The effect of apoprotein E gene polymorphism on neurocognitive functions of children with CHD.

Studies have demonstrated an association between CHD and neurodevelopmental delay. This delay is associated with many factors like reduced blood flow and oxygen, cardiac catheterisations, and genetic factors. Apo E gene polymorphism is one of these genetic factors. This study aims to show the effect of Apo E gene polymorphism on neurodevelopmental process in children having CHD. A total of 188 children having CHD were admitted to the study. Apo E gene polymorphism of these patients was determined, and psychometric evaluation was performed. The relationship between psychometric test results and gene polymorphism was evaluated. This study shows that, similar to the literature, patients having cyanotic CHD have worse scores than acyanotic patients, and the children with CHD are under risk in terms of neuropsychiatric disorders. Other novel and important findings of this study were the lower verbal scores of ε2 allele carriers than ε4 carriers in Wechsler Intelligence Scale for Children-Revised group and the worse test score of patients having VSD than other acyanotic patients. Besides, some special disorders may be seen in this patient group.

Is cervical swab an efficient method for developing a new noninvasive prenatal diagnostic test for numerical and structural chromosome anomalies?

Background/aim: Prenatal diagnosis is vital to obtain healthy generation for risky pregnancies. There have been several approaches, some of which are routinely applied in clinics to evaluate the possible prenatal deficiencies and/or diseases. In the present study, we aimed to isolate the fetal cells from endocervical samples and try to identify possible anomalies which were proved by Amniocentesis (AS) and chorionic villus sampling (CVS) methods. Materials and methods: Endoservical specimens were collected from 100 pregnant women. Cells were separated in parallel by fluorescence-activated cell sorting (FACS) and magnetic-activated cell sorting (MACS) using human leukocyte antigen (HLA) G233 and placental alkaline phosphatase (PLAP) antibodies. CMA (comprehensive meta-analysis) were carried out and male fetuses were confirmed with Sex determining region Y (SRY) amplification. Results: The percent of HLA G233 and placental and placental alkaline phosphatase (PLAP) positive cells were 4.55% and 84.59%, respectively. The percent of cells positive for both markers was 14.75%. CMA analyses were not informative. (SRY) was amplified in 67% of the samples. Conclusion: However, the success rate of the both cell sorting and scanning of DNA anomalies by aCGH and/or RT-PCR was limited, preventing the applicability of this proposal in the clinics. Still, the success of the proposed method depends on the development of the novel fetal cell-specific antibodies and the improvements in the sorting systems.

Inherited eye diseases in Turkey: Current approaches and future directions.

The aim of this review is to reveal Turkey's current status of medical practice in inherited eye diseases and the necessary steps to improve healthcare services and research activities in this area. Since consanguinity rate is high, disease burden is estimated to be high in Turkey. Universal health insurance system, easily accessible medical specialists, increasing genetic test, and counseling opportunities are the key advantages of Turkey's healthcare system. However, specialized clinics for inherited eye diseases, low-vision rehabilitation services, training of ophthalmologists about the recent developments in ocular genetics, and multidisciplinary translational research are the main headlines needed to be focused for better health services and successful research in Turkey.

Identification of small-molecule urea derivatives as novel NAMPT inhibitors via pharmacophore-based virtual screening.

