Optimisation of embryology procedures and clinical outcomes in preimplantation genetic testing of human embryos

Since the very first Preimplantation Genetic Testing (PGT) case in 1989, the field has seen many changes; in indications, in diagnosis platforms and in embryology procedures. While PGT-M (monogenic) is widely accepted as an option for couples at risk of having a child with a serious inherited condition, PGT-A (aneuploidy) and PGTSR (structural rearrangements) remain more controversial, with some still questioning the validity and utility of the data produced.

Some of the disagreement surrounding PGT-A and PGT-SR has stemmed from variation in approaches and results from different genetic laboratories and different assisted reproduction technology (ART) centres. As with all ART procedures, there are many variables that contribute to the success or failure of a treatment cycle, beyond the chromosome data produced from a trophectoderm sample. As well as technical aspects in the genetics and embryology laboratories, patient decision making contributes. Maximising clinical outcomes also depends on providing patients with

reliable data and helping them make informed choices and treatment decisions, and minimising financial and emotional cost by not subjecting them to unnecessary procedures with little chance of success.

In order to investigate and, in part, redress variability in the field of PGT, this thesis

had 7 specific aims:

• To liaise with diagnostic labs, embryology labs and medical affairs to create a unique set of guidelines for embryology labs wishing to use diagnostic services (chapter 2). Such a set of guidelines was created.

• To test the hypothesis that intracytoplasmic sperm injection is necessary as a preventive measure against paternal cell contamination in preimplantation genetic testing. Hence to ask if it is necessary for all patients undertaking PGT– even those with proven fertility – to have to undergo and pay additional fees for ICSI (chapter 3). Here, it was established that ICSI was not necessary

• To provide proof of principle that re-biopsy is, technically, a viable strategy when no result is obtained (chapter 4). This was established.

• To examine the “no result rate” in a leading group of UK PGT diagnostic laboratories since the introduction of next generation sequencing (NGS) to examine how much it varies between referring ART clinics, and assess whether re-biopsy is a viable alternative for most patients (chapter 5). It was established that re-biopsy was indeed a viable strategy.

• To perform a retrospective analysis of over 1,800 PGT-SR samples (479 cycles of 5 years) in order to: a) Provide the biggest dataset to date on PGT-SR outcomes using CCS to date; b) Test the hypothesis that the level and nature of structural chromosomal rearrangements is correlated to age, sex and time of biopsy; c) To test the hypothesis that an interchromosomal effect exists in this dataset; d) To provide a set of practical guidelines for genetic counsellors to advise patients on their likelihood of having euploid embryos for transfer based on the type of chromosome rearrangement, sex of the carrier of the rearrangement, maternal age, and any other factors that may be implicated (chapter 6). A retrospective analysis of over 1,800 PGT-SR samples (479 cycles of 5 years) was performed, to date on PGT-SR outcomes using CCS, finding no demonstrable ICE. A set of practical guidelines was put forward for genetic counsellors to advise patients on their likelihood of having euploid embryos for transfer based on the type of chromosome rearrangement, sex of the carrier of the rearrangement, maternal age, and any other factors that may be implicated

• To provide novel insight into the mechanism of twinning as a result of a PGT case (chapter 7). Here evidence is presented of twinning occurring later in development – post day 5 development, but prior to implantation - thereby shedding light into a possible novel mechanism. That is, I hypothesise that this mechanism may involve splitting of the embryo as the inner cell mass hatches, with potential involvement of altered zonal lysis and apoptosis.

The data presented in this thesis points to continued differences in practice in the field, that will continue to lead to variable PGT outcomes – as with standard IVF treatment cycles – unless further research is performed, with large data sets presented which can point toward best practice.