US20100248235A1 - Biomarkers for autism spectrum disorders - Google Patents

Biomarkers for autism spectrum disorders Download PDF

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US20100248235A1
US20100248235A1 US12/681,229 US68122908A US2010248235A1 US 20100248235 A1 US20100248235 A1 US 20100248235A1 US 68122908 A US68122908 A US 68122908A US 2010248235 A1 US2010248235 A1 US 2010248235A1
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Stephen W. Scherer
John B. Vincent
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Definitions

  • the present invention relates to genetic markers for Autism Spectrum Disorders (ASD).
  • ASSD Autism Spectrum Disorders
  • Autism is a heritable neurodevelopmental condition characterized by impairments in social communication and by a preference for repetitive activities. Autism is not a distinct categorical disorder but is the prototype of a group of conditions defined as Pervasive Developmental Disorders (PDDs) or Autism Spectrum Disorders (ASD), which include Asperger's Disorder, Childhood Disintegrative Disorder, Pervasive developmental disorder-not otherwise specified (PDD-NOS) and Rett Syndrome.
  • PDDs Pervasive Developmental Disorders
  • ASD Autism Spectrum Disorders
  • ASD is diagnosed in families of all racial, ethnic and social-economic backgrounds with incidence roughly four times higher in males compared to females. Overall population prevalence of autism has increased in recent years to a current estimate of 20 in 10,000 with incidence as high as 60 in 10,000 for all autism spectrum disorders.
  • autism spectrum disorders are among the most heritable complex disorders, the genetic risk is clearly not conferred in simple Mendelian fashion.
  • autism is part of a broader recognizable disorder (e.g. fragile X syndrome, tuberous sclerosis) or is associated with cytogenetically-detectable chromosome abnormalities.
  • a broader recognizable disorder e.g. fragile X syndrome, tuberous sclerosis
  • cytogenetically-detectable chromosome abnormalities e.g. co-morbidity of autism with microdeletion syndromes (e.g. William-Beuren and Sotos) and other genomic disorders (e.g. Prader-Willi/Angelman) suggests chromosomal imbalances are involved in the underlying etiology.
  • the most frequent cytogenetic anomaly is an interstitial, maternally-inherited duplication of 15q11-13 (1-3%) encompassing the Prader Willi/Angelman Syndrome critical region.
  • the 22q11.2 region is associated with velo-cardio-facial Syndrome and deletions at 22q13.3 appear to also represent a clinically definable syndrome. Both deletions are associated with the autistic phenotypes.
  • Other chromosome loci associated with anomalies with a higher frequency of events observed in syndromic forms of ASD include 7q (see TCAG www.chr7.org), 2q37, 5p14-15, 17p11.2.
  • reciprocal duplications overlapping the William-Beuren deletion region have been associated with the autism phenotype.
  • CNVs large scale copy number variants
  • markers have now been identified which are useful in assessing the risk of ASD in an individual, as well as being useful to diagnose the condition.
  • the markers are useful both individually and in the form of a microarray to screen individuals for risk of ASD.
  • a method of determining the risk of ASD in an individual comprising:
  • nucleic acid-containing sample obtained from the individual for a gene encoding PTCHD1, wherein a determination that the gene comprises a deletion of at least a portion of exon 1 is indicative of a risk of ASD in the individual.
  • a method of determining the risk of ASD in an individual comprising:
  • nucleic acid-containing sample obtained from the individual for a mutation that modulates the expression of at least one gene selected from the group consisting of PTCHD1, SHANK3, NFIA, DPP6, DPP10, GPR98, PQBP1, ZNF41 and FTSJ1, wherein identification of a mutation that modulates the expression of at least one of said genes is indicative of a risk of ASD.
  • a method of determining the risk of ASD in an individual comprising:
  • a method of determining the risk of ASD in an individual comprising:
  • FIG. 1 is a flow chart depicting the methodology used to identify ASD-specific CNVs
  • FIG. 2 illustrates a genome-wide distribution of ASD-specific CNVs as described in Table 3;
  • FIG. 3 illustrates the chromosome 16p11.2 region as depicted in the Autism Chromosome Rearrangement Database
  • FIG. 4 illustrates examples of CNVs observed in ASD families including probands having multiple de novo events (a); rearrangements in the SHANK3 gene (b); probands with chromosome X deletions (at PTCHD1) from female carriers (c) or inherited translocations in addition to an unrelated de novo deletion (d); overlapping events in unrelated probands either de novo (e) or inherited (f) at the DPP6 locus; and recurrent de novo events at chromosome 16p11.2 in unrelated probands either gains (h) or losses (g);
  • FIG. 5 illustrates examples of DPP6 and DPP10 ASD-related CNVs
  • FIG. 6 illustrates examples of chromosome 22q11.2 and 16p11.2 ASD-related CNVs
  • FIG. 7 illustrates the cDNA sequence (A) of the PTCHD1 gene and the corresponding amino acid sequence (B);
  • FIG. 8 illustrates ASD-related missense mutations identified in Table 7.
