Assessing the utilization of high-resolution 2-field HLA typing in solid organ transplantation.

HLA typing in solid organ transplantation (SOT) is necessary for determining HLA-matching status between donor-recipient pairs and assessing patients' anti-HLA antibody profiles. Histocompatibility has traditionally been evaluated based on serologically defined HLA antigens. The evolution of HLA typing and antibody identification technologies, however, has revealed many limitations with using serologic equivalents for assessing compatibility in SOT. The significant improvements to HLA typing introduced by next-generation sequencing (NGS) require an assessment of the impact of this technology on SOT. We have assessed the role of high-resolution 2-field HLA typing (HR-2F) in SOT by retrospectively evaluating NGS-typed pre- and post-SOT cases. HR-2F typing was highly instructive or necessary in 41% (156/385) of the cases. Several pre- and posttransplant scenarios were identified as being better served by HR-2F typing. Five different categories are presented with specific case examples. The experience of another center (Temple University Hospital) is also included, whereby 21% of the cases required HR-2F typing by Sanger sequencing, as supported by other legacy methods, to properly address posttransplant anti-HLA antibody issues.

Integrative genomics identifies LMO1 as a neuroblastoma oncogene.

Neuroblastoma is a childhood cancer of the sympathetic nervous system that accounts for approximately 10% of all paediatric oncology deaths. To identify genetic risk factors for neuroblastoma, we performed a genome-wide association study (GWAS) on 2,251 patients and 6,097 control subjects of European ancestry from four case series. Here we report a significant association within LIM domain only 1 (LMO1) at 11p15.4 (rs110419, combined P = 5.2 × 10(-16), odds ratio of risk allele = 1.34 (95% confidence interval 1.25-1.44)). The signal was enriched in the subset of patients with the most aggressive form of the disease. LMO1 encodes a cysteine-rich transcriptional regulator, and its paralogues (LMO2, LMO3 and LMO4) have each been previously implicated in cancer. In parallel, we analysed genome-wide DNA copy number alterations in 701 primary tumours. We found that the LMO1 locus was aberrant in 12.4% through a duplication event, and that this event was associated with more advanced disease (P < 0.0001) and survival (P = 0.041). The germline single nucleotide polymorphism (SNP) risk alleles and somatic copy number gains were associated with increased LMO1 expression in neuroblastoma cell lines and primary tumours, consistent with a gain-of-function role in tumorigenesis. Short hairpin RNA (shRNA)-mediated depletion of LMO1 inhibited growth of neuroblastoma cells with high LMO1 expression, whereas forced expression of LMO1 in neuroblastoma cells with low LMO1 expression enhanced proliferation. These data show that common polymorphisms at the LMO1 locus are strongly associated with susceptibility to developing neuroblastoma, but also may influence the likelihood of further somatic alterations at this locus, leading to malignant progression.

Large sample size, wide variant spectrum, and advanced machine-learning technique boost risk prediction for inflammatory bowel disease.

We performed risk assessment for Crohn's disease (CD) and ulcerative colitis (UC), the two common forms of inflammatory bowel disease (IBD), by using data from the International IBD Genetics Consortium's Immunochip project. This data set contains ~17,000 CD cases, ~13,000 UC cases, and ~22,000 controls from 15 European countries typed on the Immunochip. This custom chip provides a more comprehensive catalog of the most promising candidate variants by picking up the remaining common variants and certain rare variants that were missed in the first generation of GWAS. Given this unprecedented large sample size and wide variant spectrum, we employed the most recent machine-learning techniques to build optimal predictive models. Our final predictive models achieved areas under the curve (AUCs) of 0.86 and 0.83 for CD and UC, respectively, in an independent evaluation. To our knowledge, this is the best prediction performance ever reported for CD and UC to date.

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.

Common genetic variants on 5p14.1 associate with autism spectrum disorders.

