Products Of Conception Samples Research Articles

Chromosomal Microarray Analysis in Pregnancy Loss: Is It Time for a Consensus Approach?

ABSTRACT Affecting up to 10% to 15% of all clinically recognized pregnancies, pregnancy loss has many causes. Half of first-trimester losses come about due to chromosomal abnormalities and then decline in frequency through pregnancy, ultimately reaching a rate of 5% in stillbirths. Recurrent pregnancy loss occurs in 1% to 2% of couples, and identifying the genetic etiology of these losses can provide psychological benefits, eliminate other testing needs, and establish a risk of recurrence. As benefits of performing genetic testing on products of conception (POCs) are clear, various groups have published practice guidelines (American College of Obstetricians and Gynecologists, European Society of Human Reproduction and Embryology, American Society for Reproductive Medicine, and Royal College of Obstetricians and Gynaecologists). However, despite the consensus for the utility of testing on POCs, requirements for these tests vary. Most appropriate testing methodology, obstetric history, and gestational age are all variations that affect the benefit or utility of these tests. As there is a lack of consensus on how to use chromosomal microarray analysis (CMA) in assessment of POCs, this study assessed POC CMA data with available karyotype data and determined the positive CMA rate for POCs. The authors also included within this data set a comprehensive correlation of CMA correlations with pregnancy phenotype, gestational age, and prior prenatal screening results. Finally, the study also assessed the impact CMA results from early losses may have on clinical care. This cohort included 323 POC samples processed by the Brigham and Women's Hospital Center for Advanced Molecular Diagnostics in the Clinical Cytogenetics Laboratory over a consecutive 42-month period. Collected variables included estimated gestational age at time of pregnancy loss, maternal age, pregnancy phenotype results of any orthogonal testing on POC tissue, noninvasive prenatal screen results, CMA findings, and available parental karyotypes. All cases recorded gestational age except 1 sample lacking documentation. Categories of phenotypic data included lymphatic malformations, anatomical abnormalities of the fetus or placenta, twin or higher-level multiple gestations, environmental complications, amniotic fluid disorders, or the combined influence of any of the above. A lack of definitive phenotypic data excluded 3 cases from the analysis. Analyses of CMA results were categorized as normal, abnormal, or variants of uncertain significance (VUSs). The cases defined as normal had no identifiable clinically significant copy number variants, and abnormal cases were defined as pathogenic. Results of the study included gestational age for 322 of 323 cases, which was then used to determine the rate of fetal demise or miscarriage at different gestational ages in the cohort, with first-trimester losses at <14 weeks (n = 230) and later losses defined as occurring at >14 weeks gestational age (n = 92). The results demonstrated that frequency and genetic abnormality types (as identified by CMA) align closely with karyotype studies previously published. This means that an approximate half of the first-trimester loss cohort have a genetic abnormality, with trisomies as the most common finding. Very early losses tend to have a high positive rate of abnormalities (45.6%), which further supported CMA use in first-trimester losses. Inadvertent assessment and reporting of maternal metaphases were reduced, and yield of results improved with the implementation of CMA. Abnormal POC results may suggest balanced parental abnormalities, and follow-up parental testing in such cases is recommended. A need for additional studies exists to determine the parental follow-up studies yield. In conclusion, POC tissue genetic testing following pregnancy loss can aid in clinical management via identification of genetic etiologies, thereby refining recurrence risk. However, lack of consensus in terms of testing strategies and methodologies indicates a need for standardized methodology. This study demonstrated CMA results consistent with published karyotype studies established CMA analysis advantages over POC karyotype in all gestational ages. Even in early first-trimester losses, these results support the efficacy of CMA in POC testing.

Chromosomal microarray analysis in pregnancy loss: Is it time for a consensus approach?