  • a method of determining the risk of an autism spectrum disorder (ASD) in an individual comprising screening a biological sample obtained from the individual for a mutation that may modulate the expression of at least one gene selected from the group consisting of PTCHD1, SHANK3, NFIA, DPP6, DPP10, DPYD, GPR98, PQBP1, ZNF41 and FTSJ1.
  • ASD-associated genes are referred to herein as “ASD-associated” genes.
  • an autism spectrum disorder or “an ASD” is used herein to refer to at least one condition that results in developmental delay of an individual such as autism, Asperger's Disorder, Childhood Disintegrative Disorder, Pervasive Developmental Disorder-Not Otherwise Specified (PDD-NOS) and Rett Syndrome (APA DSM-IV 2000).
  • a biological sample obtained from the individual is utilized.
  • a suitable biological sample may include, for example, a nucleic acid-containing sample or a protein-containing sample.
  • suitable biological samples include saliva, urine, semen, other bodily fluids or secretions, epithelial cells, cheek cells, hair and the like.
  • invasively-obtained biological samples may also be used in the method, including for example, blood, serum, bone marrow, cerebrospinal fluid (CSF) and tissue biopsies such as tissue from the cerebellum, spinal cord, prostate, stomach, uterus, small intestine and mammary gland samples. Techniques for the invasive process of obtaining such samples are known to those of skill in the art.
  • the present method may also be utilized in prenatal testing for the risk of ASD using an appropriate biological sample such as amniotic fluid and chorionic villus.
  • the biological sample is screened for nucleic acid encoding selected genes in order to detect mutations associated with an ASD. It may be necessary, or preferable, to extract the nucleic acid from the biological sample prior to screening the sample.
  • Methods of nucleic acid extraction are well-known to those of skill in the art and include chemical extraction techniques utilizing phenol-chloroform (Sambrook et al., 1989), guanidine-containing solutions, or CTAB-containing buffers.
  • commercial DNA extraction kits are also widely available from laboratory reagent supply companies, including for example, the QIAamp DNA Blood Minikit available from QIAGEN (Chatsworth, Calif.), or the Extract-N-Amp blood kit available from Sigma (St. Louis, Mo.).
  • nucleic acid sample Once an appropriate nucleic acid sample is obtained, it is subjected to well-established methods of screening, such as those described in the specific examples that follow, to detect genetic mutations indicative of ASD, i.e. ASD-linked mutations.
  • Mutations such as genomic copy number variations (CNVs), which include gains and deletions of segments of DNA, for example, segments of DNA greater than about 1 kb, such as DNA segments between about 300 and 500 kb, as well as base pair mutations such as nonsense, missense and splice site mutations, including sequence mutations in both coding and regulatory regions of a gene, have been found to be indicative of ASD.
  • CNVs genomic copy number variations
  • ASD-linked mutations such as CNVs are not restricted to a single chromosome, but rather have been detected on a multiple chromosomes such as the X chromosome, chromosome 15 and chromosome 21, and on various regions of the same chromosome such as at Xp11 and Xp22.
  • Examples of CNVs that have been determined to be linked to ASD include a deletion on chromosome Xp22 including at least a portion of exon 1 of the PTCHD1 gene; a duplication on chromosome 15q11; and a deletion within the SHANK3 gene.
  • CNVs in the DPP10 gene including intronic gains, such as a 105 kb intronic gain, and exonic losses, such as a 478 kb exonic loss, both of which are more specifically identified in Table 1, have been identified; CNVs in the DPP6 gene, such as a 66 kb loss encompassing exons 2 and 3 and gains such as a CNV encompassing the entire DPP6 gene, a 270 kb exonic gain (exon 1), and a 16 kb intronic gain (see Table 1); CNVs in the SHANK3 gene such as a 276 kb loss; and CNVs in the DYPD gene such as a loss of the entire gene.
