Philippe Jonveaux

A new highly penetrant form of obesity due to deletions on chromosome 16p11.2

Obesity has become a major worldwide challenge to public health, owing to an interaction between the Western ‘obesogenic’ environment and a strong genetic contribution. Recent extensive genome-wide association studies (GWASs) have identified numerous single nucleotide polymorphisms associated with obesity, but these loci together account for only a small fraction of the known heritable component. Thus, the ‘common disease, common variant’ hypothesis is increasingly coming under challenge. Here we report a highly penetrant form of obesity, initially observed in 31 subjects who were heterozygous for deletions of at least 593 kilobases at 16p11.2 and whose ascertainment included cognitive deficits. Nineteen similar deletions were identified from GWAS data in 16,053 individuals from eight European cohorts. These deletions were absent from healthy non-obese controls and accounted for 0.7% of our morbid obesity cases (body mass index (BMI) ≥ 40 kg m-2 or BMI standard deviation score ≥ 4; P = 6.4 × 10-8, odds ratio 43.0), demonstrating the potential importance in common disease of rare variants with strong effects. This highlights a promising strategy for identifying missing heritability in obesity and other complex traits: cohorts with extreme phenotypes are likely to be enriched for rare variants, thereby improving power for their discovery. Subsequent analysis of the loci so identified may well reveal additional rare variants that further contribute to the missing heritability, as recently reported for SIM1 (ref. 3). The most productive approach may therefore be to combine the ‘power of the extreme’ in small, well-phenotyped cohorts, with targeted follow-up in case-control and population cohorts.

Key clinical features to identify girls with CDKL5 mutations

Mutations in the human X-linked cyclin-dependent kinase-like 5 (CDKL5) gene have been shown to cause infantile spasms as well as Rett syndrome (RTT)-like phenotype. To date, less than 25 different mutations have been reported. So far, there are still little data on the key clinical diagnosis criteria and on the natural history of CDKL5-associated encephalopathy. We screened the entire coding region of CDKL5 for mutations in 183 females with encephalopathy with early seizures by denaturing high liquid performance chromatography and direct sequencing, and we identified in 20 unrelated girls, 18 different mutations including 7 novel mutations. These mutations were identified in eight patients with encephalopathy with RTT-like features, five with infantile spasms and seven with encephalopathy with refractory epilepsy. Early epilepsy with normal interictal EEG and severe hypotonia are the key clinical features in identifying patients likely to have CDKL5 mutations. Our study also indicates that these patients clearly exhibit some RTT features such as deceleration of head growth, stereotypies and hand apraxia and that these RTT features become more evident in older and ambulatory patients. However, some RTT signs are clearly absent such as the so called RTT disease profile (period of nearly normal development followed by regression with loss of acquired fine finger skill in early childhood and characteristic intensive eye communication) and the characteristic evolution of the RTT electroencephalogram. Interestingly, in addition to the overall stereotypical symptomatology (age of onset and evolution of the disease) resulting from CDKL5 mutations, atypical forms of CDKL5-related conditions have also been observed. Our data suggest that phenotypic heterogeneity does not correlate with the nature or the position of the mutations or with the pattern of X-chromosome inactivation, but most probably with the functional transcriptional and/or translational consequences of CDKL5 mutations. In conclusion, our report show that search for mutations in CDKL5 is indicated in girls with early onset of a severe intractable seizure disorder or infantile spasms with severe hypotonia, and in girls with RTT-like phenotype and early onset seizures, though, in our cohort, mutations in CDKL5 account for about 10% of the girls affected by these disorders.

