Putative founder effect of Arg338* AP4M1 (SPG50) variant causing severe intellectual disability, epilepsy and spastic paraplegia: Report of three families.

Bi-allelic variants affecting one of the four genes encoding the AP4 subunits are responsible for the "AP4 deficiency syndrome." Core features include hypotonia that progresses to hypertonia and spastic paraplegia, intellectual disability, postnatal microcephaly, epilepsy, and neuroimaging features. Namely, AP4M1 (SPG50) is involved in autosomal recessive spastic paraplegia 50 (MIM#612936). We report on three patients with core features from three unrelated consanguineous families originating from the Middle East. Exome sequencing identified the same homozygous nonsense variant: NM_004722.4(AP4M1):c.1012C>T p.Arg338* (rs146262009). So far, four patients from three other families carrying this homozygous variant have been reported worldwide. We describe their phenotype and compare it to the phenotype of patients with other variants in AP4M1. We construct a shared single-nucleotide polymorphism (SNP) haplotype around AP4M1 in four families and suggest a probable founder effect of Arg338* AP4M1 variant with a common ancestor most likely of Turkish origin.

Risk factors associated with preterm birth in singletons born after IVF: a national cohort study.

RESEARCH QUESTION: What are the risk factors for prematurity other than intrauterine growth restriction in singletons after IVF? DESIGN: Data were collected from a national registry, based on an observational prospective cohort of 30,737 live births after assisted reproductive technology (fresh embryo transfers: n = 20,932 and frozen embryo transfer [FET] n = 9805) between 2014 and 2015. A population of not-small for gestational age singletons conceived after fresh embryo transfers and FET, and their parents, was selected. Data on a number of variables were collected, including type of infertility, number of oocytes retrieved and vanishing twins. RESULTS: Preterm birth occurred in 7.7% (n = 1607) of fresh embryo transfers and 6.2% (n = 611) of frozen-thawed embryo transfers (P < 0.0001; adjusted odds ratio [aOR] = 1.34 [1.21-1.49]). Endometriosis and vanishing twin increased the risk of preterm birth after fresh embryo transfer (P < 0.001; aOR 1.32 and 1.78, respectively). Polycystic ovaries or more than 20 oocytes retrieved also increased preterm birth risk (aOR 1.31 and 1.30; P = 0.003 and P = 0.02, respectively); large oocyte cohort (>20) was no longer associated with the risk of prematurity in FET. CONCLUSION: Endometriosis remains a risk for prematurity even in the absence of intrauterine growth retardation, which suggests a dysimmune effect. Large oocyte cohorts obtained by stimulation, without clinical polycystic ovary syndrome diagnosed before attempts, do not affect FET outcomes, reinforcing the idea of a phenotypic difference in the clinical presentation of polycystic ovary syndrome.

Fetal growth disorders following medically assisted reproduction: due to maternal context or techniques? A national French cohort study.

RESEARCH QUESTION: What part do maternal context and medically assisted reproduction (MAR) techniques play in the risk of fetal growth disorders? DESIGN: This retrospective nationwide cohort study uses data available in the French National Health System database and focuses on the period from 2013 to 2017. Fetal growth disorders were divided into four groups according to the origin of pregnancy: fresh embryo transfer (n = 45,201), frozen embryo transfer (FET, n = 18,845), intrauterine insemination (IUI, n = 20,179) and natural conceptions (n = 3,412,868). Fetal growth disorders were defined from the percentiles of the weight distribution according to gestational age and sex: small and large for gestational age (SGA and LGA) if <10th and >90th percentiles, respectively. Analyses were performed using univariate and multivariate logistic models. RESULTS: Compared with births following natural conception, multivariate analysis showed that the risk of SGA was higher for births following fresh embryo transfer and IUI (adjusted odds ratio [aOR] 1.26 [1.22-1.29] and 1.08 [1.03-1.12], respectively) and significantly lower following FET (aOR 0.79 [0.75-0.83]). The risk of LGA was higher for births following FET (aOR 1.32 [1.27-1.38]), especially in artificial cycles when compared with ovulatory cycles (aOR 1.25 [1.15-1.36]). In the subgroup of births without any obstetrical or neonatal morbidity, the same increased risk of SGA and LGA were observed following fresh embryo transfer or IUI and FET (aOR 1.23 [1.19-1.27] or 1.06 [1.01-1.11] and aOR 1.36 [1.30-1.43], respectively). CONCLUSIONS: An effect of MAR techniques on the risks for SGA and LGA is suggested independently from maternal context and obstetrical or neonatal morbidities. Pathophysiological mechanisms remain poorly understood and should be further evaluated, as well as the influence of embryonic stage and freezing techniques.

