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Miscarriage history association with euploid embryo transfer outcomes
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RBMO VOLUME 39 ISSUE 4 2019
ARTICLE
Miscarriage history association with euploid embryo transfer outcomes BIOGRAPHY
Ange Wang is currently an Obstetrics & Gynecology resident at Stanford Hospital. She graduated from the University of Pennsylvania with a BSE in Bioengineering and Stanford Medical School with an MD. She has published on outcomes in both assisted reproductive technology and oncology and co-authored numerous publications in the Women's Health Initiative. Ange Wang*, Jonathan Kort, Lynn Westphal KEY MESSAGE
This study suggests that a history of miscarriage prior to IVF (including a history of recurrent pregnancy loss) is not associated with frozen embryo transfer outcomes for euploid embryos. As many IVF patients have a history of pregnancy loss, this is an important finding for counselling and prognostication. ABSTRACT
Research question: Is a history of miscarriage (including recurrent pregnancy loss) associated with euploid cryopreserved embryo transfer outcomes? Design: Retrospective cohort study from 2014 to 2018 of patients at an academic medical centre, undergoing their first cycle of IVF with 24-chromosome Day 5/6 preimplantation genetic testing for aneuploidies (IVF-PGT-A). Multivariate logistic regression was used to investigate the relationship between history of miscarriage and euploid single cryopreserved embryo transfer outcomes (ongoing pregnancy, miscarriage), adjusting for an extensive list of patient and cycle confounders. Results: In the study cohort of 283 patients, the overall unadjusted positive beta human chorionic gonadotrophin (bHCG) rate was 70.0%, ongoing pregnancy rate was 52.3%, and the total pregnancy loss (biochemical and clinical pregnancy loss) rate per positive bHCG cycle was 24.7%. While 35.3% of patients had a history of at least one previous miscarriage, 14.5% of patients had a history of recurrent pregnancy loss (RPL). For patients with a history of miscarriage, it was found that the adjusted odds ratios (OR) and 95% confidence intervals (CI) for positive bHCG were 1.30 (0.51– 3.27), for ongoing pregnancy were 0.88 (0.38–2.03) and for total pregnancy loss were 1.41 (0.49–4.05), when compared with patients without a history of miscarriage. For RPL patients, OR for positive bHCG, ongoing pregnancy and total pregnancy loss also did not significantly differ when compared with patients with no history of miscarriage. Conclusions: In this cohort, there was no significant association between miscarriage history and euploid cryopreserved embryo transfer outcomes (ongoing pregnancy, total pregnancy loss) after adjustment for potential confounders. Further study in larger data sets is warranted. Department of Obstetrics and Gynecology, Stanford University School of Medicine, Stanford CA 94305, USA © 2019 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved. *Corresponding author. E-mail address: [email protected] (A Wang). https://doi.org/10.1016/j.rbmo.2019.05.011 1472-6483/© 2019 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved. LW has sponsored research through Celmatix Inc. which does not pertain to this study. No other conflicts of interest to disclose from any other authors. These data were presented in part as a poster presentation at the American Society for Reproductive Medicine (ASRM) Annual Meeting 2018 in Denver, Colorado, USA.
KEYWORDS
IVF Euploid embryos Miscarriage Preimplantation genetic testing for aneuploidies Recurrent pregnancy loss
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INTRODUCTION
M
iscarriage is estimated to occur in 8–20% of clinically recognized pregnancies, and at an even higher rate in unrecognized or subclinical pregnancies (Lohstroh et al., 2005; Wang et al., 2003; Wilcox et al., 1988). Furthermore, a smaller percentage of women (<5%) experience recurrent pregnancy loss (RPL) of two or more clinical miscarriages (Practice Committee of the American Society for Reproductive Medicine, 2012). Most early miscarriages are believed to be genetic in origin, and studies suggest that women with a history of miscarriage have increased risk of subsequent miscarriage or infertility (Lund et al., 2012; Regan et al., 1989). The literature on associations between history of miscarriage and transfer outcomes for IVF cycles is limited, particularly on the subject of euploid embryos. This association may be confounded by the fact that miscarriage history may also be linked to other factors that are poor IVF prognosticators, such as diminished ovarian reserve or increased maternal age. Existing findings on this subject have been mixed, and no studies to date have investigated this subject for euploid embryo transfers in a multivariate analysis. Some studies have reported that preimplantation genetic testing for aneuploidies (PGT-A) is a beneficial intervention in patients with history of either idiopathic pregnancy loss or miscarriage due to specific causes (Garrisi et al., 2009; Hodes-Wertz et al., 2012; Otani et al., 2006), while other studies have not found this effect (Murugappan et al., 2016). A large German study found that a previous natural conception was not associated with the outcome of an IVF cycle, although the findings were mixed on previous IVF conception (Kupka et al., 2003). Several studies have also examined general reproductive history following miscarriage and have found associations of miscarriage history with infertility, subsequent miscarriage and poor obstetric outcomes (Jivraj et al., 2001; Kling et al., 2018; Lund et al., 2012). Miscarriage has also been studied as an IVF outcome, with known associations to maternal age, ovarian reserve, embryo morphology, body mass index (BMI) and anti-Müllerian hormone (AMH) concentrations (Hill et al.,
2013; Honnma et al., 2012; Metwally et al., 2008; Padilla and Garcia, 1989; Spandorfer et al., 2004; Tarasconi et al., 2017; Toner et al., 1993; Yan et al., 2012). As chromosomal abnormalities account for the majority of early miscarriages and miscarriage history is a known prognosticator for future pregnancy outcomes as above, investigation is warranted into whether or not miscarriage history affects transfer outcomes of euploid embryos. This is the first study to investigate this question in a multivariate analysis, and a further aim of the study was to determine whether or not patients with a history of one miscarriage differed from RPL patients in this context. A larger cohort of cycles from the same patients studied in this manuscript was also used for a separate analysis on the association of AMH with euploid transfer outcomes, in investigating additional prognosticators of euploid transfer outcomes (Wang et al., 2019).
MATERIALS AND METHODS This was a retrospective cohort study from 2014 to 2018 of patients at an academic medical centre undergoing IVF with 24-chromosome Day 5/6 PGT-A. PGT-A was chosen by patients for a variety of reasons, including but not limited to advanced maternal age, diminished ovarian reserve, RPL, sex selection and single gene disorder (see Supplementary Table 1 for primary IVF indications). Inclusion criteria were single transfers of euploid embryos, autologous embryo transfer and available information on all potential patient and cycle confounders included in this study (see Statistical analysis section below). Only women undergoing their first IVF cycle at the study institution were included in the analysis, to prevent the study being confounded by previous IVF miscarriages (which may be detected at a higher rate than naturally conceived miscarriages). Institutional Review Board approval was obtained for this study from Stanford University (which is affiliated to the IVF clinic) on 30 June 2018 (reference number 6208). Ovarian stimulation was performed using one of three protocols: antagonist, long gonadotrophin-releasing hormone (GnRH) agonist (leuprolide acetate), or microdose flare. Transvaginal oocyte retrieval occurred 35–36 h after
HCG and/or Lupron trigger (given when the largest follicle measured >18 mm). Oocytes were fertilized using conventional IVF or intracytoplasmic sperm injection (ICSI). Methods for blastocyst stage culture have been previously reported in the literature (Kort et al., 2015), using SAGE™ sequential media (CooperSurgical, Trumbull, CT, U.S.A.) in low-oxygen conditions. Embryologists performed laser-assisted hatching on Day 3 after retrieval, followed by trophectoderm biopsy on either Day 5 or Day 6 depending on the rate of embryo development. Using the laser, three to five cells were removed and samples were then analysed for 24-chromosome screening using IviGen (Miami, Los Angeles, and New York) with nextgeneration sequencing (NGS). One of six senior embryologists conducted morphological assessments on Day 5 or Day 6 according to the Gardner grading system (Gardner and Schoolcraft, 1999) Using standardized laboratory protocols, blastocysts were assigned grades for expansion (1–6), inner cell mass (ICM) (A, B or C) and trophectoderm (A, B or C). A very small percentage of embryos had expansion ≤2, and therefore were unable to receive ICM and trophectoderm grades due to insufficient development. Embryos were vitrified on either Day 5 or 6 after biopsy. After receiving biopsy results, one euploid embryo was warmed and transferred in a natural or hormone replacement cycle (typically utilizing oral oestrogen and both intramuscular and vaginal progesterone supplementation). A retrospective chart review was conducted to record cycle outcome, as well as all variables included in this analysis (see below), which included a mixture of patient and cycle characteristics that may potentially affect IVF cycle outcomes. History of miscarriage was self-reported and could include either biochemical or clinical miscarriage, as patients did not always specify type of miscarriage in reports of miscarriage history. Statistical analysis Multivariable logistic regression was used to investigate the associations between history of miscarriage (prior to IVF cycle) and transfer outcomes (positive bHCG, ongoing pregnancy, miscarriage) for euploid single cryopreserved embryo transfers. Patients were also classified into
two groups: history of one miscarriage, or history of two or more miscarriages (designated as RPL). The main analysis investigated history of miscarriage in a multivariate analysis, while a sub-analysis investigated the number of miscarriages in a separate multivariate model.
