Abstract:

Objective We report a case of application of third-generation sequencing (TGS) combined with preimplantation genetic testing (PGT) for successful prevention of hereditary spastic paraplegia (HSP) caused by SPAST gene mutations and assess the value of PGT-M and TGS in managing hereditary spastic paraplegia. Methods A family affected by HSP underwent whole exon sequencing (WES), and a c.1699G>T mutation in the SPAST gene was identified. The mutation site in the proband was confirmed through Sanger sequencing. A single nucleotide polymorphism (SNP) site flanking the SPAST gene mutation was selected as the genetic linkage marker, and a SNP haplotype carrying the mutated gene was constructed. Ovarian stimulation using an antagonist regimen was performed for oocyte retrieval, followed by intracytoplasmic sperm injection (ICSI) and embryo culture. Blastocyst trophectoderm cells were biopsied for preimplantation genetic testing for monogenic disorders (PGT-M) to allow the selection of disease-free embryos for transfer. Results In this cycle, a total of 20 oocytes were retrieved, among which 18 were successfully fertilized to result in 12 blastocysts eligible for biopsy. Genetic testing revealed that all the 12 blastocysts were successfully amplified and confirmed as euploidy. Among them, 8 blastocysts did not carry paternal mutations, and a high-quality euploid embryo was selected for frozen embryo transfer (FET). Subsequent amniotic fluid testing during pregnancy confirmed the absence of paternal mutations in the fetus, resulting in the birth of a healthy baby girl. Conclusion For cases of genetic diseases with missing pedigree data, the application of third-generation sequencing and PGT-M technique can effectively block vertical transmission of SPAST gene mutation to the offspring, avoid pregnancy with an aneuploid embryo, and help families with autosomal dominant HSP obtain healthy offsprings.

Key words: hereditary spastic paraplegia, SPAST gene, preimplantation genetic diagnosis, third-generation sequencing, single nucleotide polymorphisms