Genotype-first in a cohort of 95 fetuses with multiple congenital abnormalities: when exome sequencing reveals unexpected fetal phenotype-genotype correlations.

PURPOSE: Molecular diagnosis based on singleton exome sequencing (sES) is particularly challenging in fetuses with multiple congenital abnormalities (MCA). Indeed, some studies reveal a diagnostic yield of about 20%, far lower than in live birth individuals showing developmental abnormalities (30%), suggesting that standard analyses, based on the correlation between clinical hallmarks described in postnatal syndromic presentations and genotype, may underestimate the impact of the genetic variants identified in fetal analyses. METHODS: We performed sES in 95 fetuses with MCA. Blind to phenotype, we applied a genotype-first approach consisting of combined analyses based on variants annotation and bioinformatics predictions followed by reverse phenotyping. Initially applied to OMIM-morbid genes, analyses were then extended to all genes. We complemented our approach by using reverse phenotyping, variant segregation analysis, bibliographic search and data sharing in order to establish the clinical significance of the prioritised variants. RESULTS: sES rapidly identified causal variant in 24/95 fetuses (25%), variants of unknown significance in OMIM genes in 8/95 fetuses (8%) and six novel candidate genes in 6/95 fetuses (6%). CONCLUSIONS: This method, based on a genotype-first approach followed by reverse phenotyping, shed light on unexpected fetal phenotype-genotype correlations, emphasising the relevance of prenatal studies to reveal extreme clinical presentations associated with well-known Mendelian disorders.

Obstetrical outcomes and maternal morbidities associated with COVID-19 in pregnant women in France: A national retrospective cohort study.

BACKGROUND: To the best of our knowledge, no study has exhaustively evaluated the association between maternal morbidities and Coronavirus Disease 2019 (COVID-19) during the first wave of the pandemic in pregnant women. We investigated, in natural conceptions and assisted reproductive technique (ART) pregnancies, whether maternal morbidities were more frequent in pregnant women with COVID-19 diagnosis compared to pregnant women without COVID-19 diagnosis during the first wave of the COVID-19 pandemic. METHODS AND FINDINGS: We conducted a retrospective analysis of prospectively collected data in a national cohort of all hospitalizations for births ≥22 weeks of gestation in France from January to June 2020 using the French national hospitalization database (PMSI). Pregnant women with COVID-19 were identified if they had been recorded in the database using the ICD-10 (International Classification of Disease) code for presence of a hospitalization for COVID-19. A total of 244,645 births were included, of which 874 (0.36%) in the COVID-19 group. Maternal morbidities and adverse obstetrical outcomes among those with or without COVID-19 were analyzed with a multivariable logistic regression model adjusted on patient characteristics. Among pregnant women, older age (31.1 (±5.9) years old versus 30.5 (±5.4) years old, respectively, p < 0.001), obesity (0.7% versus 0.3%, respectively, p < 0.001), multiple pregnancy (0.7% versus 0.4%, respectively, p < 0.001), and history of hypertension (0.9% versus 0.3%, respectively, p < 0.001) were more frequent with COVID-19 diagnosis. Active smoking (0.2% versus 0.4%, respectively, p < 0.001) and primiparity (0.3% versus 0.4%, respectively, p < 0.03) were less frequent with COVID-19 diagnosis. Frequency of ART conception was not different between those with and without COVID-19 diagnosis (p = 0.28). When compared to the non-COVID-19 group, women in the COVID-19 group had a higher frequency of admission to ICU (5.9% versus 0.1%, p < 0.001), mortality (0.2% versus 0.005%, p < 0.001), preeclampsia/eclampsia (4.8% versus 2.2%, p < 0.001), gestational hypertension (2.3% versus 1.3%, p < 0.03), postpartum hemorrhage (10.0% versus 5.7%, p < 0.001), preterm birth at <37 weeks of gestation (16.7% versus 7.1%, p < 0.001), <32 weeks of gestation (2.2% versus 0.8%, p < 0.001), <28 weeks of gestation (2.4% versus 0.8%, p < 0.001), induced preterm birth (5.4% versus 1.4%, p < 0.001), spontaneous preterm birth (11.3% versus 5.7%, p < 0.001), fetal distress (33.0% versus 26.0%, p < 0.001), and cesarean section (33.0% versus 20.2%, p < 0.001). Rates of pregnancy terminations ≥22 weeks of gestation, stillbirths, gestational diabetes, placenta praevia, and placenta abruption were not significantly different between the COVID-19 and non-COVID-19 groups. The number of venous thromboembolic events was too low to perform statistical analysis. A limitation of this study relies in the possibility that asymptomatic infected women were not systematically detected. CONCLUSIONS: We observed an increased frequency of pregnant women with maternal morbidities and diagnosis of COVID-19 compared to pregnant women without COVID-19. It appears essential to be aware of this, notably in populations at known risk of developing a more severe form of infection or obstetrical morbidities and in order for obstetrical units to better inform pregnant women and provide the best care. Although causality cannot be determined from these associations, these results may be in line with recent recommendations in favor of vaccination for pregnant women.

