Embryo transfer practices and assisted reproductive technology (ART) outcomes by United States census bureau region: an analysis of 46,864 fresh first autologous cycles from the sart registry

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PREGANCY LOSS IS A MAJOR RISK FOR PREMATURE IVF TREATMENT DISCONTINUATION. Karvir H,a Hunter Cohn K,a Arredondo F,b Miller BT,c Gutmann JN,d Leondires MP,e A. B. Copperman,f Yurttas Beim P.a aCelmatix Inc., New York, NY; bReproductive Medicine Associates of Texas, San Antonio, TX; cReproductive Medicine Associates of Michigan, Troy, MI; dReproductive Medicine Associates of Philadelphia, Philadelphia, PA; eReproductive Medicine Associates of Connecticut, Stamford, CT; fReproductive Medicine Associates of New York, New York, NY.

EMBRYO TRANSFER PRACTICES AND ASSISTED REPRODUCTIVE TECHNOLOGY (ART) OUTCOMES BY UNITED STATES CENSUS BUREAU REGION: AN ANALYSIS OF 46,864 FRESH FIRST AUTOLOGOUS CYCLES FROM THE SART Kelly S. Acharya, MD,a REGISTRY. Sanaz Keyhan, MD,a b a a Chaitanya R. Acharya, MS, Suheil J. Muasher, MD. Division of Reproductive Endocrinology and Infertility, Duke University Medical Center, 5704 Fayetteville Rd. Durham, NC, United States, 27713; bDepartment of Biostatistics and Bioinformatics, Duke University Medical Center, 2424 Erwin Rd. Durham, NC, United States, 27710.

BACKGROUND: Previously, we reported that one in four patients discontinue IVF treatments prematurely, before they have maximized their chances of achieving a live birth (LB). Others have linked discontinuation not only to financial access but also to stress and lack of clarity on whether continued treatment will result in a positive outcome. By better understanding why patients discontinue treatment prematurely, physicians can help drive better outcomes. OBJECTIVE: Our goal was to identify factors that increase the likelihood of patients discontinuing prematurely. MATERIALS AND METHODS: Retrospective, de-identified data from electronic medical records were obtained from five academic and private fertility treatment centers. Data included in the analysis consisted of several hundred clinical variables from 31,568 (fresh or frozen embryo transfer) autologous IVF cycles (19,047 patients) from 2002 to 2014. Factors associated with discontinuation were assessed using multivariate, time-dependent Cox proportional hazards regression models, adjusted for likelihood of LB and treatment history. RESULTS: Patients (mean age 36.4
4.7) achieved a LB or discontinued treatment after an average of 1.5 IVF cycles. Overall, patient discontinuation rates varied significantly between the five clinics (p<0.001) and ranged from 50% to 56% of patients discontinuing treatment without a LB. Advanced maternal age (hazard ratio, HR¼1.19, p<0.001) and the lack of cryopreserved embryos (HR¼1.18, p<0.001) significantly increased the risk of premature discontinuation, even after adjustment for overall prognosis. Discontinuation risk was also seen in patients who experienced a clinical (HR¼1.89, p<0.001) or a biochemical pregnancy loss (HR¼1.88, p<0.001) on the current cycle. A history of previous clinical pregnancy loss had an even more dramatic effect on the risk of discontinuation (HR¼2.66, p<0.001), even in the absence of a loss that cycle. Interestingly, higher embryo trophectoderm scores (OR¼0.95, p<0.001) led to a significant reduction in risk. CONCLUSIONS: Here we identify patients at risk for discontinuing treatment before they have maximized their chances for success. In particular, older women, women who do not produce enough embryos to freeze, and women who either experience a loss during an IVF cycle or who have a history of clinical pregnancy loss may benefit from closer support to help them manage the difficult transition after a failed cycle. Empowering patients with journey-level, cumulative-success-rate data ahead of treatment initiation and after treatment failure or pregnancy loss could have a significant impact on patient commitment and outcomes. FINANCIAL SUPPORT: Celmatix Inc.

