Interaction between Continuous Pack-Years Smoked and Polygenic Risk Score on Lung Cancer Risk: Prospective Results from the Framingham Heart Study.

BACKGROUND: Lung cancer risk attributable to smoking is dose dependent, yet few studies examining a polygenic risk score (PRS) by smoking interaction have included comprehensive lifetime pack-years smoked. METHODS: We analyzed data from participants of European ancestry in the Framingham Heart Study Original (n = 454) and Offspring (n = 2,470) cohorts enrolled in 1954 and 1971, respectively, and followed through 2018. We built a PRS for lung cancer using participant genotyping data and genome-wide association study summary statistics from a recent study in the OncoArray Consortium. We used Cox proportional hazards regression models to assess risk and the interaction between pack-years smoked and genetic risk for lung cancer adjusting for European ancestry, age, sex, and education. RESULTS: We observed a significant submultiplicative interaction between pack-years and PRS on lung cancer risk (P = 0.09). Thus, the relative risk associated with each additional 10 pack-years smoked decreased with increasing genetic risk (HR = 1.56 at one SD below mean PRS, HR = 1.48 at mean PRS, and HR = 1.40 at one SD above mean PRS). Similarly, lung cancer risk per SD increase in the PRS was highest among those who had never smoked (HR = 1.55) and decreased with heavier smoking (HR = 1.32 at 30 pack-years). CONCLUSIONS: These results suggest the presence of a submultiplicative interaction between pack-years and genetics on lung cancer risk, consistent with recent findings. Both smoking and genetics were significantly associated with lung cancer risk. IMPACT: These results underscore the contributions of genetics and smoking on lung cancer risk and highlight the negative impact of continued smoking regardless of genetic risk.

Pharmacogenetics of tuberculosis treatment toxicity and effectiveness in a large Brazilian cohort.

Background: Genetic polymorphisms have been associated with risk of anti-tuberculosis treatment toxicity. We characterized associations with adverse events and treatment failure/recurrence among adults treated for tuberculosis in Brazil. Methods: Participants were followed in Regional Prospective Observational Research in Tuberculosis (RePORT)-Brazil. We included persons with culture-confirmed drug-susceptible pulmonary tuberculosis who started treatment between 2015-2019, and who were evaluable for pharmacogenetics. Treatment included 2 months of isoniazid, rifampin or rifabutin, pyrazinamide, and ethambutol, then 4 months of isoniazid and rifampin or rifabutin, with 24 month follow-up. Analyses included 43 polymorphisms in 20 genes related to anti-tuberculosis drug hepatotoxicity or pharmacokinetics. Whole exome sequencing was done in a case-control toxicity subset. Results: Among 903 participants in multivariable genetic association analyses, NAT2 slow acetylator status was associated with increased risk of treatment-related grade 2 or greater adverse events, including hepatotoxicity. Treatment failure/recurrence was more likely among NAT2 rapid acetylators, but not statistically significant at the 5% level. A GSTM1 polymorphism (rs412543) was associated with increased risk of treatment-related adverse events, including hepatotoxicity. SLCO1B1 polymorphisms were associated with increased risk of treatment- related hepatoxicity and treatment failure/recurrence. Polymorphisms in NR1/2 were associated with decreased risk of adverse events and increased risk of failure/recurrence. In whole exome sequencing, hepatotoxicity was associated with a polymorphism in VTI1A , and the genes METTL17 and PRSS57 , but none achieved genome-wide significance. Conclusions: In a clinical cohort representing three regions of Brazil, NAT2 acetylator status was associated with risk for treatment-related adverse events. Additional significant polymorphisms merit investigation in larger study populations.

Rare genetic variation implicated in non-Hispanic white families with Alzheimer disease.

