Copy Number Variants and Fetal Growth in Stillbirths

Abstract

Background:

Fetal growth abnormalities are associated with a higher incidence of stillbirth, with small and large for gestational age infants incurring a 3 to 4, and 2 to 3-fold increased risk, respectively. Although clinical risk factors such as diabetes, hypertension, and placental insufficiency have been associated with fetal growth aberrations and stillbirth, the role of underlying genetic etiologies remains uncertain.

Objective(s):

To assess the relationship between abnormal copy number variants using chromosomal microarray and fetal growth abnormalities in stillbirths.

Study Design:

A secondary analysis utilizing a cohort study design of stillbirths from the Stillbirth Collaborative Research Network was performed. The exposure was defined as abnormal copy number variants, including aneuploidies, pathogenic copy number variants, and variants of unknown clinical significance. The outcomes were small for gestational age and large for gestational age stillbirths, defined as a birthweight less than the 10^th percentile and greater than the 90^th percentile for gestational age, respectively.

Results:

Among 393 stillbirths with chromosomal microarray and birthweight data, 16% had abnormal copy number variants. The small for gestational age outcome was more common among those with abnormal copy number variants compared to those with a normal microarray (29.5% vs. 16.5%, p=0.038). This finding was consistent after adjusting for clinically important variables. In the final model, only abnormal copy number variants and maternal age remained significantly associated with small for gestational age stillbirths with an aOR of 2.22 (95% CI: 1.12 - 4.18). While large for gestational age stillbirths were more likely to have an abnormal microarray: 6.2% vs 3.3% (p=0.275), with an OR 2.35 (95% CI: 0.70 - 7.90), this finding did not reach statistical significance.

Conclusion(s):

Genetic abnormalities are more common in the setting of small for gestational age stillborn fetuses. Abnormal copy number variants not detectable by traditional karyotype make up approximately 50% of the genetic abnormalities in this population.

Introduction:

Stillbirth affects 1 in 170 pregnancies and contributes to 60% of perinatal deaths in the United States.^1,2 Fetal growth abnormalities, such as small for gestational age (SGA), defined as a birthweight less than the 10^th percentile, or large for gestational age (LGA), defined as a birthweight greater than the 90^th percentile, are associated with a 3 to 4 and 2 to 3-times greater risk of stillbirth, respectively.^3,4 Antenatal assessments of infant birthweight, such as ultrasound biometric measurements, are imperfect proxies for SGA and LGA, however they remain the clinical standard. Fetal growth abnormalities share similar risk factors with stillbirth such as advanced maternal age, nulliparity, hypertension, diabetes, obesity, multifetal pregnancies, congenital anomalies, genetic abnormalities, and tobacco use.^5–7 In some clinical conditions such as hypertension, placental insufficiency appears to mediate the relationship between restricted fetal growth and stillbirth.^8–12 However, in many cases, there is no clear etiology for either abnormal fetal growth or stillbirth.

Genetic abnormalities have been previously estimated to contribute to 8-13% of stillbirth causes.^13,14 Well-established genetic causes of stillbirth include chromosomal aneuploidies, such as monosomy X and trisomies 21, 18, and 13.¹⁵ These same aneuploidies, as well as triploidy, are also associated with fetal growth restriction (FGR).^16,17 The interaction between genetic abnormalities and growth restriction can be caused by direct fetal effects, placental effects, or both.¹⁵ Recently, developments in molecular sequencing technologies have provided greater insight into the impact of smaller genomic changes, such as copy number variants (CNVs), monogenic mutations, and single-nucleotide polymorphisms (SNPs), that predispose to FGR and stillbirth.^13,18–21 Contemporary studies suggest that genetic alterations may contribute to 18% of stillbirth cases and 20% of FGR cases with the addition of chromosomal microarray and whole-exome or genome sequencing.^18–23 The genetic causes of fetal overgrowth or macrosomia are not as well-studied. Most known disorders are rare, like Beckwith-Wiedeman syndrome or Persistent hyperinsulinemic hypoglycemia of infancy, and influence insulin and glucose metabolism.²⁴ While some of these disorders have been described, the majority of known conditions leading to fetal overgrowth would not be identified on chromosomal microarray, which is the current standard recommendation in the genetic evaluation of a stillbirth.²⁵ The aim of this research was to test whether an association exists between microarray abnormalities and stillborn fetuses at the extremes of growth.

