnancy is highlighted.

The MassARRAY® System for Targeted SNP Genotyping.

Research to understand the genetic basis of disease, particularly complex disease, regularly involves single nucleotide polymorphism (SNP) genotyping. The use of genome-wide SNP genotyping arrays has become increasingly more commonplace for gene discovery. However, smaller-scale genotyping platforms capable of efficiently genotyping tens to hundreds of SNPs are still crucial for many aspects of this work, including replication of associations. The Agena Bioscience MassARRAY System is one such platform. Here, we provide a guide to using the MassARRAY System, from assay design, through mass spectrometry, to generation of genotype data.

Variants in CXCR4 associate with juvenile idiopathic arthritis susceptibility.

BACKGROUND: Juvenile idiopathic arthritis (JIA) is the most common chronic rheumatic disease among children, the etiology of which involves a strong genetic component, but much of the underlying genetic determinants still remain unknown. Our aim was to identify novel genetic variants that predispose to JIA. METHODS: We performed a genome-wide association study (GWAS) and replication in a total of 1166 JIA cases and 9500 unrelated controls of European ancestry. Correlation of SNP genotype and gene expression was investigated. Then we conducted targeted resequencing of a candidate locus, among a subset of 480 cases and 480 controls. SUM test was performed to evaluate the association of the identified rare functional variants. RESULTS: The CXCR4 locus on 2q22.1 was found to be significantly associated with JIA, peaking at SNP rs953387. However, this result is subjected to subpopulation stratification within the subjects of European ancestry. After adjusting for principal components, nominal significant association remained (p < 10(-4)). Because of its interesting known function in immune regulation, we carried out further analyses to assess its relationship with JIA. Expression of CXCR4 was correlated with CXCR4 rs953387 genotypes in lymphoblastoid cell lines (p = 0.014) and T-cells (p = 0.0054). In addition, rare non-synonymous and stop-gain sequence variants in CXCR4, putatively damaging for CXCR4 function, were significantly enriched in JIA cases (p = 0.015). CONCLUSION: Our results suggest the association of CXCR4 variants with JIA, implicating that this gene may be involved in the pathogenesis of autoimmune disease. However, because this locus is subjected to population stratification within the subjects of European ancestry, additional replication is still necessary for this locus to be considered a true risk locus for JIA. This cell-surface chemokine receptor has already been targeted in other diseases and may serve as a tractable therapeutic target for a specific subset of pediatric arthritis patients with additional replication and functional validation of the locus.