OBJECTIVE: To identify genetic variation influencing late-onset Alzheimer disease (LOAD), we used a large data set of non-Hispanic white (NHW) extended families multiply-affected by LOAD by performing whole genome sequencing (WGS). METHODS: As part of the Alzheimer Disease Sequencing Project, WGS data were generated for 197 NHW participants from 42 families (affected individuals and unaffected, elderly relatives). A two-pronged approach was taken. First, variants were prioritized using heterogeneity logarithm of the odds (HLOD) and family-specific LOD scores as well as annotations based on function, frequency, and segregation with disease. Second, known Alzheimer disease (AD) candidate genes were assessed for rare variation using a family-based association test. RESULTS: We identified 41 rare, predicted-damaging variants that segregated with disease in the families that contributed to the HLOD or family-specific LOD regions. These included a variant in nitric oxide synthase 1 adaptor protein that segregates with disease in a family with 7 individuals with AD, as well as variants in RP11-433J8, ABCA1, and FISP2. Rare-variant association identified 2 LOAD candidate genes associated with disease in these families: FERMT2 (p-values = 0.001) and SLC24A4 (p-value = 0.009). These genes still showed association while controlling for common index variants, indicating the rare-variant signal is distinct from common variation that initially identified the genes as candidates. CONCLUSIONS: We identified multiple genes with putative damaging rare variants that segregate with disease in multiplex AD families and showed that rare variation may influence AD risk at AD candidate genes. These results identify novel AD candidate genes and show a role for rare variation in LOAD etiology, even at genes previously identified by common variation.

Whole genome sequencing of Caribbean Hispanic families with late-onset Alzheimer's disease.

OBJECTIVE: To identify rare causal variants underlying known loci that segregate with late-onset Alzheimer's disease (LOAD) in multiplex families. METHODS: We analyzed whole genome sequences (WGS) from 351 members of 67 Caribbean Hispanic (CH) families from Dominican Republic and New York multiply affected by LOAD. Members of 67 CH and additional 47 Caucasian families underwent WGS as a part of the Alzheimer's Disease Sequencing Project (ADSP). All members of 67 CH families, an additional 48 CH families and an independent CH case-control cohort were subsequently genotyped for validation. Patients met criteria for LOAD, and controls were determined to be dementia free. We investigated rare variants segregating within families and gene-based associations with disease within LOAD GWAS loci. RESULTS: A variant in AKAP9, p.R434W, segregated significantly with LOAD in two large families (OR = 5.77, 95% CI: 1.07-30.9, P = 0.041). In addition, missense mutations in MYRF and ASRGL1 under previously reported linkage peaks at 7q14.3 and 11q12.3 segregated completely in one family and in follow-up genotyping both were nominally significant (P < 0.05). We also identified rare variants in a number of genes associated with LOAD in prior genome wide association studies, including CR1 (P = 0.049), BIN1 (P = 0.0098) and SLC24A4 (P = 0.040). CONCLUSIONS AND RELEVANCE: Rare variants in multiple genes influence the risk of LOAD disease in multiplex families. These results suggest that rare variants may underlie loci identified in genome wide association studies.

Genome-wide linkage analyses of non-Hispanic white families identify novel loci for familial late-onset Alzheimer's disease.

INTRODUCTION: Few high penetrance variants that explain risk in late-onset Alzheimer's disease (LOAD) families have been found. METHODS: We performed genome-wide linkage and identity-by-descent (IBD) analyses on 41 non-Hispanic white families exhibiting likely dominant inheritance of LOAD, and having no mutations at known familial Alzheimer's disease (AD) loci, and a low burden of APOE ε4 alleles. RESULTS: Two-point parametric linkage analysis identified 14 significantly linked regions, including three novel linkage regions for LOAD (5q32, 11q12.2-11q14.1, and 14q13.3), one of which replicates a genome-wide association LOAD locus, the MS4A6A-MS4A4E gene cluster at 11q12.2. Five of the 14 regions (3q25.31, 4q34.1, 8q22.3, 11q12.2-14.1, and 19q13.41) are supported by strong multipoint results (logarithm of odds [LOD*] ≥1.5). Nonparametric multipoint analyses produced an additional significant locus at 14q32.2 (LOD* = 4.18). The 1-LOD confidence interval for this region contains one gene, C14orf177, and the microRNA Mir_320, whereas IBD analyses implicates an additional gene BCL11B, a regulator of brain-derived neurotrophic signaling, a pathway associated with pathogenesis of several neurodegenerative diseases. DISCUSSION: Examination of these regions after whole-genome sequencing may identify highly penetrant variants for familial LOAD.