BACKGROUND: It has been shown that embryo transfer practices and multiple birth rates differ between individual states as well as mandated versus non-mandated states (1,2). However, it is less clear if there are regional differences in the United States (US) with regards to these practices and their outcomes. OBJECTIVE: The objective of this study was to determine whether there is a difference in embryo transfer practices and ART outcomes based on US region. MATERIALS AND METHODS: This was a retrospective cohort study utilizing the SART registry to identify fresh autologous IVF cycles performed between 2011 to 2012 in women under the age of 38. Only first IVF cycles were included in the analysis. Cycles were excluded if the indication for IVF was ‘‘preimplantation genetic diagnosis’’ or ‘‘embryo banking.’’ The four regions of the US (Northeast, Midwest, South, and West) were identified based on US Census Bureau. The cycles were then further stratified into stage of embryo transfer. Main outcomes were the number of embryos transferred, live birth rate (LBR) and multiple birth rate (MBR). Secondary outcomes included percentage of cycles with elective single embryo transfer (eSET) and clinical pregnancy rate (CPR). ANOVA test was used to analyze the significance of clinical variables across all regions. Post-hoc analysis was then performed using Tukey’s test and the resulting p-values were adjusted for multiple comparisons. RESULTS: Of the 46,864 cycles included in the analysis, 29.1% were performed in the Northeast, 20.6% in the Midwest, 31.6% in the South, and 18.7% in the West. Although the profiles of the patients and stimulation characteristics as shown in table 1 and 2 were statistically different due to the large sample size, these differences were not clinically relevant. The proportion of cycles with eSET was much higher with blastocyst embryo transfers than cleavage stage embryo transfers. Moreover, the Midwest had the lowest percentage of eSET cycles for blastocyst transfers while the Northeast had the highest percentage of eSET cycles for cleavage stage transfers compared to the other regions. The CPR and LBR were very similar across all regions for both cleavage and blastocyst transfers. All four regions had a high MBR for both cleavage stage (range: 24.2-28.7%) and blastocyst (range: 31.134.1%) embryos. CONCLUSIONS: Despite differences noted in embryo transfer practices and ART outcomes by state and by insurance mandate status, the same is not seen based on US region. The MBR remains high across the US for both cleavage and blastocyst transfers for patients undergoing their first fresh autologous transfer.

TABLE 1. Patient Profile, Stimulation Characteristics, and Pregnancy Outcomes for Cleavage Stage Embryo Transfer.

N Mean age Male factor (%) Tubal factor (%) Unexplained Infertility (%) PCOS (%) Diminished Ovarian reserve (%) Other Infertility (%) # Oocytes retrieved # Embryos transferred % eSET % cycles with cryopreservation # Embryos cryopreserved CPR LBR MBR

Northeast

Midwest

South

West

Anova P Value

6553 33.2
3.18 37.6 15 21.3 15.3 12.4 17.1 11.2
6.50 2.0
0.59 7 38.3 3.4
3.02 46.2 39.3 24.2

5956 32.5
3.36 45.8 12.4 14.3 18.5 13.3 19.5 10.9
6.32 2.1
0.52 1.7 20.7 4.2
3.69 45.6 39.5 26.6

9551 33.0
3.35 39.5 17.7 13.6 17.7 17.4 21.4 10.7
6.63 2.1
0.53 1.4 24.1 3.4
2.88 45.7 38.9 28.7

5696 33.5
3.22 45 14.1 12.8 11.7 23.2 21.1 11.5
7.08 2.2
0.66 2.1 17 3.7
3.26 47.7 41 25.4

<0.0001 <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 <0.05 <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 0.28 <0.05

p-value <0.05 significant.

