Hiva Fassihi

Prenatal diagnosis for severe inherited skin disorders: 25 years' experience

Backgroundu2002 Over the last 25u2003years there have been major advances in methods for prenatal testing of inherited skin disorders. Since 1979, our group at the St Johns Institute of Dermatology has performed 269 prenatal diagnoses, using a variety of approaches, including fetal skin biopsy (FSB), chorionic villus sampling (CVS) and preimplantation genetic diagnosis (PGD).

Prenatal Diagnosis of Epidermolysis Bullosa

One of the most significant benefits of translational research in dermatology has been the development of prenatal diagnosis for couples at risk of recurrence of severe inherited skin diseases. Indeed, over the last 30 years a greater understanding of the molecular basis of epidermolysis bullosa (EB), as well as technical refinements in laboratory procedures, has facilitated the development of several different approaches for prenatal diagnosis. Initial tests were based on fetal skin biopsy sampling, but these have largely been superseded by DNA based analyses, mostly using fetal DNA derived from chorionic villus sampling taken at around 10--12 weeks gestation. Further advances, however, have led to the introduction of licensed preimplantation genetic screening for some forms of EB, an approach that defines a disease-associated genotype before implantation into the uterus. Pioneering research also continues to try to develop less invasive approaches with the prospects of maternal blood sampling early during the first trimester as a feasible objective. The availability of several different options for prenatal diagnostic testing therefore has led to an increased choice for families at risk of recurrence of EB.

Development and successful clinical application of preimplantation genetic haplotyping for Herlitz junctional epidermolysis bullosa

Backgroundu2002 Herlitz junctional epidermolysis bullosa (HJEB) is a severe, life‐threatening, autosomal recessive blistering skin disease for which no cure is currently available. Prenatal diagnosis for couples at risk is feasible through fetal skin biopsy or analysis of DNA extracted from chorionic villi, but these methods can be applied only after pregnancy has been established. An alternative approach, which involves the analysis of single cells from embryos prior to establishment of pregnancy, is preimplantation genetic diagnosis (PGD). Until now, its clinical uptake has been hindered by lengthy delays in establishing mutation‐specific protocols, and by the small amount of template DNA that can be obtained from a single cell. A new method that addresses these problems, preimplantation genetic haplotyping (PGH), relies on whole genome amplification followed by haplotyping of multiple polymorphic markers using standard DNA‐based polymerase chain reaction (PCR) assays.

Oral 5, Expanding choice for prenatal testing in couples at reproductive risk of Herlitz junctional epidermolysis bullosa

Following the early demise of their first child with Herlitz junctional epidermolysis bullosa (HJEB) a couple wished to consider prenatal testing. An immediate option would be fetal skin biopsy (FSB), a technique available for over 25 years, based on transmission electron microscopy and immunohistochemical analysis of laminin-332 expression, looking for similar changes to those in the deceased childs skin. However, this test could only be conducted after 15 weeks’ gestation and was not acceptable to this family. Mutation analysis was therefore performed on the laminin-332 genes by heteroduplex analysis and direct nucleotide sequencing and the deceased child was shown to be a compound heterozygote for the LAMB3 nonsense mutation p.R635X (maternal) and splice site mutation c.3228GA (paternal). With this information, chorionic villus sampling (CVS) with DNA sequencing could be conducted at an earlier gestation (10–11 weeks). However, both FSB and CVS involve termination of an affected pregnancy. An alternative option is preimplantation genetic diagnosis (PGD), based on DNA analysis of single blastomeres extracted from late cleavage stage embryos following in vitro fertilization. We have therefore developed a novel generic PGD test for HJEB based on whole genome amplification of DNA from single blastomeres, followed by multiplex polymerase chain reactions analysing 12 linkage markers within and flanking the LAMB3 gene. This assay is now licensed for clinical use in the U.K. and its suitability for this couple is being assessed. The development of PGD broadens personal choice for couples at reproductive risk of HJEB and represents a useful translational advance from basic research into epidermolysis bullosa.