Background: The revised guidelines for the management of medullary thyroid carcinoma recommend that genetic counseling regarding reproductive options, including preimplantation genetic diagnosis (PGD), be considered for all RET mutation carriers of reproductive age to avoid the transmission of multiple endocrine neoplasia type 2 (MEN2). However, the high complexity and cost of PGD have hindered its widespread use. Thus, it is necessary to establish a simple and relatively inexpensive method to facilitate the PGD of MEN2.
Patients and methods: A customized Nimblegen EZ sequence capture array was designed to capture the targeted regions, including the RET gene, and 1 Mb range on each side of the RET gene. Targeted, capture-based next-generation sequencing of three members of one family with MEN2A (the couple and the paternal father) was conducted to identify the informative markers. The diagnosis of the embryos was achieved through haplotype analysis based on informative markers and causative mutation.
Results: Based on the sequencing results, 173 informative markers were detected, which were sufficient for the subsequent use for PGD. Seven informative markers and the causative mutation (RETC634Y) were selected and subjected to Sanger sequencing. Through haplotype analysis, four embryos without inheritance of the mutation haplotype of the RET gene were diagnosed as unaffected. One unaffected embryo was transferred, with one healthy baby born at 38 gestational weeks.
Conclusions: Targeted, capture-based next-generation sequencing for identification of informative markers together with Sanger sequencing is an easy and efficient method for the PGD of monogenic diseases such as MEN2.
Keywords: Sanger sequencing; informative marker; multiple endocrine neoplasia type 2; preimplantation genetic diagnosis; targeted sequencing.