Materials and Methods:

We performed a secondary analysis of the Stillbirth Collaborative Research Network (SCRN) study, which was a multisite case-control study with prospectively enrolled cases of stillbirths and livebirths from March 2006 to September 2008. Recruitment included a racially and ethnically diverse population from 59 hospitals in 5 geographic regions throughout the U.S.^13,26 The study was approved by the institutional review board at each hospital and the data coordinating center. An advisory board reviewed the progress and safety of the study and written informed consent was obtained from each participant.

In the present analysis, we performed a retrospective cohort study including singleton stillbirth deliveries with chromosomal microarray analysis performed on the fetus or placenta to evaluate for associations with growth abnormality outcomes (both SGA and LGA). Maternal-offspring dyads were excluded if the parents did not consent to genetic analysis, the chromosomal microarray failed, the stillbirth occurred less than 20 weeks gestational age, or if the birthweight was not recorded. Biospecimens for DNA extraction included umbilical cord blood, placental tissue, and frozen muscle and liver specimens. DNA extraction was performed with use of established methods (Puregene, Qiagen Systems). Chromosomal microarray analysis was performed at a single laboratory (Columbia University Medical Center) in 2012 using the Affymetrix Genome Wide Human Single Nucleotide Polymorphism Array 6.0m in the Chromosome Analysis Suite, version 1.0.1. CNVs of at least 500 kb in size were detected using the SNP array. Data were aligned to the Human Genome release 18 and genes annotated using the NetAffx annotation database, version 28. Microarray results of stillborn fetuses were classified into abnormal CNVs [including aneuploidies, pathogenic CNVs, and variants of unknown clinical significance (VOUS)] and normal CNVs (including no CNVs ≥ 500 kb or benign CNVs) using the American College of Medical Genetics (ACMG) standards and guidelines for interpretation and reporting.²⁷ Using ClassifyCNV tool,²⁸ we updated VOUS and pathogenic CNVs based on the 2015 ACMG guidelines.²⁹

Our primary outcomes were SGA, defined as a birthweight less than the 10^th percentile for gestational age, and LGA, defined as a birthweight greater than the 90^th percentile for gestational age. Comparison groups for the SGA and LGA analyses were appropriate for gestational age (AGA) stillbirths, or those with birthweights greater than the 10^th percentile and less than the 90^th percentile. The SGA, AGA, and LGA designations were assigned based on birthweights reported at the time of delivery of the stillborn fetus.

Gestational age can be questionable in the case of stillbirth due to the unknown time of death and paucity of obstetric ultrasounds early in pregnancy. Gestational age was determined using an algorithm based on time-of-death interval, postmortem examination, and best clinical estimate using multiple sources from the medical chart including assisted reproductive technology dating, first day of last menstrual period, and obstetric ultrasounds.³⁰ Birthweight for gestational age was defined using the Alexander population reference.³¹

