Endothelin-converting Enzyme-1b Genetic Variants Increase the Risk of Coronary Artery Ectasia.

Coronary artery ectasia (CAE), defined as a 1.5-fold or greater enlargement of a coronary artery segment compared to the adjacent normal coronary artery, is frequently associated with atherosclerotic coronary artery disease (CAD). Membrane-bound endothelin converting enzyme-1 (ECE-1) is involved in the maturation process of the most potent vasoconstrictor ET-1. Polymorphisms in the endothelin (ET) gene family have been shown associated with the development of atherosclerosis. This study aims to investigate the effects of rs213045 and rs2038089 polymorphisms in the ECE-1 gene which have been previously shown to be associated with atherosclerosis and hypertension (HT), in CAE patients. Ninety-six CAE and 175 patients with normal coronary arteries were included in the study. ECE-1b gene variations rs213045 and rs2038089 were determined by real-time PCR. The frequencies of rs213045 C > A (C338A) CC genotype (60.4% vs. 35.4%, p < 0.001) and rs2038089 T > C T allele (64.58% vs. 35.42%, p = 0.017) were higher in the CAE group compared to the control group. The multivariate regression analysis showed that the ECE-1b rs213045 CC genotype (p = 0.001), rs2038089 T allele (p = 0.017), and hypercholesterolemia (HC) (p = 0.001) are risk factors for CAE. Moreover, in nondiabetic individuals of the CAE and control groups, it was observed that the rs213045 CC genotype (p < 0.001), and rs2038089 T allele (p = 0.003) were a risk factor for CAE, but this relationship was not found in the diabetic subgroups of the study groups (p > 0.05). These results show that ECE-1b polymorphisms may be associated with the risk of CAE and this relationship may change according to the presence of type II diabetes.

Next-generation sequencing of prolidase gene identifies novel and common variants associated with low prolidase in coronary artery ectasia.

BACKGROUND: Decreased collagen biosynthesis and increased collagenolysis can cause ectasia progression in the arterial walls. Prolidase is a key enzyme in collagen synthesis; a decrease in prolidase activity or level may decrease collagen biosynthesis, which may contribute to ectasia formation. Considering that, the variations in PEPD gene encoding prolidase enzyme were evaluated by analyzing next-generation sequencing (NGS) for the first time together with known risk factors in coronary artery ectasia (CAE) patients. METHODS: Molecular analysis of the PEPD gene was performed on genomic DNA by NGS in 76 CAE patients and 76 controls. The serum levels of prolidase were measured by the sandwich-ELISA technique. RESULTS: Serum prolidase levels were significantly lower in CAE group compared to control group, and it was significantly lower in males than females in both groups (p < 0.001). On the other hand, elevated prolidase levels were observed in CAE patients in the presence of diabetes (p < 0.001), hypertension (p < 0.05) and hyperlipidemia (p < 0.05). Logistic regression analysis demonstrated that the low prolidase level (p < 0.001), hypertension (p < 0.02) and hyperlipidemia (p < 0.012) were significantly associated with increased CAE risk. We identified four missense mutations in the PEPD gene, namely G296S, T266A, P365L and S134C (novel) that could be associated with CAE. The pathogenicity of these mutations was predicted to be "damaging" for G296S, S134C and P365L, but "benign" for T266A. We also identified a novel 5'UTR variation (Chr19:34012748 G>A) in one patient who had a low prolidase level. In addition, rs17570 and rs1061338 common variations of the PEPD gene were associated with low prolidase levels in CAE patients, while rs17569 variation was associated with high prolidase levels in both CAE and controls (p < 0.05). CONCLUSIONS: Our findings indicate that the low serum prolidase levels observed in CAE patients is significantly associated with PEPD gene variations. It was concluded that low serum prolidase level and associated PEPD mutations may be potential biomarkers for the diagnosis of CAE.

Bi-allelic Pathogenic Variants in TUBGCP2 Cause Microcephaly and Lissencephaly Spectrum Disorders.

