69. IMPROVED IVF SUCCESS OF COMBINED PGT-M AND PGT-A APPLICATIONS

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 testsystems 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 testsystem (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.

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Keywords: PGT-M; single-gene disorde; retrospective analysis; PGT-A

REFERENCE Girardet, A, Ishmukhametova, A, Viart, V, Plaza, S, Claustres, M Thirteen years' experience of 893 PGD cycles for monogenic disorders in a publicly funded, nationally regulated regional hospital service. Reprod Biomed Online 2018 Feb; 36: 154–163

doi: 10.1016/j.rbmo.2019.04.121

69. IMPROVED IVF SUCCESS OF COMBINED PGT-M AND PGT-A APPLICATIONS

E. Unsal1, S. Aktuna2, M. Aydin1, L. Ozer1, V. Baltacı1 1  Mikrogen

Genetic Diagnosis Laboratory, Ankara, Turkey 2  mikrogen, ankara, Turkey

Introduction: Preimplantation Genetic Testing (PGT) has been performed since 1989 and utilized in the diagnosis of all genetic conditions at preimplantation stage (1). In the previous years, only a disorder could be diagnosed with the use of multiplex nested PCR in PGT applications. Increased use of trophectoderm biopsy and vitrification together with Whole Genome Amplification (WGA) technology enabled the detection of single gene disorders together with 24 chromosome screening. This approach allowed us to select healthy and chromosomally euploid embryos. Even though single gene disorder carrier patients are mostly young and fertile, they still have high risk of spontaneous miscarriage. In the recent years, it has been reported that when the embryos are screened for aneuploidy (PGT-A) in addition to single gene disorder testing (PGT-M), pregnancy rates can be increased from 45% to 68% and spontaneous miscarriage rates decreased from 15% to 5%. 34 single gene disorder patients, whose embryos were found to be healthy in terms of single gene disorder, were included in this study and these

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embryos were further screened for aneuploidy using Next Generation Sequencing (NGS) technology (2). Materials & Method: On the 5th and 6th days of embryonic development, WGA was performed using SurePlex DNA Amplification System (Illumina, USA) and DOPlify (PerkinElmer Health Sciences - Australia). Following multiplex nested PCR on WGA samples, fragment analysis for haplotyping and sequence analysis for mutation detection were performed. Embryos found to be suitable for transfer regarding the single gene disorder have further been screened for aneuploidy using NGS technology with VeriSeq PGS Kit (Illumina, USA). Results: This study was conducted for single gene disoder and 24 chromosome screening in 44 patients with an average age of 34. 280 embryos derived from these patients were included in PGT-M. Among 153 embryos that were found to be suitable for transfer after PGT-M, PGT-A was performed on 121 embryos upon patients' requests and 49% of these embryos were euploid. Embryo transfer could not be performed in 12 patients due to the absence of euploid embryos. According to the data we acquired, 64% pregnancy was achieved in the rest of the patients. Conclusion: While pregnancy ratio can be improved with this approach, Allele drop out (ADO) and contamination errors are still major limitation which can lead to misdiagnosis of embryos. ADO ratio was demonstrated to be 25% in PGT for blastomere cells, whereas this ratio drops down to 5% when 5-7 cells obtained by trophectoderm biopsy are tested. Thus trophectoderm cells are more appropriate for PGT-M applications combined with PGT-A. Since stable and homogenous amplification of the entire genome is a crucial step, optimised WGA

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technologies will be a key factor in these applications. Keywords: Preimplantation Genetic Testing

for Aneuploidy; Preimplantation Genetic Testing for Monogenic/Single Gene Disorders; Next Generation Sequencing; In Vitro Fertilization

REFERENCES http://www.ivf-worldwide.com/cogen/oep/pgd-pgs/ history-of-pgd-and-pgs.html Rechitsky, S, Pakhalchuk, T, San Ramos, G, Goodman, A, Zlatopolsky, Z, Kuliev, A First systematic experience of preimplantation genetic diagnosis for single-gene disorders, and/or preimplantation human leukocyte antigen typing, combined with 24-chromosome aneuploidy testing. Fertil Steril 2015 Feb; 103: 503–512

doi: 10.1016/j.rbmo.2019.04.122

70. PREIMPLANTATION GENETIC TESTING FOR YQH-AUTOSOME TRANSLOCATION CARRIERS: ANALYSIS OF GAMETE SEGREGATION

I. Trofimova1, A. Saifitdinova2, J. Loginova3, A. Kinunen4, Z. Tonyan5, O. Chiryaeva6, J. Pastukhova7, O. Leontieva7, R. Kuznetsova7, N. Bichevaya8 1  International