The maternal and fetal demographic and clinical characteristics of the stillborn fetuses were compared within CNV classifications. To compare categorical measures, we reported percentages and p-values from Wald chi-squared test. For comparisons of continuous measures, we reported means, standard deviations, and p-values from weighted ANOVA. Logistic regression models were used to test the associations of CNV classifications with the SGA and LGA outcomes. We adjusted for clinically important covariates, including smoking 3 months before pregnancy, obesity, diabetes, fetal sex, chronic hypertension, and maternal age. We excluded race and ethnicity in the multivariable analyses because baseline demographic differences between groups are likely due to independent sociologic factors such as systemic and institutional biases rather than biologic differences. Only maternal age was found to be statistically significant in these multivariable analyses and after backwards elimination was the only variable which remained in the final adjusted model. Additional exploratory analyses were performed to assess the interaction of structural anomalies and with the association of abnormal CNVs and the SGA outcome. Additionally, a sensitivity analysis was performed evaluating the association with VOUS, pathogenic CNVs, and aneuploidy separately with the SGA outcome. The analyses were completed using survey-specific procedures in SAS version 9.4 (SAS Institute Inc, Cary NC), which incorporated analytical weights that reflected SCRN probability sampling and study design. Sampling weights are applied to the analysis to incorporate differential participation rates and staggered enrollment initiation across sites.²⁶

Only pathogenic CNVs and VOUS (excluding chromosomal aneuploidies) noted in the SGA cohort are compared in Table 3. The Online Mendelian Inheritance in Man (OMIM^©), GeneReviews^®, and National Organization for Rare Disorders (NORD^®) online websites were reviewed for information related to specific genes involved in the abnormal CNVs for this cohort. The probable cause of death, as determined by the INCODE research classification tool, and any congenital structural anomalies or placental pathologic findings were included for reference in this table. The INCODE cause of death research classification tool was used for the parent SCRN study across all sites to best identify cases in which a fetal, maternal, or placental condition significantly contributed to the fetal death using a priori definitions based on the best available evidence.^30,32 Postmortem autopsies and placental histologic examinations were performed by perinatal pathologists who underwent centralized training to standardize their evaluations. Details of the postmortem and placental examinations have been previously published.^33,34

Table 3.

Abnormal copy number variants (CNVs) noted on chromosomal microarray and clinically important stillbirth variables associated with small for gestational age stillbirths. Only pathogenic CNVs and variants of unknown significance (VOUS) were described in this table (excluding aneuploidies).

Abnormal CNVa	Gestational age at stillbirth (weeks)	Birthweight percentile	Structural anomaliesb	Placental pathologyc	Probable cause of deathd	Pathogenic CNV or VOUSe
arr 21q22.13 (36,685,848-37,185,921)x3	22	<10%	Cardiac anomaly	Not availablef	Placental disease and fetal genetic causes	VOUS
arr 5p15.2 (10,908,334-11,459,739)x3	23	<10%	None	Maternal and fetal vascular lesions	Placental disease	VOUS
arr 10q23.31 (90,658,193-91,207,964)x3	24	<10%	Not availablee	Maternal vascular lesions	Not availablef	VOUS
arr Yq11.221 (18,148,539-18,999,761)x0	25	<5%	None	Fetal vascular lesions	Placental disease, maternal medical complication	VOUS
arr 4q32.3q35.2 (165,903,367-191,254,120)x1	27	<5%	Cardiac, skeletal, cranial, umbilical cord anomalies	Maternal and fetal vascular lesions, immune dysregulation	Fetal genetic causes, maternal hypertensive disorder	Pathogenic CNV
arr 16p13.11p12.3 (15,224,214-18,286,344)x3	34	<5%	None	Maternal and fetal vascular lesions	Placental disease, maternal medical complication	Pathogenic CNV
arr 7p12.3 (48,008,179-48,659,125)x3	35	<5%	None	Maternal and fetal vascular lesions, maternal inflammatory lesions	Placental disease	VOUS
arr 22q11.21q11.23 (17,256,416-22,140,054)x1	36	<5%	None	Maternal and fetal vascular lesions, fetal inflammatory lesions, immune dysregulation	Fetal genetic causes	Pathogenic CNV

CNV, copy number variant; VOUS, variant of unknown clinical significance

^aNamed using the International System for Human Cytogenetic Nomenclature (ISCN) for array chromosomal region

^bStructural anomalies based on postmortem exam with details of evaluation published previously³⁴

^cPlacental histologic and pathologic exam with details of evaluation published previously³³

