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Abstract
Objective To investigate whether there are significant differences in pregnancy outcomes between frozen-thawed D3/D5 and D3/D6 sequential embryo transfer (SeET) in women with repeated implantation failure (RIF).
Methods Women who experienced RIF and underwent frozen-thawed cycles with SeET were included and were divided into two groups: D3/D5 (n=455) and D3/D6 (n=308). Included cycles were stratified by the quality of transferred embryos and age.
Results There were 252/455 (55.4%) clinical pregnancies in the D3/D5 group and 133/308 (43.2%) in the D3/D6 group (OR: 1.63, 95% CI 1.22 to 2.18). Rates for ectopic pregnancy, early miscarriage and multiple pregnancies did not differ between the two groups. In women under 35 years, with transfers of two good-quality embryos, the clinical pregnancy rate was significantly higher in the D3/D5 group compared with the D3/D6 group (65.6% vs 42.4%, OR: 2.59, 95% CI 1.41 to 4.76). This trend persisted when analysing good-quality cleavage-stage embryo and poor-quality blastocyst (D3/D5) transfers versus two good-quality embryo (D3/D6) transfers (61.2% vs 42.4%, OR: 2.14, 95% CI 1.11 to 4.12). For women aged 35 years or older, pregnancy outcomes did not differ.
Conclusion The clinical pregnancy rate was significantly higher in the D3/D5 group compared with the D3/D6 group. When contrasting the outcomes of transferring a good-quality cleavage-stage embryo and a poor-quality blastocyst in the D3/D5 group with transferring two good-quality embryos in the D3/D6 group, the clinical pregnancy rate was significantly higher in the D3/D5 group. However, among women aged 35 years or older, there were no significant differences in pregnancy outcomes.
- Infertility
- Pregnancy Outcome
- Reproductive Medicine
- Fertilization in Vitro
Data availability statement
Data are available upon reasonable request.
This is an open access article distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited, appropriate credit is given, any changes made indicated, and the use is non-commercial. See http://creativecommons.org/licenses/by-nc/4.0/.
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WHAT IS ALREADY KNOWN ON THIS TOPIC
There is a paucity of data concerning the comparison of pregnancy outcomes between D5 and D6 blastocyst transfers in sequential embryo transfer (SeET). It is unclear whether and to what degree the prior transfer of a cleavage-stage embryo elucidates or obscures differences in pregnancy outcomes between subsequent D5 and D6 blastocyst transfers in frozen-thawed cycles.
WHAT THIS STUDY ADDS
The clinical pregnancy rate was significantly higher in the D3/D5 group compared with the D3/D6 group, particularly among women under 35 years or in cases involving transfers of two good-quality embryos.
HOW THIS STUDY MIGHT AFFECT RESEARCH, PRACTICE OR POLICY
We advocate for prioritising blastocysts available for transfer on D5 in the SeET strategy, irrespective of their quality. Delaying transfer to D6 should be discouraged, as even poor-quality blastocysts formed on D5 exhibited a higher clinical pregnancy rate compared with good-quality blastocysts formed on D6 in SeET, particularly among women younger than 35 years.