Do in vitro fertilization, intrauterine insemination or female infertility impact the risk of congenital anomalies in singletons? A longitudinal national French study.

STUDY QUESTION: Do IVF, IUI or female infertility (i.e. endometriosis, polycystic ovary syndrome [PCOS] and primary ovarian insufficiency [POI]) lead to an increased risk of congenital anomalies in singletons? SUMMARY ANSWER: After multivariable adjustments, the increased risks of congenital defects associated with IUI were no longer significant, but the underlying maternal infertility presented a potential emental risk, in addition to the risk associated with IVF. WHAT IS KNOWN ALREADY: Most epidemiological studies suggest that singletons born from ART have a higher risk of birth defects, specifically musculoskeletal, cardiovascular and urogenital disorders. However, most of these studies were established on data obtained at birth or in the neonatal period and from relatively small populations or several registries. Moreover, to our knowledge, female infertility, which is a potential confounder, has never been included in the risk assessment. STUDY DESIGN, SIZE, DURATION: Using data from the French National Health System database, we conducted a comparative analysis of all singleton births (deliveries ≥22 weeks of gestation and/or >500 g of birthweight) in France over a 5-year period (2013-2017) resulting from fresh embryo or frozen embryo transfer (fresh-ET or FET from IVF/ICSI cycles), IUI and natural conception (NC). Data were available for this cohort of children at least up to early childhood (2.5 years old). PARTICIPANTS/MATERIALS, SETTING, METHODS: A total of 3 501 495 singleton births were included (3 417 089 from NC, 20 218 from IUI, 45 303 from fresh-ET and 18 885 from FET). Data were extracted from national health databases and used to identify major birth defects. Malformations were classified according to the 10th revision of the International Classification of Disease. To analyse the effect of mode of conception, multivariable analyses were performed with multiple logistic regression models adjusted for maternal age, primiparity, obesity, smoking, history of high blood pressure or diabetes and female infertility. MAIN RESULTS AND THE ROLE OF CHANCE: In our cohort of children, the overall prevalence of congenital malformations was 3.78% after NC, 4.53% after fresh-ET, 4.39% after FET and 3.91% after IUI (132 646 children with major malformations). Compared with infants conceived naturally, children born after fresh-ET and after FET had a significantly higher prevalence of malformations, with an adjusted odds ratio (aOR) of 1.15 [95% CI 1.10-1.20, P < 0.0001] and aOR of 1.13 [95% CI 1.05-1.21, P = 0.001], respectively. Among the 15 relevant subgroups of malformations studied, we observed a significantly increased risk of eight malformations in the fresh-ET group compared with the NC group (i.e. musculoskeletal, cardiac, urinary, digestive, neurological, cleft lip and/or palate and respiratory). In the FET group, this increased risk was observed for digestive and facial malformations. The overall risk of congenital malformations, and the risk by subtype, was similar in the IUI group and the NC group (overall risk: aOR of 1.01 [95% CI 0.94-1.08, P = 0.81]). In addition, there was an overall independent increase in the risk of congenital defects when the mothers were diagnosed with endometriosis (1.16 aOR [95% CI 1.10-1.22], P < 0.0001), PCOS (1.20 aOR [95% CI 1.08-1.34], P = 0.001) or POI (1.52 aOR [95% CI 1.23-1.88], P = 0.0001). Chromosomal, cardiac and neurological anomalies were more common in the three maternal infertility groups. LIMITATIONS, REASONS FOR CAUTION: Male infertility, the in vitro fertilization method (i.e. in vitro fertilization without or with sperm injection: conventional IVF vs ICSI) and embryo stage at transfer could not be taken into account. Furthermore, residual confounding cannot be excluded as well as uncertainties regarding the diagnostic criteria used for the three female infertilities. Findings for specific malformations should be interpreted with caution because the number of cases was small in some sub-groups (potentially due to the Type I error or multiple testing). WIDER IMPLICATIONS OF THE FINDINGS: In this large study, after multivariable maternal adjustments, a moderately increased risk of defects subsisted after IVF, while those associated with IUI were no longer significant. In addition, our results showed that underlying maternal infertility could contribute to the increased risk of defects associated with IVF. These novel findings highlight the importance of taking into account the ART treatment methods and the type of infertility. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the National Agency of Biomedicine. The authors have no competing interests to disclose. TRIAL REGISTRATION NUMBER: NA.

