Dados do Trabalho

Title

Two DuoStim, two healthy babies case report of a rare non-classic congenital adrenal hyperplasia patient after preimplantation genetic test during in vitro fertilization

Objective

Non-classic congenital adrenal hyperplasia (NCAH) due to 21-hydroxilase deficiency is an autosomal recessive disorder. NCAH patients typically have 21-hidroxylase (21-OH) deficiency associated with CYP21A2 mutations. Over 200 CYP21A2 mutations have been reported, and approximately 10 of them account for the majority of affected alleles. Here, we describe a case report of two successful pregnancies of a NCAH patient presenting with one heterozygous deletion of the gene CYP21A2 plus a rare substitution in the promoter region (-102 G>A), and a partner who also has one heterozygote mutation in the same gene (Q318X).

Methods

She (31 years old) has NCAH confirmed by genetic testing with the diagnosis of a heterozygous deletion in the gene CYP21A2 plus a rare substitution in the promoter region, -102 G>A. Her husband (32 years old) was also tested and a heterozygous mutation (Q318X) was detected. After a year of infertility, the couple underwent two IVF treatments (DuoStim cycles) and Preimplantation Genetic Testing for Monogenic Diseases (PGT-M) to achieve two pregnancies. A direct diagnosis of Q318X mutation and an indirect diagnosis using informative Small Tandem Repeats (STRs) were optimized for the couple. Two independent diagnoses were applied to each embryo during PTG-M. The causative mutation detection was performed by Polymerase Chain Reaction (PCR), and the mutant site was detected using the MiniSequencing Reaction. A total of six STRs (RING3, TAP1, D6S1666, TNFa, D6S265 and D6S1683) were found surrounding the gene CYP21A2 and used as indirect diagnosis. The STRs were chosen if contained a tetranucletide repetition core, linked to the gene (upstream or downstream) and having the highest heterozygosity value. Genomic DNA was extracted from peripheral blood from the couple and both sets of parents. The STRs were classified as being informative (both couple heterozygous), partially informative (one of the couple homozygous and the other heterozygous) or not informative (both homozygous). After embryo genetic test by PGT-M, artificial endometrial preparation was performed for the transfer of frozen–thawed embryos.

Results

From the first DuoStim cycle, six top-quality embryos were biopsied and frozen. Four embryos were identified as carrier of two mutations (maternal and paternal), one embryo was carrier of only one maternal mutation, and one was carrier of both maternal mutations and the wild paternal allele. The combined PGT-A study of the embryos revealed four euploids, being two males and two females, one mosaic aneuploid and one triploid. A pregnancy was achieved after the transfer of two euploid male carrier embryos. For the second pregnancy, the couple was subjected to another DuoStim cycle resulting in eight top-quality embryos, which were genetically analyzed. Three embryos were carriers of a maternal and a paternal mutation, four were identified as carrier of only a maternal mutation and one embryo was non informative. From the four embryos carriers of a maternal mutation, three were euploid. After an implantation failure of a single carrier euploid embryo, the couple got pregnant of a second baby after a transfer of two carrier euploid embryos. Both babies were born healthy after an IVF PGT-M treatment.

Conclusion

The association of genetic counseling and assisted reproductive technologies with the use of preimplantation diagnosis can help couples to have healthy babies in a rare NCAH cases.

Keywords

NCAH, CYP21A2, PGT-M, IVF