  • intronic gains such as a 105 kb intronic gain
  • exonic losses such as a 478 kb exonic loss, both of which are more specifically identified in Table 1
  • CNVs in the DPP6 gene such as a 66 kb loss encompassing exons 2 and 3 and gains such as
  • genomic sequence variations that inhibit the expression of PTCHD1 have been linked to ASD.
  • the terminology “inhibit expression” refers broadly to sequence variations that may inhibit, or at least reduce, any one of transcription and/or translation, as well as the activity of the PTCHD1 protein.
  • a CNV in the PTCHD1 gene comprising a large deletion of the coding region which results in at least a reduction of the expression of PTCHD1 protein has been found to be indicative of ASD.
  • the CNV deletion may include, for example, at least a portion of exon 1, but may additionally include surrounding regions as well, such as intron 1, in whole or in part, or a portion or more of the upstream region thereof.
  • Genomic sequence variations other than CNVs have also been found to be indicative of ASD, including, for example, missense mutations which result in amino acid changes in a protein that may also affect protein expression.
  • missense mutations in the PTCHD1 gene have been identified which are indicative of ASD, including missense mutations resulting in the following amino acid substitutions in the Ptchd1 protein: L73F, I173V, V195I, ML336-337II and E479G.
  • genomic sequencing and profiling using well-established techniques as exemplified herein in the specific examples, may be conducted for an individual to be assessed with respect to ASD risk/diagnosis using a suitable biological sample obtained from the individual. Identification of one or more mutations associated with ASD would be indicative of a risk of ASD, or may be indicative of a diagnosis of ASD. This analysis may be conducted in combination with an evaluation of other characteristics of the individual being assessed, including for example, phenotypic characteristics.
  • a method for determining risk of ASD in an individual in which the expression or activity of a product of an ASD-linked gene mutation is determined in a biological protein-containing sample obtained from the individual.
  • Abnormal levels of the gene product or abnormal levels of the activity thereof, i.e. reduced or elevated levels, in comparison with levels that exist in healthy non-ASD individuals, are indicative of a risk of ASD, or may be indicative of ASD.
  • a determination of the level and/or activity of the gene products of one or more of PTCHD1, SHANK3, NFIA, DPP6, DPP10, DYPD, GPR98, PQBP1, ZNF41 and FTSJ1, may be used to determine the risk of ASD in an individual, or to diagnose ASD.
  • standard assays may be used to identify and quantify the presence and/or activity of a selected gene product.
  • ADI-R Autism Diagnostic Interview-Revised
  • ADOS Autism Diagnostic Observation Schedule
  • GALNACT-2 Affected brother, 4p12: 44,876,353-46,024,486 GABRG1 (breakpoint region Yes/NS MED4, NUDT15, apparently is located in intron 7)
  • SNPs For each sample, approximately 500,000 SNPs were genotyped using the combined two-chip Nspl and Styl GeneChip® Human Mapping Commercial or Early Access Arrays (Affymetrix, Inc., Santa Clara, Calif.) according to the manufacturer's instructions and as described previously (Kennedy et al. 2003 Nat Biotechnol. 21:1233-7, the contents of which are incorporated herein by reference). Briefly, 250 ng of genomic DNA was digested with Nspl and Styl restriction enzyme (New England Biolabs, Boston, Mass.), ligated to an adaptor and amplified by PCR.
  • Nspl and Styl restriction enzyme New England Biolabs, Boston, Mass.
  • PCR products were then fragmented with DNaseI to a size range of 250 bp to 2,000 bp, labelled, and hybridized to the array. After hybridization, arrays were washed on the Affymetrix fluidics stations, stained, and scanned using the Gene Chip Scanner 3000 7G and Gene Chip Operating System. Data has been submitted to the Gene Expression Omnibus database (accession GSE9222). Karyotypes were generated using standard clinical diagnostic protocols.
  • Nspl and Styl array scans were analyzed for copy number variation using a combination of DNA Chip Analyzer (dChip) (Li and Wong 2001 Genome Biology 2: 0032.1-0032.11), Copy Number Analysis for GeneChip (CNAG) (Nannya 2005 Cancer Res. 65:6071-9) and Genotyping Microarray based CNV Analysis (GEMCA) ( Komura 2006 Genome Res. 16:1575-84).
  • dChip DNA Chip Analyzer
  • CNAG Copy Number Analysis for GeneChip
  • GEMCA Genotyping Microarray based CNV Analysis
  • the reference pool was set to include all samples and performed an automatic batch pair-wise analysis using sex-matched controls. Test samples were compared to all samples within the reference pool and matched based on signal intensity standard deviations. The scan intensities for each ‘test’ sample were compared to the average intensities of the reference samples (typically the average of 5-12 samples) and used to calculate raw copy number changes. Underlying copy number changes were then inferred using a Hidden Markov Model (HMM) built into CNAG.