Targeted massively parallel sequencing of autism spectrum disorder-associated genes in a case control cohort reveals rare loss-of-function risk variants.

BACKGROUND: Autism spectrum disorder (ASD) is highly heritable, yet genome-wide association studies (GWAS), copy number variation screens, and candidate gene association studies have found no single factor accounting for a large percentage of genetic risk. ASD trio exome sequencing studies have revealed genes with recurrent de novo loss-of-function variants as strong risk factors, but there are relatively few recurrently affected genes while as many as 1000 genes are predicted to play a role. As such, it is critical to identify the remaining rare and low-frequency variants contributing to ASD. METHODS: We have utilized an approach of prioritization of genes by GWAS and follow-up with massively parallel sequencing in a case-control cohort. Using a previously reported ASD noise reduction GWAS analyses, we prioritized 837 RefSeq genes for custom targeting and sequencing. We sequenced the coding regions of those genes in 2071 ASD cases and 904 controls of European white ancestry. We applied comprehensive annotation to identify single variants which could confer ASD risk and also gene-based association analysis to identify sets of rare variants associated with ASD. RESULTS: We identified a significant over-representation of rare loss-of-function variants in genes previously associated with ASD, including a de novo premature stop variant in the well-established ASD candidate gene RBFOX1. Furthermore, ASD cases were more likely to have two damaging missense variants in candidate genes than controls. Finally, gene-based rare variant association implicates genes functioning in excitatory neurotransmission and neurite outgrowth and guidance pathways including CACNAD2, KCNH7, and NRXN1. CONCLUSIONS: We find suggestive evidence that rare variants in synaptic genes are associated with ASD and that loss-of-function mutations in ASD candidate genes are a major risk factor, and we implicate damaging mutations in glutamate signaling receptors and neuronal adhesion and guidance molecules. Furthermore, the role of de novo mutations in ASD remains to be fully investigated as we identified the first reported protein-truncating variant in RBFOX1 in ASD. Overall, this work, combined with others in the field, suggests a convergence of genes and molecular pathways underlying ASD etiology.

Age of onset of amyotrophic lateral sclerosis is modulated by a locus on 1p34.1.

Amyotrophic lateral sclerosis (ALS) is the third most common adult-onset neurodegenerative disease. Individuals with ALS rapidly progress to paralysis and die from respiratory failure within 3 to 5 years after symptom onset. Epidemiological factors explain only a modest amount of the risk for ALS. However, there is growing evidence of a strong genetic component to both familial and sporadic ALS risk. The International Consortium on Amyotrophic Lateral Sclerosis Genetics was established to bring together existing genome-wide association cohorts and identify sporadic ALS susceptibility and age at symptom onset loci. Here, we report the results of a meta-analysis of the International Consortium on Amyotrophic Lateral Sclerosis Genetics genome-wide association samples, consisting of 4243 ALS cases and 5112 controls from 13 European ancestry cohorts from across the United States and Europe. Eight genomic regions provided evidence of association with ALS, including 9p21.2 (rs3849942, odds ratio [OR] = 1.21; p = 4.41 × 10(-7)), 17p11.2 (rs7477, OR = 1.30; p = 2.89 × 10(-7)), and 19p13 (rs12608932, OR = 1.37, p = 1.29 × 10(-7)). Six genomic regions were associated with age at onset of ALS. The strongest evidence for an age of onset locus was observed at 1p34.1, with comparable evidence at rs3011225 (R(2)(partial) = 0.0061; p = 6.59 × 10(-8)) and rs803675 (R(2)(partial) = 0.0060; p = 6.96 × 10(-8)). These associations were consistent across all 13 cohorts. For rs3011225, individuals with at least 1 copy of the minor allele had an earlier average age of onset of over 2 years. Identifying the underlying pathways influencing susceptibility to and age at onset of ALS may provide insight into the pathogenic mechanisms and motivate new pharmacologic targets for this fatal neurodegenerative disease.

Evaluation of copy number variations reveals novel candidate genes in autism spectrum disorder-associated pathways.