To investigate the efficacy and outcomes of chromosomal microarray (CMA) in the cytogenomic evaluation of products of conception (POC). Over a 42-month period, 323 POC samples were tested by CMA. Results were assessed using variables including phenotype, gestational age, results from orthogonal testing, and follow-up parental analysis. CMA identified cytogenetic abnormalities in 47.4% of first trimester losses and 10.9% of second and third trimester losses. Chromosomal microarray results specifically from 5 to 7-week losses showed similar rates of abnormalities (45.6%) compared to those of all first trimester losses combined. CMA and karyotype results were discordant in 20.0% of cases, most likely due to maternal cell overgrowth in culture. The most prevalent abnormalities identified in all losses were autosomal trisomies, followed by triploidy. In 43/323 cases, the observed abnormality suggested a parental aberration that prompted follow-up studies; two of these cases indeed identified an inherited aberration. Our findings of specific types of genetic abnormalities and the respective frequencies by gestational age closely align with those of published karyotype studies, supporting the use of routine CMA testing for POCs. CMA outperforms karyotype analysis because it does not require viable, sterile cultures free of maternal admixture or admixture due to multiple gestations. Finally, CMA results can play an important role in identifying increased recurrence risks for some couples.

Stepwise molecular-genetic examination in aborted fetuses.

The evaluation of quantitative fluorescence PCR (QF-PCR) and single nucleotide polymorphism array (SNP array) analysis for the identification of chromosomal abnormalities in products of conception (POC). A total of 1,094 POC samples were processed at Gennet in the years 2018-2020. Chromosomal aneuploidies were tested by QF-PCR using a Omnibor set (STR markers 13, 18, 21, X a Y), SAB-I set (STR markers 2, 7, 15, 16, 22), SAB-II set (from November 2019, STR markers 4, 6, 14) followed by SNP array analysis (Illumina) on samples with a negative QF-PCR result. All POC samples were tested for maternal contamination. After exclusion of maternal contamination (32% samples) the total number of 742 POC samples were tested by QF-PCR. Chromosomal aneuploidies were found in 273 POC samples (36.8%). Then, 469 QF-PCR negative POC samples were tested by SNP array analysis. Normal female/male profile was confirmed in 402 samples (85.7%) and chromosomal aneuploidies and chromosomal aberrations (deletion/duplication > 10 Mb) in 51 samples (10.9%). Microdeletion/microduplication was found in 16 POC samples (3.4%), two were classified as pathogenic variants and 14 as variants of unknown significance. In a group of women > 35 years of age, statistically significant increase of the chromosomal abnormalities was confirmed. No statistically significant difference between the in vitro fertilization group and the group of spontaneous conception was found. The application of the molecular work-up based on the stepwise use of QF-PCR and SNP array clarifies the cause of the abortion in 43% POC samples. The overall detection rate in the I. trimester was 50.4%.

The genomic basis of sporadic and recurrent pregnancy loss: a comprehensive in-depth analysis of 24,900 miscarriages

Research questionWhat is the genetic cause of sporadic and recurrent pregnancy loss and does the frequency and nature of chromosomal abnormalities play a role? Types and frequency of all identifiable chromosomal abnormalities were determined to inform our understanding, medical management and recurrence risk for patients experiencing pregnancy loss. DesignGenome-wide single-nucleotide polymorphism-based chromosomal microarray (SNP-CMA) were used to evaluate 24,900 products of conception samples from various forms of pregnancy losses. ResultsSporadic miscarriage (64.7%) or recurrent pregnancy loss (RPL) (22%) were the most common referrals. Clinically significant abnormalities were observed in 55.8% (13,910) of samples, variants of uncertain significance in 1.8%, and normal results in 42.4%. In addition to autosomal trisomies (in 36% of samples), polyploidy and large segmental imbalances were identified in 7.8% and 2.8% of samples, respectively. Analysis of sequential samples from 1103 patients who had experienced RPL provided important insight into possible predispositions to RPL. ConclusionsThis expansive chromosomal microarray analyses of pregnancy loss samples illuminates our understanding of the full spectrum, relative frequencies and the role of genomic abnormalities in pregnancy loss. The empiric observations described here provide useful insight for clinicians and highlight the importance of high-resolution genomic testing for comprehensive evaluation and risk assessment of individuals experiencing pregnancy loss.

EP378: A 9-year, single-institution retrospective study of chromosomal microarray analysis data obtained from products of conception

Chromosomal microarray analysis (CMA) has been utilized in genetic testing of products of conception (POCs) due to its ability to detect submicroscopic genomic aberrations not identified by karyotype, and for its higher diagnostic yield relative to karyotyping. The aim of this study is to assess the diagnostic yield of CMA, as well as to describe the different kinds of genomic aberrations found in the POC samples received by our laboratory, including small copy number variations (CNVs) overlapping with genes involved in placental and fetal development.