Introduction
Repeated implantation failure (RIF) occurs when embryos transferred during in vitro fertilisation (IVF) procedures fail to implant despite multiple attempts.1 2 These recurring failures not only place psychological, physical and financial burdens on patients but also present substantial challenges for medical professionals.3 The sequential embryo transfer (SeET) strategy, which refers to a two-step process of transferring a cleavage-stage embryo followed by a blastocyst at different phases of the menstrual cycle within a single transfer cycle, has been proposed by researchers as a potential solution for women experiencing RIF.4 While the precise mechanisms underlying the efficacy of SeET in RIF remain to be fully elucidated, several hypotheses have been suggested. First, it is known that the human endometrium is receptive to embryo implantation only during a specific timeframe referred to as the window of implantation (WOI).5 RIF can occur due to both asynchronous (displaced) and pathological (disrupted) WOI.6 7 By transferring two embryos at different periods, there is a higher likelihood of aligning the embryos with the optimal endometrial WOI, thereby enhancing the success rate of IVF. Additionally, the catheter insertion during the first transfer may stimulate the endometrium mechanically, prompting the release of inflammatory factors that enhance embryo adhesion during the subsequent transfer.8 Second, the developmental potential of embryos is another critical consideration. Higher morphological scores were found to be linked with elevated rates of euploidy, consequently leading to improved implantation rates.9 Culturing cleavage-stage embryos in vitro for 5–6 days allows for the selection of embryos with superior implantation potential. Meanwhile, the SeET strategy decreases the likelihood of cycle cancellation resulting from the unavailability of blastocysts for transfer. In clinical practice, many studies have reported higher success rates with SeET compared with conventional one-stage embryo transfer.10–12
Embryos reach the blastocyst stage by D5 in culture, and slower development may lead to blastocysts forming by D6 or later. Blastocysts formed during the fresh embryo transfer cycle and transferred on D5 exhibit higher implantation and clinical pregnancy rates compared with those transferred on D6.13 14 This discrepancy may be attributed to reduced endometrial receptivity during implantation on D6.15 With advancements in hormonal priming for enhancing endometrial receptivity,16 similar clinical outcomes would be expected regardless of whether blastocysts are transferred on D5 or D6 in programmed frozen-thawed cycles, owing to improved endometrial-embryonic synchronisation. However, results regarding the pregnancy outcomes between D5 and D6 blastocyst transfers remain conflicting. Several studies have documented higher clinical pregnancy rates favouring D5 blastocyst transfers over D6 in frozen-thawed cycles,17–19 whereas other studies suggested similar pregnancy outcomes between D5 and D6.20 21 These contrasting findings underscore the complexity of the factors and mechanisms influencing the success of IVF. For instance, when considering the quality of transferred blastocysts, which plays a crucial role, one study reported no significant difference in pregnancy rates between high-quality D5 and D6 blastocysts.22
There is a paucity of data concerning the comparison of pregnancy outcomes between D5 and D6 blastocyst transfer in SeET. It is unclear whether and to what degree the prior transfer of a cleavage-stage embryo elucidates or obscures differences in pregnancy outcomes between subsequent D5 and D6 blastocyst transfers in frozen-thawed cycles. One recent study explored the efficiency of SeET and stratified the analysis into several aspects, including comparing the difference between D5 and D6 blastocyst transfers within the SeET group.11 It was observed that there were no significant differences in pregnancy outcomes between D5 and D6 blastocysts. Nonetheless, the study did not provide details regarding the quality of transferred embryos, and the limited sample size of the study population undermines the robustness of the conclusion.
Therefore, we designed this retrospective study to compare the pregnancy outcomes of D5 and D6 blastocyst transfers used in SeET (D3/D5 vs D3/D6) in frozen-thawed cycles. Furthermore, we stratified the analyses based on the quality of transferred embryos and maternal age. We anticipate that our findings will offer insights into selecting optimal embryo transfer strategies, ultimately aiding women experiencing RIF.
Methods
Study design and study population
This is a retrospective cohort study. Data were retrieved from the clinical database of couples who underwent frozen-thawed embryo transfer at the Reproductive Centre of Peking University Third Hospital between January 2020 and December 2023. Women who experienced RIF (RIF was defined as the inability to achieve clinical pregnancy after the transfer of at least three high-quality embryos across three fresh or frozen cycles)23 and underwent frozen-thawed cycles with SeET were included in this study and were divided into two groups: one group underwent SeET on D3 followed by D5 embryo transfer, while the other group underwent D3 followed by D6 embryo transfer. Multiple cycles could be included per woman. Exclusion criteria comprised women requiring preimplantation genetic testing (PGT) for chromosomal structural rearrangements (PGT-SR) or monogenic/single gene defects (PGT-M) or aneuploidies (PGT-A), those with endometrial thickness below 6 mm, and those diagnosed with significant endocrine disorders such as Cushing syndrome. This study was approved by the Ethics Committee of Peking University Third Hospital (No. IRB00006761-M2020004) and conducted in accordance with local legislation and institutional requirements.
Endometrial preparation
Endometrial preparation followed established protocols, using one of three methods as described earlier10: artificial cycle, natural cycle or stimulation cycle. Natural cycles were chosen for women with regular menstruation, while those with irregular menstruation and ovulation disorders were advised on either the artificial or stimulation cycle.
Women undergoing natural and stimulation cycles were given 20 mg/day of dydrogesterone tablets (Abbott Biologicals B.V., Duffton, The Netherlands) starting from the third day after ovulation. Luteal phase support for the artificial cycle consisted of 90 mg/day of vaginal progesterone (Crinone, Merk, Germany) and 20 mg/day of dydrogesterone tablets (Abbott Biologicals B.V., Duffton, The Netherlands).
