CALR gene mutational profile in myeloproliferative neoplasms with non-mutated JAK2 in Moroccan patients: A case series and germline in-frame deletion.

BACKGROUND: The discovery of somatic mutations within the gene encoding calreticulin (CALR) in 2013 represented a major milestone in the molecular diagnosis of BCR-ABL negative myeloproliferative neoplasms (MPN). In fact, exome sequencing revealed that most patients with essential thrombocythemia (ET) or primary myelofibrosis (PMF) lacking JAK2 or MPL mutations, harbor somatic insertion and/or deletion in exon 9 of CALR gene. In this study, we identified the first CALR gene mutational landscape in Moroccan patients with MPN nonmutated for the JAK2 gene. METHODS: We performed Sanger sequencing of exon 9 of CALR gene in blood samples obtained from 33 Moroccan patients with ET or PMF non-mutated for JAK2. RESULTS: Of the 33 patients analyzed, we detected eight distinct variants in 15 patients (45.4%); six indel mutations, five with type 1 recurrent 52bp deletion, four with type 2 recurrent 5bp insertion and one in frame deletion which was found to be a germline variant suggesting a very rare condition in MPN. CONCLUSION: This is the first cohort reported in CALR gene mutation analysis in Morocco. Our results were concordant with studies reported up to date and very encouraging in promoting the molecular diagnosis of myeloproliferative neoplasms in Moroccan patients. Moreover, the presence of a germline in frame deletion in a symptomatic patient should undermine the effectiveness of sizing assays without DNA sequencing in the diagnosis of CALR mutations.

Maternal Hypomethylation of KvDMR in a Monozygotic Male Twin Pair Discordant for Beckwith-Wiedemann Syndrome.

Beckwith-Wiedemann syndrome (BWS; OMIM 130650) is a heterogeneous overgrowth syndrome characterized by visceromegaly, macroglossia, tumor predisposition, and other congenital abnormalities. BWS is usually associated with abnormalities of chromosome 11p15, including (epi)genetic changes, paternal disomy and point mutations. A number of identical twin pairs, mostly female, have been reported to be clinically discordant for BWS. Studies of monozygotic twins discordant for BWS provide more information about failure in the DNA methylation maintenance machinery during very early embryonic development. Here, we report a case of monozygotic male twins discordant for BWS phenotype. Methylation analysis of the 2 imprinted domains at 11p15.5 (H19DMR and KvDMR) was performed by methylation-specific MLPA and pyrosequencing of DNA extracted from peripheral blood and buccal swabs of both twins. Hypomethylation at KvDMR was identified in both cell types of the affected twin, whereas his healthy brother presented hypomethylation only in blood cells and a normal methylation profile in buccal swab. For diagnostic purposes, it is important to remember that twins can share fetal circulation and possibly share hematopoietic stem cells early in development; therefore, the affected and unaffected twins can share an epigenotype that will resemble partial hypomethylation. If a patient is a twin, it is valuable to obtain a sample from a tissue other than blood.

Report of the First Clinical Case of a Moroccan Kabuki Patient with a Novel MLL2 Mutation.

Kabuki syndrome (also known as Niikawa-Kuroki syndrome) is a rare autosomal disorder, characterized by an unusual face, short stature, skeletal, visceral and dermatoglyphic abnormalities, cardiac anomalies, mental retardation, and immunological defects. Point mutations and large intragenic deletions and duplications of the mixed lineage leukemia 2 (MLL2) and exons deletions of lysine demethylase 6A (-KDM6A) genes have been identified as its underlying causes. We report on the first description of a Moroccan Kabuki syndrome patient with typical facial features, developmental delay, finger pads, and other anomalies carrying a novel splice site mutation in the MLL2 gene that produces a truncated and likely pathogenetic form of MLL2 protein.

Inhabitual autosomal recessive form of dentin dysplasia type I in a large consanguineous Moroccan family.

Dentin dysplasia is a rare autosomal dominant genetic disease characterized by defect of dentin development and the causal gene is DSPP (Dentin Sialophosphoprotein gene). We report in the present study a large Moroccan family in which dentin dysplasia is clearly transmitted as an autosomal recessive trait. Four males and females family members born from healthy consanguineous parents are carriers of the typical features of the dentin dysplasia type I. Polymorphic markers that span the DSPP gene, allowed us to show that this locus is not linked to dentin dysplasia in our family. We also excluded in our family the SMOC2 gene (Sparc Related Modular Calcium Binding Protein 2) which was recently identified as a causal gene in dentin dysplasia type I with microdontia and misshapen teeth. This family represents, a new description of autosomal recessive pattern of inheritance of dentin dysplasia type I. Moreover, this form of dentin dysplasia is not allelic to the autosomal dominant dentin dysplasia and the genetic cause is to be discovered.

