A high rate of consanguinity leads to a high prevalence of autosomal recessive disorders in inbred populations. One example of inbred populations is the Arab communities in Israel and the Palestinian Authority. In the Palestinian Authority in particular, due to limited access to specialized medical care, most patients do not receive a genetic diagnosis and can therefore neither receive genetic counseling nor possibly specific treatment. We used whole-exome sequencing as a first-line diagnostic tool in 83 Palestinian and Israeli Arab families with suspected neurogenetic disorders and were able to establish a probable genetic diagnosis in 51% of the families (42 families). Pathogenic, likely pathogenic or highly suggestive candidate variants were found in the following genes extending and refining the mutational and phenotypic spectrum of these rare disorders: ACO2, ADAT3, ALS2, AMPD2, APTX, B4GALNT1, CAPN1, CLCN1, CNTNAP1, DNAJC6, GAMT, GPT2, KCNQ2, KIF11, LCA5, MCOLN1, MECP2, MFN2, MTMR2, NT5C2, NTRK1, PEX1, POLR3A, PRICKLE1, PRKN, PRX, SCAPER, SEPSECS, SGCG, SLC25A15, SPG11, SYNJ1, TMCO1, and TSEN54. Further, this cohort has proven to be ideal for prioritization of new disease genes. Two separately published candidate genes (WWOX and PAX7) were identified in this study. Analyzing the runs of homozygosity (ROHs) derived from the Exome sequencing data as a marker for the rate of inbreeding, revealed significantly longer ROHs in the included families compared with a German control cohort. The total length of ROHs correlated with the detection rate of recessive disease-causing variants. Identification of the disease-causing gene led to new therapeutic options in four families.

Visual impairment due to inherited ophthalmic disorders is amongst the most common disabilities observed in populations practicing consanguineous marriages. Here we investigated the molecular genetic basis of an unselected broad range of ophthalmic disorders in 20 consanguineous families from Arab villages of Israel and the Palestinian Authority. Most patients had little or very poor prior clinical workup and were recruited in a field study. Homozygosity mapping followed by candidate gene sequencing applying conventional Sanger sequencing or targeted next generation sequencing was performed in six families. In the remaining 14 families, one affected subject per family was chosen for whole exome sequencing. We discovered likely disease-causing variants, all homozygous, in 19 of 20 independent families (95%) including a previously reported novel disease gene for congenital nystagmus associated with foveal hypoplasia. Moreover, we found a family in which disease-causing variants for two collagenopathies - Stickler and Knobloch syndrome - segregate within a large sibship. Nine of the 19 distinct variants observed in this study were novel. Our study demonstrated a very high molecular diagnostic yield for a highly diverse spectrum of rare ophthalmic disorders in Arab patients from Israel and the Palestinian Authority, even with very limited prior clinical investigation. We conclude that 'genetic testing first' may be an economic way to direct clinical care and to support proper genetic counseling and risk assessment in these families.

Introduction Glycine encephalopathy (GE), also known as non-ketotic hyperglycinemia (NKH), is a rare inborn error of glycine metabolism caused by a defect in glycine cleavage system, a multi-enzyme complex located in mitochondrial membrane. This defect results in elevated glycine concentration in plasma and cerebrospinal fluid (CSF). Clinical manifestations vary from severe lethargy, hypoactivity and apneic episodes in the neonatal form, mild or moderate psychomotor delay and seizures in the infantile form, and abnormal behaviors, ataxia and choreoathetoid movements in late onset form. More than 50 GLDC mutations were found, reflecting large heterogeneity of the gene. Methods We describe the clinical, biochemical and molecular characteristics of three Palestinian siblings who have distinct clinical phenotypes. Molecular study was performed utilizing standard Polymerase Chain Reaction (PCR) amplification then direct DNA sequencing for the affected family members. Results Their phenotypes included severe symptoms in neonatal period, infantile onset of seizure and psychomotor delay and a mild late-onset form with speech delay at age 20 months. All siblings were homozygous for a novel mutation Y164H in exon 4 of GLDC gene. The described novel homozygous variant in our study is predicted deleterious and pathogenic. Conclusions This article further expands the genetic spectrum of glycine encephalopathy and adds an evidence of the clinical heterogeneity of glycine encephalopathy even in siblings with identical mutation.