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TABLE 1 BASELINE CHARACTERISTICS OF THE PATIENTS Miscarriage history No
Yes
Overall
Total
183
100
283
Gravidity
61 (33.3)
100 (100.0)
161 (56.9)
<0.001
In terms of outcomes, positive bHCG was defined as concentration >5 mIU/ml, ongoing pregnancy was defined as continued pregnancy at 8–12 weeks with positive fetal cardiac activity, and total pregnancy loss included both biochemical and clinical miscarriage. Biochemical miscarriage was defined as loss of pregnancy with positive HCG but no ultrasound findings, while clinical miscarriage was defined as loss of a pregnancy after some ultrasound findings were present (at least a gestational sac). The analysis controlled for the following patient and cycle confounders (all of which were included in the multivariate model): primary IVF indication, history of pregnancy, history of live birth, maternal age at retrieval, maternal age at transfer, AMH, BMI, ICSI, oocyte number, number of blastocysts, morphology (expansion, ICM and trophectoderm), hormone replacement versus natural cryopreserved embryo transfer, and endometrial thickness. All tests were two-sided with significance at the alpha = 0.05 level, and Python and Microsoft Excel were used for data analysis.
Parity
46 (25.1)
38 (38.0)
84 (29.7)
0.024
RESULTS
a
Recurrent pregnancy loss – two or more miscarriages. Not applicable for group without history of miscarriage.
b
Indications may overlap, does not sum to 100%
In the study cohort of 283 patients, the overall unadjusted positive bHCG rate was 70.0% (198 patients), ongoing pregnancy rate was 52.3% (148 patients), and the total pregnancy loss rate per cycle with positive bHCG was 24.7% (49 patients) (7.6% clinical [15 patients], 17.2% biochemical [34 patients]). It was found that 35.3% of patients had a history of at least one previous miscarriage, while 14.5% of patients had a history of RPL (TABLE 1). The two groups had similar ages at retrieval and transfer, with retrieval age 35.6 years (SD 3.8) for the no miscarriage group and 35.9 years (SD 3.6) for the miscarriage group; age at transfer was 36.0 (SD 3.8) and 36.3 (SD 3.7) years, respectively. BMI (23.7 versus 24.2) and distribution of AMH categories was also similar between the two groups. The leading primary indications for IVF were male factor (25.4%), followed by diminished ovarian reserve (19.8%) and ovulatory dysfunction (11.3%)
History of
RPLa
P-value
–
41 (41.0)
41 (14.5)
–
Age at retrieval, mean (SD)
35.6 (3.8)
35.9 (3.6)
35.7 (3.7)
NS
Age at transfer, mean (SD)
36.0 (3.8)
36.3 (3.7)
36.1 (3.8)
NS
BMI, mean (SD)
23.7 (4.8)
24.2 (4.4)
23.8 (4.6)
NS
<1
31 (16.9)
18 (18.0)
49 (17.3)
NS
1 to <2
42 (23.0)
21 (21.0)
63 (22.3)
2 to <5
69 (37.7)
41 (41.0)
110 (38.9)
5+
41 (22.4)
20 (20.0)
61 (21.6)
DOR
47 (25.7)
24 (24.0)
71 (25.1)
Endometriosis
20 (10.9)
5 (5.0)
25 (8.8)
Male
75 (41.0)
25 (25.0)
100 (35.3)
Ovulatory
16 (8.7)
18 (18.0)
34 (12.0)
RPL
0 (0.0)
29 (29.0)
29 (10.2)
Tubal
9 (4.9)
12 (12.0)
21 (7.4)
Sex selection
8 (4.4)
5 (5.0)
13 (4.6)
Single gene disorder
13 (7.1)
3 (3.0)
16 (5.7)
Uterine
10 (5.5)
6 (6.0)
16 (5.7)
Unexplained
19 (10.4)
9 (9.0)
28 (9.9)
Other
11 (6.0)
1 (1.0)
12 (4.2)
AMH category (ng/ml)
IVF
indicationb <0.001
Data are presented as n (%) unless otherwise stated.
AMH = anti-Müllerian hormone; BMI = body mass index; DOR = diminished ovarian reserve; NS = not statistically significant; RPL = recurrent pregnancy loss.
(Supplementary Table 1), although many patients had multiple indications for IVF. In terms of cycle characteristics, an average of 15.5 oocytes (SD 8.3) were retrieved and the number of blastocysts obtained was 5.5 (SD 3.8) for the entire cohort (TABLE 2). The miscarriage and no miscarriage group were similar in terms of the number of oocytes retrieved, number of blastocysts obtained, expansion grades (over 80% 5 or 6 for both groups), trophectoderm grades, and type of transfer protocols. The two groups also had similar endometrial thickness (9.4, SD 1.6 and 9.0, SD 1.5 for no miscarriage and miscarriage history, respectively). Compared with the no miscarriage group, the miscarriage history group was less likely to use ICSI, and have lower ICM grade (see TABLE 2).
In this analysis, for patients with a history of miscarriage (compared with patients with no history of miscarriage), it was found that the adjusted odds ratio (OR) and 95% confidence interval (CI) for positive bHCG was 1.30 (0.51–3.27) and OR for ongoing pregnancy was 0.88 (0.38–2.03) (TABLE 3). It was also found that when compared with women without a history of miscarriage, the OR for positive HCG was 1.45 (0.54–3.85) for a history of one miscarriage and 0.89 (0.22–3.52) for RPL patients. For ongoing pregnancy, OR was 0.91 (0.38–2.19) for a history of one miscarriage, and 0.78 (0.22–2.81) for RPL patients. Depending on miscarriage category, the unadjusted positive HCG rates ranged from 65.9% to 76.3%, and ongoing pregnancy rates ranged from 43.9% to 53.6% (see TABLE 3 for full details).