Endometriosis and assisted reproductive techniques independently related to mother-child morbidities: a French longitudinal national study.

RESEARCH QUESTION: Does endometriosis increase obstetric and neonatal complications, and does assisted reproductive technology (ART) cause additional risk of maternal or fetal morbidity? DESIGN: A nationwide cohort study (2013-2018) comparing maternal and perinatal morbidities in three groups of single pregnancies: spontaneous pregnancies without endometriosis; spontaneous pregnancies with endometriosis; and ART pregnancies in women with endometriosis. RESULTS: Mean maternal ages were 30.0 (SD = 5.3), 31.7 (SD = 4.8) and 33.1 years (SD = 4.0), for spontaneous conceptions, spontaneous conceptions with endometriosis and ART pregnancies with endometriosis groups, respectively (P < 0.0001). Comparison of spontaneous conceptions with endometriosis and spontaneous conceptions: endometriosis independently increased the risk of venous thrombosis (adjusted OR [aOR] 1.51, P < 0.001), pre-eclampsia (aOR 1.29, P < 0.001), placenta previa (aOR 2.62, P < 0.001), placental abruption (aOR 1.54, P < 0.001), premature birth (aOR 1.37, P < 0.001), small for gestational age (aOR 1.05, P < 0.001) and malformations (aOR 1.06, P = 0.049). Comparison of ART pregnancies with endometriosis and spontaneous conceptions with endometriosis: ART increased the risk of placenta previa (aOR 2.43, 95% CI 2.10 to 2.82, P < 0.001), premature birth (aOR 1.42, 95% CI 1.29 to 1.55, P < 0.001) and small for gestational age (aOR 1.18, 95% CI 1.10 to 1.27, P < 0.001), independently from the effect of endometriosis. Risk of pre-eclampsia, placental abruption or congenital malformations was not increased with ART. CONCLUSION: Endometriosis is an independent risk factor for mother and child morbidities. Maternal morbidity and perinatal morbidity were significantly increased by ART in addition to endometriosis; however, some perinatal and maternal morbidity risks were increasingly linked to pathologies related to infertility.

