Empirical Studies on the Evaluation of Current Embryo Selection Techniques in Human in-vitro Fertilization

In vitro fertilization (IVF) is an assisted reproductive technology (ART) that aims to assist intended parents overcome fertility issues, including an infertility diagnosis and other barriers to conception. It involves harvesting gametes from a male and female provider to generate embryos, with the subsequent transfer of these embryos into a uterus with the goal of establishing a viable pregnancy and a healthy take home baby.

Technological advances in the field have led to drastic improvements in outcomes over the years, including selection methods of the most competent embryo for transfer within a cohort. Embryo selection tools include evaluating an embryo's chromosomal composition (preimplantation genetic testing for aneuploidy, or PGT-A) and other biomarkers/conditions that may have an impact on implantation and the ensuing pregnancy. Notably, many such methodological innovations (even some routinely used in the industry) still lack appropriate validation and are in dire need of unbiased vetting. The primary goal of this thesis is to evaluate current embryo selection techniques. This work encompasses the following items:

(1) Evaluating if current DNA sampling techniques (biopsy) of an embryo is an adequate predictor of the remaining embryo, which is a central premise of preimplantation genetic testing for aneuploidy (PGT-A); I conclude that a clinical trophectoderm (TE) biopsy correctly predicts aneuploidy in the inner cell mass (ICM) in the vast majority of cases

18

Empirical Studies on the Evaluation of Current Embryo Selection Techniques

when one or several chromosomes are abnormal, but aneuploidy of segmental (sub- chromosomal) nature is not well correlated in paired TE-ICM samples.

(2) Assessing of clinical outcomes of embryos classified as mosaic by routine clinical biopsy; I conclude that embryos classified as mosaic by PGT-A are associated with different clinical outcomes when compared to embryos diagnosed as euploid by PGT-A, namely reduced rates of implantation, ongoing pregnancy, and live birth.

(3) Defining features of mosaicism to rank embryos by their potential for a positive outcome; I conclude that mosaic level (putative percent aneuploid cells in the biopsied sample) and mosaic type (aneuploidy involved in the mosaicism) correlate with specific clinical outcomes and can be used to prospectively rank mosaic embryos in the clinical setting.

(4) Evaluating mitochondrial DNA (mtDNA) levels determined from a clinical trophectoderm biopsy as a potential predictor of implantation; I conclude that accurate calculation of mtDNA levels shows there is no association with embryo ploidy or implantation potential.

(5) Studying the health of babies from embryos reported to have high mtDNA levels; I conclude that unusually high mtDNA levels did not preclude blastocyst implantation and healthy births.

19

Empirical Studies on the Evaluation of Current Embryo Selection Techniques

(6) Performing a theoretical evaluation on the potential application of genome editing to human embryos such that abnormal embryos may be repaired and reclassified as transferable; I conclude that demand for germline genome editing (GGE) to cure heritable genetic diseases is grossly over estimated, mainly due to the role of preimplantation genetic testing, but nevertheless future applications may exist to correct chromosomal abnormalities and rescue aneuploid embryos.

(7) Exploring SARS-CoV-2 infection and its implications on embryo viability and its potential effect on strategies for embryo selection; I conclude that preimplantation embryos are vulnerable to SARS-CoV-2 infection, which can compromise their health and viability.

The data presented in this thesis empirically evaluates the utility of several embryo selection techniques on the IVF cycle. The array of methods available is certain to grow in time; to ensure that new instruments of selection or refinements to current tools continue to have a positive effect on outcomes and success rates, they must be evaluated thoroughly before being applied in the clinic in full force.