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DISCUSSION
TABLE 2 CYCLE CHARACTERISTICS Miscarriage history No
Yes
Overall
P-value
Oocytes retrieved, mean (SD)
15.3 (8.1)
16 (8.7)
15.5 (8.3)
NS
Number of blastocysts, mean (SD)*
5.4 (3.5)
5.7 (4.4)
5.5 (3.8)
NS
No
37 (20.2)
36 (36.0)
73 (25.8)
0.014
Yes – all ICSI
127 (69.4)
57 (57.0)
184 (65.0)
ICSI split
19 (10.4)
7 (7.0)
26 (9.2)
5 to 6
152 (83.1)
80 (80.0)
232 (82.0)
3 to 4
30 (16.4)
20 (20.0)
50 (17.7)
Less than 3
1 (0.5)
0 (0.0)
1 (0.4)
A
105 (57.4)
45 (45.0)
150 (53.0)
B
76 (41.5)
51 (51.0)
127 (44.9)
C
1 (0.5)
4 (4.0)
5 (1.8)
A
85 (46.4)
37 (37.0)
122 (43.1)
B
93 (50.8)
56 (56.0)
149 (52.7)
C
4 (2.2)
7 (7.0)
11 (3.9)
Medicated
64 (35.0)
44 (44.0)
108 (38.2)
Natural
119 (65.0)
56 (56.0)
175 (61.8)
Endometrial thickness (mm), mean (SD)
9.4 (1.6)
9.0 (1.5)
9.2 (1.6)
ICSI
Morphology Expansion NS
ICM 0.023
Trophectoderm NS
Type of embryo transfer NS
NS
Data are presented as n (%) unless otherwise stated. Note: One embryo was collapsed and did not receive ICM or trophectoderm grade. ICM = inner cell mass; ICSI = intracytoplasmic sperm injection; NS = not statistically significant.
For total pregnancy loss (per cycle with positive bHCG) as an outcome (TABLE 4), an unadjusted total pregnancy loss rate of 21.4% (27 patients) was found for patients in the no miscarriage group, compared with 30.6% (22 patients) for the overall miscarriage group. TABLE 4 also shows the breakdown by clinical and biochemical
pregnancy loss. In the multivariate analysis, when compared with patients with no miscarriage, the adjusted OR for total pregnancy loss outcome was 1.41 (0.49–4.05) for the overall group of miscarriage patients, 1.48 (0.49–4.43) for a history of one miscarriage, and 1.21 (0.27–5.37) for RPL patients.
In summary, in this multivariate analysis of euploid single cryopreserved embryo transfers, it was found that history of miscarriage was not associated with positive HCG, ongoing pregnancy or total pregnancy loss outcomes. Euploid embryos are estimated to implant at a rate of 60–70%, and data on euploid miscarriage is limited (Scott et al., 2013; Yang et al., 2012). The factors that make a euploid transfer result in miscarriage or fail to implant are not well understood; proposed factors have included the uterine lining, uterine microbiome or other environmental factors, genetic defects or aneuploidy in the embryo that cannot be detected through PGT-A, immunologic causes such as antiphospholipid syndrome, endocrine causes, damage to the embryos during freezing/thawing/biopsy, among others (Maxwell et al., 2016; Meldrum, 2016; Meldrum and de Ziegler, 2016). Existing literature on associations between history of miscarriage and transfer outcomes for IVF cycles is extremely limited, particularly on the subject of euploid embryos. This association may also be confounded by the fact that miscarriage history may be linked to other factors that may be poor prognosticators for an IVF cycle, such as ovarian reserve or maternal age. It is estimated that miscarriage as an outcome occurs in 8–20% of clinically recognized pregnancies, and at an even higher rate in unrecognized or subclinical pregnancies (Lohstroh et al., 2005; Wang et al., 2003; Wilcox et al., 1988). A much smaller percentage of women (<5%) experience RPL (two or more miscarriages), with an even smaller percentage (1%) experiencing three or more (Practice Committee of
TABLE 3 EUPLOID EMBRYO TRANSFER OUTCOMES BY MISCARRIAGE HISTORY No. of patients
Maternal age at retrieval (years)
PB rate (%)
Adjusted OR PB, OR (95% CI)
OP rate (%)
Adjusted OR OP, OR (95% CI)
No miscarriage
183
35.7
68.9
Reference
53.6
Reference
History of miscarriage
100
35.8
72.0
1.30 (0.51–3.27)
50.0
0.88 (0.38–2.03)
History of 1 miscarriage
59
35.5
76.3
1.45 (0.54–3.85)
54.2
0.91 (0.38–2.19)
History of 2 or more miscarriages (RPL)
41
36.1
65.9
0.89 (0.22–3.52)
43.9
0.78 (0.22–2.81)
No P-values were statistically significant. Confounders included primary IVF indication, history of pregnancy, history of live birth, maternal age at retrieval, maternal age at transfer, AMH, ICSI, oocyte number, number of blastocysts, morphology (expansion, ICM and trophectoderm), hormone replacement versus natural cryopreserved embryo transfer, and endometrial thickness. AMH = anti-Müllerian hormone; bHCG = beta human chorionic gonadotrophin; BMI = body mass index; CI = confidence interval; ICM = inner cell mass; ICSI = intracytoplasmic sperm injection; OP = ongoing pregnancy; OR = odds ratio; PB = positive bHCG; RPL = recurrent pregnancy loss.
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TABLE 4 PREGNANCY LOSS RATES FOLLOWING EUPLOID EMBRYO TRANSFER ACCORDING TO MISCARRIAGE HISTORY Biochemical pregnancy loss rate (%)
Clinical miscarriage rate (%)
TPL rate (%)a
Adjusted OR TPL, OR (95% CI)
No miscarriage
15.1
6.3
21.4
Reference
History of miscarriage
20.8
9.7
30.6
1.41 (0.49–4.05)
History of 1 miscarriage
20.0
8.9
28.9
1.48 (0.49–4.43)
History of 2 or more miscarriages (RPL)
22.2
11.1
33.3
1.21 (0.27–5.37)
No P-values were statistically significant. Confounders included primary IVF indication, history of pregnancy, history of live birth, maternal age at retrieval, maternal age at transfer, AMH, BMI, ICSI, oocyte number, number of blastocysts, morphology (expansion, ICM and trophectoderm), hormone replacement versus natural cryopreserved embryo transfer, and endometrial thickness. a Total
pregnancy loss includes both biochemical and clinical pregnancy loss. All percentages are per cycle with positive bHCG.
AMH = anti-Müllerian hormone; BMI = body mass index; BPL = biochemical pregnancy loss; CM = clinical miscarriage; ICM = inner cell mass; ICSI = intracytoplasmic sperm injection; NS = not statistically significant; OP = ongoing pregnancy; OR = odds ratio; TPL = total pregnancy loss.
the American Society for Reproductive Medicine, 2012). Known risk factors for miscarriage include previous miscarriage or RPL, advancing maternal age and smoking (Chatenoud et al., 1998; Nybo et al., 2000; Regan et al., 1989). Given that chromosomal abnormalities are believed to account for the majority of early miscarriages, and that miscarriage history is a known prognosticator for future pregnancy outcomes, further elucidation is needed into whether or not miscarriage history affects transfer outcomes of euploid embryos. Few studies have investigated the linkage between miscarriage history and IVF outcomes, and no studies to date have investigated this subject for euploid embryo transfers in a multivariate analysis. One study of 174,909 assisted reproductive technology (ART) cycles in Germany from 1998 to 2000 found that a previous natural conception (miscarriage or live birth) did not significantly affect the outcome of IVF cycles. However, a previously successful ART procedure ending in live birth had a statistically significant positive effect on IVF outcome, while an ART procedure ending in miscarriage had a significant negative effect (Kupka et al., 2003). Another study showed that the use of preimplantation genetic diagnosis improved pregnancy outcomes for the specific population of translocation carriers with recurrent miscarriages, but it was not clear whether this could be generalized to patients without translocations (Otani et al., 2006). Studies have differed on the value of PGT-A for idiopathic RPL patients; one study of 287 cycles reported a significant decrease in the miscarriage rate among RPL patients using PGT-A (Hodes-Wertz et al., 2012). A 2009 study also reported
that PGT-A of nine chromosomes decreased miscarriage rates among idiopathic RPL patients, particularly for women over age 35 (Garrisi et al., 2009). Another study of over 1800 patients found that clinical pregnancy loss rate following IVF treatment (without PGT-A) was lower than for RPL patients who conceived spontaneously (Tamhankar et al., 2015). In contrast, a study of 300 patients reported that PGT-A did not improve miscarriage, pregnancy or live birth rates among RPL patients when compared with expectant management (Murugappan et al., 2016). Studies on the subject of miscarriage history and IVF outcomes are limited as above, and there has also been literature investigating general reproductive history following miscarriage. A cohort study of 987 Danish women with recurrent miscarriage found that there was a significantly decreased chance (P < 0.01) of at least one subsequent live birth with increasing number of miscarriages (Lund et al., 2012). Other studies have reported descriptively on subsequent fertility and obstetrics outcomes in RPL patients (Kling et al., 2018), with one study suggesting that RPL patients had a significantly higher risk of preterm delivery, small for gestational age, perinatal loss, and Caesarean section compared with a control group; however, this study did not investigate fertility rates in this population (Jivraj et al., 2001). Miscarriage has also been studied as an IVF outcome, with known associations with maternal age and ovarian reserve (Padilla and Garcia, 1989; Spandorfer et al., 2004; Toner et al., 1993; Yan et al., 2012), embryo morphology (Hill et al., 2013; Honnma et al., 2012), BMI (Metwally et al., 2008; Veleva et al., 2008) and AMH concentrations (Tarasconi et al., 2017).