Correlation of Maternal Age and Chromosomal Abnormality in Products of Conception- A Single Centric Study

Purpose: The purpose of this study is to find out the types and incidence rate of chromosomal abnormalities and the relationship between maternal age and chromosomal abnormality in products of conception by retrospective analysis. Method: Karyotype study using standard GTG banding and FISH study for aneuploidy detection was done from products of conception samples. Results: A total of 513 cases of products of conception were studied retrospectively. 98 cases were studied by conventional cytogenetic technique and 415 cases were studied by a FISH method. The chromosomal abnormality was observed in 97 cases (18.91%). Trisomy was found to be a major chromosomal abnormality amongst the cases studied, followed by triploidy and monosomy. A higher incidence of chromosomal abnormality was found in women with advanced maternal age as compared to the maternal age less than 35 years.

The introduction of clinical genetic testing in Ethiopia: Experiences and lessons learned.

Limited data are available on genetic testing laboratories in low- and middle-income countries including those in sub-Saharan Africa (SSA). To characterize the need for genetic testing in SSA we describe the experience of MRC-ET Advanced Laboratory, a genetic testing laboratory in Ethiopia. Test results were analyzed based on indication(s) for testing, referral category, and diagnostic yield. A total of 1311 tests were run using the full MRC-Holland catalogue of Multiplex-Ligation Probe Amplification assays. Of all samples, 77% were postnatal samples, 15% products of conception (POC), and 8% amniotic samples. Of postnatal samples, the most common testing categories were multiple congenital anomalies (32%), disorders of sex development (17%), and Obstetrics/Gynecology (16%). Forty-three percent of postnatal samples were diagnostic, 11% were variants of uncertain significance (VUS), and 46% were normal with Trisomy 21 the most common diagnosis. Of POC samples, 10% were diagnostic, 34% revealed VUSs, and 55% were normal with Trisomy 18 the most common diagnosis. Of amniotic samples 17.5% were diagnostic, 3% revealed VUSs, and 79% were normal with Trisomy 18 the most common diagnosis. There is increasing demand for genetic testing in Ethiopia. Diagnostic genetic testing in SSA deserves increased attention as testing platforms become more affordable.

13. Revisiting centromeric polymorphisms: Implications for POC and prenatal interphase FISH studies

Interphase FISH is a routinely used rapid method to detect or confirm common prenatal aneuploidies (13, 18, 21, X, Y). Rare aneuploidies are typically observed in products of conception (POC) samples and now more recently in NIPT samples that undergo whole genome assessment. As such, FISH with expanded aneuploidy panels are being used more frequently in prenatal or POC samples to detect or confirm rare autosomal aneuploidies. We report on cases where polymorphisms in the centromeric regions have affected the interpretation of interphase FISH results.

Reducing misdiagnosis caused by maternal cell contamination in genetic testing for early pregnancy loss

ABSTRACT The analysis of products of conception (POC) is clinically important to establish the cause of early pregnancy loss. Data from such analyses can lead to specific interventions in subsequent natural or assisted conceptions. The techniques available to examine the chromosomal composition of POC have limitations and can give misleading results when maternal cell contamination (MCC) is overlooked. The aim of this study was to develop a protocol for MCC assessment and to formulate POC material handling, testing, and reporting recommendations. Using array comparative genomic hybridization, we tested 86 POC samples, of which 47 sample pairs (DNA extracted from the POC sample and maternal DNA) were assessed for the presence of MCC. MCC was evaluated using an approach we developed, which exploited the genotyping of 14 STR, AMEL, and SRY loci. POC samples showing the clear presence of villi (63.9%) did not contain any signs of the maternal genome and can therefore be reliably tested using conventional methods. The proportion of 46,XX karyotype in the unselected sample batch was 0.39, which fell to 0.23 in visually good samples and was 0.27 in samples having no signs of contamination upon MCC testing. MCC assessment can rescue visually poor samples from being discarded or wrongly genotyped. We demonstrate here that classification based on visual POC material evaluation and MCC testing leads to predictable and reliable POC genetic testing outcomes. Our formulated recommendations covering POC material collection, transportation, primary and secondary processing, as well as the array of pertinent considerations discussed here, can be implemented by laboratories to improve their POC genetic testing practices. We anticipate our protocol for MCC assessment and recommendations will help reduce the misconception regarding the etiology of miscarried fetuses and foster informed decision-making by clinicians and patients dealing with early pregnancy loss.