Assessment of chimerism and immunomodulation to prevent post-transplantation relapse in childhood acute myeloblastic leukemia: is it the right approach?

Relapse of acute myeloblastic leukemia (AML) after first allogenic hematopoietic stem-cell transplantation (allo-HSCT) is a fatal complication. Sixty-five children transplanted for AML were included in a prospective national study from June 2005 to July 2008 to explore the feasibility of preemptive immune modulation based on the monitoring of blood chimerism. Relapse occurred in 23 patients (35%). The median time between the last complete chimerism and relapse was 13.5 days (2-138). Prompt discontinuation of cyclosporin and the administration of donor lymphocyte infusions (DLIs) based on chimerism monitoring failed as a preemptive tool, either for detecting relapse or certifying long-term remission.

Autosomal-Recessive Mutations in AP3B2, Adaptor-Related Protein Complex 3 Beta 2 Subunit, Cause an Early-Onset Epileptic Encephalopathy with Optic Atrophy.

Early-onset epileptic encephalopathy (EOEE) represents a heterogeneous group of severe disorders characterized by seizures, interictal epileptiform activity with a disorganized electroencephalography background, developmental regression or retardation, and onset before 1 year of age. Among a cohort of 57 individuals with epileptic encephalopathy, we ascertained two unrelated affected individuals with EOEE associated with developmental impairment and autosomal-recessive variants in AP3B2 by means of whole-exome sequencing. The targeted sequencing of AP3B2 in 86 unrelated individuals with EOEE led to the identification of an additional family. We gathered five additional families with eight affected individuals through the Matchmaker Exchange initiative by matching autosomal-recessive mutations in AP3B2. Reverse phenotyping of 12 affected individuals from eight families revealed a homogeneous EOEE phenotype characterized by severe developmental delay, poor visual contact with optic atrophy, and postnatal microcephaly. No spasticity, albinism, or hematological symptoms were reported. AP3B2 encodes the neuron-specific subunit of the AP-3 complex. Autosomal-recessive variations of AP3B1, the ubiquitous isoform, cause Hermansky-Pudlak syndrome type 2. The only isoform for the δ subunit of the AP-3 complex is encoded by AP3D1. Autosomal-recessive mutations in AP3D1 cause a severe disorder cumulating the symptoms of the AP3B1 and AP3B2 defects.

Familial pancreatic adenocarcinoma: A retrospective analysis of germline genetic testing in a French multicentre cohort.

The rate of genetic diagnosis of French patients with familial pancreatic ductal adenocarcinoma (PDAC) is not known. We report germline genetic testing data from 133 index cases meeting criteria for familial pancreatic cancer (FPC) as well as 87 'FPC-like' index cases who did not fulfilled strict FPC definition but were evocative for a PDAC predisposition. The overall rate of genetic diagnosis (in BRCA1, BRCA2, CDKN2A, and ATM genes) was 8.3% in FPC patients and 4.6% in FPC-like patients, consistent with the literature in other populations. Genetic variants were also identified in FANCA and BAP1 genes, as well as in the CDKN2A p12 transcript. This pancreas-specific transcript is a known key player in driving pancreatic oncogenesis. This might be the first described case of a PDAC genetic predisposition due to a variant in this specific transcript.

Risk estimation of uniparental disomy of chromosome 14 or 15 in a fetus with a parent carrying a non-homologous Robertsonian translocation. Should we still perform prenatal diagnosis?

OBJECTIVE: Uniparental disomy (UPD) testing is currently recommended during pregnancy in fetuses carrying a balanced Robertsonian translocation (ROB) involving chromosome 14 or 15, both chromosomes containing imprinted genes. The overall risk that such a fetus presents a UPD has been previously estimated to be around ~0.6-0.8%. However, because UPD are rare events and this estimate has been calculated from a number of studies of limited size, we have reevaluated the risk of UPD in fetuses for whom one of the parents was known to carry a nonhomologous ROB (NHROB). METHOD: We focused our multicentric study on NHROB involving chromosome 14 and/or 15. A total of 1747 UPD testing were performed in fetuses during pregnancy for the presence of UPD(14) and/or UPD(15). RESULT: All fetuses were negative except one with a UPD(14) associated with a maternally inherited rob(13;14). CONCLUSION: Considering these data, the risk of UPD following prenatal diagnosis of an inherited ROB involving chromosome 14 and/or 15 could be estimated to be around 0.06%, far less than the previous estimation. Importantly, the risk of miscarriage following an invasive prenatal sampling is higher than the risk of UPD. Therefore, we do not recommend prenatal testing for UPD for these pregnancies and parents should be reassured.

