P–158 Assisted Hatching on D + 3 in order to facilitate trophectoderm biopsy in blastocyst for PGT-A is not advisable in all patients
Authors: Belchin P , Cabello Y , Sanche. d. Burgos M , Guerrero J , Riva M D , Garcia-Enguidanos A , Izquierdo E , Ordonez D 
Published: August 2021

Abstract

Study question
Is it useful or beneficial to perform Assisted Hatching (AH) on D + 3 previously to biopsy for PGT-A on blastocyst stage on D + 5?

Summary answer
The routine use of AH on D + 3 to facilitate the embryo biopsy on D + 5 could negatively influence the development of the embryos to blastocyst stage.

What is known already
The blastocyst stage is the optimal stage for performing biopsies for PGT-A, which has been reported as a key factor determining the growing clinical application of this strategy worldwide. For trophectoderm (TE) biopsy, laser-assisted drilling is used to create a zona opening on D + 3 or D + 5 of development. The method of zona opening on D + 3 allows some of the TE cells to herniate during blastocyst formation and expansion, which facilitates the biopsy process. However, this method may result in herniation of inner cell mass cells instead of TE or maybe could affect the development of the embryo to blastocyst stage.

Study design, size, duration
A total of 100 PGT-A cycles were performed in 2019 and 2020. In 78 of them laser-assisted drilling was used to create a zona opening on D + 5 only in those embryos which arrived to blastocyst stage for TE biopsy (Group No-AH). In 22 cycles the same drilling was achieved on D + 3 in all embryos, independently of their quality (Group AH). The average of embryos per cycle in each group was 5 and 4.3 respectively.

Participants/materials, setting, methods
A total of 100 PGT-A cycles coming from 65 patients were studied. The average of the age of the patients was 40.83 (SD 3.45) in the group No-AH vs 42.18 (SD 3.42) in the Group AH (p = 0.108), so the age was not a determining factor for the development of the embryos. We analyzed by χ 2 test differences between groups on fertilization rates, number of embryos, development to blastocyst stage, euploidy and pregnancy rates.

Main results and the role of chance
The fertilization rate was 74.79% (No-AH group) and 68.53% (AH group) with no significative statistical differences (p = 0.12).
In the No-AH group, the TE biopsy was performed on D + 5 in 63 cycles (81%). In the AH group, 41% of cycles didn’t reach the blastocyst stage, obtaining statistical differences between groups (p = 0.035). We found also significant differences in the number of cycles with biopsied blastocyst when we had 1 to 6 embryos/cycle on D + 3 between groups (p = 0.002), without obtaining any blastocyst to be diagnosed in 53% of the cycles in AH group vs 27% in No-AH group. When the number of embryos on D + 3 per cycle was &gt; 6, at least 1 embryo reached the blastocyst stage in both groups, although this number was higher in No-AH group. The rate of biopsied blastocysts was significantly higher in the No-AH group compared to the AH group (46.61 vs 34.69) with a p = 0.031.
The rate of euploid embryos analyzed was 23.30% in the No-AH group compared to 29.41% in the AH group, although no significant differences were found (p = 0.44) between groups.
In the No-AH group, a clinical pregnancy rate of 52.94% was obtained (n = 34) vs 50% in the AH group (n = 4) (p = 0.91).

Limitations, reasons for caution
We have recently started to perform AH on D + 3, so the number of cases is smaller than No-AH group. We use a time lapse incubator in all cases, so in the No-AH the culture dish is changed, disturbing the stable incubation environment, while in the other group it is not.
Wider implications of the findings: The use of AH on D + 3 in order to facilitate the TE biopsy on D + 5 could affect negatively the development of the embryos to blastocyst stage. Its routine use should be avoided based on laboratory workload, mainly if the patient has less than 7 embryos at D + 3.

Trial registration number
Not applicable