^dProbable cause of death using INCODE system,^30,32 when two codes were of equal highest probability, both causes were reported

^ePathogenic CNV and VOUS designation assigned by the American College of Medical Genetics (ACMG)²⁷ and updated to the most recent 2015 ACMG guidelines

^fNot available: autopsy or placental pathology declined by family

Results:

Among 663 stillbirth maternal-offspring dyads, 43 with multifetal pregnancies, 186 without chromosomal microarray data, and 41 without data to calculate population birthweight percentiles were excluded (Figure 1). Among the 393 stillbirths that were included in the present analysis, 16% had abnormal CNVs (Table 1). The abnormal CNVs included 10 stillbirths affected by pathogenic CNVs (15.9%), 24 VOUS (38.1%), and 29 aneuploidies (46.0%). Maternal-offspring dyads with abnormal CNVs were more likely to be of Hispanic ethnicity and less likely to be Black compared to those with normal CNVs: 48.1% vs 33.4%, and 9.5% vs 24.0%, respectively (p=0.014). Fetal structural abnormalities were more common in the abnormal CNV group compared to stillbirths with a normal chromosomal microarray: 48.1% vs 20.2% (p=<0.001).

Figure 1.

Study subjects from original Stillbirth Collaborative Research Network cohort included in this secondary analysis.

Table 1.

Pregnancy demographic characteristics among pregnancies affected by stillbirth with abnormal copy number variants (CNVs) on chromosomal microarray analysis compared to and those with a normal microarray.

Maternal demographic variables
Characteristic	Value	Abnormal CNVs	Normal Microarray	p-value
Unweighted		N=63	N=330
Weighted		N=62	N=326
Age	Mean ± SD	27.3 ± 8.0	27.3 ± 6.2	0.943
BMI	Mean ± SD	27.9 ± 7.9	27.5 ± 6.5	0.720
Race/Ethnicity	White non-Hispanic	22 (35.0)	122 (37.4)	0.014
	Black, non-Hispanic	6 (9.5)	78 (24.0)
	Hispanic	30 (48.1)	109 (33.4)
	Other	5 (7.3)	17 (5.2)
Maternal education	Less than college	34 (59.9)	157 (51.5)	0.250
	College or higher	23 (40.1)	148 (48.5)
Insurance status	Any assistance	36 (58.4)	186 (57.0)	0.838
	Commercial, HMO, or military	26 (41.6)	140 (43.0)
Parity	Nulliparous	26 (42.1)	145 (44.8)	0.700
	Multiparous	36 (57.9)	179 (55.2)
Smoking	3 months before pregnancy	8 (14.2)	60 (19.5)	0.314
Hypertension		9 (14.4)	34 (10.6)	0.434
Preeclampsia		8 (13.3)	33 (10.5)	0.582
Pre-gestational diabetes		5 (8.5)	17 (5.4)	0.417
Gestational diabetes		5 (8.0)	16 (5.0)	0.424
Fetal demographic variables
Characteristic	Value	Abnormal CNVs	Normal Microarray	p-value
Unweighted		N=63	N=330
Weighted		N=62	N=326
Mean gestational age	Weeks ± SD	28.2 ± 6.4	28.8 ± 6.8	0.489
Fetal sexa	Male	36 (57.0)	159 (49.1)	0.256
Any structural anomaly		26 (48.1)	59 (20.2)	<.001
Aneuploidy		28 (45.0)	-	-

CNV, copy number variant; BMI, body mass index; SD, standard deviation. ^aTwo instances of undetermined sex

Table 2 and figure 2 demonstrate our main findings. Abnormal CNVs noted on chromosomal microarray were significantly associated with SGA birthweights: 29.5% vs 16.5% (p=0.038), with an OR of 2.23 (95% CI: 1.18 - 4.18). This result remained significant when adjusting for maternal age, smoking, diabetes, hypertension, obesity, and fetal sex. A final model included only maternal age and CNV classification with an aOR of 2.22 (95% CI: 1.12 - 4.18). The presence of abnormal CNVs was more common among LGA stillbirths, 6.2% vs 3.3% (p=0.275) with an OR of 2.35 (95% CI: 0.70 - 7.90), however this result was not statistically significant. Adjusted modeling of the LGA outcome was not performed given the sample size (n=15).