  • HMM Hidden Markov Model
  • CNVs were merged if they were detected in the same individual by more than one algorithm using the outside probe boundaries.
  • Control samples consisted of (i) CNVs observed in 500 Europeans from the from the German PopGen project (Krawczak et al. Community Genet 2006; 9 (1):55-61), and CNVs found in a cohort of 1000 Caucasian non-disease controls from the Ontario population (ref. 24).
  • the ACRD that had 834 putative CNVs or breakpoints mapped to the genome was established.
  • a CNV was considered ASD-specific if it was >10 kb, contained at least three probes and at least 20% of its total length was unique when compared to controls.
  • PCR validation of CNV calls was performed using Quantitative Multiplex PCR of short fluorescent fragments (QMPSF) (Redon et al. Nature. 444:444-54) or SYBR-Green 1 based real-time quantitative PCR (qPCR) using controls at the ACCN1, CFTR or FOXP2 loci (PMID: 14552656).
  • QMPSF Quantitative Multiplex PCR of short fluorescent fragments
  • qPCR real-time quantitative PCR
  • genomic sequences 140-220 bp within putative CNVs were PCR amplified using dye-labelled primers corresponding to unique sequences. Each reaction also included co-amplified control amplicons corresponding to either ACCN1 or CFTR located at 17q11.2 and 7q31.2, respectively. Briefly, 40 ng of genomic DNA was amplified by PCR in a final volume of 25 ⁇ l using AmpliTaq® DNA polymerase (manufactured for Applied Biosystems by Roche Molecular Systems, Inc.) After an initial step of denaturation at 95° C. for 5 minutes conditions were as follows: 25 PCR cycles of 94° C. for 30 seconds, annealing at 60° C.
  • SYBR Green I-based real-time qPCR amplification was performed using a Mx3005P quantitative PCR system (Stratagene, La Jolla, USA). Non-fluorescent primers were designed to amplify short genomic fragments ( ⁇ 140 bp) in putative CNV loci. Each assay also included amplification of a control amplicon corresponding to FOXP2 at 7q31.1 for comparison.
  • test samples were assayed in 15 ⁇ l reaction mixtures in 96-well plates containing: 7.5 ⁇ l of reaction mix, 1.8 ⁇ l of primer, 6.0 ng of genomic DNA at 1.2 ng/ ⁇ l, 0.225 ⁇ l of reference dye with 1:500 dilution, and 0.475 ⁇ l of water.
  • PCR conditions consisted of 10 minutes of polymerase activation at 95° C., followed by 40 cycles of: 95° C. for 15 seconds and a single step at 60° C. for 1 minute for annealing and elongation. These steps were then followed by a final cycle of 95° C.
  • a total of 426 ASD index cases were tested for CNV content including 394 typical idiopathic cases and 32 others that were enrolled based on prior knowledge of having a cytogenetic abnormality.
  • the Affymetrix 500k SNP array was used because it provided the highest resolution screen available for both SNP genotype and CNV data.
  • the ancestry of each sample was categorized (to guide selection of controls). Backgrounds of the samples were found to be: 90.3%, 4.5%, 4.5%, and 0.7%, European, European/mixed, Asian, or Yoruban, respectively.
  • This ‘stringent’ dataset contained 1312 CNVs ( ⁇ 3 CNVs per genome, mean size 603 kb). Using q-PCR, 48% (12/26) and 96% (48/50) of random CNVs were validated in the full and stringent collections, respectively.
  • Karyotyping also provided the ability to characterize the chromosomal context (e.g. ring chromosomes) of some of the CNV regions, something not possible using microarrays alone. Therefore, 313 unbiased idiopathic cases where blood was available were examined and 5.8% (18/313) cases were found to have balanced (11) or unbalanced (7) karyotypes (all unbalanced karyotypic changes (7) were also found by microarray analysis and are included in the CNV statistics).
  • chromosomal context e.g. ring chromosomes
  • Structural variants found in ASD cases were initially prioritized to possibly be etiologic if they were not in controls and, (i) de novo in origin (25 cases) (see Table 5 below), (ii) overlapping (27 cases at 13 loci) in two or more unrelated samples (see Table 7 below), (iii) recurrent (same breakpoints) in two or more unrelated samples (four cases at two loci), (iv) or inherited (the remainder).
