Preimplantation genetic testing for four families with severe combined immunodeficiency: Three unaffected livebirths

Abstract

Purpose: Severe combined immunodeficiency (SCID) is a set of rare monogenic inherited diseases that together represent the most severe form of the primary immunodeficiency disease phenotype. Preimplantation genetic testing for monogenic defects (PGT-M) is an effective reproductive technology strategy to prevent disease-causing gene mutations from being transmitted to offspring. The aim of this study was to report the use of PGT-M strategy based on karyomapping in four families to avoid the birth of SCID children.

Methods: Four couples underwent the PGT-M strategy due to SCID. The strategy of PGT-M started with a biopsy of the trophectoderm cells of embryos, and the whole genome was amplified by multiple replacement amplification (MDA). Then, the single nucleotide polymorphisms (SNPs) in the region upstream and downstream of the mutation site were subsequently identified via karyomapping, and the results were analyzed via SNPs linkage analysis. The aneuploids of the embryos were identified simultaneously. Finally, prenatal amniocentesis was used to verify the validity of the PGT-M results.

Results: We identified three novel variants (case1: IL2RG c.720_726delGAGCCAC; case 3: RAG2 c.770 C > T; and case 4: LIG4 c.1347 A > T). All four couples with SCID pathogenic gene mutations were subjected to karyomapping linkage analysis, and embryos with the pathogenic gene mutation were successfully identified. Euploid blastocysts without pathogenic alleles were transplanted, and healthy offspring were ultimately born. Prenatal diagnosis also confirmed the validity of our results.

Conclusion: This study revealed that karyomapping is an efficient approach for identifying SCID. Through PGT-M with karyomapping linkage analysis, healthy babies were born to families carrying mutations in the SCID pathogenic gene.

Keywords: Karyomapping; PGT-M; Rare disease; SCID.

Fig. 1

Workflow of PGT for severe combined immunodeficiency disease by karyomapping. ICSI, intracytoplasmic sperm injection; TE, trophoblastic ectodermal; MDA, multiple replacement amplification; PGT, preimplantation genetic testing for monogenic defects; E, embryo; CNV, copy number variation

Fig. 2

Pedigree of the four families carring SCID pathogenic genes. The upper right box shows the symbols and their meanings. The genotypes are displayed below the family members. WT, wild type. n/a, not available

Fig. 3

PGT-M strategy for severe combined immunodeficiency disease. (A) Pedigree diagram and linkage analysis identified for the disease-carring allele. We sequenced the amplified genomes from each embryo, couple, and reference. This is the pedigree diagram of case 2 in the PGT-M cycle. The SNPs in the upstream and downstream regions flanking the IL2RG gene within a range of 2 Mb are shown on the left. The heterozygous SNPs (AB) for the female and homozygous SNPs (AA or BB) for the male and the female’s mother were applied in the linkage analyses. The yellow bars indicate the pathogenic allele in the female’s mother passed by the female to embryos E1 and E2. (B) Results of CNV detection in abnormal embryos. Top: CNVs of abnormal embryos E1 by karyomapping microarray; Middle: CNVs on the chromosome 16 of abnormal E1 embryos. Bottom: CNVs of normal E5 embryos identified via a karyomapping microarray. M, male; F, female; R, reference, female’s mother; E, embryo; SNP, single nucleotide polymorphism; CNV, copy number variation