center for reproductive medicine, Federal State Budget military educational institution of higher professional education “Military Medical Academy named after S.M. Kirov” under the Ministry of Defe, 53k1 Komendantskij prospectAlmazov National Medical Research Centre2 Akkuratova street, SaintPetersburg 197350197341, Russian Federation 2  International center for reproductive medicine, Herzen State Pedagogical University of Russia, 48 Moyka Embankment, 53k1 Komendantskij prospect, Saint-Petersburg 197350191186, Russian Federation 3  DiaCarta, Inc, 2600 Hilltop Drive, RichmondCalifornia, CA 94806, United States 4  International center for reproductive medicine, St. Petersburg Centre for Medical Genetics, 53k1 Komendantskij prospect5 Tobolskaya street, SaintPetersburg 197350, Russian Federation 5  Almazov National Medical Research Centre, 2 Akkuratova street, Saint-Petersburg 197341, Russian Federation 6  Federal State Budget military educational institution of higher professional education “Military Medical Academy named after S.M. Kirov” under the Ministry of Defense of the Russian Federation, D.O. Ott Research Institute of Obstetrics Gynecology and Reproductology, 6 Lebedeva street3 Mendeleev line, Sint-Petersburg 194044199034, Russian Federation 7  International center for reproductive medicine, 53k1 Komendantskij prospect, Saint-Petersburg 197350, Russian Federation 8  International center for reproductive medicine, 53k1 Komendantskij prospect, Saint-Petersburg 197350, Romania

Introduction: Y-autosome translocations occur in

approximately 1:2000 in population and represent special group of reciprocal exchanges. Due to a balanced nature they do not influence on phenotype in general. However, Y-autosome translocation can dislocate sex bivalent formation and chromosome synapsis in the first meiotic division prophase that led to gamete aneuploidy or pachytene arrest. More often Y-derived DNA, predominantly Yq12 band, is translocated onto the short arm of acrocentric chromosomes, mainly chromosomes 15 and 22 due to a strong correlation in their satellite DNA sequences. In women inheritance of such derivative acrocentric chromosomes could be associated with the risk of ovarian malignancy, but the particular mechanism of this pathological effect is not entirely clear since the Yq12 band consists of tandemly organized satellite DNA sequences. It can be associated with the transcription of satellite DNAwhich occurs during carcinogenesis. Material & methods: Two families with Yq12/15p and one – with Yq12/22p translocations were analyzed during IVF cycle with PGT in International Centre for Reproductive Medicine. The mean age of men and women was 36±2.8 and 34.25±1.4 years respectively. Karyotyping analysis was performed using GTG banding technique on metaphase chromosomes from peripheral blood lymphocytes. Fluorescence in situ hybridization (FISH) was used to clarify the breakpoint on chromosomes and confirm the chromosomes-specificity of the probes. Additionally, seven chromosomes (13, 14, 15 or 22, 16, 18, 21, X), which are not involved in rearrangement, were analyzed. Results: FISH with chromosome specific probes was performed on blastomere from cleavagestage embryo. A total of 42 blastomeres were analyzed. In 43% of the blastomeres «adjacent-1»

segregationwith derivate autosome and X or Y chromosomes was revealed. Segregation 3:1 was observed in 10% of the cells, with no predominant inheritance of the derivative autosome. In 21% of cells the type of chromosome segregation could not be determined due to mosaicismor polyploidy. 45% of the cells were aneuploid for autosomes, which are not involved in the translocation, which may be connected with interchromosomal effect. Conclusions: Carriers of Yq12/15p or Yq12/22p translocations have a predominant inheritance of the derivative autosome, which can be associated with the disruption ofsex bivalent segregation. This research was supported by Russian Foundation for Basic Research (grant #18-34-00279). Keywords: PGT-SR; translocation; satellite DNA; meiosis

doi: 10.1016/j.rbmo.2019.04.123

71. PREIMPLANTATION GENETIC DIAGNOSIS OF NEURODEGENERATIVE DISEASES

C.H. Liao1, M.Y. Chang2, G.C. Ma3, C.F. Lin4, S.P. Chang3, W.H. Lin3, H.F. Chen5, S.U. Chen5, Y.C. Lee6, C.C. Chao7, M. Chen8, S.T. Hsieh7 1  Department

of Pediatrics, National Taiwan University Children's Hospital, Taipei, Taiwan 2  Division of Pediatric Neurology, Department of Pediatrics, Changhua Christian Children's Hospital, Changhua, Taiwan 3  Department of Genomic Medicine and Center for Medical Genetics, Changhua Christian Hospital, Changhua, Taiwan 4  Department of Medical Genetics, National Taiwan University Hospital, Taipei, Taiwan 5  Department of Obstetrics and Gynecology, College of Medicine and Hospital, National Taiwan University, Taipei, Taiwan 6  Department of Neurology, Taipei Veterans General Hospital, Taipei, Taiwan 7  Department of Neurology, National Taiwan University Hospital, Taipei, Taiwan 8  Department of Genomic Medicine, Changhua Christian Hospital, Changhua, Taiwan

Introduction: Preimplantation genetic diagnosis (PGD) has become a crucial approach to help carriers of inherited disorders give birth to