Diversity of genetic events associated with MLH1 promoter methylation in Lynch syndrome families with heritable constitutional epimutation.

PURPOSE: Constitutional epimutations are an alternative to genetic mutations in the etiology of genetic diseases. Some of these epimutations, termed secondary, correspond to the epigenetic effects of cis-acting genetic defects transmitted to the offspring following a Mendelian inheritance pattern. In Lynch syndrome, a few families with such apparently heritable MLH1 epimutations have been reported so far. METHODS: We designed a long-range polymerase chain reaction next-generation sequencing strategy to screen MLH1 entire gene and applied it to 4 French families with heritable epimutations and 10 additional patients with no proven transmission of their epimutations. RESULTS: This strategy successfully detected the insertion of an Alu element in MLH1 coding sequence in one family. Two previously unreported MLH1 variants were also identified in other epimutation carriers: a nucleotide substitution within intron 1 and a single-nucleotide deletion in the 5'-UTR. Detection of a partial MLH1 duplication in another family required multiplex ligation-dependent probe amplification technology. We demonstrated the segregation of these variants with MLH1 methylation and studied the functional consequences of these defects on transcription. CONCLUSION: This is the largest cohort of patients with MLH1 secondary epimutations associated with a broad spectrum of genetic defects. This study provides further insight into the complexity of molecular mechanisms leading to secondary epimutations.

Mutation of SLC9A1, encoding the major Na⁺/H⁺ exchanger, causes ataxia-deafness Lichtenstein-Knorr syndrome.

Lichtenstein-Knorr syndrome is an autosomal recessive condition that associates sensorineural hearing loss and cerebellar ataxia. Here, we report the first identification of a gene involved in Lichtenstein-Knorr syndrome. By using a combination of homozygosity mapping and whole-exome sequencing, we identified the homozygous p.Gly305Arg missense mutation in SLC9A1 that segregates with the disease in a large consanguineous family. Mutant glycine 305 is a highly conserved amino acid present in the eighth transmembrane segment of all metazoan orthologues of NHE1, the Na(+)/H(+) exchanger 1, encoded by SLC9A1. We demonstrate that the p.Gly305Arg mutation causes the near complete de-glycosylation, mis-targeting and loss of proton pumping activity of NHE1. The comparison of our family with the phenotypes of spontaneous and knockout Slc9a1 murine models demonstrates that the association between ataxia and hearing loss is caused by complete or near complete loss of function of NHE1 and altered regulation of pHi in the central nervous system.

WWOX-related encephalopathies: delineation of the phenotypical spectrum and emerging genotype-phenotype correlation.

BACKGROUND: Homozygous mutations in WWOX were reported in eight individuals of two families with autosomal recessive spinocerebellar ataxia type 12 and in two siblings with infantile epileptic encephalopathy (IEE), including one who deceased prior to DNA sampling. METHODS: By combining array comparative genomic hybridisation, targeted Sanger sequencing and next generation sequencing, we identified five further patients from four families with IEE due to biallelic alterations of WWOX. RESULTS: We identified eight deleterious WWOX alleles consisting in four deletions, a four base-pair frameshifting deletion, one missense and two nonsense mutations. Genotype-phenotype correlation emerges from the seven reported families. The phenotype in four patients carrying two predicted null alleles was characterised by (1) little if any psychomotor acquisitions, poor spontaneous motility and absent eye contact from birth, (2) pharmacoresistant epilepsy starting in the 1st weeks of life, (3) possible retinal degeneration, acquired microcephaly and premature death. This contrasted with the less severe autosomal recessive spinocerebellar ataxia type 12 phenotype due to hypomorphic alleles. In line with this correlation, the phenotype in two siblings carrying a null allele and a missense mutation was intermediate. CONCLUSIONS: Our results obtained by a combination of different molecular techniques undoubtedly incriminate WWOX as a gene for recessive IEE and illustrate the usefulness of high throughput data mining for the identification of genes for rare autosomal recessive disorders. The structure of the WWOX locus encompassing the FRA16D fragile site might explain why constitutive deletions are recurrently reported in genetic databases, suggesting that WWOX-related encephalopathies, although likely rare, may not be exceptional.