Autism spectrum disorders (ASDs) represent a group of childhood neurodevelopmental and neuropsychiatric disorders characterized by deficits in verbal communication, impairment of social interaction, and restricted and repetitive patterns of interests and behaviour. To identify common genetic risk factors underlying ASDs, here we present the results of genome-wide association studies on a cohort of 780 families (3,101 subjects) with affected children, and a second cohort of 1,204 affected subjects and 6,491 control subjects, all of whom were of European ancestry. Six single nucleotide polymorphisms between cadherin 10 (CDH10) and cadherin 9 (CDH9)-two genes encoding neuronal cell-adhesion molecules-revealed strong association signals, with the most significant SNP being rs4307059 (P = 3.4 x 10(-8), odds ratio = 1.19). These signals were replicated in two independent cohorts, with combined P values ranging from 7.4 x 10(-8) to 2.1 x 10(-10). Our results implicate neuronal cell-adhesion molecules in the pathogenesis of ASDs, and represent, to our knowledge, the first demonstration of genome-wide significant association of common variants with susceptibility to ASDs.

Autism genome-wide copy number variation reveals ubiquitin and neuronal genes.

Autism spectrum disorders (ASDs) are childhood neurodevelopmental disorders with complex genetic origins. Previous studies focusing on candidate genes or genomic regions have identified several copy number variations (CNVs) that are associated with an increased risk of ASDs. Here we present the results from a whole-genome CNV study on a cohort of 859 ASD cases and 1,409 healthy children of European ancestry who were genotyped with approximately 550,000 single nucleotide polymorphism markers, in an attempt to comprehensively identify CNVs conferring susceptibility to ASDs. Positive findings were evaluated in an independent cohort of 1,336 ASD cases and 1,110 controls of European ancestry. Besides previously reported ASD candidate genes, such as NRXN1 (ref. 10) and CNTN4 (refs 11, 12), several new susceptibility genes encoding neuronal cell-adhesion molecules, including NLGN1 and ASTN2, were enriched with CNVs in ASD cases compared to controls (P = 9.5 x 10(-3)). Furthermore, CNVs within or surrounding genes involved in the ubiquitin pathways, including UBE3A, PARK2, RFWD2 and FBXO40, were affected by CNVs not observed in controls (P = 3.3 x 10(-3)). We also identified duplications 55 kilobases upstream of complementary DNA AK123120 (P = 3.6 x 10(-6)). Although these variants may be individually rare, they target genes involved in neuronal cell-adhesion or ubiquitin degradation, indicating that these two important gene networks expressed within the central nervous system may contribute to the genetic susceptibility of ASD.

Evaluating the role of the FUS/TLS-related gene EWSR1 in amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease affecting motor neurons. Mutations in related RNA-binding proteins TDP-43, FUS/TLS and TAF15 have been connected to ALS. These three proteins share several features, including the presence of a bioinformatics-predicted prion domain, aggregation-prone nature in vitro and in vivo and toxic effects when expressed in multiple model systems. Given these commonalities, we hypothesized that a related protein, EWSR1 (Ewing sarcoma breakpoint region 1), might also exhibit similar properties and therefore could contribute to disease. Here, we report an analysis of EWSR1 in multiple functional assays, including mutational screening in ALS patients and controls. We identified three missense variants in EWSR1 in ALS patients, which were absent in a large number of healthy control individuals. We show that disease-specific variants affect EWSR1 localization in motor neurons. We also provide multiple independent lines of in vitro and in vivo evidence that EWSR1 has similar properties as TDP-43, FUS and TAF15, including aggregation-prone behavior in vitro and ability to confer neurodegeneration in Drosophila. Postmortem analysis of sporadic ALS cases also revealed cytoplasmic mislocalization of EWSR1. Together, our studies highlight a potential role for EWSR1 in ALS, provide a collection of functional assays to be used to assess roles of additional RNA-binding proteins in disease and support an emerging concept that a class of aggregation-prone RNA-binding proteins might contribute broadly to ALS and related neurodegenerative diseases.

A genome-wide meta-analysis of six type 1 diabetes cohorts identifies multiple associated loci.

