Genetic Disorders and the Fetus. Группа авторов

will not predict the eventual genotype of the oocytes if crossover occurs, because the primary oocyte in this case will be heterozygous for the abnormal gene. The frequency of crossover varies with the distance between the locus and the centromere, approaching as much as 50 percent for telomeric genes, for which the first polar body approach would be of only limited value, unless the oocytes can be tested further (Figure 2.1). Therefore, the second polar body analysis is required to detect hemizygous normal oocytes resulting after the second meiotic division. In fact, the accumulated experience shows that the most accurate diagnosis can be achieved in cases where the first polar body is heterozygous, so the detection of the normal or mutant gene in the second polar body predicts the opposite mutant or normal genotype of the resulting maternal contribution to the embryo after fertilization.⁴

Figure 2.1 Scheme demonstrating the principle of preimplantation genetic analysis by sequential DNA analysis of the first and second polar body, using the cystic fibrosis (CF) locus as an example.

      Source: Verlinsky Y, Kuliev AMA. Preimplantation genetic diagnosis. In: Milunsky A, Milunsky JM, eds. Genetic disorders and the fetus: diagnosis, prevention and treatment, 6th edn. Oxford, UK: John Wiley & Sons, 2010.

To study a possible detrimental effect of the procedure, micromanipulated oocytes were followed and evaluated at different stages of development.^{3, 4, 35} The absence of any deleterious effect of polar body removal on fertilization, preimplantation, and, possibly, postimplantation development made it possible to consider the polar body approach as a nondestructive test for genotyping the oocytes before fertilization and implantation. In another study, to assess the effect of the second polar body sampling on the viability and developmental potential of the resulting embryo, 343 biopsied and 445 nonbiopsied mouse embryos were compared for the percentage of embryos reaching the blastocyst stage.³⁶

The results of PGT‐M performed by polar body biopsy, representing the world's largest series, is shown in Table 2.2. A total of 1,016 PGT‐M cycles were performed, for 538 autosomal recessive, 191 autosomal dominant, and 287 X‐linked disorders. Of 1,016 cycles initiated, 838 (82.5 percent) resulted in transfer of 1,656 embryos (1.98 embryos per transfer on the average), 349 (41.6 percent) clinical pregnancies, and 385 babies born. Only two misdiagnoses were observed in the case of PGT for fragile‐X syndrome and muscular dystrophy, which were due to consented transfer of additional embryo with insufficient marker analysis to exclude the probability of allele dropout (ADO) (see later). The example of PGT‐M by polar body sampling is shown in Figure 2.2.

Table 2.2 Clinical outcome of PGT‐M performed by polar body approach.

Conditions/mode of inheritance/sampling type	Patient	Cycles	Embryo transfer	No. embryos	Pregnancy	Spontaneous abortions	Baby
Autosomal recessive
Polar bodies	76	131	99	204	36	10	36
Polar bodies + blastomere/blastocyst	254	407	344	701	143	21	168
Subtotal	330	538	443	905	179	31	204
Autosomal dominant
Polar bodies	29	52	40	84	22	4	21
Polar bodies + blastomere/blastocyst	79	139	122	233	49	7	61
Subtotal	108	191	162	317	71	11	82
X‐linked
Polar bodies	39	86	63	110	22	4	20
Polar bodies + blastomere/blastocyst	116	201	170	324	77	12	79
Subtotal	155