  • CNVs were found at known ASD loci: NLGN4 and 22q, 15q, SHANK3 and NRXN1 in categories i, ii, iii, and iv, respectively.
  • ASD structural variants found in controls eg. NRXN1 could also be involved.
  • Probands with abnormal karyotypes (1-14) are separated from probands belonging to simplex (SPX) and multiplex (MPX) families with normal karyotypes (15-25).
  • SPX simplex
  • MPX multiplex
  • 3 De novo event detected by either karyotype (k) or microarray (a)
  • k karyotype
  • a microarray
  • CNV size is based on array results. The breakpoints have not been accurately defined, and CNVs may be smaller or larger than posted.
  • New ASD candidates identified were those with a structural change (either de novo or found in two or more unrelated ASD cases, or for the X chromosome an allele being transmitted maternally from an unaffected carrier) specific to that gene, including ANKRD11, DLGAP2, DPP6, DPP10, DPYD, PCDH9 and PTCHD1 (Tables 5 and 6).
  • ANKRD11, DLGAP2, DPP6, DPP10, DPYD, PCDH9 and PTCHD1 Tables 5 and 6
  • NLGN4, SHANK3 and NRXN1 were also identified.
  • the PCDH9 and NRXN1 genes are also found as CNVs in controls in the DGV (Database of Genomic Variants).
  • Additional positional candidate genes identified were those found interrupted by balanced cytogenetic breakpoints including NEGR1, PIP5K1B, GABRG1, KLHL3, STK3, ST7, SATB2 (Table 1). Moreover, 77 CNVs in the stringent dataset overlapped with the Autism Chromosome Rearrangement Database providing a second line of evidence for involvement ( FIG. 2 ). For example, a 4.6 Mb de novo duplication at Xp11.23-11.22 was detected in a female SK0306-004 (Table 5) and a male in the database.
  • DPP6 and DPP10 emerge as being positional and functional candidates.
  • DPP6 ⁇ 1.5 Mb in size at 2q14.1
  • DPP10 ⁇ 1.3 Mb at 7q36.2 code for accessory trans-membrane dipeptidyl peptidase-like subunits that affect the expression and gating of Kv4.2 channels (KCND2).
  • Kv4.2 channels function in regulation of neurotransmitter release and neuronal excitability in the glutamatergic synapse at the same sites where SHANK3 and the NLGN gene products are found.
  • autism balanced breakpoints have been mapped near KCND2 at 7q31.
  • Structural variants overlapping loci involved in medical genetic conditions including Waardenburg Type IIA (3p14.1), speech and language disorder (7q31), mental retardation (MR) (15q23-q24, 16p11.2) and velocardialfacial syndrome (VCFS) (22q13) were identified (Table 5), amongst others. Identification of the structural variant at these loci led to clinical re-assessment and either identification or refinement of the diagnosis, for additional syndromic features. Other instances (eg. SK0186-PTCHD1 deletion) ( FIG. 4 c ) prompted re-testing of the entire family and eventually a diagnosis of mild-ASD in a previously undiagnosed sibling. This family was then redesignated multiplex as opposed to simplex.
  • FIGS. 4 and 5 A recurrent ⁇ 500 kb duplication at 16p11.2 in two ASD families (SK0102 and NA0133) was also discovered. As with DPP6/DPP10 and 22q11.2, there were carriers of these structural variants without ASD. In a third family (MM0088), the proband has a larger 676 kb de novo deletion and it is only detected in one of two ASD siblings. ( FIG. 4 g ).
  • ASD loci include (i) those that contain genes functioning in the PSD, (ii) and/or chromosomal regions previously shown to be involved in mental retardation, and (iii) involve dysregulation of gene expression.
  • CNVs that implicate ASD loci include the SHANK3, NLGN, and NRXN1-PSD genes and also identify novel loci at DPP6 and DPP10 (amongst others including PCDH9, RPS6KA2, RET from the full dataset) were identified.
  • a genome scan with Affymetrix 500K SNP Arrays was used to identify a CNV deletion on chromosome Xp22.11 that spans exon 1 of the PTCHD1 gene. Exon 1 is shown bolded in FIG. 7 spanning nucleotide positions 1-359.
  • the Cdna sequence of the PTCHD1 gene (NM — 173495) as well as the amino acid sequence of the corresponding encoded protein is illustrated in FIG. 7 which illustrates a genomic size of:: 59325, an exon/coding exon count of 3 encoding a protein of 783 amino acids.