Mutations of the P53 gene in acute myeloid leukaemia

Summary. In a previous report we found point mutations in exons 5–8 of the P53 gene in five of 46 patients with acute myeloid leukaemia (AML), with a predominance of mutations in the 10 patients with 17p monosomy. In this report we extended our findings studying such mutations in 66 unselected additional cases of AML, using polymerase chain reaction single strand conformation polymorphism (SSCP) analysis and nucleotide sequencing. Three of the 66 new cases had a point mutation, leading to a change in one encoded amino acid. Thus, eight of the 112 AML studied had P53 mutations in exons 5–8, suggesting that the incidence of P53 mutation is relatively low in AML. A predominance of mutations in exon 8 (5/8) was found. Six of the eight patients with mutations were older than 60 years of age, and all eight cases had a short survival. All seven mutated cases karyotyped showed complex cytogenetic findings, especially monosomy 5 and/or 7, thus questioning the pathogenic importance of P53 mutations in a context of multiple genetic abnormalities. However, five of them also had 17p monosomy, and in the remaining two cases SSCP and sequence analysis also suggested loss of the normal P53 allele. This supported a role for the P53 gene mutations in leukaemogenesis in the relatively small number of AML patients in whom they were found, through loss of tumour suppressive activity of both normal P53 alleles, as reported in solid tumours.

Delineation of 15q13.3 microdeletions

Masurel‐Paulet A, Andrieux J, Callier P, Cuisset JM, Le Caignec C, Holder M, Thauvin‐Robinet C, Doray B, Flori E, Alex‐Cordier MP, Beri M, Boute O, Delobel B, Dieux A, Vallee L, Jaillard S, Odent S, Isidor B, Beneteau C, Vigneron J, Bilan F, Gilbert‐Dussardier B, Dubourg C, Labalme A, Gautier A, Pernes P, Bidon C, Pinoit JM, Huet F, Mugneret F, Aral B, Jonveaux P, Sanlaville D, Faivre L. Delineation of 15q13.3 microdeletions.

A retrospective study by oligonucleotide array‐CGH analysis in 50 fetuses with multiple malformations

To retrospectively define the frequency and the nature of submicroscopic chromosomal imbalances among fetuses with multiple congenital anomalies (MCA).

Mutational spectrum of CDKL5 in early-onset encephalopathies: a study of a large collection of French patients and review of the literature.

The CDKL5 gene has been implicated in the molecular etiology of early‐onset intractable seizures with infantile spasms (IS), severe hypotonia and atypical Rett syndrome (RTT) features. So far, 48 deleterious alleles have been reported in the literature. We screened the CDKL5 gene in a cohort of 177 patients with early‐onset seizures, including 30 men and 10 girls with Aicardi syndrome. The screening was negative for all men as well as for women with Aicardi syndrome, excluding the CDKL5 gene as a candidate for this neurodevelopmental disorder. We report 11 additional de novo mutations in CDKL5 in female patients. For the first time, the MLPA approach allowed the identification of a partial deletion encompassing the promoter and the first two exons of CDKL5. The 10‐point mutations consist of five missenses (with recurrent amino acid changes at p.Ala40 and p.Arg178), four splicing variants and a 1‐base pair duplication. We present a review of all mutated alleles published in the literature. In our study, the overall frequency of mutations in CDKL5 in women with early‐onset seizures is around 8.6%, a result comparable with previous reports. Noteworthy, the CDKL5 mutation rate is high (28%) in women with early‐onset seizures and IS.

Clinical phenotype of germline RUNX1 haploinsufficiency: from point mutations to large genomic deletions

Germline RUNX1 mutations result in a rare autosomal dominant condition characterized by qualitative and quantitative platelet defects and predisposition to the development of myeloid malignancies (familial platelet disorder with propensity to acute myeloid leukaemia, FPD/AML). Only 13 pedigrees have previously been described so far. We report on two novel germline RUNX1 mutations: (1) an out-of-frame 8 bp heterozygous deletion (c.442_449del) in an FPD/AML pedigree and (2) a de novo 3.5 Mb deletion in the 21q22.11.21q22.12 region encompassing the RUNX1 gene in a mentally retarded female patient with short stature and thrombocytopenia. Interestingly, a similar de novo submicroscopic deletion has been recently reported in the literature in a mentally retarded patient. Mental retardation is one of the most common disorders and primary causes of thrombocytopenia are rare. When occurring together, these features should prompt to test for 21q22 deletion for comprehensive genetic counselling and clinical management.