Non-association of Crohn's disease with NOD2 gene variants in Moroccan patients.

UNLABELLED: Crohn's disease is a chronic inflammatory bowel disease, with multifactorial traits, that can involve any part of the gastrointestinal tract. In recent years, a dozen genome-wide association scan and meta-analysis were published bringing the number of susceptibility alleles to more than 30 variations. However, the major susceptibility gene for Crohn's disease is NOD2, located on proximal 16q, which is involved in the innate immune response. Three main variants of this gene: two single nucleotide polymorphisms p.Arg702Trp and p.Gly908Arg substitutions and frameshift polymorphism p.Leu1007fsinsC are involved in susceptibility to Crohn's disease. There is no data about the frequency of these allelic variants in Moroccan patients with Crohn's disease. The aim of our study is to genotype the NOD2 gene to assess the involvement of these three variants in susceptibility to Crohn's disease for Moroccans. METHODS: We carried out genotyping for the three variants p.Arg702Trp, p.Gly908Arg and p.Leu1007fsinsC of NOD2 gene using PCR-sequencing among 101 Moroccan patients with Crohn's disease and 107 healthy controls. RESULTS: The three main variants of NOD2 gene were present in Moroccan patients with no significant difference compared to controls. CONCLUSION: This preliminary study shows no evidence association of NOD2 gene with Crohn's disease in the Moroccan population.

The first mutations in the MYH gene reported in Moroccan colon cancer patients.

BACKGROUND: Biallelic germline mutations in the MYH gene cause MYH-associated polyposis (MAP) disease, an autosomal recessive form of inherited colorectal cancer. People with MAP tend to develop attenuated multiple adenomatous colon polyps during their lifetime and will have an increased risk of colorectal cancer. Contrary to familial adenomatous polyposis, the number of adenomas is often lower in MAP (from 5 to 100), and even some patients have recently been reported with no identified adenomas. There have been many investigations into MAP that have been conducted in many different countries. Currently there is limited data on MAP in Morocco, and it is reasonable to think, that the prevalence of this form of genetic predisposition is as high as other autosomal recessive genetic diseases found in countries with high rates of consanguinity. The aim of this study is to examine the frequency of MYH mutations in colorectal cancer and/or attenuated polyposis in Moroccan patients. PATIENTS AND METHODS: The study population consisted of 62 patients; 52 with colorectal cancer, three of them had attenuated polyposis (2 to 99 adenomatous polyps). 10 other patients were referred to our department for polyposis without colorectal cancer. We carried out DNA analysis in 62 patients to screen for the three recurrent mutations c.494A>G (p.Tyr165Cys), c.1145G>A (p.Gly382Asp) and c.1185_1186dup, p.Glu396GlyfsX43, whereas 40 subjects were screened for germline MYH mutations in the whole coding sequence of the MYH gene by direct DNA sequencing. All these 40 patients, except two, had colorectal cancer without polyposis. RESULTS: Three patients with colorectal cancer and attenuated polyposis carried biallelic mutations in the MUTYH gene one with the c.494 A>G mutation, one with the c.1105delC mutation, one with the c.1145G>A mutation. One patient with 25 adenomas without colorectal cancer carried the c.1145G>A mutation at a homozygote state and one patient with 3 polyps was heterozygote for the mutation c.1145G>A. No biallelic mutations of MYH gene were detected in colorectal cancer patients and in patients with small number (<5) of polyps without colorectal cancer. CONCLUSION: We report the first biallelic MYH mutations in four Moroccan patients with clinical criteria of MAP; three of them had colorectal cancer with attenuated polyposis. No MYH mutations were found in colorectal patients without polyposis. Despite the relatively small sample size of the current study, our findings suggest that the MAP is not a frequent cause of colon cancer in Morocco as we had expected, and the molecular analysis of MYH gene should be restricted to patients displaying the classical phenotype of MAP.

Prevalence of MYH-associated polyposis related to three recurrent mutations in Morocco.