Lissencephaly comprises a spectrum of malformations of cortical development. This spectrum includes agyria, pachygyria, and subcortical band heterotopia; each represents anatomical malformations of brain cortical development caused by neuronal migration defects. The molecular etiologies of neuronal migration anomalies are highly enriched for genes encoding microtubules and microtubule-associated proteins, and this enrichment highlights the critical role for these genes in cortical growth and gyrification. Using exome sequencing and family based rare variant analyses, we identified a homozygous variant (c.997C>T [p.Arg333Cys]) in TUBGCP2, encoding gamma-tubulin complex protein 2 (GCP2), in two individuals from a consanguineous family; both individuals presented with microcephaly and developmental delay. GCP2 forms the multiprotein γ-tubulin ring complex (γ-TuRC) together with γ-tubulin and other GCPs to regulate the assembly of microtubules. By querying clinical exome sequencing cases and through GeneMatcher-facilitated collaborations, we found three additional families with bi-allelic variation and similarly affected phenotypes including a homozygous variant (c.1843G>C [p.Ala615Pro]) in two families and compound heterozygous variants consisting of one missense variant (c.889C>T [p.Arg297Cys]) and one splice variant (c.2025-2A>G) in another family. Brain imaging from all five affected individuals revealed varying degrees of cortical malformations including pachygyria and subcortical band heterotopia, presumably caused by disruption of neuronal migration. Our data demonstrate that pathogenic variants in TUBGCP2 cause an autosomal recessive neurodevelopmental trait consisting of a neuronal migration disorder, and our data implicate GCP2 as a core component of γ-TuRC in neuronal migrating cells.

CDK6 is an essential direct target of NUP98 fusion proteins in acute myeloid leukemia.

Fusion proteins involving Nucleoporin 98 (NUP98) are recurrently found in acute myeloid leukemia (AML) and are associated with poor prognosis. Lack of mechanistic insight into NUP98-fusion-dependent oncogenic transformation has so far precluded the development of rational targeted therapies. We reasoned that different NUP98-fusion proteins deregulate a common set of transcriptional targets that might be exploitable for therapy. To decipher transcriptional programs controlled by diverse NUP98-fusion proteins, we developed mouse models for regulatable expression of NUP98/NSD1, NUP98/JARID1A, and NUP98/DDX10. By integrating chromatin occupancy profiles of NUP98-fusion proteins with transcriptome profiling upon acute fusion protein inactivation in vivo, we defined the core set of direct transcriptional targets of NUP98-fusion proteins. Among those, CDK6 was highly expressed in murine and human AML samples. Loss of CDK6 severely attenuated NUP98-fusion-driven leukemogenesis, and NUP98-fusion AML was sensitive to pharmacologic CDK6 inhibition in vitro and in vivo. These findings identify CDK6 as a conserved, critical direct target of NUP98-fusion proteins, proposing CDK4/CDK6 inhibitors as a new rational treatment option for AML patients with NUP98-fusions.

Peroxisome Proliferator-Activated Receptor Gamma Pro12Ala/C161T Genotypes and Risky Haplotype Altering Risk of Breast Cancer: A Turkish Case-Control Study.

Breast cancer (BC) has a high incidence rate among women worldwide, and the mechanisms and etiology of this disease are not yet fully understood. The peroxisome proliferator-activated receptor gamma (PPARgamma), a nuclear hormone receptor that plays important roles in energy metabolism and cellular differentiation, is also suggested to be effective in cancer development. However, the results of studies investigating the cancer association with PPARgamma are inconsistent, creating a need for further investigation of the effects of this transcription factor on BC risk. We have examined the Pro12Ala-(rs1801282) and C161T-(rs3856806) polymorphisms of the PPARgamma gene in Turkish patients with BC in this case-control study. A total of 95 women diagnosed with BC as cases and 119 controls were genotyped for PPARgamma polymorphisms by polymerase chain reaction and restriction fragment length polymorphism techniques. The ProPro genotype and T161 allele were associated with an increased risk of BC comparing with the Ala12 allele and CC161 genotype, respectively (p < 0.001). The multivariate regression analysis confirmed that the ProPro genotype (p < 0.011), T161 allele (p < 0.001), smoking (p = 0.019), and advanced age (> 60 years) (p = 0.007) are risk factors for breast cancer. We also found that the PPARgamma Pro12Ala and C161T polymorphisms were in linkage disequilibrium (D':0.511, r2:0.099). It was determined that carrying ProPro-T161 risky PPARgamma haplotype was associated with a higher risk of BC compared to protective Ala12-CC161 haplotype (p < 0.01, OR:7.797, 95% CI:3.521-17.263). We concluded that PPARgamma Pro12Ala and C161T polymorphisms are associated with increased BC risk, and ProPro-T161 risky haplotype, which is in linkage disequilibrium, increases this effect.