Cytogenetic signatures of recurrent pregnancy losses.

To investigate the incidence of chromosomal abnormalities in the products of conception (POC) of patients with spontaneous miscarriages (SM) and with recurrent pregnancy losses (RPL) and to determine biological mechanisms contributing to RPL. During a 20-year period, 12 096 POC samples underwent classical chromosome analysis. Cytogenetic findings were compared between the SM and RPL cohorts. Analysis of RPL cohort has identified an increased incidence of inherited and de novo structural chromosome abnormalities, recurrent polyploid conceptions, and complex mosaic alterations. These abnormalities are the signature of genomic instability, posing a high risk of genetic abnormalities to offspring independent of maternal age. Predominance of male conceptions in the RPL cohort points toward an X-linked etiology and gender-specific intolerance for certain genetic abnormalities. Our study showed several possible genetic etiologies of RPL, including parental structural chromosome rearrangements, predisposition to meiotic nondisjunction, and genomic instability. Loss of karyotypically normal fetuses might be attributed to defects in genes essential for fetal development, as well as aberrations affecting the X chromosome. Molecular studies of parental and POC genomes will help to identify inherited defects in genes involved in meiotic divisions and DNA repair to confirm our hypotheses, and to discover novel fetal-essential genes.

1002: Importance of single chromosome uniparental disomy in pregnancy loss and gene discovery

Report on products of conception (POC) samples with uniparental disomy (UPD) of a single chromosome, estimate the incidence of single UPD in miscarriage, and highlight single UPD cases associated with genetic conditions or placental insufficiency that may relate to the loss. Retrospective analysis of POC and maternal (mat) blood samples shipped to a lab for genotyping using Illumina CytoSNP-12b microarrays with informatics. Of 47,394 samples, 40,980 (86.5%) had fetal results: 113 (0.3%) had single UPD, including 29 (25.7%) with additional findings. 16 chromosomes had at least 1 case of single UPD (Table 1). The majority had heterozygous (htz) UPD (87, 77.0%). 38 cases (33.6%) involved chromosomes with a known phenotype due to imprinting or placental insufficiency (Table 1). The average mat age of single UPD cases was 35.5 years and the average gestational age was 62 days (range 35-240 days) vs. 36.8 years and 55 days (range 35–83 days) for cases with single UPD and aneuploidy. Four cases of single UPD miscarried past the first trimester, 3 involved chromosomes with known phenotypes. Single UPD of specific chromosomes is associated with pregnancy loss and/or genetic disease through imprinting defects, unmasking recessive alleles, and late trisomy rescue. This study identified 84 cases of single UPD without additional findings. 38 involved chromosomes known to cause an abnormal phenotype due to imprinting or placental insufficiency which may explain the loss. As 28 miscarriages had maternal UPD of chromosome 16, performing POC analysis that can identify UPD in miscarriages after diagnosis of intrauterine growth restriction is critical.

Genetic analysis of products of conception using a HLPA/SNP-array strategy

BackgroundFetal chromosomal abnormalities was the most frequent cause of miscarriage, and the traditional testing method G-banded karyotyping has limitations. Then high-throughput ligation-dependent probe amplification (HLPA) and single nucleotide polymorphism array (SNP-array) were introduced for genetic analysis on products of conception (POC).MethodsHLPA and SNP-array analysis were combined. POC samples were initially tested using HLPA, followed by SNP-array analysis on samples that were found to be normal by HLPA.ResultsOf the 326 POC samples tested, the overall abnormality rate was 54.6% (178/326), including 44.8% (146/326) chromosomal abnormalities identified by HLPA and 9.8% (32/326) additional chromosomal abnormalities further detected by SNP-array.ConclusionsThe combination of HLPA and SNP-array analysis is an efficient and cost-effective strategy for genetic analysis of POC.