This study is novel compared with previous studies by using a multivariate analysis to investigate IVF outcomes for euploid embryos in relation to miscarriage history (both overall miscarriage as well as RPL). A multivariate analysis is very important for this subject to control for confounders, as miscarriage history has been established to be linked to poor prognosticators of IVF cycles as detailed above. Similar to the large German study, it was found that a previous miscarriage (non-IVF) did not significantly affect the results of IVF cycles in terms of either miscarriage or ongoing pregnancy. Although patients with a history of miscarriage had non-significant elevated OR for miscarriage as an outcome in this study, no directionality was observed for RPL patients compared with patients with history of one miscarriage, providing additional confidence in these results. As the majority of miscarriages are due to genetic causes, the findings are not unexpected. In combination with previous literature, this study suggests that a history of miscarriages, either isolated or recurrent, is not a poor prognosticator for IVF/PGT-A cycles. Although some studies have suggested that miscarriage history (particularly RPL) may be negatively associated with subsequent natural conception, this study supports IVF/PGT-A as an intervention for RPL patients to consider. However, findings must be interpreted with caution given the sample size, retrospective nature of the study and limited literature on the subject. The strengths of this study include the novel subject, detailed information on potential patient and cycle confounders which were used for a multivariate analysis, information on both miscarriage
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and ongoing pregnancy as outcomes, and single institution format, which allows for more standardized data collection. In particular, this is the first study to investigate the association between miscarriage history and IVF outcomes of euploid embryos in a multivariate analysis. Given the high prevalence of miscarriage (particularly among patients seeking IVF treatment), this study is very applicable clinically. Limitations of the study include the retrospective format, the relatively small sample size, self-report of miscarriage outcomes (which may be subject to recall bias) and lack of information on live birth outcomes. Some OR had wide CI and it is possible that the sample size limited the ability to find a difference in some associations. The study is also limited by the fact that the findings cannot be generalized to all patients with a history of miscarriage, but only those who were able to produce euploid embryos for transfer, which likely represents a better prognosis group. The study included patients with a history of miscarriage prior to starting IVF treatment at the study institution, so the findings are not necessarily generalizable to patients with a previous IVF miscarriage (which may be detected at a higher rate than spontaneous miscarriages); this area should be studied given mixed findings previously reported in the literature. There was no information available on type of miscarriage (biochemical versus clinical), and additional study is warranted on whether history of miscarriage type is a prognosticator. Also, it was not possible to differentiate between idiopathic miscarriage and miscarriage with a known cause such as antiphospholipid syndrome, although these aetiologies of miscarriages are relatively rare. This cohort was also probably better than average in terms of prognosis based on a higher number of oocytes retrieved, AMH and average maternal age of 35–36. Therefore, patient counselling on miscarriage history in relation to euploid embryo transfer must include these important caveats. In summary, using a multivariate analysis, it was found that history of miscarriage was not associated with IVF outcomes (positive bHCG, ongoing pregnancy, total pregnancy loss) for single embryo transfer of euploid embryos. These findings are encouraging for patients with miscarriage or RPL history who have
euploid embryos available to transfer. However, further study is warranted in larger cohorts and in a prospective format, particularly as OR and absolute rates for miscarriage outcome were elevated but not significant for patients with history of miscarriage or RPL. Other areas for future investigation (in terms of euploid embryo transfer outcomes) include whether or not previous IVF miscarriage is a different prognosticator than previous natural conception miscarriage, or if associations vary based on type of miscarriage history.
ACKNOWLEDGEMENTS The authors would like to acknowledge Arthur Chiang for guidance on data analysis, and Qianyang Zhao for assistance with procuring data for analysis.
SUPPLEMENTARY MATERIALS Supplementary material associated with this article can be found in the online version at doi:10.1016/j.rbmo.2019.05.011.
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comparing the rate of aneuploidy within presumed euploid embryos that resulted in miscarriage or live birth using next-generation sequencing. Fertility and sterility 2016; 106 Meldrum, D.R. Introduction: Examining the many potential reasons why euploid blastocysts do not always result in viable pregnancies: Part 1. Fertility and sterility 2016; 105: 545–547 Meldrum, D.R., de Ziegler, D. Introduction: Examining the many potential reasons why euploid blastocysts do not always result in viable pregnancies (and deliveries): Part 2. Fertility and sterility 2016; 105: 841–843 Metwally, M., Ong, K.J., Ledger, W.L., Li, T.C. Does high body mass index increase the risk of miscarriage after spontaneous and assisted conception? A meta-analysis of the evidence. Fertility and sterility 2008; 90: 714–726 Murugappan, G., Shahine, L.K., Perfetto, C.O., Hickok, L.R., Lathi, R.B. Intent to treat analysis of in vitro fertilization and preimplantation genetic screening versus expectant management in patients with recurrent pregnancy loss. Human reproduction 2016; 31: 1668–1674 Nybo, A.A., Wohlfahrt, J., Christens, P., Olsen, J., Melbye, M. Is maternal age an independent risk factor for fetal loss? The Western journal of medicine 2000; 173: 331 Otani, T., Roche, M., Mizuike, M., Colls, P., Escudero, T., Munne, S. Preimplantation genetic diagnosis significantly improves the pregnancy outcome of translocation carriers with a history of recurrent miscarriage and unsuccessful pregnancies. Reproductive biomedicine online 2006; 13: 869–874 Padilla, S.L., Garcia, J.E. Effect of maternal age and number of in vitro fertilization
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procedures on pregnancy outcome. Fertility and sterility 1989; 52: 270–273 Practice Committee of the American Society for Reproductive Medicine. Evaluation and treatment of recurrent pregnancy loss: A committee opinion. Fertility and sterility 2012; 98: 1103–1111 Regan, L., Braude, P.R., Trembath, P.L. Influence of past reproductive performance on risk of spontaneous abortion. Bmj 1989; 299: 541–545 Scott, R.T.Jr., Upham, K.M., Forman, E.J., Hong, K.H., Scott, K.L., Taylor, D., Tao, X., Treff, N.R. Blastocyst biopsy with comprehensive chromosome screening and fresh embryo transfer significantly increases in vitro fertilization implantation and delivery rates: A randomized controlled trial. Fertility and sterility 2013; 100: 697–703 Spandorfer, S.D., Davis, O.K., Barmat, L.I., Chung, P.H., Rosenwaks, Z. Relationship between maternal age and aneuploidy in in vitro fertilization pregnancy loss. Fertility and sterility 2004; 81: 1265–1269 Tamhankar, V.A., Liu, B., Yan, J., Li, T.C. A comparison of pattern of pregnancy loss in women with infertility undergoing ivf and women with unexplained recurrent miscarriages who conceive spontaneously. Obstetrics and gynecology international 2015; 2015: 989454 Tarasconi, B., Tadros, T., Ayoubi, J.M., Belloc, S., de Ziegler, D., Fanchin, R. Serum antimullerian hormone levels are independently related to miscarriage rates after in vitro fertilizationembryo transfer. Fertility and sterility 2017; 108: 518–524 Toner, J.P., Veeck, L.L., Muasher, S.J. Basal folliclestimulating hormone level and age affect