Nicotinamide phosphoribosyltransferase (NAMPT) catalyzes the condensation of nicotinamide (NAM) with 5-phosphoribosyl-1-prophosphate (PRPP) to yield nicotinamide mononucleotide (NMN), a rate limiting enzyme in a mammalian salvage pathway of nicotinamide adenine dinucleotide (NAD+) synthesis. Recently, intracellular NAD+ has received substantial attention due to the recent discovery that several enzymes including poly(ADP-ribose) polymerases (PARPs), mono(ADP-ribose) transferases (ARTs), and sirtuins (SIRTs), use NAD+ as a substrate, suggesting that intracellular NAD+ level may regulate cytokine production, metabolism, and aging through these enzymes. NAMPT is found to be upregulated in various types of cancer, and given its importance in the NAD+ salvage pathway, NAMPT is considered as an attractive target for the development of new cancer therapies. In this study, the reported NAMPT inhibitors bearing amide, cyanoguanidine, and urea scaffolds were used to generate pharmacophore models and pharmacophore-based virtual screening studies were performed against ZINC database. Following the filtering steps, ten hits were identified and evaluated for their in vitro NAMPT inhibitory effects. Compounds GF4 (NAMPT IC50 = 2.15 ± 0.22 µM) and GF8 (NAMPT IC50 = 7.31 ± 1.59 µM) were identified as new urea-typed inhibitors of NAMPT which also displayed cytotoxic activities against human HepG2 hepatocellular carcinoma cell line with IC50 values of 15.20 ± 1.28 and 24.28 ± 6.74 µM, respectively.

Genetic analysis of CHST6 and TGFBI in Turkish patients with corneal dystrophies: Five novel variations in CHST6.

PURPOSE: To identify pathogenic variations in carbohydrate sulfotransferase 6 (CHST6) and transforming growth factor, beta-induced (TGFBI) genes in Turkish patients with corneal dystrophy (CD). METHODS: In this study, patients with macular corneal dystrophy (MCD; n = 18), granular corneal dystrophy type 1 (GCD1; n = 12), and lattice corneal dystrophy type 1 (LCD1; n = 4), as well as 50 healthy controls, were subjected to clinical and genetic examinations. The level of antigenic keratan sulfate (AgKS) in the serum samples of patients with MCD was determined with enzyme-linked immunosorbent assay (ELISA) to immunophenotypically subtype the patients as MCD type I and MCD type II. DNA was isolated from venous blood samples from the patients and controls. Variations were analyzed with DNA sequencing in the coding region of CHST6 in patients with MCD and exons 4 and 12 in TGFBI in patients with LCD1 and GCD1. Clinical characteristics and the detected variations were evaluated to determine any existing genotype-phenotype correlations. RESULTS: The previously reported R555W mutation in TGFBI was detected in 12 patients with GCD1, and the R124C mutation in TGFBI was detected in four patients with LCD1. Serum AgKS levels indicated that 12 patients with MCD were in subgroup I, and five patients with MCD were in subgroup II. No genetic variation was detected in the coding region of CHST6 for three patients with MCD type II. In other patients with MCD, three previously reported missense variations (c. 1A>T, c.738C>G, and c.631 C>T), three novel missense variations (c.164 T>C, c.526 G>A, c. 610 C>T), and two novel frameshift variations (c.894_895 insG and c. 462_463 delGC) were detected. These variations did not exist in the control chromosomes, 1000 Genomes, and dbSNP. CONCLUSIONS: This is the first molecular analysis of TGFBI and CHST6 in Turkish patients with different types of CD. We detected previously reported, well-known hot spot mutations in TGFBI in the patients with GCD1 and LCD1. Eight likely pathogenic variations in CHST6, five of them novel, were reported in patients with MCD, which enlarges the mutational spectrum of MCD.

The Role of FOXP3 Polymorphisms in Graves' Disease with or without Ophthalmopathy in a Turkish Population.