Characteristics of first trimester miscarriages assesed by chromosomal analysis of products of conception with next generation sequencing

Chromosomal abnormalities are the major cause of early pregnancy loss. Chromosome testing of products of conception (POC) provides valuable information for counseling and clinical managing of patients. We previously showed that next generation sequencing (NGS) can be utilized as a technique demanding lesser specimen with a lower failure rate, higher resolution, and shorter turnaround time than conventional karyotyping which is requiring labor-intensive and time-consuming cell culture with possible maternal cell contamination. We aimed to assess the efficacy of NGS method for chromosomal analysis of POC. In addition, we attempted to identify any associations between the incidence of chromosomal abnormalities and the profile of patients as well as fetal development in an assisted reproductive technology (ART) program. Retrospective study with a single reference genetic laboratory. Total of 131 consenting patients with first trimester miscarriages after vitrified-warmed embryo transfer were involved. POC samples were obtained bydilation and curettage between 7 to 10 gestational weeks. Chorionic villi were isolated under a dissecting microscopy, subsequently processed for NGS chromosomal analysis. Incidence of each chromosomal abnormality was reported and evaluated according to the patient profile, such as maternal age, previous history of miscarriage and fetal development. Finally, frequency of mosaics was also assessed. After NGS analysis, 28 cases (21.4%) were found to be normal, and the remaining 103 (78.6%) were abnormal, including 10 (7.6%) mosaics. Among normal karyotypes, ratio of female to male was 1.15 (15/13). Trisomies were the most common abnormalities except for the chromosome X monosomy (10.7%). Aneuploidy of chromosome 22 (20/113, 17.7%), 15 (16/113, 14.2%), 16 (16/113, 14.2%), X (13/113, 11.5%) and 21 (11/113, 9.7%) including overlaps, were most frequently involved. Mean maternal age of chromosomally abnormal cases was significantly higher than that of normal karyotypes (39.0 ± 18.5 vs 36.9 ± 16.5 years, P<0.05). Patients with more than equal 3 previous miscarriages showed a significantly lower rate of abnormalities than those with <3 miscarriages (28.6% vs 81.5%, P<0.01). Rate of abnormalities with positive fetal cardiac activity was not different from that of anembryonic pregnancies (80.0% vs 76.1%), although fetal cardiac activity was detected in all the 45,XO cases. Interestingly, however, mosaic abnormalities were significantly more often detected in anembryonic pregnancies than the other (15.2% vs 3.5%, P<0.05). With more conclusive and accurate results and higher resolution by NGS, we were able to characterize early pregnancy loss after ART, demonstrating relatively high rate of abnormalities with gender ratio being close to 1. Patients with repeated pregnancy loss showed lower chromosomal abnormalities indicating other causes for miscarriages in this group of patients. A higher incidence of mosaics detected in anembryonic pregnancies warrants further investigation.

Single-nucleotide polymorphism microarray detects molar pregnancies in 3% of miscarriages

To determine the frequency of molar pregnancy in miscarriage cases based on single-nucleotide polymorphism (SNP) microarray testing on products of conception (POC) tissue and to estimate the sensitivity of ultrasound and histopathologic evaluation for cases identified to be at risk for gestational trophoblastic disease. Retrospective cohort analysis. Reference laboratory. POC specimens from 26,101 consecutive miscarriages. POC samples were sent to a single laboratory for analysis. Maternal age, gestational age, egg donor use, indication for testing, and additional clinical information were obtained from the requisition form and a survey sent to ordering providers. Rates of full paternal uniparental disomy (UPD), indicative of complete molar pregnancy, and triploidy of paternal origin, indicative of partial molar pregnancy, in POC samples. Paternal triploidy was detected in 638 cases (2.8%) and full paternal UPD in 72 cases (0.3%). Of the cases with complete clinical information (224/710; 31.5%), histopathology and/or ultrasound did not detect 71% of partial molar pregnancies and 30% of complete molar pregnancies. Molar pregnancy, detected in 3.1% of all miscarriages in this study with the use of 24-chromosome SNP microarray testing, occurred significantly more frequently than previously estimated with the use of ultrasound and/or histopathology.

Utility and performance of bacterial artificial chromosomes-on-beads assays in chromosome analysis of clinical prenatal samples, products of conception and blood samples.