Diabetes impacts approximately 200 million people worldwide, of whom approximately 10% are affected by type 1 diabetes (T1D). The application of genome-wide association studies (GWAS) has robustly revealed dozens of genetic contributors to the pathogenesis of T1D, with the most recent meta-analysis identifying in excess of 40 loci. To identify additional genetic loci for T1D susceptibility, we examined associations in the largest meta-analysis to date between the disease and ∼2.54 million SNPs in a combined cohort of 9,934 cases and 16,956 controls. Targeted follow-up of 53 SNPs in 1,120 affected trios uncovered three new loci associated with T1D that reached genome-wide significance. The most significantly associated SNP (rs539514, P = 5.66×10⁻¹¹) resides in an intronic region of the LMO7 (LIM domain only 7) gene on 13q22. The second most significantly associated SNP (rs478222, P = 3.50×10⁻9 resides in an intronic region of the EFR3B (protein EFR3 homolog B) gene on 2p23; however, the region of linkage disequilibrium is approximately 800 kb and harbors additional multiple genes, including NCOA1, C2orf79, CENPO, ADCY3, DNAJC27, POMC, and DNMT3A. The third most significantly associated SNP (rs924043, P = 8.06×10⁻9 lies in an intergenic region on 6q27, where the region of association is approximately 900 kb and harbors multiple genes including WDR27, C6orf120, PHF10, TCTE3, C6orf208, LOC154449, DLL1, FAM120B, PSMB1, TBP, and PCD2. These latest associated regions add to the growing repertoire of gene networks predisposing to T1D.

A genome-wide study reveals copy number variants exclusive to childhood obesity cases.

The prevalence of obesity in children and adults in the United States has increased dramatically over the past decade. Genomic copy number variations (CNVs) have been strongly implicated in subjects with extreme obesity and coexisting developmental delay. To complement these previous studies, we addressed CNVs in common childhood obesity by examining children with a BMI in the upper 5(th) percentile but excluding any subject greater than three standard deviations from the mean in order to reduce severe cases in the cohort. We performed a whole-genome CNV survey of our cohort of 1080 defined European American (EA) childhood obesity cases and 2500 lean controls (< 50(th) percentile BMI) who were genotyped with 550,000 SNP markers. Positive findings were evaluated in an independent African American (AA) cohort of 1479 childhood obesity cases and 1575 lean controls. We identified 17 CNV loci that were unique to at least three EA cases and were both previously unreported in the public domain and validated via quantitative PCR. Eight of these loci (47.1%) also replicated exclusively in AA cases (six deletions and two duplications). Replicated deletion loci consisted of EDIL3, S1PR5, FOXP2, TBCA, ABCB5, and ZPLD1, whereas replicated duplication loci consisted of KIF2B and ARL15. We also observed evidence for a deletion at the EPHA6-UNQ6114 locus when the AA cohort was investigated as a discovery set. Although these variants may be individually rare, our results indicate that CNVs contribute to the genetic susceptibility of common childhood obesity in subjects of both European and African ancestry.

Variants of DENND1B associated with asthma in children.

BACKGROUND: Asthma is a complex disease that has genetic and environmental causes. The genetic factors associated with susceptibility to asthma remain largely unknown. METHODS: We carried out a genomewide association study involving children with asthma. The sample included 793 North American children of European ancestry with persistent asthma who required daily inhaled glucocorticoid therapy and 1988 matched controls (the discovery set). We also tested for genomewide association in an independent cohort of 917 persons of European ancestry who had asthma and 1546 matched controls (the replication set). Finally, we tested for an association between 20 single-nucleotide polymorphisms (SNPs) at chromosome 1q31 and asthma in 1667 North American children of African ancestry who had asthma and 2045 ancestrally matched controls. RESULTS: In our meta-analysis of all samples from persons of European ancestry, we observed an association, with genomewide significance, between asthma and SNPs at the previously reported locus on 17q21 and an additional eight SNPs at a novel locus on 1q31. The SNP most strongly associated with asthma was rs2786098 (P=8.55x10(-9)). We observed replication of the association of asthma with SNP rs2786098 in the independent series of persons of European ancestry (combined P=9.3x10(-11)). The alternative allele of each of the eight SNPs on chromosome 1q31 was strongly associated with asthma in the children of African ancestry (P=1.6x10(-13) for the comparison across all samples). The 1q31 locus contains the 1q31 locus contains DENND1B, a gene expressed by natural killer cells and dendritic cells. DENND1B protein is predicted to interact with the tumor necrosis factor α receptor [corrected]. CONCLUSIONS: We have identified a locus containing DENND1B on chromosome 1q31.3 that is associated with susceptibility to asthma.

