68. PREIMPLANTATION GENETIC TESTING OF MONOGENIC DISEASE: EXPERIENCE IN RUSSIA

Abstract

Introduction Preimplantation genetic testing of monogenic disease (PGT-M) is technically challenging, because extremely small amount of biomaterial increases allele dropout (ADO) rates, contamination significance, failed amplification (FA) events. We report here our experience of PGT-M in “Genetico” center in Russia. Material & methods A retrospective analysis of all requests and cycles of PGT-M referred to our center was performed. Embryo biopsies from 29 medical centers were sent for PGT-M. Embryos were mostly biopsied on day 5-6. PGT-M assays combine direct diagnosis of the pathogenic variants and linkage analysis of highly heterozygous STRs. The nested PCR was used for DNA amplification from different sources: single cells or whole genome amplification (WGA) products of embryo biopsy, total DNA. PGT-A by NGS or aCGH was performed for unaffected embryos upon patient's request. Results Of 109 couples referred to our center for PGT-M, 92 completed preliminary test for PGT-M for 42 genetic condition and HLA gaplotyping: 24 autosomal-dominant (AD) requests, 54 autosomal recessive (AR), 13 X-linked requests. The most frequent indication was spinal muscular atrophy (17 couples) and cystic fibrosis (8 couples). All these couples referred for PGT-M not after preconception screening, but because of affected child birth. We performed 85 PGT-M cycles with 413 embryos. Almost all of these samples undergo WGA for possibility to combine PGT-M with PGT-A, also each test-system was validated for single-cell either. The WGA failed in 16 cases (3,9%). The reliability was decreased for 12 embryo results (3%), because of increased number of ADO or marker FA events. Median number of markers included in test-systems was 12 and for embryo analysis it was 10. These highly informative test-systems contributed to low number of inconclusive results – only for 7 samples (1,8%) ( Girardet et al. 2018 ). The recombination events are not the reason to decrease the reliability if sufficient number of informative markers employed. The recombination event was revealed in 39 samples (9,4%). Median distance between markers of test-system (2,97 Mb) suggests lower recombination rate 3,75%. Of all analyzed embryos 131 (31,7%) did not inherited any of pathogenic alleles (AD – 61; AR – 48; X-linked – 11), 135 (49%) were carriers (AR – 122; X-linked – 11), 136 (32,9%) were affected (AD – 49; AR – 67; X-linked – 10). For 156 (58,6%) unaffected embryos PGT-A was performed and 91 (34,1%) were suitable for transfer. Chromosomal abnormalities for three embryos were revealed at PGT-M stage and confirmed by PGT-A. At the moment we have information about 43 transfers, 19 pregnancies and 7 healthy births and no affected pregnancy or birth. Conclusions Highly informative test system and accurate analysis of results can lead to both - high accuracy of obtained results and decreased number of embryos, that were rejected because of inconclusive results. PGT-M appears to be more robust analysis than PGT-A.