Chromosome analysis of prenatal samples and products of conception (POC) has conventionally been done by karyotyping (KT). Shortcomings of KT like high turnaround time and culture failure led to technology innovations, such as the bacterial artificial chromosomes (BAC)s-on-Beads (BoBs)-based tests, Prenatal BoBs (prenatal samples) and KaryoLite BoBs (POC samples). In the present study, we validated and evaluated the utility of each test on prenatal, POC and blood samples. Study A (n = 305; 259 prenatal + 46 blood/POC) and Study B (n = 176; 146 POC/chorionic vill + 30 blood/amniotic fluid) samples were analyzed using Prenatal and KaryoLite BoBs kits, respectively. KT, array-based Comparative Genomic Hybridization (arrayCGH) and fluorescence in situ hybridization (FISH) were used for comparison of results. Ability of KaryoLite BoBs to identify ring chromosomes was tested. Prenatal BoBs had zero test failure rate and results of all samples were concordant with KT results. Totally four microdeletions were identified by Prenatal BoBs but not by KT. In Study B, all but two POC samples (one triploid and one tetraploid) were concordant with KT and arrayCGH. Partial chromosomal imbalance detection rate was ~64% and KaryoLite BoBs indicated the presence of a ring chromosome in all four cases. The failure rate of KaryoLite BoBs was 3%. We conclude that Prenatal BoBs (common aneuploidies and nine microdeletions) together with KT constitutes more comprehensive prenatal testing compared to FISH and KT. KaryoLite BoBs for aneuploidies of all chromosomes is highly successful in POC analysis and the ability to indicate presence of ring chromosomes improves its clinical sensitivity. Both tests are robust and could also be used for different specimens.

A feasible diagnostic approach for the translocation carrier from the indication of products of conception

BackgroundChromosome translocations are rare but frequently associated with infertility. The objective of this study is to investigate the feasibility of using chromosomal microarray analysis (CMA) on products of conception (POC) samples as an indicator of parental balanced translocation. From January 2011 to December 2016, CMA using Affymetrix Cytoscan™750K array was performed on 1294 POC samples in our hospital. Karyotyping and fluorescence in situ hybridization (FISH) using parental blood samples were performed to validate the origin of subchromosomal copy number variations (CNVs).ResultsIn the 1294 cases of POCs, we detected CNVs of terminal duplication and deletion that imply unbalanced translocation derivatives in 16 cases, and accurate diagnosis with the parental study was made in all the cases by karyotyping and/or FISH. In 10/16 (62.5%) of these cases, CNVs were inherited from one carrier parent of balanced translocation (Cases 1 to 10), while 6/16 (37.5%) cases occurred de novo (Cases 11 to 16).ConclusionThis study clearly illustrated the importance of the utilization of CMA on POC, followed by parental karyotyping and FISH to better characterize CNVs. This approach is especially useful for couples in whom one partner carries a cryptic/submicroscopic balanced translocation but has an apparently normal karyotype.

Exploring the cause of early miscarriage with SNP-array analysis and karyotyping

Objective: The aim of this study is to explore the cause of miscarriage, providing risk assessment to guide the next pregnancy.Methods: Four hundred eighty-four products-of-conception (POC) samples were analyzed by single nucleotide polymorphism (SNP) array, and peripheral blood samples of couples were collected for karyotyping or fluorescence in situ hybridization (FISH) analysis.Results: Four hundred sixty-eight of the 484 (96.7%) fresh POC samples were successfully analyzed using SNP-array. The rate of clinically significant chromosomal abnormalities were 58.3% (274/468), in which rates of aneuploidy, polyploidy, partial aneuploidy, uniparental isodisomy (isoUPD), and pathogenic microdeletion/microduplication were 43.4% (203/468), 8.8% (41/468), 3.6% (17/468), 1.9% (9/48), and 0.9% (4/468), respectively. The percentage of embryonic chromosomal abnormalities significantly increased with maternal age of patients older than 35 years old. Among 468 couples, 12 major chromosomal rearrangements were detected by G-banding, including nine reciprocal translocations, two Robertsonian translocations, and one superfemale.Conclusions: Chromosome abnormality is the main causes of early miscarriage, and aneuploidies are the most common type of chromosomal abnormalities. Application of SNP array and karyotyping in early miscarriage can provide more genetic information about miscarriage, providing risk assessment to guide the next pregnancy.