Objectives: Forkhead box P3 (FOXP3) gene polymorphisms have been evaluated in many autoimmune diseases, including Graves' disease (GD), in different populations. However, those polymorphisms have not been analyzed in GD or Graves' ophthalmopathy (GO) in the Turkish population. In this study, we aimed to evaluate the frequency of FOXP3 polymorphisms in GD with or without ophthalmopathy in a Turkish population. Materials and Methods: The study included 100 patients with GO, 74 patients with GD without ophthalmopathy, and 100 age- and sex-matched healthy controls. In all study participants, rs3761547 (-3499 A/G), rs3761548 (-3279 C/A), and rs3761549 (-2383 C/T) single nucleotide polymorphisms (SNPs) were detected using the polymerase chain reaction-restriction fragment length polymorphism method. The chi-square test was used to evaluate genotype and allele frequencies. Odds ratios and 95% confidence intervals were calculated for genotype and allele risks. Results: In the patient group (including GD with or without ophthalmopathy), the rs3761548 AC and AA genotype and rs3761549 CT genotype were significantly more frequent than in the control group (all p<0.05). No genotypic and allelic differences were observed for rs3761547 between the patient and control groups (all p>0.05). There was no statistically significant difference between the GO and GD without ophthalmopathy groups concerning the allele and genotype frequencies of all three FOXP3 SNPs (all p>0.05). Conclusion: The AC and AA genotypes of rs3761548 (-3279) and CT genotype of rs3761549 (-2383 C/T) were shown to be possible risk factors for GD development in the Turkish population. However, none of the three SNPs was shown to be associated with the development of GO in patients with GD.

Vicinal Diaryl-Substituted Isoxazole and Pyrazole Derivatives with In Vitro Growth Inhibitory and In Vivo Antitumor Activity.

The vicinal diaryl heterocyclic framework has been widely used for the development of compounds with significant bioactivities. In this study, a series of diaryl heterocycles were designed and synthesized based on an in-house diaryl isoxazole derivative (9), and most of the newly synthesized derivatives demonstrated moderate to good antiproliferative activities against a panel of hepatocellular carcinoma and breast cancer cells, exemplified with the diaryl isoxazole 11 and the diaryl pyrazole 85 with IC50 values in the range of 0.7-9.5 µM. Treatments with both 11 and 85 induced apoptosis in these tumor cells, and they displayed antitumor activity in vivo in the Mahlavu hepatocellular carcinoma and the MDA-MB-231 breast cancer xenograft models, indicating that these compounds could be considered as leads for further development of antitumor agents. Important structural features of this compound class for the antitumor activity have also been proposed, which warrant further exploration to guide the design of new and more potent diaryl heterocycles.

EGF-SNX3-EGFR axis drives tumor progression and metastasis in triple-negative breast cancers.

Epidermal growth factor receptor (EGFR) has critical roles in epithelial cell physiology. Over-expression and over-activation of EGFR have been implicated in diverse cancers, including triple-negative breast cancers (TNBCs), prompting anti-EGFR therapies. Therefore, developing potent therapies and addressing the inevitable drug resistance mechanisms necessitates deciphering of EGFR related networks. Here, we describe Sorting Nexin 3 (SNX3), a member of the recycling retromer complex, as a critical player in the epidermal growth factor (EGF) stimulated EGFR network in TNBCs. We show that SNX3 is an immediate and sustained target of EGF stimulation initially at the protein level and later at the transcriptional level, causing increased SNX3 abundance. Using a proximity labeling approach, we observed increased interaction of SNX3 and EGFR upon EGF stimulation. We also detected colocalization of SNX3 with early endosomes and endocytosed EGF. Moreover, we show that EGFR protein levels are sensitive to SNX3 loss. Transient RNAi models of SNX3 downregulation have a temporary reduction in EGFR levels. In contrast, long-term silencing forces cells to recover and overexpress EGFR mRNA and protein, resulting in increased proliferation, colony formation, migration, invasion in TNBC cells, and increased tumor growth and metastasis in syngeneic models. Consistent with these results, low SNX3 and high EGFR mRNA levels correlate with poor relapse-free survival in breast cancer patients. Overall, our results suggest that SNX3 is a critical player in the EGFR network in TNBCs with implications for other cancers dependent on EGFR activity.

Identification of an mRNA isoform switch for HNRNPA1 in breast cancers.