BACKGROUND: MYH-associated polyposis (MAP) is an autosomal recessive inherited disease. People with MAP tend to develop multiple adenomatous colon polyps during their lifetime and have an increased risk of colorectal cancer. MAP has only recently been described and there is much to be learned about the condition. Recessively inherited mutations in the base excision repair gene MYH have recently been associated with predisposition to colorectal adenomas and cancer. The epidemiology of MYH-associated polyposis (MAP) is poorly known in populations with high levels of consanguinity like North African populations, in particular in Morocco, and the MAP carrier frequency in the general Moroccan population has never been evaluated. The present study was carried out among the Moroccan population, using molecular epidemiology methods, to estimate the prevalence of homozygote or compound heterozygote genotype conferring MAP due to three mutations reported as recurrent in MAP: c.494A>G (Y165C), c.1145G>A (G382D) and c.1186_1187insGG (p.Glu396fsX42). METHODS: To estimate the prevalence of MYH mutations in Morocco, DNA extracted from blood samples of 400 healthy Moroccans was tested for recurrent MYH mutations using real-time PCR or DNA fragment analysis. Heterozygotes profiles were confirmed by direct sequencing. We searched for the mutations c.494A>G and c.1145G>A in 400 subjects, and the mutation c.1186_1187insGG in 250 subjects. RESULTS: One subject was heterozygous for c.494A>G (1/400 or 0.25%), three others for c.1145G>A (3/400 or 0.75%) and one was heterozygous for p.Glu396fsX42 (1/250 or 0.4%). The carrier frequency of one of these three mutations in the Moroccan population was calculated to be 1.4% and the frequency of homozygous or compound heterozygote for these three recurrent mutations is 1/10 000.These figures allowed one to estimate at 3500 the number of Moroccans with high risk of developing colon cancer due only to these three recurrent mutations. CONCLUSION: This preliminary study shows that the Moroccan population is at risk for MAP. This could help to define diagnosis strategies and patient care and may also have implications for genetic counselling.

[First North African observation of Leber congenital amaurosis secondary to CEP290 gene mutation]. / Première observation maghrébine d'amaurose congénitale de Leber secondaire à une mutation du gène CEP290.

Leber congenital amaurosis (LCA) is a the earliest and most severe form of retinal dystrophy responsible for congenital blindness. LCA has genetic heterogeneity and the study of this disease is elucidating the genetics and molecular interactions involved in the development of the retina. To date, 11 LCA genes have been mapped, ten of which have been identified. The CEP290 gene has been shown to account for Joubert and Senior-Loken syndromes and to be a frequent cause of nonsyndromic LCA. We report here the first Arab patient, born to consanguineous parents, with Leber congenital amaurosis attributable to mutation of the CEP290 gene.

Comparative clinical and transcriptomal profiles of breast cancer between French and South Mediterranean patients show minor but significative biological differences.

BACKGROUND: In Western countries, breast cancer incidence and mortality are higher than in Mediterranean countries. These differences have been ascribed to environmental factors but also to late-stage diagnostic and biological specific characteristics. PATIENTS AND METHODS: Between September 2002 and September 2005, we collected clinical data by phone counselling 180 French and Mediterranean breast cancer patients and performed microarray experiments. RESULTS: Characteristics of breast cancer in patients from Lebanon, Tunisia and Morocco were more aggressive (more SBR grade III and positive node invasion) and patients were 10 years younger at diagnosis. Sixteen differentially expressed genes such as MMP9, VEGF, PHB1, BRCA1, TFAP2C, GJA1 and TFF1 were also found. Additionally, an up-regulation of cytokeratins KRT8 and KRT18 may indicate a luminal B subtype in "South" (Lebanon, Tunisia and Morocco) tumors while "North" (France) tumors may more frequently be luminal A type. CONCLUSION: This study allowed the identification of specific clinical and transcriptomic parameters in patients from South Mediterranean countries.

Autosomal dominant anhidrotic ectodermal dysplasias at the EDARADD locus.