Development of a Chromosomal Microarray Test for the Detection of Abnormalities in Formalin-Fixed, Paraffin-Embedded Products of Conception Specimens

Testing the products of conception (POCs) provides information about the cause of fetal loss and helps determine the recurrence risk of future losses and chromosome abnormalities in subsequent pregnancies. Historically, the Mayo Clinic Cytogenetics Laboratory performed targeted fluorescent in situ hybridization (FISH) testing to identify aneuploidy of only certain chromosomes in formalin-fixed, paraffin-embedded (FFPE) POC samples. Chromosomal microarray studies using the Affymetrix OncoScan FFPE Assay can detect copy number changes across the genome. We validated the utility of the OncoScan assay using 25 archival FFPE POC specimens with previous FISH results (five normal, 12 trisomy, six triploidy, two monosomy). Of the five normal samples, four had no clinically relevant findings, and one sample was found to have trisomy 9, which is not detectable by the FISH test. For the 20 samples with abnormal FISH results, the OncoScan assay identified all reported abnormalities along with additional findings. A sample with trisomy 22 was also found to have trisomy 7. Another sample reported as triploidy was found to have four copies of chromosome 16. In conclusion, we verified the performance characteristics of OncoScan on FFPE POC specimens and found it acceptable for clinical use. Additional information was identified in 3 of 25 cases (12%) that would explain the pregnancy loss or provide recurrence risk for the family.

36 - Development of a Chromosomal Microarray Test for the Detection of Abnormalities in Formalin-Fixed Paraffin-Embedded Products of Conception Specimens

Testing the products of conception (POCs) provides information about the cause of fetal loss and helps determine the recurrence risk of future losses and chromosome abnormalities in subsequent pregnancies. Historically, the Mayo Clinic Cytogenetics Laboratory performed targeted fluorescent in situ hybridization (FISH) testing to identify aneuploidy of only certain chromosomes in formalin-fixed, paraffin-embedded (FFPE) POC samples. Chromosomal microarray studies using the Affymetrix OncoScan FFPE Assay can detect copy number changes across the genome. We validated the utility of the OncoScan assay using 25 archival FFPE POC specimens with previous FISH results (five normal, 12 trisomy, six triploidy, two monosomy). Of the five normal samples, four had no clinically relevant findings, and one sample was found to have trisomy 9, which is not detectable by the FISH test. For the 20 samples with abnormal FISH results, the OncoScan assay identified all reported abnormalities along with additional findings. A sample with trisomy 22 was also found to have trisomy 7. Another sample reported as triploidy was found to have four copies of chromosome 16. In conclusion, we verified the performance characteristics of OncoScan on FFPE POC specimens and found it acceptable for clinical use. Additional information was identified in 3 of 25 cases (12%) that would explain the pregnancy loss or provide recurrence risk for the family.

Spectrum of Cytogenomic Abnormalities Revealed by Array Comparative Genomic Hybridization on Products of Conception Culture Failure and Normal Karyotype Samples

Approximately 30% of pregnancies after implantation end up in spontaneous abortions, and 50% of them are caused by chromosomal abnormalities. However, the spectrum of genomic copy number variants (CNVs) in products of conception (POC) and the underlying gene-dosage-sensitive mechanisms causing spontaneous abortions remain largely unknown. In this study, array comparative genomic hybridization (aCGH) analysis was performed as a salvage procedure for 128 POC culture failure (POC-CF) samples and as a supplemental procedure for 106 POC normal karyotype (POC-NK) samples. Chromosomal abnormalities were detected in 10% of POC-CF and pathogenic CNVs were detected in 3.9% of POC-CF and 5.7% of POC-NK samples. Compiled results from this study and relevant case series through a literature review demonstrated an abnormality detection rate (ADR) of 35% for chromosomal abnormalities in POC-CF samples, 3.7% for pathogenic CNVs in POC-CF samples, and 4.6% for pathogenic CNVs in POC-NK samples. Ingenuity Pathway Analysis (IPA) was performed on the genes from pathogenic CNVs found in POC samples. The denoted primary gene networks suggested that apoptosis and cell proliferation pathways are involved in miscarriage. In summary, a similar spectrum of cytogenomic abnormalities was observed in POC culture success and POC-CF samples. A threshold effect correlating the number of dosage-sensitive genes in a chromosome with the observed frequency of autosomal trisomy is proposed. A rationalized approach using firstly fluorescence in situ hybridization (FISH) testing with probes of chromosomes X/Y/18, 13/21, and 15/16/22 for common aneuploidies and polyploidies and secondly aCGH for other cytogenomic abnormalities is recommended for POC-CF samples.