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the chance for and outcome of pre-embryo cryopreservation. Fertility and sterility 1993; 59: 664–667 Veleva, Z., Tiitinen, A., Vilska, S., Hyden-Granskog, C., Tomas, C., Martikainen, H., Tapanainen, J.S. High and low bmi increase the risk of miscarriage after ivf/icsi and fet. Human reproduction 2008; 23: 878–884 Wang, X., Chen, C., Wang, L., Chen, D., Guang, W., French, J. Conception, early pregnancy loss, and time to clinical pregnancy: A population-based prospective study. Fertility and sterility 2003; 79: 577–584 Wang, A., Lathi, R., Kort, J., Westphal, L. Antimüllerian hormone in association with euploid embryo transfer outcomes. Reprod. Biomed. Online 2019; 39: 609–616. https://doi. org/10.1016/j.rbmo.2019.05.006 Wilcox, A.J., Weinberg, C.R., O'Connor, J.F., Baird, D.D., Schlatterer, J.P., Canfield, R.E., Armstrong, E.G., Nisula, B.C. Incidence of early loss of pregnancy. The New England journal of medicine 1988; 319: 189–194 Yan, J., Wu, K., Tang, R., Ding, L., Chen, Z.J. Effect of maternal age on the outcomes of in vitro fertilization and embryo transfer (ivf-et). Science China Life sciences 2012; 55: 694–698 Yang, Z., Liu, J., Collins, G.S., Salem, S.A., Liu, X., Lyle, S.S., Peck, A.C., Sills, E.S., Salem, R.D. Selection of single blastocysts for fresh transfer via standard morphology assessment alone and with array cgh for good prognosis ivf patients: Results from a randomized pilot study. Molecular cytogenetics 2012; 5: 24 Received 22 December 2018; received in revised form 13 May 2019; accepted 16 May 2019.
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ARTICLE
Miscarriage history association with euploid embryo transfer outcomes BIOGRAPHY
Ange Wang is currently an Obstetrics & Gynecology resident at Stanford Hospital. She graduated from the University of Pennsylvania with a BSE in Bioengineering and Stanford Medical School with an MD. She has published on outcomes in both assisted reproductive technology and oncology and co-authored numerous publications in the Women's Health Initiative. Ange Wang*, Jonathan Kort, Lynn Westphal KEY MESSAGE
This study suggests that a history of miscarriage prior to IVF (including a history of recurrent pregnancy loss) is not associated with frozen embryo transfer outcomes for euploid embryos. As many IVF patients have a history of pregnancy loss, this is an important finding for counselling and prognostication. ABSTRACT
Research question: Is a history of miscarriage (including recurrent pregnancy loss) associated with euploid cryopreserved embryo transfer outcomes? Design: Retrospective cohort study from 2014 to 2018 of patients at an academic medical centre, undergoing their first cycle of IVF with 24-chromosome Day 5/6 preimplantation genetic testing for aneuploidies (IVF-PGT-A). Multivariate logistic regression was used to investigate the relationship between history of miscarriage and euploid single cryopreserved embryo transfer outcomes (ongoing pregnancy, miscarriage), adjusting for an extensive list of patient and cycle confounders. Results: In the study cohort of 283 patients, the overall unadjusted positive beta human chorionic gonadotrophin (bHCG) rate was 70.0%, ongoing pregnancy rate was 52.3%, and the total pregnancy loss (biochemical and clinical pregnancy loss) rate per positive bHCG cycle was 24.7%. While 35.3% of patients had a history of at least one previous miscarriage, 14.5% of patients had a history of recurrent pregnancy loss (RPL). For patients with a history of miscarriage, it was found that the adjusted odds ratios (OR) and 95% confidence intervals (CI) for positive bHCG were 1.30 (0.51– 3.27), for ongoing pregnancy were 0.88 (0.38–2.03) and for total pregnancy loss were 1.41 (0.49–4.05), when compared with patients without a history of miscarriage. For RPL patients, OR for positive bHCG, ongoing pregnancy and total pregnancy loss also did not significantly differ when compared with patients with no history of miscarriage. Conclusions: In this cohort, there was no significant association between miscarriage history and euploid cryopreserved embryo transfer outcomes (ongoing pregnancy, total pregnancy loss) after adjustment for potential confounders. Further study in larger data sets is warranted. Department of Obstetrics and Gynecology, Stanford University School of Medicine, Stanford CA 94305, USA © 2019 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved. *Corresponding author. E-mail address: [email protected] (A Wang). https://doi.org/10.1016/j.rbmo.2019.05.011 1472-6483/© 2019 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved. LW has sponsored research through Celmatix Inc. which does not pertain to this study. No other conflicts of interest to disclose from any other authors. These data were presented in part as a poster presentation at the American Society for Reproductive Medicine (ASRM) Annual Meeting 2018 in Denver, Colorado, USA.
KEYWORDS
IVF Euploid embryos Miscarriage Preimplantation genetic testing for aneuploidies Recurrent pregnancy loss
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INTRODUCTION
M
iscarriage is estimated to occur in 8–20% of clinically recognized pregnancies, and at an even higher rate in unrecognized or subclinical pregnancies (Lohstroh et al., 2005; Wang et al., 2003; Wilcox et al., 1988). Furthermore, a smaller percentage of women (<5%) experience recurrent pregnancy loss (RPL) of two or more clinical miscarriages (Practice Committee of the American Society for Reproductive Medicine, 2012). Most early miscarriages are believed to be genetic in origin, and studies suggest that women with a history of miscarriage have increased risk of subsequent miscarriage or infertility (Lund et al., 2012; Regan et al., 1989). The literature on associations between history of miscarriage and transfer outcomes for IVF cycles is limited, particularly on the subject of euploid embryos. This association may be confounded by the fact that miscarriage history may also be linked to other factors that are poor IVF prognosticators, such as diminished ovarian reserve or increased maternal age. Existing findings on this subject have been mixed, and no studies to date have investigated this subject for euploid embryo transfers in a multivariate analysis. Some studies have reported that preimplantation genetic testing for aneuploidies (PGT-A) is a beneficial intervention in patients with history of either idiopathic pregnancy loss or miscarriage due to specific causes (Garrisi et al., 2009; Hodes-Wertz et al., 2012; Otani et al., 2006), while other studies have not found this effect (Murugappan et al., 2016). A large German study found that a previous natural conception was not associated with the outcome of an IVF cycle, although the findings were mixed on previous IVF conception (Kupka et al., 2003). Several studies have also examined general reproductive history following miscarriage and have found associations of miscarriage history with infertility, subsequent miscarriage and poor obstetric outcomes (Jivraj et al., 2001; Kling et al., 2018; Lund et al., 2012). Miscarriage has also been studied as an IVF outcome, with known associations to maternal age, ovarian reserve, embryo morphology, body mass index (BMI) and anti-Müllerian hormone (AMH) concentrations (Hill et al.,
2013; Honnma et al., 2012; Metwally et al., 2008; Padilla and Garcia, 1989; Spandorfer et al., 2004; Tarasconi et al., 2017; Toner et al., 1993; Yan et al., 2012). As chromosomal abnormalities account for the majority of early miscarriages and miscarriage history is a known prognosticator for future pregnancy outcomes as above, investigation is warranted into whether or not miscarriage history affects transfer outcomes of euploid embryos. This is the first study to investigate this question in a multivariate analysis, and a further aim of the study was to determine whether or not patients with a history of one miscarriage differed from RPL patients in this context. A larger cohort of cycles from the same patients studied in this manuscript was also used for a separate analysis on the association of AMH with euploid transfer outcomes, in investigating additional prognosticators of euploid transfer outcomes (Wang et al., 2019).
