Paulo A. Otto

Fragile X Premutation Is a Significant Risk Factor for Premature Ovarian Failure: The International Collaborative POF in Fragile X Study—Preliminary Data

The preliminary results of an international collaborative study examining premature menopause in fragile X carriers are presented. A total of 760 women from fragile X families was surveyed about their fragile X carrier status and their menstrual and reproductive histories. Among the subjects, 395 carried a premutation, 128 carried a full mutation, and 237 were noncarriers. Sixty-three (16%) of the premutation carriers had experienced menopause prior to the age of 40 compared with none of the full mutation carriers and one (0.4%) of the controls. Based on these preliminary data, there is a significant association between fragile X premutation carrier status and premature menopause.

Array-CGH detection of micro rearrangements in mentally retarded individuals: Clinical significance of imbalances present both in affected children and normal parents

Background: The underlying causes of mental retardation remain unknown in about half the cases. Recent array-CGH studies demonstrated cryptic imbalances in about 25% of patients previously thought to be chromosomally normal. Objective and methods: Array-CGH with approximately 3500 large insert clones spaced at ∼1 Mb intervals was used to investigate DNA copy number changes in 81 mentally impaired individuals. Results: Imbalances never observed in control chromosomes were detected in 20 patients (25%): seven were de novo, nine were inherited, and four could not have their origin determined. Six other alterations detected by array were disregarded because they were shown by FISH either to hybridise to both homologues similarly in a presumptive deletion (one case) or to involve clones that hybridised to multiple sites (five cases). All de novo imbalances were assumed to be causally related to the abnormal phenotypes. Among the others, a causal relation between the rearrangements and an aberrant phenotype could be inferred in six cases, including two imbalances of the X chromosome, where the associated clinical features segregated as X linked recessive traits. Conclusions: In all, 13 of 81 patients (16%) were found to have chromosomal imbalances probably related to their clinical features. The clinical significance of the seven remaining imbalances remains unclear. The limited ability to differentiate between inherited copy number variations which cause abnormal phenotypes and rare variants unrelated to clinical alterations currently constitutes a limitation in the use of CGH-microarray for guiding genetic counselling.

Whole-genome array-CGH screening in undiagnosed syndromic patients: old syndromes revisited and new alterations

We report array-CGH screening of 95 syndromic patients with normal G-banded karyotypes and at least one of the following features: mental retardation, heart defects, deafness, obesity, craniofacial dysmorphisms or urogenital tract malformations. Chromosome imbalances not previously detected in normal controls were found in 30 patients (31%) and at least 16 of them (17%) seem to be causally related to the abnormal phenotypes. Eight of the causative imbalances had not been described previously and pointed to new chromosome regions and candidate genes for specific phenotypes, including a connective tissue disease locus on 2p16.3, another for obesity on 7q22.1→q22.3, and a candidate gene for the 3q29 deletion syndrome manifestations. The other causative alterations had already been associated with well-defined phenotypes including Sotos syndrome, and the 1p36 and 22q11.21 microdeletion syndromes. However, the clinical features of these latter patients were either not typical or specific enough to allow diagnosis before detection of chromosome imbalances. For instance, three patients with overlapping deletions in 22q11.21 were ascertained through entirely different clinical features, i.e., heart defect, utero-vaginal aplasia, and mental retardation associated with psychotic disease. Our results demonstrate that ascertainment through whole-genome screening of syndromic patients by array-CGH leads not only to the description of new syndromes, but also to the recognition of a broader spectrum of features for already described syndromes. Furthermore, on the technical side, we have significantly reduced the amount of reagents used and costs involved in the array-CGH protocol, without evident reduction in efficiency, bringing the method more within reach of centers with limited budgets.

Genomic imbalances associated with müllerian aplasia

Background: Aplasia of the müllerian ducts leads to absence of the uterine corpus, uterine cervix, and upper (superior) vagina. Patients with müllerian aplasia (MA) often exhibit additional clinical features such as renal, vertebral and cardiac defects. A number of different syndromes have been associated with MA, and in most cases its aetiology remains poorly understood. Objective and methods: 14 syndromic patients with MA and 46,XX G-banded karyotype were screened for DNA copy number changes by ∼1 Mb whole genome bacterial artificial chromosome (BAC) array based comparative genomic hybridisation (CGH). The detected alterations were validated by an independent method and further mapped by high resolution oligo-arrays. Results: Submicroscopic genomic imbalances affecting the 1q21.1, 17q12, 22q11.21, and Xq21.31 chromosome regions were detected in four probands. Presence of the alterations in the normal mother of one patient suggests incomplete penetrance and/or variable expressivity. Conclusion: 4 of the 14 patients (29%) were found to have cryptic genomic alterations. The imbalances on 22q11.21 support recent findings by us and others that alterations in this chromosome region may result in impairment of müllerian duct development. The remaining imbalances indicate involvement of previously unknown chromosome regions in MA, and point specifically to LHX1 and KLHL4 as candidate genes.