Embryo quality assessment
Cleavage-stage embryo quality classification was mainly based on the size, shape and fragment ratio in blastomeres.24 High-quality cleavage-stage embryos were defined as those with 6–10 cells on day 3 and <20% embryo fragmentation. Poor-quality cleavage-stage embryos were defined as those with uneven cell size, irregular cell morphology and embryo fragmentation ranging from 21% to 49%. Blastocysts were graded according to the number and morphology of the inner cell mass (ICM) and the trophectoderm (TE).24 The development of the ICM was assessed as follows: A, tightly packed, many cells; B, loosely grouped, several cells; or C, very few cells. The TE was assessed as follows: A, many cells forming a cohesive epithelium; B, few cells forming a loose epithelium; or C, very few large cells. High-quality blastocysts were graded as AA, AB, BA and BB, and poor-quality blastocysts were AC, CA, BC, CB and CC. The embryos were transferred using the Cook Sydney IVF catheter (k-jets-7019-SIVF).
Outcome measures
The primary outcome assessed in this analysis was clinical pregnancy, defined as a pregnancy confirmed through ultrasonographic visualisation of one or more gestational sacs.25 The secondary outcomes included human chorionic gonadotropin (hCG) positivity, ectopic pregnancy, miscarriage and multiple births. hCG positivity was determined by hCG levels ≥25 IU/L measured 14–21 days postembryo transfer. Ectopic pregnancy was identified when foetal tissue was implanted outside the uterus or attached to an abnormal or scarred area within the uterus. Miscarriage refers to the loss of a pregnancy subsequent to clinical pregnancy confirmation. Multiple pregnancies were characterised by the presence of two or more gestational sacs observed on ultrasound.
Statistical analysis
The analysis was conducted using SPSS V.25.0 (IBM, New York, USA). A significance level of p<0.05 was employed to denote statistical significance. Baseline characteristics were presented as mean±SD or median with IQR. Categorical variables were expressed as proportions (percentages). Normality testing was carried out using histograms, normal probability plots (Q–Q plots) and the Kolmogorov-Smirnov test. A comparison of baseline characteristics between the study and control groups was conducted using the χ2 test for categorical variables and either the Student’s t-test or Mann-Whitney U test for continuous variables. If any baseline variables exhibited significant differences between the two groups, they would have been incorporated into the model exploring pregnancy outcomes. To compare outcomes between the two groups, a generalised estimation equations (GEE) model with an exchangeable correlation matrix was used to account for the dependency of cycles from the same couple. OR and their corresponding 95% CI were reported for the assessed pregnancy outcomes. Further stratified analysis was performed based on the quality of transferred embryos (in addition to stratifying the analyses by ‘two good-quality embryos’ and ‘two poor-quality embryos’, we investigated the impact of the quality of D5 and D6 blastocysts following the transfer of high-quality D3 embryos on pregnancy outcomes in SeET) and maternal age (age <35 and ≥35 years). In addition, we conducted a multiple variable regression analysis (variables included in the model were female age, female body mass index (BMI), years of infertility, infertility type, previous failed cycles, fertilisation method, endometrial preparation, endometrial thickness, transferred embryo quality and day of blastocyst in SeET) using the GEE model to identify the significant parameters associated with clinical pregnancy outcome.
Results
Study population
There were 829 frozen-thawed cycles with SeET recorded at the Reproductive Centre of Peking University Third Hospital between January 2020 and December 2023. Among these cycles, 763 fulfilled our inclusion criteria. There were 455 cycles with D3/D5 embryo transfers and 308 cycles with D3/D6 embryo transfers. The distribution of cycles based on embryo quality and maternal age is outlined in detail in figure 1.
Flow chart of the current study showing the recruitment and groups.
Baseline characteristics
The baseline characteristics of the cycles included in the study are displayed in table 1. The mean±SD maternal age and BMI were 33.9±4.2 years and 22.7±3.5 kg/m2 in the D3/D5 group and 34.4±4.1 years and 22.5±3.5 kg/m2 in the D3/D6 group (p=0.08 for maternal age and p=0.55 for BMI). No significant differences were observed in baseline parameters such as duration of infertility, infertility type, previous failed cycles and endometrial thickness. The distribution of transferred embryos based on quality, categorised as two good-quality embryos, two poor-quality embryos, a good-quality cleavage-stage embryo and a poor-quality blastocyst, and a poor-quality cleavage-stage embryo and a good-quality blastocyst, displayed significant differences between the two groups (p<0.001). As we stratified the analysis based on embryo quality, it was not adjusted in the analysis exploring pregnancy outcomes for the overall group.