Autism spectrum disorders (ASDs) are highly heritable, yet relatively few associated genetic loci have been replicated. Copy number variations (CNVs) have been implicated in autism; however, the majority of loci contribute to <1% of the disease population. Therefore, independent studies are important to refine associated CNV regions and discover novel susceptibility genes. In this study, a genome-wide SNP array was utilized for CNV detection by two distinct algorithms in a European ancestry case-control data set. We identify a significantly higher burden in the number and size of deletions, and disrupting more genes in ASD cases. Moreover, 18 deletions larger than 1 Mb were detected exclusively in cases, implicating novel regions at 2q22.1, 3p26.3, 4q12 and 14q23. Case-specific CNVs provided further evidence for pathways previously implicated in ASDs, revealing new candidate genes within the GABAergic signaling and neural development pathways. These include DBI, an allosteric binder of GABA receptors, GABARAPL1, the GABA receptor-associated protein, and SLC6A11, a postsynaptic GABA transporter. We also identified CNVs in COBL, deletions of which cause defects in neuronal cytoskeleton morphogenesis in model vertebrates, and DNER, a neuron-specific Notch ligand required for cerebellar development. Moreover, we found evidence of genetic overlap between ASDs and other neurodevelopmental and neuropsychiatric diseases. These genes include glutamate receptors (GRID1, GRIK2 and GRIK4), synaptic regulators (NRXN3, SLC6A8 and SYN3), transcription factor (ZNF804A) and RNA-binding protein FMR1. Taken together, these CNVs may be a few of the missing pieces of ASD heritability and lead to discovering novel etiological mechanisms.

Evidence of novel fine-scale structural variation at autism spectrum disorder candidate loci.

BACKGROUND: Autism spectrum disorders (ASD) represent a group of neurodevelopmental disorders characterized by a core set of social-communicative and behavioral impairments. Gamma-aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the brain, acting primarily via the GABA receptors (GABR). Multiple lines of evidence, including altered GABA and GABA receptor expression in autistic patients, indicate that the GABAergic system may be involved in the etiology of autism. METHODS: As copy number variations (CNVs), particularly rare and de novo CNVs, have now been implicated in ASD risk, we examined the GABA receptors and genes in related pathways for structural variation that may be associated with autism. We further extended our candidate gene set to include 19 genes and regions that had either been directly implicated in the autism literature or were directly related (via function or ancestry) to these primary candidates. For the high resolution CNV screen we employed custom-designed 244 k comparative genomic hybridization (CGH) arrays. Collectively, our probes spanned a total of 11 Mb of GABA-related and additional candidate regions with a density of approximately one probe every 200 nucleotides, allowing a theoretical resolution for detection of CNVs of approximately 1 kb or greater on average. One hundred and sixty-eight autism cases and 149 control individuals were screened for structural variants. Prioritized CNV events were confirmed using quantitative PCR, and confirmed loci were evaluated on an additional set of 170 cases and 170 control individuals that were not included in the original discovery set. Loci that remained interesting were subsequently screened via quantitative PCR on an additional set of 755 cases and 1,809 unaffected family members. RESULTS: Results include rare deletions in autistic individuals at JAKMIP1, NRXN1, Neuroligin4Y, OXTR, and ABAT. Common insertion/deletion polymorphisms were detected at several loci, including GABBR2 and NRXN3. Overall, statistically significant enrichment in affected vs. unaffected individuals was observed for NRXN1 deletions. CONCLUSIONS: These results provide additional support for the role of rare structural variation in ASD.

An X chromosome-wide association study in autism families identifies TBL1X as a novel autism spectrum disorder candidate gene in males.