Table 2.

Fetal growth parameters at time of stillbirth compared to microarray findings.

Characteristic	Value	Abnormal CNVs	Normal Microarray	p-value
Unweighted		N=63	N=330
Weighted		N=62	N=326
SGAa	<10th percentile	18 (29.5)	54 (16.5)	0.038
AGAa	10th – 90th percentile	40 (64.2)	261 (80.2)	0.275
LGAa	>90th percentile	4 (6.2)	11 (3.3)	0.365
Weight at delivery	<2500g	53 (84.7)	244 (75.0)	0.065
	≥2500g	10 (15.3)	81 (25.0)
Birthweight (grams)	Mean ±SD	1324 ±997	1462 ±1179	0.330

CNV, copy number variant; SGA, small for gestational age; AGA, appropriate for gestational age; LGA, large for gestational age; SD, standard deviation.

^aBirthweight percentiles based on Alexander population reference.³¹

Figure 2.

Association between abnormal chromosomal microarray and fetal growth outcomes in stillbirth fetuses. Multivariable model adjusted for smoking 3 months before pregnancy, obesity, diabetes, fetal sex, hypertension, and maternal age with variables removed through backwards selection. Final model includes only maternal age.

Through exploratory sensitivity analyses, we assessed effect modification of aneuploidies on the association of abnormal CNVs with the SGA outcome. The overall association was not statistically significant (p=0.055), although the association of stillbirths affected by aneuploidies alone was of a greater magnitude OR 2.5 (95% CI: 1.07 - 5.65) than was observed for the primary analysis. Similarly, when measuring the association between SGA and aneuploidies combined with pathogenic CNVs excluding VOUS, the magnitude is also greater OR 2.4 (95% CI: 1.15 – 4.98) than that of all genetic abnormalities combined. The results from this sensitivity analysis are depicted in Supplementary Table 1. We tested for an interaction with the association of abnormal CNVs and SGA in the presence [OR 1.1 (95% CI: 0.38 – 3.02), and absence [OR 1.9 (95% CI: 0.72 – 5.25)] of structural anomalies and did not detect a statistically significant difference (p=0.423) through additional exploratory analyses.

Among the SGA stillbirths with abnormal CNVs, 53% were aneuploidies, including seven instances of trisomy 18, and two involving monosomy X. The remaining 47% of abnormalities involved pathogenic CNVs and VOUS within autosomes and one deletion on the Y chromosome, all described in Table 3. Table 3 additionally outlines the gestational age at time of stillbirth, the birthweight percentile, associated congenital anomalies and placental pathology, the probable cause of death, protein-coding genes involved in the CNV, and clinical associations among liveborn carriers described in the literature. The overarching probable cause of death in SGA stillbirths with abnormal CNVs was placental disease (62.5%), with the majority affected by pathologic maternal and fetal vascular lesions.

Comment:

Principle findings:

Our study demonstrates that genetic abnormalities are found in approximately 30% of SGA stillbirths. The odds of a stillborn fetus having a birthweight less than the 10^th percentile is two times higher with abnormal chromosomal microarray results as opposed to a normal microarray. There were no statistically significant differences between abnormal CNVs and LGA stillbirths. This may have been due to a limited occurrence of the LGA outcome (n=15) or due to the absence of an association.