Baseline characteristics of two groups with sequential cleavage and blastocyst embryo transfer
Pregnancy outcomes for the overall D3/D5 and D3/D6 groups
Pregnancy outcomes for the overall D3/D5 and D3/D6 groups are shown in table 2. The proportion of hCG positives was statistically significantly higher in the D3/D5 group than in the D3/D6 group (62.2% vs 51.9%, OR: 1.52, 95% CI: 1.14 to 2.04). There were 252/455 (55.4%) clinical pregnancies in the D3/D5 group and 133/308 (43.2%) in the D3/D6 group, with statistically significant differences (OR: 1.63, 95% CI: 1.22 to 2.18). Rates for ectopic pregnancy (0.7% vs 1.0%, OR and 95% CI: 0.67, 0.14 to 3.30), early miscarriage (10.7% vs 14.3%, OR and 95% CI: 1.82, 0.82 to 4.04), and multiple pregnancies (22.6% vs 16.5%, OR and 95% CI: 1.48, 0.85 to 2.55) did not differ between the D3/D5 group and the D3/D6 group.
Pregnancy outcomes of the overall group
Pregnancy outcomes based on embryo quality
The pregnancy outcomes for the D3/D5 and D3/D6 groups, classified by the quality of transferred embryos, are illustrated in table 3. When comparing a good-quality cleavage-stage embryo and a poor-quality blastocyst in the D3/D5 group versus two good-quality embryos in the D3/D6 group, the clinical pregnancy rate (56.0% vs 43.5%, OR and 95% CI: 1.65, 1.01 to 2.70) was significantly higher in the D3/D5 group than the D3/D6 group.
Pregnancy outcomes according to the quality of the transferred embryo
Pregnancy outcomes based on maternal age
Pregnancy outcomes for the D3/D5 and D3/D6 groups, categorised by age (<35 years and ≥35 years), are presented in table 4. In women under 35 years, the rates of hCG positivity (68.4% vs 53.4%, OR and 95% CI: 1.87, 1.27 to 2.77) and clinical pregnancy (62.4% vs 45.4%, OR and 95% CI: 2.00, 1.36 to 2.95) were higher in the D3/D5 group compared with the D3/D6 group. Such differences were not observed in women aged 35 years or older. Early miscarriage occurred more frequently in the D3/D5 group compared with the D3/D6 group (15.1% vs 13.0%, OR and 95% CI: 2.87, 1.20 to 6.85). Rates of ectopic pregnancy and multiple pregnancy did not differ between D3/D5 and D3/D6 either in women under 35 years or in women aged 35 years or older.
Pregnancy outcomes according to maternal age
Pregnancy outcomes based on maternal age and embryo quality
We further explored the pregnancy outcomes, taking into account both embryo quality and maternal age, and the results are shown in table 5. In women under 35 years with transfers of two good-quality embryos, the clinical pregnancy rate was significantly higher in the D3/D5 group compared with the D3/D6 group (65.6% vs 42.4%, OR and 95% CI: 2.59, 1.41 to 4.76). This trend persisted when analysing a good-quality cleavage-stage embryo and poor-quality blastocyst (D3/D5) transfers versus two good-quality embryos (D3/D6) transfers (61.2% vs 42.4%, OR and 95% CI: 2.14, 1.11 to 4.12). For women aged 35 years or older and with transfers of two good-quality embryos, there were more early miscarriages in the D3/D5 group compared with the D3/D6 group (OR and 95% CI: 3.40, 1.25 to 9.21). Other pregnancy outcomes did not differ.
Pregnancy outcomes according to the age and quality of transferred embryo
Multiple variable regression on clinical pregnancy
The day of blastocyst development in SeET was the only parameter demonstrating a significant association with clinical pregnancy (OR for D3/D5 vs D3/D6: 1.73, 95% CI: 1.07 to 2.81). Although there was a trend indicating a significant association between maternal age and clinical pregnancy, it did not reach statistical significance (β: −0.05, 95% CI: −0.11 to 0.01). The effects of other parameters are detailed in online supplemental table 1.
