Clinical re-biopsy of segmental gains-the primary source of preimplantation genetic testing false positives

Abstract

Purpose: Does re-biopsy of blastocysts classified as abnormal (ABN) due to segmental aneuploidy (SA) have clinical utility?

Methods: The live birth (LB) outcomes of mosaic SAs, compared to other categories, were determined after transfer of 3084 PGT-A tested blastocysts. An initial 12-month trial thawed 111 blastocysts classified as ABN due to a SA for clinical re-biopsy, with an additional 58 from a subsequent 16-month revised protocol. Where re-biopsy failed to corroborate the original classification, blastocysts were reported as mosaic and suitable for clinical use.

Results: Segmental mosaics had a LB rate (54.1%) which was indistinguishable from that of euploid (53.7%). Numeric mosaics had statistically significant (P < 0.05) reduced LB rates compared to euploid, with high-level numerics (19.2%) also exhibiting a significant reduction compared to low level (42.3%). Of the initial 111 blastocysts with SAs, 85 could be re-biopsied. Segmental gains became suitable for re-biopsy at a high rate (90.9%), with 84.2% (16/19) of these reclassified as mosaic. Only 73.0% of deletions and complex changes were suitable for re-biopsy, of which 73.0% (46/63) were confirmed ABN. The subsequent 16-month period primarily focused on gains, confirming the high rate at which they can be reclassified as clinically useable.

Conclusions: Blastocysts harboring mosaic segmental duplications, rather than SAs in general, are the primary source of false-positive PGT-A results and represent a category with a LB rate similar to that of euploid. A high degree of confidence in the reliability of PGT-A results can be maintained by performing confirmatory clinical TE biopsies.

Keywords: Clinical re-biopsy; False positives; Mosaicism; Preimplantation genetic testing; Segmental aneuploidy.

Fig. 1

Abnormal and mosaic results from PGT-A embryos. a Percentage of 14,705 PGT-A tested embryos that were classified as ABNn (one or more numeric abnormalities), ABNs (one or more SAs), ABNb (numeric + SAs), A (NAD; no abnormality detected), B1 (noisy/mosaicism < 20%), B2 (mosaic SAs 20– < 80%), B3 (numeric mosaicism 20–40%), C (numeric mosaicism > 40– < 80%), or UNK (no result). The percentages of B2 samples with single or multiple SAs are indicated. For the ABNs and B2 categories, “both” refers to the percentage of samples that contained at least one gain as well as at least one loss. b For the ABN category, the number of SAs detected (black) is generally in proportion to the size of each chromosome (grey), with the relative chromosome size derived from the number of Mbs for which VeriSeq NGS produces mappable data

Fig. 2

Clinical re-biopsy of embryos harboring SAs. a Examples of SAs, illustrating a simple gain on chromosome 2 (top), complex losses on chromosome 5 (middle), and both (a full gain and loss) on chromosome 8 (bottom). b Proportion of thawed embryos for which a re-biopsy result was not obtained (stripped), were confirmed ABN (stippled), or were reclassified as mosaic (white), stratified by TE grade and day of the original biopsy, which indicated poor survival and a high rate of concordance for day 6 TE3 embryos in particular. The number of embryos in each category is indicated. c Thawing/clinical re-biopsy results demonstrating the high percentage of simple gains reclassified as mosaic (white) and the small proportion confirmed ABN (stippled) or discarded (stripped) compared to the other four SA subgroups. d The number of thawed SAs and those reclassified as mosaic (white) or confirmed ABN (stippled) is broadly in proportion to the size of each chromosome