Roughly 50% of the genetic abnormalities found within this cohort of SGA stillbirths were aneuploidies historically found in the setting of miscarriage or stillbirth (trisomy 18 or monosomy X). The remainder were abnormal CNVs or VOUS widespread throughout the genome without a predominant pattern (Table 3). While our exploratory analysis evaluating the effect modification of aneuploidy results in combination with pathogenic CNVs and VOUS was not statistically significant, the magnitude of the association between genetic abnormalities and SGA birthweight was greatest when aneuploidy results were evaluated independently. Additionally, the magnitude of the association between SGA and microarray abnormalities excluding VOUS was also greater. These results from the sensitivity analysis may be explained by the fact that VOUS, and pathogenic CNVs to a lesser extent, are likely a heterogenous group of genetic differences. Some of the pathogenic CNVs or VOUS may have been identified because of true associations between fetal growth abnormalities and stillbirth, while others might never prove to be clinically significant.

While congenital anomalies are commonly associated with stillbirths affected by genetic abnormalities (~30% in most studies)¹³, only two fetuses (25%) in our SGA cohort were affected by congenital anomalies (cardiac, skeletal, cranial, and umbilical cord anomalies). Through additional exploratory analyses, we did not find a significant interaction between the presence of anomalies and the association with genetic abnormalities and SGA birthweights. Rather, the predominant cause of death noted by the INCODE classification system for these SGA stillbirths associated with abnormal CNVs was placental disease or pathology.

One CNV associated with our SGA cohort was a duplication involving the p arm of chromosome 21. Two genes involved in this CNV, CHAF1B and HLCS, are both important in histone modification prior to DNA replication,^35,36 which could lead to downstream effects in rapidly dividing tissues in the embryo and placenta. While rarely described in the literature, a duplication in the p arm of chromosome 5 was seen in our cohort with only one involved gene: CTNND2, which encodes a transcription factor for a protein localized to the neuromuscular junction.³⁷ This could result in direct fetal effects that alter neuromuscular mobility and growth. A duplication at the q arm of chromosome 10 was also noted. This is a relatively conserved region in the mammalian genome, including genes involved in smooth muscle contractility, susceptibility to viruses, and the development of autoimmune disease.^38–41 This CNV is also one of the few described in our study to be previously associated with growth restriction in postnatal life.⁴² One microdeletion in the Y chromosome was noted. Clinical associations with placental trophoblastic tumors and Y chromosome microdeletions have been documented, which raises the suspicion that Y chromosome gene products play a role in placental function.^43,44 Another CNV noted was a large (6 x10⁷ base pair) deletion in the q arm of chromosome 4, which involves several genes important for DNA repair and cell motility.^45,46 One of these well-studied genes, NEK1, is particularly important in cilia function.⁴⁷ Mutations in NEK1 have been implicated in a form of short-rib polydactyly syndrome, which is associated with significant growth disturbances.⁴⁸ TLL1 encodes a proteinase that is important in forming collagen which is vital in ongoing fetal development.⁴⁹ A small but gene-rich duplication in the p arm of chromosome 16 was another CNV discovered in our SGA cohort involving NDE1, which encodes a protein important for mitosis in rapidly dividing cells.^50–52 The genes involved in these CNVs deserve further study to assess their relationship to normal and abnormal fetal and placental growth.

Results in the context of what is known: Previous work regarding the genetics of stillbirth has demonstrated that common chromosomal aneuploidies, especially monosomy X and trisomies 21, 18, and 13 account for approximately 6-8% of overall stillbirth risk.¹⁴ Modem molecular sequencing techniques including chromosomal microarray or whole-exome sequencing raises the detection rate to approximately 13-20%, with the discovery of pathogenic CNVs, monogenic mutations, and SNPs.^13,18,19,21

Genetic causes of fetal growth abnormalities in live births have also been studied.^16,22,23 Triploidy remains the most common genetic cause of isolated FGR in fetuses below 26 weeks, and trisomy 18 afterwards. When chromosomal microarray is utilized, the detection rate of possible causitive genetic abnormalities increases from 7.4% to 18.5% among fetuses with structural anomalies, and from 15.1% to 18.9% among fetuses without structural anomalies.²² Our study adds novelty to the current published literature in quantifying the proportion of genetic abnormalities found among SGA stillborn fetuses that would not be detected by traditional karyotype. We found that 47% of genetic abnormalities in this cohort were the result of pathogenic CNVs or VOUS, which were only detectable by chromosomal microarray.

