Non-invasive prenatal diagnosis (NIPD) of cystic fibrosis: an optimized protocol using MEMO fluorescent PCR to detect the p.Phe508del mutation.

BACKGROUND: Analysis of cell-free foetal DNA (cff-DNA) in maternal plasma is very promising for early diagnosis of monogenic diseases; in particular, cystic fibrosis (CF). However, NIPD of single-gene disorders has been limited by the availability of suitable technical platforms and the need to set up patient or disease-specific custom-made approaches. METHODS: To make research applications more readily accessible to the clinic, we offer a simple assay combining two independent methods to determine the presence or absence of paternally inherited foetal allele p.Phe508del (the most frequent mutation in CF patients worldwide). The first method detects the presence or absence of a p.Phe508del allele by Mutant Enrichment with 3'-Modified Oligonucleotide PCR coupled to Fragment Length Analysis (MEMO-PCR-FLA). The second method detects the p.Phe508del allele with classical Multiplex Fluorescent PCR including five intragenic and extragenic STR markers of the CFTR locus and a specific SRY sequence. RESULTS: We collected 24 plasma samples from 23 women carrying foetuses at risk for CF and tested each sample using both methods. Our new procedures were successfully applied to 10 couples where fathers carried the p.Phe508del mutation and mothers were carrying a different mutation in the CFTR gene. These simple tests provided clear positive or negative results from the maternal plasma of the pregnant women. We confirmed the presence of cff-DNA in the studied samples by the identification of a tri-allelic DNA profile using a miniSTR kit. All results were correlated with chorionic villus sampling or amniocentesis analyses. CONCLUSIONS: This NIPD approach, easily set up in any clinical laboratory where prenatal diagnosis is routinely performed, offers many advantages over current methods: it is simple, rapid, and cost-effective. It opens up the possibility for testing a large number of couples with offspring at risk for CF.

Extreme oral manifestations in a Marfan-type syndrome.

A 12-year-old girl with an otherwise typical Marfan syndrome (Ghent criteria fulfilled) presented with highly unusual oral manifestations consisting of supernumerary teeth and severe dental crowding. Pathological examination of the supernumerary teeth revealed an elevated number of pulpoliths. No mutation in the FBN1, TGFBR1 and TGFBR2 genes was identified despite exhaustive screening, suggesting that another gene defect could explain this association of marfanoid features with dental abnormalities.

Mazabraud syndrome in two patients: clinical overlap with McCune-Albright syndrome.

Mazabraud syndrome is a rare sporadic disorder, mainly characterized by bone fibrous dysplasia and intramuscular myxomas. We report here two new cases of Mazabraud syndrome. One of our patients (Patient 1) also had café-au-lait spots and multinodular goiter suggestive of McCune-Albright syndrome. We review the 37 previously reported cases with Mazabraud syndrome and discuss the 6/37 patients with criteria of Mazabraud and McCune-Albright syndromes. Based on the clinical overlap between the two syndromes, we tested the GNAS1 gene in blood leukocytes and skin fibroblasts of Patient 1, but found no evidence of an activating mutation in the GNAS1 gene.