Pregnancy following exclusion testing for SCA3 on polar bodies in a fresh cycle: a comparison of STR-based targeted haplotyping and genome-wide SNP analysis and meiomapping

Abstract

Introduction Preimplantation genetic testing of monogenic disease (PGT-M) is allowed in Germany only with approval from the federal ethics committee and autosomal dominant disorders are often rejected. Polar body analysis, however, does not require ethical approval. A 34-year-old asymptomatic patient with a family history of spinocerebellar ataxia type 3 (SCA3) applied for PGT-M on trophectoderm samples but was rejected. The patient's father has a repeat expansion in the ATXN3 gene and is affected with SCA3, a progressive autosomal dominantly inherited neurological disease characterized by ataxia, spasticity and ocular movement abnormalities. As the patient does not want to know her status, we offered exclusion testing based on polar body analysis in a fresh cycle and have compared targeted haplotyping by STR analysis with genome-wide SNP genotyping and meiomap analysis. Material and methods The patient and her father were genotyped for 12 STR markers within 0.44 Mb of the ATXN3 gene on 14q32. 11 STRs were heterozygous (9 fully informative) and used for phasing high and low risk haplotypes. Eggs were collected (Day 0) and the first polar body (PB1) biopsied from mature MII oocytes. ICSI was then performed and PB2 biopsied from three fertilised zygotes on Day 1. After whole genome amplification (WGA) (Repli-g Single Cell Kit; Qiagen, Venlo, Netherlands) multiplex PCR was performed and the products run on a capillary sequencer. Haplotypes were constructed based on the pre-case workup. In addition, genome-wide SNP genotyping of the DNAs from the patient, her father and the WGA products from the polar bodies was carried out (HumanKaryomap-12 SNP array; Illumina, San Diego, USA) for maternal haplotyping and meiomap analysis. Results WGA and multiplex PCR was successful for all of the polar bodies except for the PB2 of Oocyte 1. As all 11 closely linked informative STRs were homozygous for the high-risk haplotype in PB1 from this oocyte, the corresponding PB2 and oocyte was deduced to have the low risk haplotype assuming normal segregation during meiosis. For Oocytes 2 and 3, the PB1 was heterozygous for most STRs and the PB2 of these oocytes were homozygous for the high and low risk haplotypes, respectively. Therefore, the embryos derived from Oocytes 1 and 2 were deduced to have the low risk maternal haplotype while the embryo from Oocyte 3 should have the high-risk haplotype. On Day 5, the blastocysts arising from Oocytes 1 and 2 were transferred, resulting in an ongoing pregnancy (currently week 10). Genome-wide SNP genotyping and meiomap analysis was concordant with STR analysis in all cases. Conclusions We report a pregnancy following targeted STR-based haplotyping and exclusion testing of an autosomal dominant disease by polar body analysis in a fresh cycle. Genome-wide SNP genotyping and meiomap analysis was fully concordant with the STR results but had the advantage that it also demonstrated that segregation of the maternal chromosome 14 was normal and predicted that the embryos from the two oocytes in which both polar bodies were analysed were euploid for all maternal chromosomes.