BACKGROUND: Autism spectrum disorder (ASD) is a complex neurodevelopmental disorder with a strong genetic component. The skewed prevalence toward males and evidence suggestive of linkage to the X chromosome in some studies suggest the presence of X-linked susceptibility genes in people with ASD. METHODS: We analyzed genome-wide association study (GWAS) data on the X chromosome in three independent autism GWAS data sets: two family data sets and one case-control data set. We performed meta- and joint analyses on the combined family and case-control data sets. In addition to the meta- and joint analyses, we performed replication analysis by using the two family data sets as a discovery data set and the case-control data set as a validation data set. RESULTS: One SNP, rs17321050, in the transducin ß-like 1X-linked (TBL1X) gene [OMIM:300196] showed chromosome-wide significance in the meta-analysis (P value = 4.86 × 10-6) and joint analysis (P value = 4.53 × 10-6) in males. The SNP was also close to the replication threshold of 0.0025 in the discovery data set (P = 5.89 × 10-3) and passed the replication threshold in the validation data set (P = 2.56 × 10-4). Two other SNPs in the same gene in linkage disequilibrium with rs17321050 also showed significance close to the chromosome-wide threshold in the meta-analysis. CONCLUSIONS: TBL1X is in the Wnt signaling pathway, which has previously been implicated as having a role in autism. Deletions in the Xp22.2 to Xp22.3 region containing TBL1X and surrounding genes are associated with several genetic syndromes that include intellectual disability and autistic features. Our results, based on meta-analysis, joint analysis and replication analysis, suggest that TBL1X may play a role in ASD risk.

Copy number variants in extended autism spectrum disorder families reveal candidates potentially involved in autism risk.

Copy number variations (CNVs) are a major cause of genetic disruption in the human genome with far more nucleotides being altered by duplications and deletions than by single nucleotide polymorphisms (SNPs). In the multifaceted etiology of autism spectrum disorders (ASDs), CNVs appear to contribute significantly to our understanding of the pathogenesis of this complex disease. A unique resource of 42 extended ASD families was genotyped for over 1 million SNPs to detect CNVs that may contribute to ASD susceptibility. Each family has at least one avuncular or cousin pair with ASD. Families were then evaluated for co-segregation of CNVs in ASD patients. We identified a total of five deletions and seven duplications in eleven families that co-segregated with ASD. Two of the CNVs overlap with regions on 7p21.3 and 15q24.1 that have been previously reported in ASD individuals and two additional CNVs on 3p26.3 and 12q24.32 occur near regions associated with schizophrenia. These findings provide further evidence for the involvement of ICA1 and NXPH1 on 7p21.3 in ASD susceptibility and highlight novel ASD candidates, including CHL1, FGFBP3 and POUF41. These studies highlight the power of using extended families for gene discovery in traits with a complex etiology.

A de novo 1.5 Mb microdeletion on chromosome 14q23.2-23.3 in a patient with autism and spherocytosis.

Autism is a neuro-developmental disorder characterized by deficits in social interaction and communication as well as restricted interests or repetitive behaviors. Cytogenetic studies have implicated large chromosomal aberrations in the etiology of approximately 5-7% of autism patients, and the recent advent of array-based techniques allows the exploration of submicroscopic copy number variations (CNVs). We genotyped a 14-year-old boy with autism, spherocytosis and other physical dysmorphia, his parents, and two non-autistic siblings with the Illumina Human 1M Beadchip as part of a study of the molecular genetics of autism and determined copy number variants using the PennCNV algorithm. We identified and validated a de novo 1.5 Mb microdeletion of 14q23.2-23.3 in our autistic patient. This region contains 15 genes, including spectrin beta (SPTB), encoding a cytoskeletal protein previously associated with spherocytosis, methylenetetrahydrofolate dehydrogenase 1 (MTHFD1), a folate metabolizing enzyme previously associated with bipoloar disorder and schizophrenia, pleckstrin homology domain-containing family G member 3 (PLEKHG3), a guanide nucleotide exchange enriched in the brain, and churchill domain containing protein 1 (CHURC1), homologs of which regulate neuronal development in model organisms. While a similar deletion has previously been reported in a family with spherocytosis, severe learning disabilities, and mild mental retardation, this is the first implication of chr14q23.2-23.3 in the etiology of autism and points to MTHFD1, PLEKHG3, and CHURC1 as potential candidate genes contributing to autism risk.

A noise-reduction GWAS analysis implicates altered regulation of neurite outgrowth and guidance in autism.