Phenotypic variability in Rett syndrome associated with FOXG1 mutations in females

Background The FOXG1 gene has been recently implicated in the congenital form of Rett syndrome (RTT). It encodes the fork-head box protein G1, a winged-helix transcriptional repressor with expression restricted to testis and brain, where it is critical for forebrain development. So far, only two point mutations in FOXG1 have been reported in females affected by the congenital form of RTT. Aim To assess the involvement of FOXG1 in the molecular aetiology of classical RTT and related disorders. Methods The entire multi-exon coding sequence of FOXG1 was screened for point mutations and large rearrangements in a cohort of 35 MECP2/CDKL5 mutation-negative female patients including 31 classical and four congenital forms of RTT. Results Two different de novo heterozygous FOXG1-truncating mutations were identified. The subject with the p.Trp308X mutation presented with a severe RTT-like neurodevelopmental disorder, whereas the p.Tyr400X allele was associated with a classical clinical RTT presentation. Conclusions These new cases give additional support to the genetic heterogeneity in RTT and help to delineate the clinical spectrum of the FOXG1-related phenotypes. FOXG1 screening should be considered in the molecular diagnosis of RTT.

Complex Segmental Duplications Mediate a Recurrent dup(X)(p11.22-p11.23) Associated with Mental Retardation, Speech Delay, and EEG Anomalies in Males and Females

Submicroscopic copy-number variations make a considerable contribution to the genetic etiology of human disease. We have analyzed subjects with idiopathic mental retardation (MR) by using whole-genome oligonucleotide-based array comparative genomic hybridization (aCGH) and identified familial and de novo recurrent Xp11.22-p11.23 duplications in males and females with MR, speech delay, and a peculiar electroencephalographic (EEG) pattern in childhood. The size of the duplications ranges from 0.8-9.2 Mb. Most affected females show preferential activation of the duplicated X chromosome. Carriers of the smallest duplication show X-linked recessive inheritance. All other affected individuals present dominant expression and comparable clinical phenotypes irrespective of sex, duplication size, and X-inactivation pattern. The majority of the rearrangements are mediated by recombination between flanking complex segmental duplications. The identification of common clinical features, including the typical EEG pattern, predisposing genomic structure, and peculiar X-inactivation pattern, suggests that duplication of Xp11.22-p11.23 constitutes a previously undescribed syndrome.

Microdeletion at chromosome 4q21 defines a new emerging syndrome with marked growth restriction, mental retardation and absent or severely delayed speech

Background Genome-wide screening of large patient cohorts with mental retardation using microarray-based comparative genomic hybridisation (array-CGH) has recently led to identification several novel microdeletion and microduplication syndromes. Methods Owing to the national array-CGH network funded by the French Ministry of Health, shared information about patients with rare disease helped to define critical intervals and evaluate their gene content, and finally determine the phenotypic consequences of genomic array findings. Results In this study, nine unrelated patients with overlapping de novo interstitial microdeletions involving 4q21 are reported. Several major features are common to all patients, including neonatal muscular hypotonia, severe psychomotor retardation, marked progressive growth restriction, distinctive facial features and absent or severely delayed speech. The boundaries and the sizes of the nine deletions are different, but an overlapping region of 1.37 Mb is defined; this region contains five RefSeq genes: PRKG2, RASGEF1B, HNRNPD, HNRPDL and ENOPH1. Discussion Adding new individuals with similar clinical features and 4q21 deletion allowed us to reduce the critical genomic region encompassing two genes, PRKG2 and RASGEF1B. PRKG2 encodes cGMP-dependent protein kinase type II, which is expressed in brain and in cartilage. Information from genetically modified animal models is pertinent to the clinical phenotype. RASGEF1B is a guanine nucleotide exchange factor for Ras family proteins, and several members have been reported as key regulators of actin and microtubule dynamics during both dendrite and spine structural plasticity. Conclusion Clinical and molecular delineation of 4q21 deletion supports a novel microdeletion syndrome and suggests a major contribution of PRKG2 and RASGEF1B haploinsufficiency to the core phenotype.