MATERIALS AND METHODS This was a retrospective cohort study from 2014 to 2018 of patients at an academic medical centre undergoing IVF with 24-chromosome Day 5/6 PGT-A. PGT-A was chosen by patients for a variety of reasons, including but not limited to advanced maternal age, diminished ovarian reserve, RPL, sex selection and single gene disorder (see Supplementary Table 1 for primary IVF indications). Inclusion criteria were single transfers of euploid embryos, autologous embryo transfer and available information on all potential patient and cycle confounders included in this study (see Statistical analysis section below). Only women undergoing their first IVF cycle at the study institution were included in the analysis, to prevent the study being confounded by previous IVF miscarriages (which may be detected at a higher rate than naturally conceived miscarriages). Institutional Review Board approval was obtained for this study from Stanford University (which is affiliated to the IVF clinic) on 30 June 2018 (reference number 6208). Ovarian stimulation was performed using one of three protocols: antagonist, long gonadotrophin-releasing hormone (GnRH) agonist (leuprolide acetate), or microdose flare. Transvaginal oocyte retrieval occurred 35–36 h after
HCG and/or Lupron trigger (given when the largest follicle measured >18 mm). Oocytes were fertilized using conventional IVF or intracytoplasmic sperm injection (ICSI). Methods for blastocyst stage culture have been previously reported in the literature (Kort et al., 2015), using SAGE™ sequential media (CooperSurgical, Trumbull, CT, U.S.A.) in low-oxygen conditions. Embryologists performed laser-assisted hatching on Day 3 after retrieval, followed by trophectoderm biopsy on either Day 5 or Day 6 depending on the rate of embryo development. Using the laser, three to five cells were removed and samples were then analysed for 24-chromosome screening using IviGen (Miami, Los Angeles, and New York) with nextgeneration sequencing (NGS). One of six senior embryologists conducted morphological assessments on Day 5 or Day 6 according to the Gardner grading system (Gardner and Schoolcraft, 1999) Using standardized laboratory protocols, blastocysts were assigned grades for expansion (1–6), inner cell mass (ICM) (A, B or C) and trophectoderm (A, B or C). A very small percentage of embryos had expansion ≤2, and therefore were unable to receive ICM and trophectoderm grades due to insufficient development. Embryos were vitrified on either Day 5 or 6 after biopsy. After receiving biopsy results, one euploid embryo was warmed and transferred in a natural or hormone replacement cycle (typically utilizing oral oestrogen and both intramuscular and vaginal progesterone supplementation). A retrospective chart review was conducted to record cycle outcome, as well as all variables included in this analysis (see below), which included a mixture of patient and cycle characteristics that may potentially affect IVF cycle outcomes. History of miscarriage was self-reported and could include either biochemical or clinical miscarriage, as patients did not always specify type of miscarriage in reports of miscarriage history. Statistical analysis Multivariable logistic regression was used to investigate the associations between history of miscarriage (prior to IVF cycle) and transfer outcomes (positive bHCG, ongoing pregnancy, miscarriage) for euploid single cryopreserved embryo transfers. Patients were also classified into
two groups: history of one miscarriage, or history of two or more miscarriages (designated as RPL). The main analysis investigated history of miscarriage in a multivariate analysis, while a sub-analysis investigated the number of miscarriages in a separate multivariate model.
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TABLE 1 BASELINE CHARACTERISTICS OF THE PATIENTS Miscarriage history No
Yes
Overall
Total
183
100
283
Gravidity
61 (33.3)
100 (100.0)
161 (56.9)
<0.001
In terms of outcomes, positive bHCG was defined as concentration >5 mIU/ml, ongoing pregnancy was defined as continued pregnancy at 8–12 weeks with positive fetal cardiac activity, and total pregnancy loss included both biochemical and clinical miscarriage. Biochemical miscarriage was defined as loss of pregnancy with positive HCG but no ultrasound findings, while clinical miscarriage was defined as loss of a pregnancy after some ultrasound findings were present (at least a gestational sac). The analysis controlled for the following patient and cycle confounders (all of which were included in the multivariate model): primary IVF indication, history of pregnancy, history of live birth, maternal age at retrieval, maternal age at transfer, AMH, BMI, ICSI, oocyte number, number of blastocysts, morphology (expansion, ICM and trophectoderm), hormone replacement versus natural cryopreserved embryo transfer, and endometrial thickness. All tests were two-sided with significance at the alpha = 0.05 level, and Python and Microsoft Excel were used for data analysis.
Parity
46 (25.1)
38 (38.0)
84 (29.7)
0.024
RESULTS
a
Recurrent pregnancy loss – two or more miscarriages. Not applicable for group without history of miscarriage.
b
Indications may overlap, does not sum to 100%
In the study cohort of 283 patients, the overall unadjusted positive bHCG rate was 70.0% (198 patients), ongoing pregnancy rate was 52.3% (148 patients), and the total pregnancy loss rate per cycle with positive bHCG was 24.7% (49 patients) (7.6% clinical [15 patients], 17.2% biochemical [34 patients]). It was found that 35.3% of patients had a history of at least one previous miscarriage, while 14.5% of patients had a history of RPL (TABLE 1). The two groups had similar ages at retrieval and transfer, with retrieval age 35.6 years (SD 3.8) for the no miscarriage group and 35.9 years (SD 3.6) for the miscarriage group; age at transfer was 36.0 (SD 3.8) and 36.3 (SD 3.7) years, respectively. BMI (23.7 versus 24.2) and distribution of AMH categories was also similar between the two groups. The leading primary indications for IVF were male factor (25.4%), followed by diminished ovarian reserve (19.8%) and ovulatory dysfunction (11.3%)
History of
RPLa
P-value
–
41 (41.0)
41 (14.5)
–
Age at retrieval, mean (SD)
35.6 (3.8)
35.9 (3.6)
35.7 (3.7)
NS
Age at transfer, mean (SD)
36.0 (3.8)
36.3 (3.7)
36.1 (3.8)
NS
BMI, mean (SD)
23.7 (4.8)
24.2 (4.4)
23.8 (4.6)
NS
<1
31 (16.9)
18 (18.0)
49 (17.3)
NS
1 to <2
42 (23.0)
21 (21.0)
63 (22.3)
2 to <5
69 (37.7)
41 (41.0)
110 (38.9)
5+
41 (22.4)
20 (20.0)
61 (21.6)
DOR
47 (25.7)
24 (24.0)
71 (25.1)
Endometriosis
20 (10.9)
5 (5.0)
25 (8.8)
Male
75 (41.0)
25 (25.0)
100 (35.3)
Ovulatory
16 (8.7)
18 (18.0)
34 (12.0)
RPL
0 (0.0)
29 (29.0)
29 (10.2)
Tubal
9 (4.9)
12 (12.0)
21 (7.4)
Sex selection
8 (4.4)
5 (5.0)
13 (4.6)
Single gene disorder
13 (7.1)
3 (3.0)
16 (5.7)
Uterine
10 (5.5)
6 (6.0)
16 (5.7)
Unexplained
19 (10.4)
9 (9.0)
28 (9.9)
Other
11 (6.0)
1 (1.0)
12 (4.2)
AMH category (ng/ml)
IVF
indicationb <0.001
Data are presented as n (%) unless otherwise stated.