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PCRS Abstracts

Vol. 105, No. 2, Supplement, February 2016

TABLE 2. Patient Profile, Stimulation Characteristics, and Pregnancy Outcomes for Blastocyst Embryo Transfer.

N Mean age Male factor (%) Tubal factor (%) Unexplained Infertility (%) PCOS (%) Diminished Ovarian reserve (%) Other Infertility (%) # Oocytes retrieved # Embryos transferred % eSET % cycles with cryopreservation # Embryos cryopreserved CPR LBR MBR

Northeast

Midwest

South

West

Anova P Value

7081 35.5
3.28 37.4 15.4 18.8 20.3 4.9 23 16.5
7.63 1.7
0.53 24.3 23.4 4.1
2.96 60 51.3 31.1

3698 31.9
3.34 40.4 15.4 17.5 23.7 6.6 20.3 15.6
7.34 1.8
0.45 17.3 20.4 4.2
3.25 60.4 53.5 34.1

5257 32.3
3.35 40.3 17.6 14.2 21.3 7.1 21.7 16.2
7.86 1.7
0.50 23.4 34.2 4.3
3.13 59.8 52.3 31.2

3072 32.6
3.37 44.5 13 13.6 18.1 10.6 21.9 17.8
8.66 1.8
0.52 22.4 22 4.6
3.36 61.8 54.1 31.9

<0.0001 <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 <0.05 <0.05 <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 <0.05 <0.05

p-value <0.05 significant. FINANCIAL SUPPORT: None. References: 1. Sunderam, S., et al., Assisted Reproductive Technology SurveillanceUnited Sates, 2012. MMWR Surveill Summ, 2015. 64(6): p. 1-29. 2. Boulet, SL., et al, Embryo transfer practices and perinatal outcomes by insurance mandate status, Fertil Steril, 2015. 104(2): p. 402-9.

P-32 ANALYTICAL VALIDATION OF A NOVEL NEXT-GENERATION SEQUENCING BASED PREIMPLANTATION GENETIC SCREENING TECHNOLOGY. J. Gole,a T. Mullen,a G. Celia,b C. Wagner,c B. Kaplan,d M. Katz-Jaffe,e W. Schoolcraft,e M. Umbarger.a aGood Start Genetics Inc., Cambridge, MA, USA; bDominion Fertility, Arlington, VA, USA; cGlobal Genetics Institute LLC, Highland Park, IL, USA; dFertility Centers of Illinois, Highland Park, IL, USA; eColorado Center for Reproductive Medicine, Lone Tree, CO, USA. BACKGROUND: Current data indicate that preimplantation genetic screening (PGS) increases live birth and decreases miscarriage rates. However, high cost has limited its adoption. This may be overcome by utilizing more efficient and streamlined workflows employing next-generation DNA sequencing (NGS). We have developed a new method for PGS that couples NGS with a technology termed FAST-SeqS which utilizes a PCR reaction to amplify >10,000 repetitive elements from all chromosomes while simultaneously attaching sequencing adapters and sample specific molecular barcodes, thereby representing a substantially more streamlined workflow than that of previously reported NGS-based PGS technologies. OBJECTIVE: To evaluate the analytical accuracy of the targeted FASTSeqS PGS technology using well-characterized cell lines and trophectoderm (TE) biopsies obtained from blastocyst stage embryos. MATERIALS AND METHODS: Cells from fibroblast cell lines and TE biopsies of embryos were tested using FAST-SeqS, and the resulting molecular karyotype calls were compared to either G-banding-derived karyotypes (cell lines) or array comparative genomic hybridization (aCGH)-derived molecular karyotypes (TE biopsies). MATERIALS AND METHODS: Six aneuploid [47,XY,+9; 47,XY,+13; 47,XX,+18; 47,XY,+21; 47,XXY; 47,XYY] and two euploid [46,XX; 46,XY] cell lines were micromanipulated to yield samples containing 5 cells that were tested using our FAST-SeqS assay. In the embryo studies, paired TE biopsies were obtained from previously cryopreserved blastocysts. One biopsy was tested using FAST-SeqS and the other was tested using Illumina 24sure aCGH (with blinded independent calling by two operators). Discrepancies between the platforms were resolved by re-analyzing the 24sure whole genome amplification product using the Illumina VeriSeq PGS workflow. RESULTS: For cell lines, 168 aneuploid and 55 euploid samples were correctly called, yielding 100% sensitivity and specificity. TE biopsies from a total of 265 embryos yielded a concordance rate in line with the previously reported accuracy of aCGH (Gutierrez-Mateo et al., 2011) and previously reported blastocyst diploid-aneuploid mosaicism rates(Capalbo et al., 2013; Taylor et al., 2015). Out of a total of 6509 chromosomes analyzed,