Roles of HNRNPA1 are beginning to emerge in cancers; however, mechanisms causing deregulation of HNRNPA1 function remain elusive. Here, we describe an isoform switch between the 3'-UTR isoforms of HNRNPA1 in breast cancers. We show that the dominantly expressed isoform in mammary tissue has a short half-life. In breast cancers, this isoform is downregulated in favor of a stable isoform. The stable isoform is expressed more in breast cancers, and more HNRNPA1 protein is synthesized from this isoform. High HNRNPA1 protein levels correlate with poor survival in patients. In support of this, silencing of HNRNPA1 causes a reversal in neoplastic phenotypes, including proliferation, clonogenic potential, migration, and invasion. In addition, silencing of HNRNPA1 results in the downregulation of microRNAs that map to intragenic regions. Among these miRNAs, miR-21 is known for its transcriptional upregulation in breast and numerous other cancers. Altogether, the cancer-specific isoform switch we describe here for HNRNPA1 emphasizes the need to study gene expression at the isoform level in cancers to identify novel cases of oncogene activation.

Intelligent Ratio: A New Method for Carrier and Newborn Screening in Spinal Muscular Atrophy.

Aim: Spinal muscular atrophy (SMA) is an inherited, autosomal recessive neuromuscular disease that causes high morbidity and mortality. The prevalence is 1-2/100,000, while the incidence is 1/6000-1/10,000 among live births. Due to the high carrier frequency (1/40-1/60) of SMA-associated alleles, screening can prevent new cases. The aim of the current study was to present the development of a new, quantitative, real-time, polymerase chain reaction (PCR)-based screening test that uses an intelligent ratio (IR) for analyses, as well as a comparison of the results with the gold standard. Materials and Methods: Included in the study were 100 patients with various risk genotypes for survivor motor neuron 1 (SMN1) and SMN2 genes whose genetics had been previously investigated using multiplex ligation probe amplification (MLPA). A combination of the 5' nuclease assay and allele-specific PCR was used to quantify the SMN1 deletion mutation with real-time PCR using the FII gene as a reference. All of the optimized standards were adapted to software that provided automated analyses. The approval number of the institutional ethics committee for the study is 2012-KAEK-15/1497. Results: The results of the screening test were completely compatible with the MLPA results; it achieved 100% sensitivity and specificity compared with the gold standard. The use of the IR in the analyses provided a user-independent method that quickly and accurately provided results, regardless of the amount of DNA used of the extraction method. Conclusion: Carrier or newborn screening of SMA is essential in countries that have high rates of consanguineous marriages. The screening test presented in this study that uses FII as a reference gene proved to be low-cost, reliable, applicable, accurate, and amenable to use in an automated system for SMA screening.

Pharmacophore modeling and virtual screening studies to identify novel selective SIRT2 inhibitors.

Sirtuins (SIRTs) are a class of NAD+-dependent protein histone deacetylases (HDACs) that catalyse the reversible deacetylation of lysine residues in the histones or non-histone substrates. Mammalian sirtuins consist of seven isoforms (SIRT1-7), which show different subcellular localizations and enzymatic functions. Among the seven human sirtuins, SIRT2 predominantly located in the cytoplasm but is enriched in the nucleus during mitosis. Its activity has been found to be modulate the pathophysiology of various diseases such as cancer, metabolic and neurodegenerative disorders. Therefore, selective SIRT2 inhibitors are of growing interest as potentially candidate therapeutic agents to treat SIRT2-driven pathologies as well as valuable tools to investigate and define the biological roles of SIRT2. Herein, in order to identify potent leads against SIRT2, a multi-step pharmacophore based-virtual screening campaign was performed and 31 predicted compounds were subjected to in vitro biological evaluation. Finally, compound 2 and 3 showing better SIRT2 inhibition potency were selected for further in vitro cytotoxic assays against a panel of three human cancer cell lines. This study will hopefully provide a basis for developing potent and selective SIRT2 inhibitors.

Single-Nucleotide Polymorphisms in IL23R-IL12RB2 (rs1495965) Are Highly Prevalent in Patients with Behcet's Uveitis and Vary Between Populations.