Additionally, we note that abnormal CNVs were associated with placental causes of stillbirth, supporting the hypothesis that genetic abnormalities may lead to placental dysfunction and insufficiency, as evidenced by both maternal and fetal vascular lesions in this cohort. Finally, the majority of the abnormal CNVs described in our study have not been previously associated with growth restriction or stillbirth.

Clinical implications:

Though the phenotypes of FGR and stillbirth have overlapping clinical etiologies, the findings from our research suggest that these abnormal CNVs may result in disruptions to common biologic pathways important for both fetal growth and survival. Most of the SGA stillbirths affected by abnormal CNVs in our cohort were associated with maternal and fetal placental vascular lesions, which could be the primary etiology for FGR and resultant stillbirth. Our findings support the recent recommendations for chromosomal microarray in the evaluation of a stillbirth as opposed to karyotype, ^13,19,25 especially in the setting of growth restriction,^14,53 since roughly half of the genetic abnormalities associated with SGA stillbirths would not have been discovered with conventional karyotype.

Research implications:

While some of the abnormal CNVs associated with SGA stillbirths in this study are known to be associated with growth restriction, an association with stillbirth was not previously noted. The abnormal CNVs reviewed here involve many autosomes and even the Y chromosome. Genes involved in these CNVs deserve further investigation for their role in placental function and fetal growth and survival.

Strengths and limitations:

Our secondary analysis was performed using the SCRN database, which derives from one of the largest thoroughly investigated cohorts involving prospectively collected data in pregnancies affected by stillbirth.^13,26,30,54 This cohort arose from a geographically diverse, multi-institutional network with racial and ethnic diversity reflective of the U.S. population. Abnormal CNVs were updated to reflect the most recent designations for pathogenic CNVs and VOUS per ACMG standards. One major limitation includes the small numbers of LGA fetuses, which limited our ability to form inferences about an association with abnormal chromosomal microarray findings. Microarray-based analyses are limited in identifying balanced chromosomal rearrangements, single-gene disorders, and sometimes entire duplications of the genome (i.e. triploidy or tetraploidy). Also, the detection of CNVs restricted to those ≥500 kb may have limited our ability to detect smaller pathogenic CNVs that could have been associated with our outcomes. Additionally, including VOUS in our abnormal CNV designation could lead to misclassification of our exposure, especially if the specific VOUS in our study prove to be clinically insignificant in the future. Lastly, there is the possibility of confined placental mosaicism, or genetic abnormalities found only in the placenta and discrepant from the fetal genotype, which may have gone undetected depending on the tissue sampled for genetic analysis. An optimal future study would include both placental and fetal tissue for genetic analysis.

Conclusions:

Genetic abnormalities are more common in SGA stillborn fetuses, half of which are pathogenic CNVs or VOUS. Further research is needed to understand the common biologic pathways underlying fetal growth restriction, placental insufficiency, and stillbirth.

Condensation page.

1. Condensation: Abnormal copy number variants on chromosomal microarray are more common in small for gestational age stillbirths than those with normal growth.

2. Short Title: Copy Number Variants and Fetal Growth in Stillbirths

3. AJOG at a Glance:

   1. Why was this study conducted: To understand the association between genetic abnormalities in stillbirths at the extremes of fetal growth (small and large for gestational age).
   2. Key findings: Small for gestational age stillbirths are two times more likely to be affected by genetic abnormalities than those with a normal growth pattern. Half of these genetic abnormalities are abnormal copy number variants that would not have been detected by karyotype.
   3. What does this study add to what is known? Genetic abnormalities are more common in small for gestational age stillborn fetuses, 50% of which are the result of abnormal copy number variants (deletions or duplications). Further research is needed to understand the genetic etiologies underlying fetal growth restriction, placental insufficiency, and stillbirth.