A genome-wide association study identifies KIAA0350 as a type 1 diabetes gene.

Type 1 diabetes (T1D) in children results from autoimmune destruction of pancreatic beta cells, leading to insufficient production of insulin. A number of genetic determinants of T1D have already been established through candidate gene studies, primarily within the major histocompatibility complex but also within other loci. To identify new genetic factors that increase the risk of T1D, we performed a genome-wide association study in a large paediatric cohort of European descent. In addition to confirming previously identified loci, we found that T1D was significantly associated with variation within a 233-kb linkage disequilibrium block on chromosome 16p13. This region contains KIAA0350, the gene product of which is predicted to be a sugar-binding, C-type lectin. Three common non-coding variants of the gene (rs2903692, rs725613 and rs17673553) in strong linkage disequilibrium reached genome-wide significance for association with T1D. A subsequent transmission disequilibrium test replication study in an independent cohort confirmed the association. These results indicate that KIAA0350 might be involved in the pathogenesis of T1D and demonstrate the utility of the genome-wide association approach in the identification of previously unsuspected genetic determinants of complex traits.

Strong synaptic transmission impact by copy number variations in schizophrenia.

Schizophrenia is a psychiatric disorder with onset in late adolescence and unclear etiology characterized by both positive and negative symptoms, as well as cognitive deficits. To identify copy number variations (CNVs) that increase the risk of schizophrenia, we performed a whole-genome CNV analysis on a cohort of 977 schizophrenia cases and 2,000 healthy adults of European ancestry who were genotyped with 1.7 million probes. Positive findings were evaluated in an independent cohort of 758 schizophrenia cases and 1,485 controls. The Gene Ontology synaptic transmission family of genes was notably enriched for CNVs in the cases (P = 1.5 x 10(-7)). Among these, CACNA1B and DOC2A, both calcium-signaling genes responsible for neuronal excitation, were deleted in 16 cases and duplicated in 10 cases, respectively. In addition, RET and RIT2, both ras-related genes important for neural crest development, were significantly affected by CNVs. RET deletion was exclusive to seven cases, and RIT2 deletions were overrepresented common variant CNVs in the schizophrenia cases. Our results suggest that novel variations involving the processes of synaptic transmission contribute to the genetic susceptibility of schizophrenia.

In silico replication of the genome-wide association results of the Type 1 Diabetes Genetics Consortium.

Recently, the Type 1 Diabetes Genetics Consortium (T1DGC) reported 22 novel type 1 diabetes (T1D)-associated loci identified by meta-analysis of three genome-wide association studies (GWASs) with a case-control design. However, the association of 10 of these 22 reported loci was not confirmed in the T1DGC family cohort (P > 0.1). To address concerns about potential bias from population stratification, this study aims to replicate the association in three independent GWAS cohorts, one of which was based on the stratification-proof transmission disequilibrium analysis. Three European-descent population samples were included in this study: 483 cases and both parents, a case-control cohort of 514 cases and 2027 controls, and an additional cohort of 1078 cases and 341 controls from the dbGaP database. Among the 22 SNPs reported by the T1DGC, we had high-quality genotypes for 15; the remaining were imputed. T1D association was replicated in seven loci after Bonferroni correction for 22 independent hypotheses. An additional eight loci had nominal (one-sided) significance of P < 0.1 in the same direction, giving a false discovery rate of 3.35%. The genetic susceptibility conferred by non-HLA loci in our family cohort with one affected offspring was higher than the T1DGC multiplex families. Reciprocally, the frequency of strongly predisposing HLA alleles in the multiplex families was higher. This study replicated T1D association with at least as many of these novel loci as expected from the power of our sample size, thus supporting the validity of the new discoveries.

Comparative genetic analysis of inflammatory bowel disease and type 1 diabetes implicates multiple loci with opposite effects.