Follow-up of post-transplant minimal residual disease and chimerism in childhood lymphoblastic leukaemia: 90 d to react.

Relapse after transplantation is a major cause of treatment failure in paediatric acute lymphoblastic leukaemia (ALL). Here, we report the findings of a prospective national study designed to investigate the feasibility of immune intervention in children in first or subsequent remission following myeloablative conditioning. This study included 133 children who received a transplant for ALL between 2005 and 2008. Minimal Residual Disease (MRD) based on T cell receptor/immunoglobulin gene rearrangements was measured on days -30, 30, 90 and 150 post-transplantation. Ciclosporin treatment was rapidly discontinued and donor lymphocyte infusions (DLI) were programmed for patients with a pre- or post-transplant MRD status ≥10(-3) . Only nine patients received DLI. Pre- and post-transplant MRD status, and the duration of ciclosporin were independently associated with 5-year overall survival (OS), which was 62·07% for the whole cohort. OS was substantially higher in patients cleared of MRD than in those with persistent MRD (52·3% vs. 14·3%, respectively). Only pre-transplant MRD status (Hazard Ratio 2·57, P = 0·04) and duration of ciclosporin treatment (P < 0·001) were independently associated with relapse. The kinetics of chimerism were not useful for predicting relapse, whereas MRD monitoring up to 90 d post-transplantation was a valuable prognostic tool to guide therapeutic intervention.

WWOX and severe autosomal recessive epileptic encephalopathy: first case in the prenatal period.

WWOX has been recently implicated in autosomal recessive spinocerebellar ataxia type 12 (SCAR12) and severe early-onset epileptic encephalopathy (EOEE). By array comparative genomic hybridization, we identified a 0.6 Mb homozygous deletion in 16q23.1 in a fetus presenting with brain anomalies. His older sister who died at the age of 22 months from an EOEE was also homozygous for the copy number variations in 16q23.1. This deletion includes the first six exons of WWOX and results in a null genotype in homozygous patients. This family gives additional support for the implication of WWOX in severe EOEEs. We report for the first time prenatal ultrasound findings in a fetus with a WWOX-null genotype. Our study expands the range of brain abnormalities in WWOX-related EOEEs. This additional family confirms the genotype-phenotype correlation with WWOX-null alleles associated with the most severe form of WWOX-related epileptic encephalopathy with premature death.

A pure familial 6q15q21 split duplication associated with obesity and transmitted with partial reduction.

Familial transmission of chromosome 6 duplications is rare. We report on the first observation of a maternally-inherited pure segmental 6q duplication split into two segments, 6q15q16.3 and 6q16.3q21, and associated with obesity. Obesity has previously been correlated to chromosome 6 q-arm deletion but has not yet been assessed in duplications. The aim of this study was to characterize the structure of these intrachromosomal insertional translocations by classic cytogenetic banding, array-CGH, FISH, M-banding and genotyping using microsatellites and SNP array analysis, in a mother and four offspring. The duplicated 6q segments, 9.75 Mb (dup 1) and 7.05 Mb (dup 2) in size in the mother, were inserted distally into two distinct chromosome 6q regions. They were transmitted to four offspring. A son and a daughter inherited the two unbalanced insertions and displayed, like the mother, an abnormal phenotype with facial dysmorphism, intellectual disability, and morbid obesity. Curiously, two daughters with a normal phenotype inherited only the smaller segment, 6q16.3q21. The abnormal phenotype was associated with the larger proximal 6q15q16.3 duplication. We hypothesize a mechanism for this exceptional phenomenon of recurrent reduction and transmission of the duplication during meiosis in a family. We expect the interpretation of our findings to be useful for genetic counseling and for understanding the mechanisms underlying these large segmental 6q duplications and their evolution.

