Demographic Data:
For quality control, we removed twelve individuals with a diagnosis of Down syndrome, five individuals with more than two large CNVs, and one individual with a CNV larger than 45 MB. After quality control, genotype data was available for 997 ASD individuals and WES data for 808 ASD individuals (Table 1). Of these probands, 70% were male, and males tended to have a lower age of ASD diagnosis (Table 1).
Table 1
Genetic characterization of probands with ASD
| Genotyped Probands | | Sequenced Probands | | Intersection |
Total (n) | 997 | | 808 | | 689 |
Average diagnosis age (SD) | 8.2 (8.5) | | 7.9 (8.3) | | 8 (8.5) |
Females (n, (%)) | 299 (30%) | | 229 (28%) | | 189 (28%) |
Average diagnosis age (SD) | 8.9 (9.3) | | 8.6 (8.9) | | 9 (9.4) |
Males (n, (%)) | 698 (70%) | | 579 (72%) | | 500 (72%) |
Average diagnosis age (SD) | 7.9 (8.1) | | 7.6 (8) | | 7.6 (8.1) |
Of the comorbidities and birth characteristics for the population of ASD probands (Table 2), ID was most common (46%), and epilepsy was second (31%). Individuals with congenital anomalies had the lowest age of ASD diagnosis, while individuals with SZ had the highest age of ASD diagnosis. Comorbidities and birth characteristics of the probands which were not genotyped or sequenced are presented in Table S4.
Table 2
Demographics of Comorbidities and Birth Characteristics
| Genotyped Probands (N = 997) | | Sequenced Probands (N = 808) |
Comorbidities and Birth Characteristics | n (%) | Average diagnosis age for AD (SD) | | n (%) | Average diagnosis age for AD (SD) |
ID | 474 (47%) | 6.5 (7) | | 381 (47%) | 6.5 (7.1) |
Epilepsy | 309 (31%) | 5.5 (7.2) | | 225 (31%) | 5.5 (7.2) |
ADHD | 203 (20%) | 8.4 (7.4) | | 160 (20%) | 7.7 (7.2) |
Scholastic skill disorders | 155 (15%) | 5.1 (5.6) | | 121 (15%) | 5.2 (6.2) |
Speech/language disorders | 133 (13%) | 4.8 (5.8) | | 102 (13%) | 4.6 (5.1) |
OCD + anxiety disorder | 100 (10%) | 9.2 (7.3) | | 85 (10%) | 9.7 (7.7) |
HC – small | 87 (11%) | 4.6 (5.9) | | 73 (11%) | 4 (4.4) |
SZ | 86 (9%) | 13.9(10.2) | | 67 (8%) | 11.7 (9.8) |
Congenital anomalies | 74 (11%) | 2.9 (4.9) | | 57 (10%) | 3.1 (4.7) |
Motor function disorders | 55 (5%) | 4.8 (5) | | 43 (5%) | 4.6 (4.8) |
HC – large | 40 (5%) | 7.9 (7.3) | | 30 (5%) | 8.4 (7.7) |
Large for gestational age | 38 (5%) | 7.6 (7.7) | | 30 (4%) | 5.2 (5.7) |
Small for gestational age | 37 (5%) | 4.1 (6.2) | | 35 (5%) | 4.3 (6.3) |
Missing values for congenital anomalies (Genotyped Probands /Sequenced Probands) 336/260, large for gestational age 207/139, small for gestational age 207/139, head – small 220/153, head – large 220/153 individuals. ADHD: attention-deficit/hyperactivity disorder, SZ: schizophrenia (SZ), OCD: obsessive-compulsive disorder, ID: intellectual disability. |
Genetic findings
Of the 997 individuals who were genotyped, 104 (10.4%) carried one or more likely pathogenic CNVs (Table 3, Table S2). Fourteen probands had two likely pathogenic CNVs each, for a total of 118 likely pathogenic CNVs in the population sample; 65 of these were deletions and 53 were duplications, and they ranged in size from 217 kb to 43 Mb (median 3.6 Mb). There were 53 CNVs considered to be recurrent genomic disorders. Those loci with more than one overlapping likely pathogenic CNV/genomic disorder in different probands are described in Table 3; see Table S2 for full results.