Inflammatory bowel disease, including Crohn's disease (CD) and ulcerative colitis (UC), and type 1 diabetes (T1D) are autoimmune diseases that may share common susceptibility pathways. We examined known susceptibility loci for these diseases in a cohort of 1689 CD cases, 777 UC cases, 989 T1D cases and 6197 shared control subjects of European ancestry, who were genotyped by the Illumina HumanHap550 SNP arrays. We identified multiple previously unreported or unconfirmed disease associations, including known CD loci (ICOSLG and TNFSF15) and T1D loci (TNFAIP3) that confer UC risk, known UC loci (HERC2 and IL26) that confer T1D risk and known UC loci (IL10 and CCNY) that confer CD risk. Additionally, we show that T1D risk alleles residing at the PTPN22, IL27, IL18RAP and IL10 loci protect against CD. Furthermore, the strongest risk alleles for T1D within the major histocompatibility complex (MHC) confer strong protection against CD and UC; however, given the multi-allelic nature of the MHC haplotypes, sequencing of the MHC locus will be required to interpret this observation. These results extend our current knowledge on genetic variants that predispose to autoimmunity, and suggest that many loci involved in autoimmunity may be under a balancing selection due to antagonistic pleiotropic effect. Our analysis implies that variants with opposite effects on different diseases may facilitate the maintenance of common susceptibility alleles in human populations, making autoimmune diseases especially amenable to genetic dissection by genome-wide association studies.

Diverse genome-wide association studies associate the IL12/IL23 pathway with Crohn Disease.

Previous genome-wide association (GWA) studies typically focus on single-locus analysis, which may not have the power to detect the majority of genuinely associated loci. Here, we applied pathway analysis using Affymetrix SNP genotype data from the Wellcome Trust Case Control Consortium (WTCCC) and uncovered significant association between Crohn Disease (CD) and the IL12/IL23 pathway, harboring 20 genes (p = 8 x 10(-5)). Interestingly, the pathway contains multiple genes (IL12B and JAK2) or homologs of genes (STAT3 and CCR6) that were recently identified as genuine susceptibility genes only through meta-analysis of several GWA studies. In addition, the pathway contains other susceptibility genes for CD, including IL18R1, JUN, IL12RB1, and TYK2, which do not reach genome-wide significance by single-marker association tests. The observed pathway-specific association signal was subsequently replicated in three additional GWA studies of European and African American ancestry generated on the Illumina HumanHap550 platform. Our study suggests that examination beyond individual SNP hits, by focusing on genetic networks and pathways, is important to unleashing the true power of GWA studies.

High-resolution mapping and analysis of copy number variations in the human genome: a data resource for clinical and research applications.

We present a database of copy number variations (CNVs) detected in 2026 disease-free individuals, using high-density, SNP-based oligonucleotide microarrays. This large cohort, comprised mainly of Caucasians (65.2%) and African-Americans (34.2%), was analyzed for CNVs in a single study using a uniform array platform and computational process. We have catalogued and characterized 54,462 individual CNVs, 77.8% of which were identified in multiple unrelated individuals. These nonunique CNVs mapped to 3272 distinct regions of genomic variation spanning 5.9% of the genome; 51.5% of these were previously unreported, and >85% are rare. Our annotation and analysis confirmed and extended previously reported correlations between CNVs and several genomic features such as repetitive DNA elements, segmental duplications, and genes. We demonstrate the utility of this data set in distinguishing CNVs with pathologic significance from normal variants. Together, this analysis and annotation provides a useful resource to assist with the assessment of CNVs in the contexts of human variation, disease susceptibility, and clinical molecular diagnostics.

Chromosome 6p22 locus associated with clinically aggressive neuroblastoma.