Supplemental material
Discussion
In this retrospective cohort study, we compared the pregnancy outcomes in SeET between D3/D5 and D3/D6. We observed that the clinical pregnancy rate was significantly higher in the D3/D5 group than the D3/D6 group, especially in women under 35 years or in transfers of two good-quality embryos. When comparing a good-quality cleavage-stage embryo and a poor-quality blastocyst in the D3/D5 group versus two good-quality embryos in the D3/D6 group, the clinical pregnancy rate was significantly higher in the D3/D5 group than the D3/D6 group. Advanced maternal age and poor quality of transferred embryos appeared to eliminate the efficacy of D3/D5 SeET compared to D3/D6 SeET. Additionally, the day of blastocyst development in SeET was the only parameter demonstrating a significant association with clinical pregnancy.
SeET is recognised as an effective strategy for women with RIF, as numerous studies have reported improved pregnancy outcomes with SeET compared with conventional embryo transfer methods.10–12 In our centre, the strategy of SeET has been increasingly adopted for RIF women.10 However, selecting the embryos with the greatest potential for successful implantation remains an ongoing challenge. Many studies indicate that the timing of blastocyst transfer during IVF significantly influences clinical outcomes.26 Thus, a dilemma faced by physicians and couples is the decision between selecting D5 or D6 blastocysts for SeET. Data regarding the comparison of D5 blastocysts with D6 blastocysts within SeET is scarce. One recent retrospective study indicated no significant differences in pregnancy outcomes between D5 (n=216) and D6 blastocysts (n=144) within the SeET group.11 The result is in contrast with our study with a larger sample size, revealing a significantly higher clinical pregnancy rate in the D3/D5 group compared with the D3/D6 group. Notably, Li et al11 also reported a higher clinical pregnancy rate in the D3/D5 group compared with the D3/D6 group (33.8% vs 28.5%), although without reaching statistical significance. Given the relatively small sample size,11 it might be a false-negative result. On the other hand, differences in baseline characteristics of included patients and other confounding factors, such as variations in laboratory practices, may also contribute to the inconsistency between these two study findings. Meanwhile, both Li et al’s study11 and our study observed similar rates of miscarriage and multiple pregnancies between the D3/D5 and D3/D6 embryo transfers. The mechanism behind the higher clinical pregnancy rate observed in our study with D3/D5 SeET is unknown. One conceivable explanation is that earlier development of blastocysts is inherently linked to a greater probability of embryonic euploidy, thereby resulting in increased pregnancy rates.27 Another potential reason is that the speed of blastocyst development correlates with the occurrence of abnormal spindle morphology, consequently impacting implantation rates and pregnancy outcomes negatively.28 Moreover, our study suggested that prior transfer of a cleavage-stage embryo does not mitigate differences in subsequent D5 and D6 blastocyst transfers in frozen-thawed cycles.
As embryo quality plays a crucial role in pregnancy outcomes in IVF treatment,29 we explored the differences in pregnancy outcomes between the D3/D5 group and the D3/D6 group based on embryo quality. We observed a significant increase in the clinical pregnancy rate in the D3/D5 group compared with the D3/D6 group when both transferred embryos were of good quality, but not when both were of poor quality. However, given the limited sample size of transferred poor-quality embryos in either D3/D5 or D3/D6, the wide interval of effect size undermines the robustness of the results. Importantly, even when comparing a good-quality cleavage-stage embryo and a poor-quality blastocyst in the D3/D5 group to two good-quality embryos in the D3/D6 group, the clinical pregnancy rate remained significantly higher in the D3/D5 group than in the D3/D6 group. To the best of our knowledge, there may not yet be studies comparing the impact of the quality of D5 and D6 blastocysts following the transfer of high-quality D3 embryos on pregnancy outcomes in SeET. However, we can catch a glimpse of recent research examining the disparities in pregnancy outcomes between the transfer of a single good-quality blastocyst on D6 versus a single poor-quality blastocyst on D5 in frozen-thawed cycles,30 which reported a trend of a higher live birth rate of D5 poor-quality blastocyst transfer than that of D6 high-quality blastocyst transfer. Both studies pointed in the same direction: blastocysts available for transfer on D5 should be prioritised irrespective of their quality. Delaying transfer to day 6 should be avoided, whether in the conventional embryo transfer strategy or the SeET strategy. Still, additional studies are required to thoroughly investigate the impact of embryo quality on SeET.