BACKGROUND: Genome-wide Association Studies (GWAS) have proved invaluable for the identification of disease susceptibility genes. However, the prioritization of candidate genes and regions for follow-up studies often proves difficult due to false-positive associations caused by statistical noise and multiple-testing. In order to address this issue, we propose the novel GWAS noise reduction (GWAS-NR) method as a way to increase the power to detect true associations in GWAS, particularly in complex diseases such as autism. METHODS: GWAS-NR utilizes a linear filter to identify genomic regions demonstrating correlation among association signals in multiple datasets. We used computer simulations to assess the ability of GWAS-NR to detect association against the commonly used joint analysis and Fisher's methods. Furthermore, we applied GWAS-NR to a family-based autism GWAS of 597 families and a second existing autism GWAS of 696 families from the Autism Genetic Resource Exchange (AGRE) to arrive at a compendium of autism candidate genes. These genes were manually annotated and classified by a literature review and functional grouping in order to reveal biological pathways which might contribute to autism aetiology. RESULTS: Computer simulations indicate that GWAS-NR achieves a significantly higher classification rate for true positive association signals than either the joint analysis or Fisher's methods and that it can also achieve this when there is imperfect marker overlap across datasets or when the closest disease-related polymorphism is not directly typed. In two autism datasets, GWAS-NR analysis resulted in 1535 significant linkage disequilibrium (LD) blocks overlapping 431 unique reference sequencing (RefSeq) genes. Moreover, we identified the nearest RefSeq gene to the non-gene overlapping LD blocks, producing a final candidate set of 860 genes. Functional categorization of these implicated genes indicates that a significant proportion of them cooperate in a coherent pathway that regulates the directional protrusion of axons and dendrites to their appropriate synaptic targets. CONCLUSIONS: As statistical noise is likely to particularly affect studies of complex disorders, where genetic heterogeneity or interaction between genes may confound the ability to detect association, GWAS-NR offers a powerful method for prioritizing regions for follow-up studies. Applying this method to autism datasets, GWAS-NR analysis indicates that a large subset of genes involved in the outgrowth and guidance of axons and dendrites is implicated in the aetiology of autism.

Identification of chromosome 7 inversion breakpoints in an autistic family narrows candidate region for autism susceptibility.

Chromosomal breaks and rearrangements have been observed in conjunction with autism and autistic spectrum disorders. A chromosomal inversion has been previously reported in autistic siblings, spanning the region from approximately 7q22.1 to 7q31. This family is distinguished by having multiple individuals with autism and associated disabilities. The region containing the inversion has been strongly implicated in autism by multiple linkage studies, and has been particularly associated with language defects in autism as well as in other disorders with language components. Mapping of the inversion breakpoints by FISH has localized the inversion to the region spanning approximately 99-108.75 Mb of chromosome 7. The proximal breakpoint has the potential to disrupt either the coding sequence or regulatory regions of a number of cytochrome P450 genes while the distal region falls in a relative gene desert. Copy number variant analysis of the breakpoint regions detected no duplication or deletion that could clearly be associated with disease status. Association analysis in our autism data set using single nucleotide polymorphisms located near the breakpoints showed no significant association with proximal breakpoint markers, but has identified markers near the distal breakpoint ( approximately 108-110 Mb) with significant associations to autism. The chromosomal abnormality in this family strengthens the case for an autism susceptibility gene in the chromosome 7q22-31 region and targets a candidate region for further investigation.

A genome-wide association study of autism reveals a common novel risk locus at 5p14.1.

Although autism is one of the most heritable neuropsychiatric disorders, its underlying genetic architecture has largely eluded description. To comprehensively examine the hypothesis that common variation is important in autism, we performed a genome-wide association study (GWAS) using a discovery dataset of 438 autistic Caucasian families and the Illumina Human 1M beadchip. 96 single nucleotide polymorphisms (SNPs) demonstrated strong association with autism risk (p-value < 0.0001). The validation of the top 96 SNPs was performed using an independent dataset of 487 Caucasian autism families genotyped on the 550K Illumina BeadChip. A novel region on chromosome 5p14.1 showed significance in both the discovery and validation datasets. Joint analysis of all SNPs in this region identified 8 SNPs having improved p-values (3.24E-04 to 3.40E-06) than in either dataset alone. Our findings demonstrate that in addition to multiple rare variations, part of the complex genetic architecture of autism involves common variation.