Phenotypic variability in Angelman syndrome: comparison among different deletion classes and between deletion and UPD subjects

Angelman syndrome (AS) can result from either a 15q11–q13 deletion (del), paternal uniparental disomy (UPD), imprinting, or UBE3A mutations. Here, we describe the phenotypic and behavioral variability detected in 49 patients with different classes of deletions and nine patients with UPD. Diagnosis was made by methylation pattern analysis of exon 1 of the SNRPN-SNURF gene and by microsatellite profiling of loci within and outside the 15q11–q13 region. There were no major phenotypic differences between the two main classes (BP1–BP3; BP2–BP3) of AS deletion patients, except for the absence of vocalization, more prevalent in patients with BP1–BP3 deletions, and for the age of sitting without support, which was lower in patients with BP2–BP3 deletions. Our data suggest that gene deletions (NIPA1, NIPA2, CYF1P1, GCP5) mapped to the region between breakpoints BP1 and BP2 may be involved in the severity of speech impairment, since all BP1–BP3 deletion patients showed complete absence of vocalization, while 38.1% of the BP2–BP3 deletion patients were able to pronounce syllabic sounds, with doubtful meaning. Compared to UPD patients, deletion patients presented a higher incidence of swallowing disorders (73.9% del × 22.2% UPD) and hypotonia (73.3% del × 28.57% UPD). In addition, children with UPD showed better physical growth, fewer or no seizures, a lower incidence of microcephaly, less ataxia and higher cognitive skills. As a consequence of their milder or less typical phenotype, AS may remain undiagnosed, leading to an overall underdiagnosis of the disease.

Analysis of a novel functional polymorphism within the promoter region of the serotonin transporter gene (5-HTT) in Brazilian patients affected by bipolar disorder and schizophrenia.

It has been suggested that the serotonin transporter (5-hydroxytryptamine-transporter or 5-HTT) may be involved in the pathogenesis of affective disorders. Recently, Collier et al. (1996) found that the frequency of the low-activity short variant (s) of the 5-HTT-linked polymorphic region (5-HTTLPR) was higher among patients with affective disorders than in normal controls. However, since the observed level of significance was not high, they suggest that these findings should be replicated in independent samples. We have analyzed 86 unrelated patients (47 with bipolar disorder and 39 with schizophrenia) and 98 normal controls from the Brazilian population for the 5-HTTLPR. Statistical analysis revealed that the genotypes (LL, Ls, ss) as well as the estimated allele frequencies (L,s) did not differ significantly among the three studied groups or between bipolar and normal controls. In addition, although not statistically significant, the genotype ss in our sample was less frequent among our bipolar patients than in our normal controls (12.8% versus 16.3%) which is the opposite of what was found by Collier et al. (24% versus 18%) in the European study. Although it will be important to extend the present analysis in a larger sample, our preliminary results suggest that the 5-HTTLPR does not seem to play a major role in the genetics of bipolar and schizophrenic disorders at least in this group of Brazilian psychiatric patients.

The short variant of the polymorphism within the promoter region of the serotonin transporter gene is a risk factor for late onset Alzheimer's disease

We analyzed a deletion/insertion polymorphism within the promoter region of the serotonin transporter gene (5-HTTPLR) in 81 patients with late onset Alzheimers (AD) disease (mean age 70.02 ± 8.13 years). Control groups included 81 normal subjects with comparable age (mean age 75.6 ± 10.2) and 82 younger normal subjects (mean age 37.4 ± 9.1). Statistical analysis showed a significant difference in the genotype and gene frequencies between the AD group and normal controls (χ2 = 9.021; 2 d.f. and χ2 = 5.59, 1 d.f., respectively, P<0.05) due to the higher frequency of the l allele and the lower frequency of the s allele in controls than among ad patients. however, no differences were found in the genotype frequencies in older as compared to younger normal control groups (χ2 = 0.337, 2 d.f. and P>0.05). The present study confirms, in a different population, that the short variant of the 5-HTTPLR polymorphism may be a risk factor for late onset AD.

Prevalence of GJB2 (connexin-26) and GJB6 (connexin-30) mutations in a cohort of 300 Brazilian hearing-impaired individuals: implications for diagnosis and genetic counseling.