Table 3
Known genomic disorders with more than one case identified, based on CNV findings
Chromosomal Disorder | Number Identified |
15q11q13 PWS BP1-BP3 class 1 | 8 (6 dup, 2 del) |
15q13.3 microdeletion syndrome BP4 - BP5 | 5 (2 dup, 3 del) |
16p13.11 syndrome MYH11 | 5 (2 dup, 3 del) |
22q11.2 DGS typical AD 3 MB | 5 (1 dup, 4 del) |
16p11.2 typical BP5-BP5 TBX6 | 4 (1 dup, 3 del) |
1q21 recurrent region | 3 (dup) |
2q37 monosomy | 3 (del) |
Phelan-McDermid Syndrome (22q13 deletion syndrome) | 2 (del) |
16p12.2 recurrent region (EEF2K) | 2 (del) |
17q12 RCAD syndrome (HNF1B) | 2 (dup) |
Of the 808 individuals for whom WES was performed, 69 (8.8%) had a likely pathogenic variant. and no individuals had more than one (Table S3). Of these SNVs, 34 were predicted loss of function (frameshift, nonsense, splice acceptor or donor), and the remaining 35 were deleterious missense variants. Genes with the highest frequency of suspected likely pathogenic findings were GRIN2B (n = 6), POGZ (n = 5), SATB1 (n = 4), DYNC1H1 (n = 4), and CREBBP (n = 3). Two individuals had likely significant variants in each of the following genes: CACNA1E, CHD8, DIP2A, FOXP1, RORB, SETD5, STXBP1, SUV420H1 (also referred to as KMT5B), and SYNGAP1. One individual had likely significant variants in each of the following genes: ADNP, ANK2, ARID1B, ASXL3, CELF4, CHD2, DNMT3A, DSCAM, FOXP2, GABRB2, GIGYF1, HECTD4, KCNMA1, KDM6B, LRRC4C, MYT1L, NACC1, NR3C2, NUP155, PHF21A, PPP1R9B, PPP2R5D, SCN1A, SCN2A, SHANK3, SIN3A, TCF4, TEK, and ZMYND8. In total, 168 individuals had a likely pathogenic CNV or SNV (5 individuals had both a CNV and SNV).
Comorbidities and Birth Characteristics of the Probands
Evaluating medical and psychiatric comorbidities among individuals with ASD (Tables 4 and 5), the likely pathogenic CNV and SNV groups showed slightly different average ages of ASD diagnosis by group and by sex, although none of these differences were significant (P value > 0.05). Of the phenotypes of individuals with likely pathogenic CNVs and SNVs (Table 5), ID was the most common disorder.