Age emerges as another pivotal factor to consider, as it exerts a significant influence on the success of IVF treatment. We set the cut-off age at 35 years for conducting stratified analysis. The results indicated that women under 35 years experiencing RIF may derive greater benefit in terms of clinical pregnancy with D3/D5 SeET compared with D3/D6 SeET. This preference may stem from the higher prevalence of oocyte aneuploidy and reduced uterine receptivity among older women.31 Thus, the influence of D3/D5 and D3/D6 strategies on pregnancy outcomes may be overshadowed by advanced maternal age. When considering both embryo quality and maternal age, similar findings emerged: no significant differences in pregnancy outcomes were observed in women aged 35 or older, regardless of whether two good-quality or two poor-quality embryos were transferred. This suggested that the impact of age outweighs that of embryo quality in SeET in IVF treatment. A more thorough and comprehensive evaluation is essential to determine the most suitable embryo transfer strategy for women with RIF at an advanced age.
Notably, among the parameters examined, the stage of blastocyst development in SeET emerged as the sole factor showing a substantial association with clinical pregnancy in the multivariable model. This underscores the significant impact of blastocyst development in SeET on pregnancy outcomes. Our study contributes to the limited existing evidence regarding the comparison of pregnancy outcomes between D3/D5 and D3/D6 SeET, especially the comparison of the subsequent transfer of poor-quality D5 blastocysts and good-quality D6 blastocysts after a good-quality D3 cleavage-stage embryo. These findings advocate for prioritising blastocysts available for transfer on D5 in the SeET strategy, irrespective of their quality. These data will help guide the selection of embryonic strategies for SeET in women with RIF.
However, there are some limitations to this study. The retrospective study design is susceptible to inherent confounding and bias. Additionally, the sample size may still be insufficient for conducting stratified analyses based on the quality of transferred embryos, particularly within the category of poor-quality embryos. Due to the lower occurrence rates, the results regarding comparisons between the two groups regarding early miscarriage, ectopic pregnancy and multiple pregnancies lacked robustness. Furthermore, the absence of routine screening with PGT prevents us from determining the euploidy rate and exploring its potential role. Last, we have not finalised the follow-up of these women in terms of healthy live birth rate, which is a more concerning endpoint than clinical pregnancy32 and could not be analysed in this study.
Conclusion
The clinical pregnancy rate was significantly higher in the D3/D5 group compared with the D3/D6 group, particularly among women under 35 years or in cases involving transfers of two good-quality embryos. When contrasting the outcomes of transferring a good-quality cleavage-stage embryo and a poor-quality blastocyst in the D3/D5 group with transferring two good-quality embryos in the D3/D6 group, the clinical pregnancy rate was significantly higher in the D3/D5 group. However, among women aged 35 years or older, there were no significant differences in pregnancy outcomes.
Data availability statement
Data are available upon reasonable request.
Ethics statements
Patient consent for publication
Acknowledgments
We thank all participants involved in this study.
References
Supplementary materials
Supplementary Data
This web only file has been produced by the BMJ Publishing Group from an electronic file supplied by the author(s) and has not been edited for content.
Footnotes
ZW and JG are joint first authors.
XL and RL contributed equally.
Contributors XL and RL conceived the study; ZW and JG collected and analysed the original data; XL, RL, ZW and JG contributed to data interpretation and manuscript editing. RL accepts full responsibility for the work and/or the conduct of the study, had access to the data, and controlled the decision to publish.
Funding The research was supported by Capital’s Funds for Health Improvement and Research (CFH2024-2-40910).
Competing interests RL and XL have served as the editorial member of GOCM. There are no competing interests for other authors.
Patient and public involvement Patients and/or the public were not involved in the design, or conduct, or reporting, or dissemination plans of this research.
Provenance and peer review Not commissioned; externally peer reviewed.
Supplemental material This content has been supplied by the author(s). It has not been vetted by BMJ Publishing Group Limited (BMJ) and may not have been peer-reviewed. Any opinions or recommendations discussed are solely those of the author(s) and are not endorsed by BMJ. BMJ disclaims all liability and responsibility arising from any reliance placed on the content. Where the content includes any translated material, BMJ does not warrant the accuracy and reliability of the translations (including but not limited to local regulations, clinical guidelines, terminology, drug names and drug dosages), and is not responsible for any error and/or omissions arising from translation and adaptation or otherwise.