We report the first Palestinian child manifesting with 3-methylglutaconic aciduria psychomotor delay, muscle hypotonia, sensori-neural deafness, and Leigh-like lesions on brain magnetic resonance imaging (MRI), a clinical phenotype that is characteristic of MEGDEL syndrome. MEGDEL syndrome was recently found to be caused by mutations in SERAC1, encoding a protein essential for mitochondrial function, phospholipid remodeling, and intracellular cholesterol trafficking. We identified a novel homozygous mutation in SERAC1 gene (c.1018delT) that generates frame shift and premature termination of protein translation. Plasma and cerebrospinal fluid lactate, plasma alanine, and respiratory chain complexes in fresh muscle were normal. This report further expands the genetic spectrum of MEGDEL syndrome and adds to the evidence that it is associated with variable patterns of respiratory chain abnormalities.

Clopidogrel is an antiplatelet drug used to prevent recurrent ischemic events after acute coronary syndrome and/or coronary stent implantation. Single nucleotide polymorphisms (SNPs) such as CYP2C19*2 and ABCB1 C3435T have been found to play a role in different individual responses to clopidogrel. Since the prevalence of these SNPs is generally known to differ from one population to another, the aim of this study was to examine their prevalence in both a Palestinian and Turkish population. One hundred unrelated Palestinian subjects and 100 unrelated Turkish subjects were analyzed for CYP2C19*2 and ABCB1 C3435T polymorphisms by the amplification refractory mutation system (ARMS).Results showed an ABCB1 3435 T allele frequency of 0.46 (95% CI 0.391 to 0.529) in the Palestinian sample and 0.535 (95% CI 0.4664 to 0.6036) in the Turkish sample. CYP2C19*2 allele frequency was 0.095 (95% CI 0.0558 to 0.134) in the Palestinian sample and 0.135 (95% CI 0.088 to 0.182) in the Turkish sample.Our results provide information about the prevalence of the polymorphisms related to clopidogrel response in both the Palestinian and Turkish populations, in order to improve the safety and efficacy of clopidogrel through use of genetically guided, individualized treatment. The prevalence of these clinically significant alleles shed light on the importance of testing them before prescribing clopidogrel. (C) 2014 The Authors. Published by Elsevier B.V.

Autoinflammatory diseases are a group of disorders characterized by seemingly unprovoked inflammation in the absence of high-titer autoantibodies or antigen-specific T-cells. Familial Mediterranean fever (FMF) is the archetypal hereditary periodic fever syndrome and autoinflammatory disorder. It is characterized by recurrent self-limiting episodes of fever and painful polyserositis. FMF is an autosomal recessive disorder, with considerable prevalence in specific ethnic groups, namely, non-Ashkenazi Jews, Armenians, Turks and Arabs and the FMF carrier rate can be as high as one in four. The gene responsible for FMF, MEFV, is located on the short arm of human chromosome 16, and was independently identified by two positional cloning consortia. Mutations, as well as, polymorphisms in MEFV are continuously identified. In Arabic FMF patients the spectrum and distribution of MEFV mutations are distinctive and the portion of unidentified mutations is undoubtedly the highest amongst the groups commonly affected by FMF. The comprehensive identification of MEFV mutant alleles among FMF patients is needed for the efficient examination of specific genotype – phenotype correlation patterns and for the development of molecular tools to support the clinical diagnosis.