Investigation of potential gene-gene interactions between APOE and RELN contributing to autism risk.

BACKGROUND: Several candidate gene studies support RELN as susceptibility gene for autism. Given the complex inheritance pattern of autism, it is expected that gene-gene interactions will exist. A logical starting point for examining potential gene-gene interactions is to evaluate the joint effects of genes involved in a common biological pathway. RELN shares a common biological pathway with APOE, and Persico et al. have observed transmission distortion of the APOE-2 allele in autism families. OBJECTIVE: We evaluated RELN and APOE for joint effects in autism susceptibility. METHODS: A total of 470 Caucasian autism families were analyzed (265 multiplex; 168 trios with no family history; 37 positive family history but only one sampled affected). These families were genotyped for 11 RELN polymorphisms, including the 5' untranslated region repeat previously associated with autism, as well as for the APOE functional allele. We evaluated single locus allelic and genotypic association with the pedigree disequilibrium test and geno-PDT, respectively. Multilocus effects were evaluated using the extended version of the multifactorial dimensionality reduction method. RESULTS: For the single locus analyses, there was no evidence for an effect of APOE in our data set. Evidence for association with RELN (rs2,073,559; trio subset P=0.07 PDT; P=0.001 geno-PDT; overall geno-PDT P=0.05), however, was found. For multilocus geno-PDT analysis, the joint genotype of APOE and RELN rs2,073,559 was highly significant (trio subset, global P=0.0001), probably driven by the RELN single locus effect. Using the extended version of the multifactorial dimensionality reduction method to detect multilocus effects, there were no statistically significant associations for any of the n-locus combinations involving RELN or APOE in the overall or multiplex subset. In the trio subset, 1-locus and 2-locus models selected only markers in RELN as best models for predicting autism case status. CONCLUSION: Thus, we conclude that there is no main effect of APOE in our autism data set, nor is there any evidence for a joint effect of APOE with RELN. RELN, however, remains a good candidate for autism susceptibility.

Lack of association between autism and SLC25A12.

OBJECTIVE: Autism has a strong, complex genetic component, most likely involving several genes. Multiple genomic screens have shown evidence suggesting linkage to chromosome 2q31-q33, which includes the SLC25A12 gene. Recently, an association between autism risk and two single nucleotide polymorphisms (SNPs) in SLC25A12 was reported. This study aimed to test for association in SLC25A12 in an independent data set of 327 families with autistic offspring. METHOD: The authors analyzed two SNPs that were significant in the previous study group, as well as seven additional SNPs within the gene. Association analyses for individual SNPs as well as haplotypes were performed. RESULTS: There was no evidence of an association between SLC25A12 and autism. CONCLUSIONS: These results suggest that SLC25A12 is not a major contributor to autism risk in these families.

Analysis of the autism chromosome 2 linkage region: GAD1 and other candidate genes.

Autism has a strong and complex genetic component, involving several genes. Genomic screens, including our own, have shown suggestive evidence for linkage over a 20-30 cM region on chromosome 2q31-q33. Two subsequent reports showed that the linkage evidence increased in the subset of families with phrase speech delay (PSD), defined as onset of phrase speech later than 3 years of age. To further investigate the linkage in the presumptive candidate region, microsatellite markers in a 2 cM grid covering the interval from 164 to 203 cM were analyzed in 110 multiplex (2 or more sampled autism patients) families. A maximum heterogeneity LOD (HLOD) score of 1.54 was detected at D2S1776 (173 cM) in the overall dataset (dominant model), increasing to 1.71 in the PSD subset. While not conclusive, these data continue to provide suggestive evidence for linkage, particularly considering replication by multiple independent groups. Positive LOD scores extended over the entire region, continuing to define a broad candidate interval. Association studies were performed on several functional candidates mapping within the region. These included GAD1, encoding GAD67, whose levels are reduced in autopsy brain material from autistic subjects, and STK17B, ABI2, CTLA4, CD28, NEUROD1, PDE1A, HOXD1 and DLX2. We found no evidence for significant allelic association between autism and any of these candidates, suggesting that they do not play a major role in the genetics of autism or that substantial allelic heterogeneity at any one of these loci dilutes potential disease-allele association.