Severe sex differentiation disorder in a boy with a 3.8 Mb 10q25.3-q26.12 microdeletion encompassing EMX2.

The molecular basis of male disorders of sex development (DSD) remains unexplained in a large number of cases. EMX2 has been proposed to play a role in the masculinization process for the past two decades, but formal evidence for this causal role is scarce. The aim of this study is to yield additional support to this hypothesis by reporting on a male patient who presented with 46,XY DSD, a single kidney, intellectual disability, and the smallest microdeletion including EMX2 reported to date. EMX2 haploinsufficiency is likely to explain the masculinization defect observed in our patient, similar to what has been described in the mouse. In the case of cytogenetically diagnosed cases, deletions of EMX2 have been associated with a wide range of DSD, ranging from hypospadias to complete sex reversal.

De novo complex X chromosome rearrangement unmasking maternally inherited CSF2RA deletion in a girl with pulmonary alveolar proteinosis.

We report on a 3-year-old girl with a de novo complex X chromosome rearrangement associated with congenital pulmonary alveolar proteinosis (PAP) and short stature. Array comparative genome hybridization and FISH analyses contributed to characterize the complex rearrangement consisting of a 7.37 Mb terminal deletion of Xp22.33p22.2, a 17.3 Mb interstitial inverted duplication of Xp22.2p21.3, and a 10.14 Mb duplication of Xq27.3q28. PCR analysis of microsatellite markers supported a paternal origin of the X chromosome rearrangement. A pre-meiotic two-step mechanism may explain the occurrence of this complex X rearrangement: an inverted duplication deletion event on Xp, and duplication of the Xq27.3qter region through a telomere capture event stabilizing the broken chromosome Xp end. The girl has also inherited from her healthy mother an X chromosome with a colony stimulating factor 2 receptor, alpha (CSF2RA) gene deletion. Consistent with the recessive mode of inheritance, the de novo paternal Xp22.33p22.2 deletion combined to the maternally inherited CSF2RA gene deletion led to homozygous deletion of CSF2RA and PAP diagnosis in the girl. The Xp deletion encompasses the pseudoautosomal region 1 (PAR1) which contains genes that escape X inactivation. Short stature homeobox (SHOX) haploinsufficiency explains growth retardation. Absence of other symptoms in relation to the X deletion/amplification is most probably due to skewed X inactivation. Finally, inherited deletions may unmask rare pathogenic genomic rearrangement and contribute to clinical phenotypes by a recessive mode of gene action.

Mutations in WNT10A are frequently involved in oligodontia associated with minor signs of ectodermal dysplasia.

Ectodermal dysplasias (ED) are a clinically and genetically heterogeneous group of hereditary disorders that have in common abnormal development of ectodermal derivatives. Hypohidrotic ectodermal dysplasia (HED) is characterized by abnormal development of eccrine sweat glands, hair, and teeth. The X-linked form of the disease, caused by mutations in the EDA gene, represents the majority of patients with the hypohidrotic form. Autosomal dominant and autosomal recessive forms are occasionally seen, and result from mutations in at least three genes (WNT10A, EDAR, or more rarely EDARADD). We have screened for mutations in EDAR (commonly involved in the hypohidrotic form) and WNT10A (involved in a wide spectrum of ED and in isolated hypodontia) in a cohort of 36 patients referred for EDA molecular screening, which failed to identify any mutation. We identified eight EDAR mutations in five patients (two with homozygous mutations, one with compound heterozygous mutations, and two with heterozygous mutation), four of which were novel variants. We identified 28 WNT10A mutations in 16 patients (5 with homozygous mutations, 7 with compound heterozygous mutations, and 4 with heterozygous mutations), seven of which were novel variants. Our study allows a more precise definition of the phenotypic spectrum associated with EDAR and WNT10A mutations and underlines the importance of the implication of WNT10A among patients with ED.