Table 4
Characteristics of individuals with significant CNVs and SNVs
| Genotyped Probands Individuals with likely pathogenic CNV n (%) | | Sequenced Probands Individuals with likely pathogenic SNV n (%) | | Intersection Individuals with likely pathogenic SNV or CNV n (%) |
Total (%) | 104 (10.4%) | | 69 (8.5%) | | 121 (17.6%) |
Average diagnosis age (SD) | 7.1 (8.1) | | 7.3 (7.3) | | 6.7 (7.4) |
Females (n, (%)) | 37 (35%) | | 20 (30%) | | 35 (29%) |
Average diagnosis age (SD) | 7.8 (9.2) | | 8.2 (7) | | 6.2 (6.6) |
Males (n, (%)) | 67 (65%) | | 49 (70%) | | 86 (71%) |
Average diagnosis age (SD) | 6.8 (7.6) | | 6.9 (7.5) | | 7 (7.7) |
Table 5
Comorbidities and birth characteristics of individuals with ASD
Phenotypes | Genotyped Probands Individuals with likely pathogenic CNV (N = 104) n (%) | Sequenced Probands Individuals with likely pathogenic SNV (N = 69) n (%) | Intersection Individuals with likely pathogenic CNV/SNV (N = 121) n (%) |
ID | 58 (55%) | 47 (68%) | 75 (50%) |
Epilepsy | 34 (32%) | 30 (43%) | 49 (32%) |
Scholastic skill disorders | 25 (24%) | 11 (16%) | 33 (11%) |
ADHD | 21 (20%) | 11 (16%) | 23 (15%) |
Speech/language disorders | 18 (17%) | 7 (10%) | 16 (7%) |
Congenital anomalies | 18 (17%) | 3 (4%) | 15 (10%) |
HC - small | 13 (12%) | 4 (5.7%) | 14 (9%) |
SZ | 12 (11%) | 4 (6%) | 14 (9%) |
OCD + Anxiety | 10 (10%) | 4 (6%) | 10 (8%) |
Motor function disorders | 8 (8%) | 2 (3%) | 7 (5%) |
Small for gestational age | 7 (7%) | 0 (0%) | 5 (3%) |
HC - big | 4 (4%) | 4 (5.7%) | 6 (4%) |
Large for gestational age | 2 (2%) | 3 (2%) | 1 (1%) |
Missing values for congenital anomalies (Genotyped Probands /Sequenced Probands) 336/260, large for gestational age 207/139, small for gestational age 207/139, head – small 220/153, head – large 220/153 individuals. ADHD: attention-deficit/hyperactivity disorder, SZ: schizophrenia (SZ), OCD: obsessive-compulsive disorder, ID: intellectual disability. |
Congenital anomalies and scholastic skill disorders were associated with having likely pathogenic CNVs (Table 6) with an odds ratio (OR) of 1.85 (95%CI, 1.12,2.97), and 3.05 (95%CI, 1.64,5.47), respectively (Table 6). For probands carrying a damaging SNV, versus those who did not, ID and epilepsy showed a significant positive association (Table 6). In general, these patterns were similar when carrier status for either a CNV or SNV was assessed (Table 6).
Table 6
Odds ratios for comorbidities and birth characteristics of individuals with ASD
Phenotypes | Genotyped Probands Individuals with Likely pathogenic CNV | | Sequenced Probands Individuals with Likely pathogenic SNV | | Intersection Individuals with Likely pathogenic CNV/SNV |
| OR | 95% CI | P value | | OR | 95% CI | P value | | OR | 95% CI | P value |
Congenital anomalies | 3.05 | (1.64,5.47) | 0.0002* | | 0.57 | (0.14,1.64) | 0.36 | | 1.63 | (0.84,3.03) | 0.13 |
Small for gestational age | 2.10 | (0.82,4.71) | 0.09 | | 0 | - | - | | 1.01 | (0.33,2.52) | 0.98 |
Scholastic skill disorders | 1.85 | (1.12,2.97) | 0.01* | | 1.08 | (0.52,2.05) | 0.82 | | 1.98 | (1.19,3.21) | 0.007* |
HC - small | 1.63 | (0.82,3.01) | 0.14 | | 0.67 | (0.20,1.73) | 0.46 | | 1.36 | (0.70,2.52) | 0.34 |
Motor function disorders | 1.51 | (0.64,3.12) | 0.3 | | 0.51 | (0.08,1.70) | 0.36 | | 1.33 | (0.52,3.00) | 0.51 |
ID | 1.45 | (0.96,2.18) | 0.07 | | 2.6 | (1.55,4.48) | 0.0003* | | 2.31 | (1.55,3.47) | 5e-05* |