Objective: Hereditary nonsyndromic deafness is an autosomal recessive condition in about 80% of cases, and point mutations in the GJB2 gene (connexin 26) and two deletions in the GJB6 gene (connexin 30), del(GJB6-D13S1830) and del(GJB6-D13S1854), are reported to account for 50% of recessive deafness. Aiming at establishing the frequencies of GJB2 mutations and GJB6 deletions in the Brazilian population, we screened 300 unrelated individuals with hearing impairment, who were not affected by known deafness related syndromes. Methods: We firstly screened the most frequently reported mutations, c.35delG and c.167delT in the GJB2 gene, and del(GJB6-D13S1830) and del(GJB6-D13S1854) in the GJB6 gene, through specific techniques. The detected c.35delG and c.167delT mutations were validated by sequencing. Other mutations in the GJB2 gene were screened by single-strand conformation polymorphism and the coding region was sequenced when abnormal patterns were found. Results: Pathogenic mutations in GJB2 and GJB6 genes were detected in 41 individuals (13.7%), and 80.5% (33/41) presented these mutations in homozygosis or compound heterozygosis, thus explaining their hearing defect. The c.35delG in the GJB2 gene was the most frequent mutation (37/300; 12.4%), detected in 23% familial and 6.2% the sporadic cases. The second most frequent mutation (1%; 3/300) was the del(GJB6-D13S1830), always found associated with the c.35delG mutation. Nineteen different sequence variations were found in the GJB2 gene. In addition to the c.35delG mutation, nine known pathogenic alterations were detected c.167delT, p.Trp24X, p.Val37Ile, c.176_191del16, c.235delC, p.Leu90Pro, p.Arg127His, c.509insA, and p.Arg184Pro. Five substitutions had been previously considered benign polymorphisms: c.-15C>T, p.Val27Ile, p.Met34Thr, p.Ala40Ala, and p.Gly160Ser. Two previously reported mutations of unknown pathogenicity were found (p.Lys168Arg, and c.684C>A), and two novel substitutions, p.Leu81Val (c.G241C) and p.Met195Val (c.A583G), both in heterozygosis without an accompanying mutation in the other allele. None of these latter four variants of undefined status was present in a sample of 100 hearing controls. Conclusions: The present study demonstrates that mutations in the GJB2 gene and del(GJB6 D13S1830) are important causes of hearing impairment in Brazil, thus justifying their screening in a routine basis. The diversity of variants in our sample reflects the ethnic heterogeneity of the Brazilian population.

Report of a del22q11 in a patient with Mayer-Rokitansky-Küster-Hauser (MRKH) anomaly and exclusion of WNT-4, RAR-gamma, and RXR-alpha as major genes determining MRKH anomaly in a study of 25 affected women.

How to cite this article: Cheroki C, Krepischi-Santos AC, Rosenberg C, Jehee FS, Mingroni-Netto RC, Filho IP,Filho SZ, Kim CA, Bagnoli VR, Mendonc¸a BB, Szuhai K, Otto PA. 2006. Report of a del22q11 in a patientwith Mayer-Rokitansky-Ku¨ster-Hauser (MRKH) anomaly and exclusion of WNT-4, RAR-gamma, andRXR-alpha as major genes determining MRKH anomaly in a study of 25 affected women.Am J Med Genet Part A 140A:1339–1342.

Clinical characterization of autosomal dominant and recessive variants of Robinow syndrome

Robinow syndrome is a genetically heterogeneous condition characterized by mesomelic limb shortening associated with facial and genital anomalies that can be inherited in an autosomal dominant or recessive mode. We characterized these two variants clinically, with the aim of establishing clinical criteria to enhance the differential diagnosis between them or other similar conditions. The frequencies of clinical signs considered important for the discrimination of the dominant or recessive variants were estimated in a sample consisting of 38 patients personally examined by the authors and of 50 affected subjects from the literature. Using the presence of rib fusions as diagnostic of the recessive variant, and also based on the inheritance pattern in familial cases, we classified 37 patients as having the recessive form and other 51 as having the dominant form. The clinical signs present in more than 75% of patients with either form, and therefore the most important for the characterization of this syndrome were hypertelorism, nasal features (large nasal bridge, short upturned nose, and anteverted nares), midface hypoplasia, mesomelic limb shortening, brachydactyly, clinodactyly, micropenis, and short stature. Hemivertebrae and scoliosis were present in more than 75% of patients with the recessive form, but in less than 25% of patients with the dominant form. Umbilical hernia (32.3%) and supernumerary teeth (10.3%) were found exclusively in patients with the dominant form.