FERTILITY & STERILITYÒ

16 false positive and 5 false negative calls were made. Most of the chromosome-level discrepancies (10 false positive and 4 false negative) occurred in five samples that were correctly classified as aneuploid, indicating that these samples were likely aneuploid-aneuploid mosaic, a relatively common form of mosaicism(Capalbo et al., 2013; Taylor et al., 2015). CONCLUSIONS: Our NGS-based PGS approach yields molecular karyotype calls that are both highly accurate and consistent with traditional aCGH calls. The cost effectiveness of the technology should enable expanded access to PGS. SUPPORT: This study was funded by Good Start Genetics. References: 1. Capalbo, A. et al. FISH reanalysis of inner cell mass and trophectoderm samples of previously array-CGH screened blastocysts shows high accuracy of diagnosis and no major diagnostic impact of mosaicism at the blastocyst stage. Hum Reprod, v. 28, n. 8, p. 2298-307, Aug 2013. 2. Gutierrez-Mateo, C. et al. Validation of microarray comparative genomic hybridization for comprehensive chromosome analysis of embryos. Fertil Steril, v. 95, n. 3, p. 953-8, Mar 2011. 3. Taylor, T. H. et al. Mosaicism in the trophectoderm is present albeit at a low rate of clinical significance. Fertility and Sterility, v. 104, n. 3, p. e282-e283. P-33 THE EFFECT OF PYRROLOQUINOLINE QUINONE ON MEIOTIC PROGRESSION AND METABOLIC PARAMETERS IN DIABETIC MOUSE OOCYTES. Violet E. Klenov, Christina E. Boots, Wendy Zhang, Kelle H. Moley. OBGYN, Washington University, St. Louis, MO, United States. BACKGROUND: Diabetes produces oocyte defects secondary to meiotic spindle abnormalities leading to embryonic aneuploidy and functional abnormalities of oocyte mitochondria[1]. We have previously demonstrated that supplementation of in vitro maturation media (IVM) with pyrroloquinoline quinone (PQQ, an antioxidant) improves meiotic progression in oocytes from mice fed a high-fat-diet as compared to controls. Given the continued epidemic of diabetes and metabolic syndrome, this study is particularly relevant. OBJECTIVE: To determine whether maternal diabetes and its resulting metabolic insults to the oocyte are reversible after supplementation of IVM media with PQQ. MATERIALS AND METHODS: B6SJLF1 mice (age 21-28d) received streptozotocin (200mg/kg). After four days, mice with a random glucose >300 mg/dl were injected with pregnant mare serum gonadotropin (5IU), and germinal vesicle (GV) stage oocytes were collected and matured in vitro with PQQ-supplemented (50uM) vs. control media. Mature oocytes were stained with anti-a tubulin antibody and DAPI in order to visualize spindles and chromosomes. Fifteen mature oocytes from each group were analyzed for concentrations of adenosine triphosphate (ATP), phosphocreatine (PCr) and citrate. Imaging was performed on a Leica confocal microscope and analysis performed blindly with ImageJ software. ANOVA, students t-tests, and chi-square analysis were used for statistical analysis.

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