SZ | 1.42 | (0.71,2.62) | 0.29 | | 0.66 | (0.20,1.66) | 0.43 | | 1.48 | (0.76,2.72) | 0.23 |
Speech/language disorders | 1.39 | (0.79,2.35) | 0.23 | | 0.77 | (0.31,1.62) | 0.52 | | 1.14 | (0.62,2.00) | 0.66 |
Epilepsy | 1.08 | (0.69,1.65) | 0.73 | | 1.76 | (1.06,2.89) | 0.03* | | 1.63 | (1.08,2.44) | 0.02* |
HC - big | 1.01 | (0.30,2.62) | 0.99 | | 0.86 | (0.53,5.02) | 0.27 | | 1.39 | (0.50,3.36) | 0.49 |
ADHD | 0.99 | (0.58,1.61) | 0.97 | | 0.75 | (0.37,1.41) | 0.4 | | 1.05 | (0.62,1.70) | 0.86 |
OCD + Anxiety | 0.93 | (0.44,1.77) | 0.84 | | 0.5 | (0.15,1.25) | 0.19 | | 0.74 | (0.35,1.43) | 0.4 |
Large for Gestational age | 0.47 | (0.08,1.58) | 0.31 | | 0 | - | - | | 0.16 | (0.01,0.77) | 0.07 |
Missing values for congenital anomalies (Genotyped Probands /Sequenced Probands) 336/260, large for gestational age 207/139, small for gestational age 207/139, head – small 220/153, head – large 220/153 individuals. ADHD: attention-deficit/hyperactivity disorder, SZ: schizophrenia (SZ), OCD: obsessive-compulsive disorder, ID: intellectual disability. |
We next compared the effect of carrier status for ASD subjects who do or do not manifest ID for the largest group of genetically characterized subjects, those who were genotyped (Table 7). Although ASD subjects who had a likely pathogenic CNV were more likely to have ID, ASD subjects with and without ID showed no significant differences in the association of carrier status with other, potentially associated phenotypes (P value > 0.05 for all tests). Thus, conditioning on ID status does not explain much of the variation for other CNV-related associations. For instance, the risk for congenital abnormalities are similar for likely pathogenic CNV carriers whether or not they meet criteria for ID, 2.46 versus 3.87 (Table 7).
Table 7
Comparison of individuals with and without ID
Phenotypes | Genotyped Probands without ID (N = 523) Individuals with Likely pathogenic CNV (N = 46, 8.8%) | | Genotyped Probands with ID (N = 474) Individuals with Likely pathogenic CNV (N = 58, 12.2%) |
| OR | 95% CI | P value | | OR | 95% CI | P value |
Congenital anomalies | 3.87 | (1.41,9.66) | 0.005* | | 2.46 | (1.09,5.26) | 0.02* |
Speech/language disorders | 2.19 | (0.94,4.66) | 0.05* | | 0.92 | (0.41,1.89) | 0.83 |
Small for gestational age | 2.17 | (0.48,7.08) | 0.24 | | 2.01 | (0.55,5.81) | 0.23 |
Scholastic skill disorders | 2.06 | (0.92,4.25) | 0.06 | | 1.61 | (0.83,3.00) | 0.14 |
HC - big | 1.89 | (0.43,5.99) | 0.33 | | 0.42 | (0.02,2.11) | 0.4 |
OCD + Anxiety | 1.58 | (0.65,3.41) | 0.27 | | 0.39 | (0.06,1.33) | 0.2 |
SZ | 1.24 | (0.41,3.09) | 0.66 | | 1.63 | (0.63,3.68) | 0.27 |
Epilepsy | 1.23 | (0.58,2.45) | 0.57 | | 0.86 | (0.49,1.51) | 0.61 |
ADHD | 1.14 | (0.54,2.26) | 0.72 | | 0.89 | (0.41,1.78) | 0.76 |
Large for gestational age | 0.92 | (0.14,3.31) | 0.91 | | 0 | - | - |
Motor function disorders | 0.86 | (0.13,3.02) | 0.84 | | 1.99 | (0.71,4.86) | 0.15 |
HC - small | 0.26 | (0.01,1.25) | 0.19 | | 2.85 | (1.31,5.94) | 0.006* |
Data for sleeping disorders, hypotonia, birth defects, prenatal growth rate, gestational age in weeks, weight and height at birth, and Apgar scores were underpowered due to a high number of missing values. ADHD: attention-deficit/hyperactivity disorder, SZ: schizophrenia (SZ), OCD: obsessive-compulsive disorder, ID: intellectual disability. |