  • the deletion was determined to be an ⁇ 156 kb deletion on Xp22.11 on a male proband.
  • the physical position of this CNV is chrX:22,962,800-23,119,000 (UCSC 2004 Assembly).
  • the deletion is flanked by SNP probes rs7055928 and rs1918560 (at 22.956 and 23.133 Mb from the Xp terminus, respectively).
  • the most proximal and distal SNPs (from the Affymetrix SNP microarrays) within the deleted region, as determined by the SNP microarray analysis, are rs7879064 (23.119 Mb) and rs4828958 (22.972 Mb).
  • PCR amplicons from within the deleted region were used to confirm the deletion by Qper (PCR primers and locations are given below). This deletion spans the entire exon 1 of the PTCHD1 gene (NM — 173495). Analysis of both Sty and Nsp chips data identified this event and was further validated using PCR and QPCR techniques. The following primers were used:
  • PTCHD-CNV1F ATTCGCAGTTCCTTCGTCTT (SEQ ID NO: 1)
  • PTCHD-CNV1R AAAGTGGATTGATCGGTTCC (SEQ ID NO: 2)
  • PTCHD-CNV2F GCTTGAGGACGTGTTTCTCC (SEQ ID NO: 3)
  • PTCHD-CNV2R CTAGGAGAGGTGGCGCTCT (SEQ ID NO: 4)
  • This CNV is autism specific as it was not present in the Database of Genomic Variants (DGV) and in other controls. Furthermore, the segregation of this deletion was characterized in family and it was identified that the deletion was transmitted from a heterozygous mother. A male sibling also had language deficits.
  • PTCHD1-x1F AGCGTGCGCCTCGCCCT (SEQ ID NO: 5) PTCHD1-x1R TCCTTGTCCAGGAGGCTGGGA (SEQ ID NO: 6) PTCHD1-x1Bf GCGCCCGCTCTGCTCTA (SEQ ID NO: 7) PTCHD1-x1Br TCCTTGTCCAGGAGGCTGGGA (SEQ ID NO: 8) PTCHD1-x2-F GAATGTCCACCCTCTCCAAA (SEQ ID NO: 9) PTCHD1-x2-R AAGGCTACTCCTGGCCTTTT (SEQ ID NO: 10) PTCHD1-x3a-F CTTTGACCCAGTAGTCCCTCA (SEQ ID NO: 11) PTCHD1-x3a-R GCACAAACCCCTTGGTGTA (SEQ ID NO: 12) PTCHD1-x3b-F TGTGATTGGGTTTTACATATATGAGTC (SEQ ID NO: 13) PTCHD1-x3
  • the mutation screening revealed an I173V mutation.
  • ADI No family history of PDD FRX and head and ADOS-1 criteria for diagnosis of autism. Severe expressive and CT scan was receptive language delay. No dysmorphology observed. normal Family 5 M ML336-7II Meet ADI and ADOS-1 criteria for diagnosis of autism.
  • Controls (439) were tested for the I173V and V195I mutations, 500 controls for ML336-337II, and 282 controls for L73F and E479G. None of these mutations were present in controls. Furthermore, the fact that these mutations were all maternally inherited to male probands, and were not observed in our control populations, indicates that the mutations are associated with ASD. In turn, it is reasonable to assume that these mutations contribute to the etiology of autism, and perhaps in-combination with other disease-related loci, give rise to the ASD phenotype.
  • DPD dihydropyrimidine dehydrogenase

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WO2012173809A2 (en) * 2011-06-02 2012-12-20 Ehli Erik Method of identifying de novo copy number variants (cnv) using mz twins discordant for attention problems/disorders
WO2012173809A3 (en) * 2011-06-02 2013-04-04 Ehli Erik Method of identifying de novo copy number variants (cnv) using mz twins discordant for attention problems/disorders
WO2016109449A1 (en) * 2014-12-29 2016-07-07 The Board Of Trustees Of The Leland Stanford Junior University Methods of diagnosing autism spectrum disorders
WO2018183525A1 (en) * 2017-03-28 2018-10-04 The Regents Of The University Of California Methods for assessing risk of or diagnosing genetic defects by identifying de novo mutations or somatic mosaic variants in sperm or somatic tissues
CN117025758A (zh) * 2023-09-19 2023-11-10 华北理工大学 环状rna在制备诊断孤独症的生物标志物中的应用

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US10526653B2 (en) 2020-01-07
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US11254984B2 (en) 2022-02-22
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