Purpose: To test the frequency of single-nucleotide polymorphisms in the IL-10, IL23R-IL12RB2 genes in patients with Behcet's uveitis. Methods: Blood samples were collected from 89 Israeli and Turkish patients, and from healthy control subjects of different origins. Genomic DNA was extracted from peripheral blood leukocytes and genotyped. Results: The risk allele, A, in rs1800871, of IL-10 gene was highly prevalent in Behcet's uveitis and healthy control samples alike; highest among the Turkish groups. Prevalence of G allele, in rs1495965, in the IL23R-IL12RB2 gene was high in Behcet's uveitis patients, and among healthy Turkish and Israelis of Middle Eastern origin, while lower among the other Israeli control group (77.9%, 78.9%, 27.8%, respectively, P < 0.001). Conclusion: Our findings highlight the differences between populations and may account for the increased prevalence of the disease among Turkish and Israelis of Middle Eastern origin. Further studies are required to map other healthy and affected populations.

The relation between isolated micropenis in childhood with CAG and GGN repeat polymorphisms in the androgen receptor gene

Background/aim: In micropenis cases accompanied by external genital abnormalities such as hypospadias and cryptorchidism, infertility and spermatogenic failures have been reported to correlate with androgen receptor ( AR ) gene CAG and GGN repeat polymorphisms. While there is one study on isolated micropenis and CAG repeats, no study related to GGN repeats has been reported. We investigated the relation between CAG and GGN repeats in the AR gene with development of penis length in boys with isolated micropenis. Materials and methods: A total of 24 Turkish boys with isolated micropenis (<­2.5 SD) and 64 healthy controls who had normal basal serum gonadotropin levels were examined. Genotyping was performed by DNA sequencing of the patients and controls. Results: The distribution of CAG and GGN repeat lengths in our patients and controls was within the normal range and did not significantly differ between the patients and the controls. Conclusion: CAG repeat length in the AR constitutes one of multiple genetic factors relevant to the development of isolated micropenis, and the expansion of this repeat can be detected as a likely modifying factor. Moreover, the interactions of other genes that may be involved in the etiology of isolated micropenis with CAG and GGN repeats have to be taken into consideration.

Influence of Folate-Related Gene Polymorphisms on High-Dose Methotrexate-Related Toxicity and Prognosis in Turkish Children with Acute Lymphoblastic Leukemia.

OBJECTIVE: High-dose methotrexate (HD-MTX) is widely used in the consolidation phase of childhood acute lymphoblastic leukemia (ALL), but the roles that polymorphisms in folate-related genes (FRGs) play in HD-MTX toxicity and prognosis in children with ALL are not understood. The aims of this study were to investigate the frequencies of polymorphisms in the genes for thymidylate synthase (TS), methionine synthase reductase (MTRR), and methylene tetrahydrofolate reductase (MTHFR) in Turkish children with ALL and to assess associations between these polymorphisms and HD-MTX-related toxicity and leukemia prognosis in this patient group. MATERIALS AND METHODS: FRG polymorphisms were assessed by real-time polymerase chain reaction. Survival status, MTX levels, and toxicity data were retrieved from 106 patients' charts. RESULTS: The allele frequencies for the FRG polymorphisms were as follows: TS 2R 41.0%, 3R 57.0%, and 4R 2.0%; MTRR 66A 42.4% and 66G 57.6%; MTHFR 677C 59.3% and 677T 40.7%; and MTHFR 1298A 58.1% and 1298C 41.9%. At the 48th hour of HD-MTX infusion, serum MTX was significantly higher in patients who had TS 2R/3R/4R variants as compared to those with wild-type TS (p<0.05). No significant differences were detected with respect to event-free survival or toxicity between wild-type and other FRG variants. CONCLUSION: The frequencies of FRG polymorphisms in Turkish children with ALL are similar to those reported in other Caucasian populations. This is the first published finding of the TS 3R/4R variant in the Turkish population. The results indicate that HD-MTX can be tolerated by leukemic children with some polymorphic variants of FRG; thus, it may prevent future risk of leukemic relapse.