BACKGROUND: Neuroblastoma is a malignant condition of the developing sympathetic nervous system that most commonly affects young children and is often lethal. Its cause is not known. METHODS: We performed a genomewide association study by first genotyping blood DNA samples from 1032 patients with neuroblastoma and 2043 control subjects of European descent using the Illumina HumanHap550 BeadChip. Samples from three independent groups of patients with neuroblastoma (a total of 720 patients) and 2128 control subjects were then genotyped to replicate significant associations. RESULTS: We observed a significant association between neuroblastoma and the common minor alleles of three consecutive single-nucleotide polymorphisms (SNPs) at chromosome band 6p22 and containing the predicted genes FLJ22536 and FLJ44180 (P=1.71x10(-9) to 7.01x10(-10); allelic odds ratio, 1.39 to 1.40). Homozygosity for the at-risk G allele of the most significantly associated SNP, rs6939340, resulted in an increased likelihood of the development of neuroblastoma (odds ratio, 1.97; 95% confidence interval, 1.58 to 2.45). Subsequent genotyping of the three 6p22 SNPs in three independent case series confirmed our observation of an association (P=9.33x10(-15) at rs6939340 for joint analysis). Patients with neuroblastoma who were homozygous for the risk alleles at 6p22 were more likely to have metastatic (stage 4) disease (P=0.02), amplification of the MYCN oncogene in the tumor cells (P=0.006), and disease relapse (P=0.01). CONCLUSIONS: A common genetic variation at chromosome band 6p22 is associated with susceptibility to neuroblastoma.

The role of height-associated loci identified in genome wide association studies in the determination of pediatric stature.

BACKGROUND: Human height is considered highly heritable and correlated with certain disorders, such as type 2 diabetes and cancer. Despite environmental influences, genetic factors are known to play an important role in stature determination. A number of genetic determinants of adult height have already been established through genome wide association studies. METHODS: To examine 51 single nucleotide polymorphisms (SNPs) corresponding to the 46 previously reported genomic loci for height in 8,184 European American children with height measurements. We leveraged genotyping data from our ongoing GWA study of height variation in children in order to query the 51 SNPs in this pediatric cohort. RESULTS: Sixteen of these SNPs yielded at least nominally significant association to height, representing fifteen different loci including EFEMP1-PNPT1, GPR126, C6orf173, SPAG17, Histone class 1, HLA class III and GDF5-UQCC. Other loci revealed no evidence for association, including HMGA1 and HMGA2. For the 16 associated variants, the genotype score explained 1.64% of the total variation for height z-score. CONCLUSION: Among 46 loci that have been reported to associate with adult height to date, at least 15 also contribute to the determination of height in childhood.

A genome-wide association study identifies a locus for nonsyndromic cleft lip with or without cleft palate on 8q24.

OBJECTIVE: To identify, in a non-hypothesis manner, novel genetic factors associated with nonsyndromic cleft lip with or without cleft palate (NSCL/P). STUDY DESIGN: We performed a genome-wide association study in a pediatric cohort of European decent consisting of 111 NSCL/P cases and 5951 control subjects. All subjects were consecutively recruited from the Greater Philadelphia area from 2006 to 2009. High throughput genome-wide single nucleotide polymorphism genotyping was carried out with the Illumina Infinium II HumanHap550 BeadChip technology. RESULTS: We observed association at the genome-wide significance level with SNP rs987525 at a locus on 8q24, which harbors no characterized genes to date (P = 9.18 x 10(-8); odds ratio = 2.09, 95% confidence interval = 1.59 to 2.76). While searching for a replication cohort, the same genetic determinant was established through a genome-wide association study of NSCL/P in Germany, so this previous report acts as a de novo replication for our independent observation outlined here. CONCLUSIONS: These results strongly suggest that a locus on 8q24 is involved in the pathogenesis of NSCL/P.

Novel, complex interruptions of the GAA repeat in small, expanded alleles of two affected siblings with late-onset Friedreich ataxia.

Friedreich ataxia (FA) is an autosomal recessive disorder associated with expanded GAA repeats in intron 1 of the FRDA gene. Two siblings presented with a mild form of FA at >60 years of age. Both had a large expansion (>600 repeats) and a small expansion (120 repeats) by long-range PCR. Sequence analysis of the small allele revealed multiple, complex interruptions in the GAA repeat. These 2 patients presented later than predicted from their allele size alone, when compared with a large cohort of FA patients. Accounting for the interruptions in the GAA repeat, though, did not make the age of onset consistent with that noted in other patients. Three additional patients with late onset FA and small expanded alleles also exhibited interrupted GAA repeats that were not associated with inappropriately late onset. Our observations suggest that interrupted GAA repeats do not clearly impact the age of onset in FA.