Anhidrotic ectodermal dysplasia (EDA) is a disorder of ectodermal differentiation characterized by sparse hair, abnormal or missing teeth, and inability to sweat. X-linked EDA is the most common form, caused by mutations in the EDA gene, which encodes ectodysplasin, a member of the tumor necrosis factor (TNF) family. Autosomal dominant and recessive forms of EDA have been also described and are accounted for by two genes. Mutations in EDAR, encoding a TNF receptor (EDAR) cause both dominant and recessive forms. In addition, mutations in a recently identified gene, EDARADD, encoding EDAR-associated death domain (EDARADD) have been shown to cause autosomal recessive EDA. Here, we report a large Moroccan family with an autosomal dominant EDA. We mapped the disease gene to chromosome 1q42.2-q43, and identified a novel missense mutation in the EDARADD gene (c.335T>G, p.Leu112Arg). Thus, the EDARADD gene accounts for both recessive and dominant EDA. EDAR is activated by its ligand, ectodysplasin, and uses EDARADD to build an intracellular complex and activate nuclear factor kappa B (NF-kB). We compared the functional consequences of the dominant (p.Leu112Arg) and recessive mutation (p.Glu142Lys), which both occurred in the death domain (DD) of EDARADD. We demonstrated that the p.Leu112Arg mutation completely abrogated NF-kB activation, whereas the p.Glu142Lys retained the ability to significantly activate the NF-kB pathway. The p.Leu112Arg mutation is probably a dominant negative form as its cotransfection impaired the wild-type EDARADD's ability to activate NF-kB. Our results confirm that NF-kB activation is impaired in EDA and support the role of EDARADD DD as a downstream effector of EDAR signaling.

[Haemangioma of the right atrium revealed by cardiogenic shock]. / Hémangiome de l'oreillette droite révélé par un choc cardiogénique.

Intracardiac haemangioma is a very rare primary benign tumour. A 20 year old female patient, with no significant previous medical history, presented to the emergency department with cardiovascular collapse and vague abdominal pains, with no peripheral signs of cardiac failure. The electrocardiogram showed sinus rhythm with diffuse reploarisation disturbances. Chest radiography revealed cardiomegaly (cardiothoracic index of 0.67) with a right paracardial opacity. Abdominal ultrasound showed a moderate peritoneal effusion and transthoracic ultrasound showed a tumour occupying the right atrial cavity but sparing the interatrial septum. The patient underwent emergency open heart surgery for tumour resection and right atrial wall repair with autologous pericardium. Histology confirmed a haemangioma. Follow-up at one month was uneventful. The clinical, diagnostic and therapeutic features of this case are underlined.

Homozygosity mapping of a Dyggve-Melchior-Clausen syndrome gene to chromosome 18q21.1.

Dyggve-Melchior-Clausen syndrome (DMC) is an autosomal recessive condition characterised by short trunk dwarfism, scoliosis, microcephaly, coarse facies, mental retardation, and characteristic radiological features. X rays show platyspondyly with double vertebral hump, epiphyseal dysplasia, irregular metaphyses, and a characteristic lacy appearance of the iliac crests. Electron microscopy of chondrocytes have shown widened cisternae of rough endoplasmic reticulum and biochemical analyses have shown accumulation of glucosaminoglycan in cartilage, but the pathogenesis of DMC remains unexplained. Here, we report on the homozygosity mapping of a DMC gene to chromosome 18q21.1 in seven inbred families (Zmax=9.65 at theta=0 at locus D18S1126) in the genetic interval (1.8 cM) defined by loci D18S455 and D18S363. Despite the various geographical origins of the families reported here (Morocco, Tunisia, Portugal, and Lebanon), this condition was genetically homogeneous in our series. Continuing studies will hopefully lead to the identification of the disease causing gene.

Autosomal recessive anhidrotic ectodermal dysplasia in a large Moroccan family.

We studied a large Moroccan family in which anhidrotic ectodermal dysplasia is transmitted as an autosomal recessive trait. Fourteen family members, both males and females, were affected and they all had a common ancestor. Linkage analysis by homozygosity mapping in this family will permit the gene localisation of this rare form of anhidrotic ectodermal dysplasia.

Linkage analysis of families with severe childhood autosomal recessive muscular dystrophy in Morocco indicates genetic homogeneity of the disease in north Africa.

It has been previously shown in Tunisian and Algerian families that the locus for SCARMD maps to the proximal part of 13q, and in Algerian families that the disease is associated with deficiency of the 50 kDa dystrophin associated glycoprotein (50DAG). We have tested this linkage in six families from Morocco where this disease is also prevalent. In one family the 50DAG was tested and found to be negative in a muscle biopsy. Our results showed similar linkage in this country, with statistical tests indicating genetic homogeneity between the three Maghreb countries.

Spondyloepiphyseal dysplasia tarda: linkage with genetic markers from the distal short arm of the X chromosome.

A linkage study of six families with spondyloepiphyseal dysplasia tarda (SEDL) has been performed. A linkage to site DXS41 (theta = 0.08; z = 3.07) and DXS92 (theta = 0.05; z = 2.95) has been established. We propose that the SEDL locus lies on the distal part of the short arm of the X chromosome.