AMH = anti-Müllerian hormone; BMI = body mass index; DOR = diminished ovarian reserve; NS = not statistically significant; RPL = recurrent pregnancy loss.
(Supplementary Table 1), although many patients had multiple indications for IVF. In terms of cycle characteristics, an average of 15.5 oocytes (SD 8.3) were retrieved and the number of blastocysts obtained was 5.5 (SD 3.8) for the entire cohort (TABLE 2). The miscarriage and no miscarriage group were similar in terms of the number of oocytes retrieved, number of blastocysts obtained, expansion grades (over 80% 5 or 6 for both groups), trophectoderm grades, and type of transfer protocols. The two groups also had similar endometrial thickness (9.4, SD 1.6 and 9.0, SD 1.5 for no miscarriage and miscarriage history, respectively). Compared with the no miscarriage group, the miscarriage history group was less likely to use ICSI, and have lower ICM grade (see TABLE 2).
In this analysis, for patients with a history of miscarriage (compared with patients with no history of miscarriage), it was found that the adjusted odds ratio (OR) and 95% confidence interval (CI) for positive bHCG was 1.30 (0.51–3.27) and OR for ongoing pregnancy was 0.88 (0.38–2.03) (TABLE 3). It was also found that when compared with women without a history of miscarriage, the OR for positive HCG was 1.45 (0.54–3.85) for a history of one miscarriage and 0.89 (0.22–3.52) for RPL patients. For ongoing pregnancy, OR was 0.91 (0.38–2.19) for a history of one miscarriage, and 0.78 (0.22–2.81) for RPL patients. Depending on miscarriage category, the unadjusted positive HCG rates ranged from 65.9% to 76.3%, and ongoing pregnancy rates ranged from 43.9% to 53.6% (see TABLE 3 for full details).
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DISCUSSION
TABLE 2 CYCLE CHARACTERISTICS Miscarriage history No
Yes
Overall
P-value
Oocytes retrieved, mean (SD)
15.3 (8.1)
16 (8.7)
15.5 (8.3)
NS
Number of blastocysts, mean (SD)*
5.4 (3.5)
5.7 (4.4)
5.5 (3.8)
NS
No
37 (20.2)
36 (36.0)
73 (25.8)
0.014
Yes – all ICSI
127 (69.4)
57 (57.0)
184 (65.0)
ICSI split
19 (10.4)
7 (7.0)
26 (9.2)
5 to 6
152 (83.1)
80 (80.0)
232 (82.0)
3 to 4
30 (16.4)
20 (20.0)
50 (17.7)
Less than 3
1 (0.5)
0 (0.0)
1 (0.4)
A
105 (57.4)
45 (45.0)
150 (53.0)
B
76 (41.5)
51 (51.0)
127 (44.9)
C
1 (0.5)
4 (4.0)
5 (1.8)
A
85 (46.4)
37 (37.0)
122 (43.1)
B
93 (50.8)
56 (56.0)
149 (52.7)
C
4 (2.2)
7 (7.0)
11 (3.9)
Medicated
64 (35.0)
44 (44.0)
108 (38.2)
Natural
119 (65.0)
56 (56.0)
175 (61.8)
Endometrial thickness (mm), mean (SD)
9.4 (1.6)
9.0 (1.5)
9.2 (1.6)
ICSI
Morphology Expansion NS
ICM 0.023
Trophectoderm NS
Type of embryo transfer NS
NS
Data are presented as n (%) unless otherwise stated. Note: One embryo was collapsed and did not receive ICM or trophectoderm grade. ICM = inner cell mass; ICSI = intracytoplasmic sperm injection; NS = not statistically significant.
For total pregnancy loss (per cycle with positive bHCG) as an outcome (TABLE 4), an unadjusted total pregnancy loss rate of 21.4% (27 patients) was found for patients in the no miscarriage group, compared with 30.6% (22 patients) for the overall miscarriage group. TABLE 4 also shows the breakdown by clinical and biochemical
pregnancy loss. In the multivariate analysis, when compared with patients with no miscarriage, the adjusted OR for total pregnancy loss outcome was 1.41 (0.49–4.05) for the overall group of miscarriage patients, 1.48 (0.49–4.43) for a history of one miscarriage, and 1.21 (0.27–5.37) for RPL patients.
In summary, in this multivariate analysis of euploid single cryopreserved embryo transfers, it was found that history of miscarriage was not associated with positive HCG, ongoing pregnancy or total pregnancy loss outcomes. Euploid embryos are estimated to implant at a rate of 60–70%, and data on euploid miscarriage is limited (Scott et al., 2013; Yang et al., 2012). The factors that make a euploid transfer result in miscarriage or fail to implant are not well understood; proposed factors have included the uterine lining, uterine microbiome or other environmental factors, genetic defects or aneuploidy in the embryo that cannot be detected through PGT-A, immunologic causes such as antiphospholipid syndrome, endocrine causes, damage to the embryos during freezing/thawing/biopsy, among others (Maxwell et al., 2016; Meldrum, 2016; Meldrum and de Ziegler, 2016). Existing literature on associations between history of miscarriage and transfer outcomes for IVF cycles is extremely limited, particularly on the subject of euploid embryos. This association may also be confounded by the fact that miscarriage history may be linked to other factors that may be poor prognosticators for an IVF cycle, such as ovarian reserve or maternal age. It is estimated that miscarriage as an outcome occurs in 8–20% of clinically recognized pregnancies, and at an even higher rate in unrecognized or subclinical pregnancies (Lohstroh et al., 2005; Wang et al., 2003; Wilcox et al., 1988). A much smaller percentage of women (<5%) experience RPL (two or more miscarriages), with an even smaller percentage (1%) experiencing three or more (Practice Committee of
TABLE 3 EUPLOID EMBRYO TRANSFER OUTCOMES BY MISCARRIAGE HISTORY No. of patients
Maternal age at retrieval (years)
PB rate (%)
Adjusted OR PB, OR (95% CI)
OP rate (%)
Adjusted OR OP, OR (95% CI)
No miscarriage
183
35.7
68.9
Reference
53.6
Reference
History of miscarriage
100
35.8
72.0
1.30 (0.51–3.27)
50.0
0.88 (0.38–2.03)
History of 1 miscarriage
59
35.5
76.3
1.45 (0.54–3.85)
54.2
0.91 (0.38–2.19)
History of 2 or more miscarriages (RPL)
41
36.1
65.9
0.89 (0.22–3.52)
43.9
0.78 (0.22–2.81)
No P-values were statistically significant. Confounders included primary IVF indication, history of pregnancy, history of live birth, maternal age at retrieval, maternal age at transfer, AMH, ICSI, oocyte number, number of blastocysts, morphology (expansion, ICM and trophectoderm), hormone replacement versus natural cryopreserved embryo transfer, and endometrial thickness. AMH = anti-Müllerian hormone; bHCG = beta human chorionic gonadotrophin; BMI = body mass index; CI = confidence interval; ICM = inner cell mass; ICSI = intracytoplasmic sperm injection; OP = ongoing pregnancy; OR = odds ratio; PB = positive bHCG; RPL = recurrent pregnancy loss.