Objective . This study aimed at analyzing the association between recurrent pregnancy loss (RPL) and factor V G1691A (FVL), prothrombin G20210 (FII); and MTHFR C677T (MTHFR) in Palestinian women. Method . We studied 329 Palestinian women with RPL and/or stillbirth (SB); and compared them to 402 healthy reproductive Palestinian women. Cases and controls were tested for the above mutations. Odds ratio (OR) at confidence interval (CI) of 95% was used as a measure of association between the mutations and RPL. Results . Our statistical analysis showed a slightly increased association, which was not significant between FVL and RPL (OR 1.32, 95% CI 0.90–1.94), and no association between FII (OR 0.84, 95% CI 0.38–1.92), MTHFR (OR 0.58, 95% CI 0.32–1.03), and RPL. Further analysis of RPL subgroups revealed an association between FVL and first-trimester loss (OR 1.33, 95% CI 0.892–1.989), and second-trimester loss (OR 1.13, 95% CI 0.480–2.426), both were not statistically significant. Furthermore, the only statistically significant association was between FVL and SB (OR 2.0, 95% CI 1.05–3.70). Conclusion . Our analysis had failed to find a significant association between FVL, FII, MTHFR; and RPL in either the first or second trimester. FVL was significantly associated with fetal loss if the loss was a stillbirth.

Vladimir Sorin1,*, Aya Mizrahi2, Patricia Ohana2, Suhail Ayesh2, Tatiana Birman2, Abraham Hochberg2, Abraham Czerniak1 Department of Surgery “A”, E.Wolfson Medical Center, Holon, Israel Department of Biological Chemistry, Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel __________________________________________________________________________________ *Correspondence: Dr. Vladimir Sorin, Department of Surgery, E.Wolfson Medical Center, Ha Lochamim 62, Holon, Israel, 58100; Fax: +97225610250; e-mail: vladimir.sorin@gmail.com

Sickle cell disease is an inherited autosomal recessive disorder of the beta-globin chain. In Palestine it is accompanied by a low level of Hb F (mean 5.14%) and a severe clinical presentation. In this study, 59 Palestinian patients, homozygotes for Hb S were studied for their haplotype background. Eight polymorphic sites in the beta-globin gene cluster were examined. The Benin haplotype was predominant with a frequency of 88.1%, followed by a frequency of 5.1% for the Bantu haplotype. One chromosome was found to carry the Cameroon haplotype (0.85%). Three atypical haplotypes were also found (5.95%). Heterogeneity was observed in Hb F production, ranging between 1.5 and 17.0%, whereas the (G)gamma ratio was homogeneous among all haplotypes with a normal amount of about 41%. Our results are in agreement with previous reports of the Benin haplotype origin in the Mediterranean.

Summary H19 is a maternally expressed oncofetal gene, which is expressed at substantial levels in embryonic tissues and various human tumor types, but marginally or not expressed in the corresponding tissue of the adult. Dramatic increase in H19 mRNA was found in hepatocytes following the initiation of their proliferation by hepatectomy. H19 is a target gene for HGF, which also sharply increases after hepatectomy. The growth of liver metastases after hepatectomy and it's correlation with levels of the H19 and HGF RNA were evaluated on an orthotopic model of rat colon liver metastases. Significant increase in tumor growth was found after 70% hepatectomy. HGF and H19 RNAs levels in tumors were substantially elevated after hepatectomy compared to those observed in tumors with no hepatectomy. A direct positive correlation between the level of H19 RNA and the extent of liver resection was found. HGF has an important role in overexpression of H19 resulting in a rapid growth of metastases in the remaining liver after hepatectomy. These findings may have important clinical applications, when decisions regarding partial hepatectomy, two stage hepatectomy procedure or staged radiofrequency ablation prior to hepatectomy has to be taken.

Objectives: The aim of our study was to determine the prevalence of Mediterranean fever gene (MEFV) mutations among Palestinian patients with Behcet's disease (BD).Methods: We screened 42 BD patients from the West Bank and Jerusalem for most of the MEFV mutations known to date. Patients diagnosed clinically according to the International Study Group (ISG) criteria were recruited from Makassed Islamic Charitable Hospital and private clinics. We performed the DNA testing using direct DNA sequencing of exon 10 of the MEFV gene and using the amplification refractory mutation system (ARMS) technique for mutations located in other exons.Results: We found that 40.5% of the samples had nine different MEFV mutations and one polymorphism. E148Q was the most prevalent mutation, found in 38.1% of the mutated alleles. M694V, V726A, M694I, A744S, P369S, R408Q, and F479L were each detected in 4.8% of the mutated alleles studied. The polymorphism P706 was detected in 9.5% of the mutated alleles. The mutations A744S, P369S, R408Q, and F479L were reported for the first time in BD patients. V722M, a novel MEFV mutation that has not been reported before in either FMF or BD patients, was identified in this study.Conclusion: This study is the first genetic analysis of MEFV mutations among Palestinian BD patients. It reflects their mutations profile, providing further data that MEFV mutations are an additional genetic susceptibility factor in BD.

