Jean-Michel Dupont

Loss-of-function mutations in FGFR1 cause autosomal dominant Kallmann syndrome

We took advantage of overlapping interstitial deletions at chromosome 8p11–p12 in two individuals with contiguous gene syndromes and defined an interval of roughly 540 kb associated with a dominant form of Kallmann syndrome, KAL2. We establish here that loss-of-function mutations in FGFR1 underlie KAL2 whereas a gain-of-function mutation in FGFR1 has been shown to cause a form of craniosynostosis. Moreover, we suggest that the KAL1 gene product, the extracellular matrix protein anosmin-1, is involved in FGF signaling and propose that the gender difference in anosmin-1 dosage (because KAL1 partially escapes X inactivation) explains the higher prevalence of the disease in males.

Biallelic mutations in the prokineticin-2 gene in two sporadic cases of Kallmann syndrome

Kallmann syndrome is a developmental disease that combines hypogonadotropic hypogonadism and anosmia. Putative loss-of-function mutations in PROKR2 or PROK2, encoding prokineticin receptor-2 (a G protein-coupled receptor), and one of its ligands, prokineticin-2, respectively, have recently been reported in approximately 10% of Kallmann syndrome affected individuals. Notably, given PROKR2 mutations were found in the heterozygous, homozygous, or compound heterozygous state in patients, thus raising the question of a possible digenic inheritance of the disease in heterozygous patients. Indeed, one of these patients was also carrying a missense mutation in KAL1, the gene responsible for the X chromosome-linked form of Kallmann syndrome. Mutations in PROK2, however, have so far been found only in the heterozygous state. Here, we report on the identification of PROK2 biallelic mutations, that is, a missense mutation, p.R73C, and a frameshift mutation, c.163delA, in two out of 273 patients presenting as sporadic cases. We conclude that PROK2 mutations in the homozygous state account for a few cases of Kallmann syndrome. Moreover, since the same R73C mutation was previously reported in the heterozygous state, and because Prok2 knockout mice exhibit an abnormal phenotype only in the homozygous condition, we predict that patients carrying monoallelic mutations in PROK2 have another disease-causing mutation, presumably in still undiscovered Kallmann syndrome genes.

Non-random, individual-specific methylation profiles are present at the sixth CTCF binding site in the human H19/IGF2 imprinting control region

Expression of imprinted genes is classically associated with differential methylation of specific CpG-rich DNA regions (DMRs). The H19/IGF2 locus is considered a paradigm for epigenetic regulation. In mice, as in humans, the essential H19 DMR—target of the CTCF insulator—is located between the two genes. Here, we performed a pyrosequencing-based quantitative analysis of its CpG methylation in normal human tissues. The quantitative analysis of the methylation level in the H19 DMR revealed three unexpected discrete, individual-specific methylation states. This epigenetic polymorphism was confined to the sixth CTCF binding site while a unique median-methylated profile was found at the third CTCF binding site as well as in the H19 promoter. Monoallelic expression of H19 and IGF2 was maintained independently of the methylation status at the sixth CTCF binding site and the IGF2 DMR2 displayed a median-methylated profile in all individuals and tissues analyzed. Interestingly, the methylation profile was genetically transmitted. Transgenerational inheritance of the H19 methylation profile was compatible with a simple model involving one gene with three alleles. The existence of three individual-specific epigenotypes in the H19 DMR in a non-pathological situation means it is important to reconsider the diagnostic value and functional importance of the sixth CTCF binding site.

Fluorescence in situ hybridization on methylcellulose cultured hematopoietic stem cells from myelodysplastic syndromes