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TABLE 4 PREGNANCY LOSS RATES FOLLOWING EUPLOID EMBRYO TRANSFER ACCORDING TO MISCARRIAGE HISTORY Biochemical pregnancy loss rate (%)
Clinical miscarriage rate (%)
TPL rate (%)a
Adjusted OR TPL, OR (95% CI)
No miscarriage
15.1
6.3
21.4
Reference
History of miscarriage
20.8
9.7
30.6
1.41 (0.49–4.05)
History of 1 miscarriage
20.0
8.9
28.9
1.48 (0.49–4.43)
History of 2 or more miscarriages (RPL)
22.2
11.1
33.3
1.21 (0.27–5.37)
No P-values were statistically significant. Confounders included primary IVF indication, history of pregnancy, history of live birth, maternal age at retrieval, maternal age at transfer, AMH, BMI, ICSI, oocyte number, number of blastocysts, morphology (expansion, ICM and trophectoderm), hormone replacement versus natural cryopreserved embryo transfer, and endometrial thickness. a Total
pregnancy loss includes both biochemical and clinical pregnancy loss. All percentages are per cycle with positive bHCG.
AMH = anti-Müllerian hormone; BMI = body mass index; BPL = biochemical pregnancy loss; CM = clinical miscarriage; ICM = inner cell mass; ICSI = intracytoplasmic sperm injection; NS = not statistically significant; OP = ongoing pregnancy; OR = odds ratio; TPL = total pregnancy loss.
the American Society for Reproductive Medicine, 2012). Known risk factors for miscarriage include previous miscarriage or RPL, advancing maternal age and smoking (Chatenoud et al., 1998; Nybo et al., 2000; Regan et al., 1989). Given that chromosomal abnormalities are believed to account for the majority of early miscarriages, and that miscarriage history is a known prognosticator for future pregnancy outcomes, further elucidation is needed into whether or not miscarriage history affects transfer outcomes of euploid embryos. Few studies have investigated the linkage between miscarriage history and IVF outcomes, and no studies to date have investigated this subject for euploid embryo transfers in a multivariate analysis. One study of 174,909 assisted reproductive technology (ART) cycles in Germany from 1998 to 2000 found that a previous natural conception (miscarriage or live birth) did not significantly affect the outcome of IVF cycles. However, a previously successful ART procedure ending in live birth had a statistically significant positive effect on IVF outcome, while an ART procedure ending in miscarriage had a significant negative effect (Kupka et al., 2003). Another study showed that the use of preimplantation genetic diagnosis improved pregnancy outcomes for the specific population of translocation carriers with recurrent miscarriages, but it was not clear whether this could be generalized to patients without translocations (Otani et al., 2006). Studies have differed on the value of PGT-A for idiopathic RPL patients; one study of 287 cycles reported a significant decrease in the miscarriage rate among RPL patients using PGT-A (Hodes-Wertz et al., 2012). A 2009 study also reported
that PGT-A of nine chromosomes decreased miscarriage rates among idiopathic RPL patients, particularly for women over age 35 (Garrisi et al., 2009). Another study of over 1800 patients found that clinical pregnancy loss rate following IVF treatment (without PGT-A) was lower than for RPL patients who conceived spontaneously (Tamhankar et al., 2015). In contrast, a study of 300 patients reported that PGT-A did not improve miscarriage, pregnancy or live birth rates among RPL patients when compared with expectant management (Murugappan et al., 2016). Studies on the subject of miscarriage history and IVF outcomes are limited as above, and there has also been literature investigating general reproductive history following miscarriage. A cohort study of 987 Danish women with recurrent miscarriage found that there was a significantly decreased chance (P < 0.01) of at least one subsequent live birth with increasing number of miscarriages (Lund et al., 2012). Other studies have reported descriptively on subsequent fertility and obstetrics outcomes in RPL patients (Kling et al., 2018), with one study suggesting that RPL patients had a significantly higher risk of preterm delivery, small for gestational age, perinatal loss, and Caesarean section compared with a control group; however, this study did not investigate fertility rates in this population (Jivraj et al., 2001). Miscarriage has also been studied as an IVF outcome, with known associations with maternal age and ovarian reserve (Padilla and Garcia, 1989; Spandorfer et al., 2004; Toner et al., 1993; Yan et al., 2012), embryo morphology (Hill et al., 2013; Honnma et al., 2012), BMI (Metwally et al., 2008; Veleva et al., 2008) and AMH concentrations (Tarasconi et al., 2017).
This study is novel compared with previous studies by using a multivariate analysis to investigate IVF outcomes for euploid embryos in relation to miscarriage history (both overall miscarriage as well as RPL). A multivariate analysis is very important for this subject to control for confounders, as miscarriage history has been established to be linked to poor prognosticators of IVF cycles as detailed above. Similar to the large German study, it was found that a previous miscarriage (non-IVF) did not significantly affect the results of IVF cycles in terms of either miscarriage or ongoing pregnancy. Although patients with a history of miscarriage had non-significant elevated OR for miscarriage as an outcome in this study, no directionality was observed for RPL patients compared with patients with history of one miscarriage, providing additional confidence in these results. As the majority of miscarriages are due to genetic causes, the findings are not unexpected. In combination with previous literature, this study suggests that a history of miscarriages, either isolated or recurrent, is not a poor prognosticator for IVF/PGT-A cycles. Although some studies have suggested that miscarriage history (particularly RPL) may be negatively associated with subsequent natural conception, this study supports IVF/PGT-A as an intervention for RPL patients to consider. However, findings must be interpreted with caution given the sample size, retrospective nature of the study and limited literature on the subject. The strengths of this study include the novel subject, detailed information on potential patient and cycle confounders which were used for a multivariate analysis, information on both miscarriage
62 2
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and ongoing pregnancy as outcomes, and single institution format, which allows for more standardized data collection. In particular, this is the first study to investigate the association between miscarriage history and IVF outcomes of euploid embryos in a multivariate analysis. Given the high prevalence of miscarriage (particularly among patients seeking IVF treatment), this study is very applicable clinically. Limitations of the study include the retrospective format, the relatively small sample size, self-report of miscarriage outcomes (which may be subject to recall bias) and lack of information on live birth outcomes. Some OR had wide CI and it is possible that the sample size limited the ability to find a difference in some associations. The study is also limited by the fact that the findings cannot be generalized to all patients with a history of miscarriage, but only those who were able to produce euploid embryos for transfer, which likely represents a better prognosis group. The study included patients with a history of miscarriage prior to starting IVF treatment at the study institution, so the findings are not necessarily generalizable to patients with a previous IVF miscarriage (which may be detected at a higher rate than spontaneous miscarriages); this area should be studied given mixed findings previously reported in the literature. There was no information available on type of miscarriage (biochemical versus clinical), and additional study is warranted on whether history of miscarriage type is a prognosticator. Also, it was not possible to differentiate between idiopathic miscarriage and miscarriage with a known cause such as antiphospholipid syndrome, although these aetiologies of miscarriages are relatively rare. This cohort was also probably better than average in terms of prognosis based on a higher number of oocytes retrieved, AMH and average maternal age of 35–36. Therefore, patient counselling on miscarriage history in relation to euploid embryo transfer must include these important caveats. In summary, using a multivariate analysis, it was found that history of miscarriage was not associated with IVF outcomes (positive bHCG, ongoing pregnancy, total pregnancy loss) for single embryo transfer of euploid embryos. These findings are encouraging for patients with miscarriage or RPL history who have
euploid embryos available to transfer. However, further study is warranted in larger cohorts and in a prospective format, particularly as OR and absolute rates for miscarriage outcome were elevated but not significant for patients with history of miscarriage or RPL. Other areas for future investigation (in terms of euploid embryo transfer outcomes) include whether or not previous IVF miscarriage is a different prognosticator than previous natural conception miscarriage, or if associations vary based on type of miscarriage history.
ACKNOWLEDGEMENTS The authors would like to acknowledge Arthur Chiang for guidance on data analysis, and Qianyang Zhao for assistance with procuring data for analysis.
SUPPLEMENTARY MATERIALS Supplementary material associated with this article can be found in the online version at doi:10.1016/j.rbmo.2019.05.011.
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