Background Mutations and epigenetic aberrant signaling of growth factors pathways contribute to carcinogenesis. Recent studies reveal that non-coding RNAs are controllers of gene expression. H19 is an imprinted gene that demonstrates maternal monoallelic expression without a protein product; although its expression is shut off in most tissues postnatally, it is re-activated during adult tissue regeneration and tumorigenesis. Moreover, H19 is highly expressed in liver metastasis derived from a range of carcinomas. The objective of this study is to explore the role of H19 in carcinogenesis, and to determine its identification as an anti-tumor target. Methodology/ Principle Findings By controlling oxygen pressure during tumor cell growth and H19 expression levels, we investigated the role of H19 expression in vitro and in vivo in hepatocellular (HCC) and bladder carcinoma. Hypoxia upregulates the level of H19 RNA. Ablations of tumorigenicity of HCC and bladder carcinomas in vivo are seen by H19 knockdown which also significantly abrogates anchorage-independent growth after hypoxia recovery, while ectopic H19 expression enhances tumorigenic potential of carcinoma cells in vivo. Knocking-down H19 message in hypoxic stress severely diminishes p57kip2 induction. We identified a number of potential downstream targets of H19 RNA, including angiogenin and FGF18. Conclusions H19 RNA harbors pro-tumorigenic properties, thus the H19 gene behaves as an oncogene and may serve as a potential new target for anti-tumor therapy.

Real-time PCR technology is highly advantageous for gene studies based on the genetic nature of the transferred material. Urine and blood samples were collected before and after treatment. Treatment of bladder carcinoma patients with plasmid constructs expressing the diphtheria toxin gene was monitored. Detection range from 5×106 copies to ≤10 copies was observed with repeatability and reproducibility. No trace amount of the plasmid was detected in the patient's bloodstream, and a continuous clearance in urine, as well, was observed 72h post-injection. These results show the possibility to design a follow-up method to monitor quantitatively the amount of the plasmid DNA in bladder cancer patients.

OBJECTIVES:Carcinogen-induced bladder cancer in rodents is a key model for evaluation of novel therapies for bladder cancer because of its similarity to the clinical disease. The major drawback of the model is the difficulty in assessing tumor burden in living animals and at necropsy. The objective of this work was to present simple and accurate solutions for this problem. METHODS:Sixty female Wistar rats were given N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN) at a concentration of 0.05% in the drinking water for 35 weeks. Periodic evaluation of tumorigenesis was done by ultrasonography of the anesthetized animals. The tumor burden was evaluated after killing the rats by weighing the bladder, digital measurement of the tumor dimensions, and histologic examination. RESULTS:Focal urothelial hyperplasia was noted by the 5th BBN week, severe dysplasia by the 15th BBN week, and transitional cell carcinoma from the 20th week on. Carcinoma was seen on digital photographs taken from the 20th week on. Tumors as small as 1 mm could be easily measured. A poor correlation (R2 = 0.33) was found between bladder weight and the digital photographic measurements of small tumors (20th BBN week). However, when larger tumors were considered (30th BBN week), a good correlation was found (R2 = 0.81). CONCLUSIONS:Tumor progress in the rat BBN model was accurately monitored by ultrasonography in living animals. Digital measurement of tumor dimensions provided a precise method for evaluation of tumor burden at necropsy.