Myelodysplastic syndromes (MDS) are clonal malignancies characterized by peripheral blood pancytopenia and signs of maturation disturbances of one or several cell lineages in bone marrow. MDS present as chimeras associating normal polyclonal and malignant monoclonal progenitors cells in various proportions. Numerous cytogenetic abnormalities have been reported in MDS and can be detected by fluorescence in situ hybridization (FISH) on interphase cells. We have used this technique on methylcellulose cultured hematopoietic progenitors obtained from three patients suffering from MDS and exhibiting informative karyotypic features. Hematopoietic cells were cultured for 14 days, and individual clones (BFU-E, CFU-GM) were picked up and then cytocentrifuged for FISH analysis. We used centromeric probes realized and labeled in our laboratory by PCR to detect aneuploidies for chromosomes 7 and 11 in two patients. Furthermore, we could detect a 5q partial deletion on interphase cells from the third patient using a 5q31 specific probe visualized with the HNPP Fluorescent Detection Set from Boehringer Mannheim. In conclusion, FISH is a helpful method to detect malignant clones in hematopoietic progenitor cultures and hence to study the relative growth of normal vs. leukemic cells in MDS.

Maternal serum screening in cases of mosaic and translocation Down syndrome

To determine if the second‐trimester maternal serum markers (MSM) screening for Down syndrome (DS) is efficient in DS mosaicism or structural rearrangement cases.

Sonographic Measurement of the Fetal Iliac Angle Cannot Be Used Alone as a Marker for Trisomy 21

The fetal iliac wings angle was studied in 255 fetuses before amniocentesis at 16.7 weeks (± 1.3), using a sonographic axial view of the fetal pelvis. The measurement could be performed in 208 fetuses (81.6%), of whom 4 had trisomy 21 (T 21). The mean iliac angle was greater in fetuses with T 21 than in normal fetuses (69.8° vs. 88.7°; p = 0.03). This measurement is subject to significant intra- and interexaminer variability (interclass correlation coefficient: 0.65 and 0.23, respectively). When a 90° value is used as a threshold, specificity, sensitivity, positive and negative predictive values are, respectively, 80, 75, 7.0 and 99.4%. The 20% rate of false-positives rules out the use of this measurement as the sole criterion for the indication of amniocentesis for T 21 antenatal diagnosis.

Optimized criteria for using fluorescence in situ hybridization in the prenatal diagnosis of common aneuploidies.

To evaluate the medical and economic performance of three strategies for selecting patients eligible for interphase FISH in the prenatal diagnosis of common aneuploidies.

Mosaic Isochromosome 20q and Normal Outcome: A New Case Ascertained by Fluorescence in situ Hybridization and a Review of the Literature

We describe a new case of mosaic isochromosome 20q revealed by amniocentesis. A 46,XX/46,XX,i(20q) chromosomic complement was indirectly confirmed by fluorescent in situ hybridization. Since control chromosome analysis performed on cord blood showed a normal karyotype, pregnancy was continued and resulted in the birth of a normal female infant.

aCGH may be useful in Case of Dominant Disorders Known to be Caused by Gene Mutations: Two Case Reports

We describe two cases where clinical diagnoses were carried out (Rett syndrome-like and Nicolaides-Baraitser syndrome) with no identified mutation respectively in the CDKL5 and the SMARCA2 genes. Conversely a chromosomal microdeletion with contiguous deletion of a part of these two genes was found by arrayCGH in each corresponding case. The aim of this report is then to highlight the possible implication of chromosomal microdeletions with contiguous gene deletions in dominant pathologies where no mutation is found in the causative gene.

Unusual isochromosome 5p marker chromosome.

Unusual Isochromosome 5p Marker Chromosome Fanny Roulet-Coudrier, Amine Rouibi, Clotilde Thuillier, Sylvie Bourthoumieu, Aziza Lebbar, Jean-Michel Dupont, and Catherine Yardin* Service d’Histologie, Cytologie, Biologie Cellulaire et Cytogénétique, Hôpital de la Mère et de l’Enfant (HME), CHU (Centre Hospitalier Universitaire) Dupuytren, Limoges, France Service de Pédiatrie, Hôpital de la Mère et de l’Enfant (HME), CHU (Centre Hospitalier Universitaire) Dupuytren, Limoges, France CHREC (Comité de Recherche Clinique du HME), CHU (Centre Hospitalier Universitaire) Dupuytren, Limoges, France Service de Cytogénétique, Groupe Hospitalier Cochin-Saint Vincent de Paul, APHP (Assistance Publique Hôpitaux de Paris), Paris, France