OBJECTIVE:To investigate the spectrum of mutations and genotypes in the pyrin gene in familial Mediterranean fever (FMF) patients. METHODS:Blood samples of 511 suspected FMF patients, received from the Molecular Genetics Laboratory, Makassed Islamic Charitable Hospital, Mount Olives, Jerusalem during the period from June 1999 to August 2004, were investigated by genotyping 24 different MEFV mutations. RESULTS:Our work revealed the presence of 14 different mutations from the identified 24 mutations in the gene which are assembled in 6 homozygous, 9 heterozygous and 16 compound heterozygous genotypes. The homozygous genotypes represent the predominant format among our patients representing approximately 38% of the revealed genotypes. Interestingly, in 94 (31.4%) of the tested subjects, only one mutation in the pyrin gene could be identified while the other mutant allele remains unidentified. Moreover, the genotype of 3 (1%) patients revealed the presence of triplet mutations in the pyrin gene. CONCLUSION:The results of our study clearly suggest that the origin of FMF among the Palestinian population is mostly homozygous. The identification of a significant number of patients with one known mutation indicates potentially the presence of new mutations in the gene which will be investigated in the future.

β-Thalassemia (thal) is an autosomal recessive disorder that results in hypochromic hemolytic anemia in affected patients. In the West Bank area of Palestine, the prevalence of β-thal trait is approximately 3.5% among the population, with an estimated 120,000 carriers. Seventeen β-globin gene mutations could be identified in 148 patients using polymerase chain reaction (PCR), amplification refractory mutation system (ARMS)-PCR and direct sequencing. The predominant mutations included: IVS-I-6 (T→C) (28.7%), IVS-I-110 (G→A) (17.6%), codon 37 (G→A) (10.4%), IVS-I-1 (G→A) (9%), codons 106/107 (+ G) (6.8%) and codon 39 (C→T) (4.6%). Other less frequent and rare mutations included: IVS-II-1 (G→A), codon 5 (–CT), IVS-II-848 (C→A), –30 (T→A), codons 8/9 (+ G), IVS-I-5 (G→C), –28 (A→C), IVS-II-745 (C→G), codon 6 (–A), codon 27 (G→T) and codon 30 (AGG→ACG). Most patients (62.2%) were homozygous for one type of mutation, while the rest (27.3%) were compound heterozygotes. Some patients were heterozygous for β-thal and sickle cell anemia traits. No mutations could be detected in both alleles of eight patients, while in seven patients only one mutant allele could be detected. Further investigations are needed to resolve the corresponding genotypes of these patients. This study represents a comprehensive investigation of the type, frequency, and distribution of thalassemia mutations among the Palestinian population in the West Bank region of Palestine. A degree of similarity and significant variations was evident in the type and frequency of mutations when the present mutations profile was compared with similar ones among various Arab and non Arab populations. The association between the identified mutations and the corresponding genotypes of our patients with specific polymorphism frameworks in the β-globin gene was performed and the results revealed linkage disequilibrium.

Aims: To investigate the expression of the imprinted oncofetal H19 gene in hepatic metastases derived from a range of human carcinomas and assess its prognostic value with the view of developing a DNA based treatment for such metastases. Methods: Non-radioactive in situ hybridisation for H19 RNA was performed on paraffin wax embedded sections of liver biopsies or partial hepatectomy specimens, taken from 80 patients with hepatic metastases derived from carcinomas from several medical centres in Israel. The degree of expression was graded qualitatively according to the number of cells expressing H19 and the intensity of staining. The medical files were searched for demographic data and survival times before and after diagnosis of hepatic metastases. Results: H19 expression was found in the hepatic metastases of 64 of 80 patients. High expression (higher staining grades) of H19 in the metastases was found in 43 of 80 patients. However, H19 expression status in the hepatic metastases did not correlate with either the length of time to development of metastasis or overall survival. Conclusions: H19 is highly expressed in more than half of hepatic metastases derived from a range of carcinomas. Thus, these metastases may be suitable candidates for H19 DNA based treatment. Further studies are needed to determine whether H19 expression has prognostic value in metastatic liver disease using larger numbers of specific subtypes of primary carcinomas.