Interactive effects of interferon-gamma functional single nucleotid polymorphism (+874 T/A) with cardiovascular risk factors in coronary heart disease and early myocardial infarction risk.

Atherosclerosis is an inflammatory disease characterized by extensive lipid accumulation in the artery wall. Throughout the atherosclerotic process, interferon-gamma (IFN-γ), which is an important pro-inflammatory cytokine, plays a central role in atherosclerotic plaque instability and the occurrence of myocardial infarction (MI). In this study, we aimed to investigate the relationship between IFN-γ +874 T/A (rs2430561) polymorphism and coronary heart disease (CHD) as well as its effects on MI and CHD. Three hundred and ninety patients with CHD (229 with MI, 161 without MI) and 233 healthy controls were screened by the amplification refractory mutation system (ARMS) PCR method for IFN-γ +874 T/A polymorphism. For MI risk, early adult age was important risk factors and the risk was increased with IFN-γ +874 T/A polymorphism. IFN-γ T allele was significantly increased in the CHD patients with age≤45 (p = 0.048) and patients with history of MI (p = 0.007). As IFN-γ is an inflammatory cytokine with an emerging role in the atherosclerotic process, it was suggested that inhibition of IFN-γ activity could be a therapeutic strategy to stabilize human atherosclerotic plaque. Our findings support the association between MI risk and IFN-γ +874 T/A polymorphism in the Turkish population, particularly by increasing the level of IFN-γ in young patients, thereby causing rupture of vulnerable plaques in atherosclerotic lesions. Identification of the IFN-γ +874 T/A gene variants as risk factors for early CHD and MI development may be a practical biomarker to guide the MI risk process and determine the ideal therapeutic approach.

Receptor for advanced glycation end products polymorphisms in coronary artery ectasia.

BACKGROUND: Although the implication of receptor of advanced glycation endproducts (RAGE) has been reported in coronary artery disease, its roles in coronary artery ectasia (CAE) have remained undetermined. Furthermore, the effect of RAGE polymorfisms were not well-defined in scope of soluble RAGE (sRAGE) levels. Thus, we aimed to investigate the influence of the functional polymorphisms of RAGE -374T > A (rs1800624) and G82S (rs2070600) in CAE development. METHODS: This prospective observational study was conducted in 2 groups selected of 2452 patients who underwent elective coronary angiography (CAG) for evaluation after positive noninvasive heart tests. Group-I included 98 patients with non-obstructive coronary artery disease and CAE, and Group-II (control) included 100 patients with normal coronary arteries. SNPs were genotyped by real-time PCR using Taqman® genotyping assay. Serum sRAGE and soluble lectin-like oxidized receptor-1 (sOLR1) were assayed by ELISA and serum lipids were measured enzymatically. RESULTS: The frequencies of the RAGE -374A allele and -374AA genotype were significantly higher in CAE patients compared to controls (p < 0.001). sRAGE levels were not different between study groups, while sOLR1 levels were elevated in CAE (p = 0.004). In controls without systemic disease, -374A allele was associated with low sRAGE levels (p < 0.05), but this association was not significant in controls with HT. Similarly, sRAGE levels of CAE patients with both HT and T2DM were higher than those no systemic disease (p = 0.02). The -374A allele was also associated with younger patient age and higher platelet count in the CAE group in both total and subgroup analyses. In the correlation analyses, the -374A allele was also negatively correlated with age and positively correlated with Plt in all of these CAE groups. In the total CAE group, sRAGE levels also showed a positive correlation with age and a negative correlation with HDL-cholesterol levels. On the other hand, a negative correlation was observed between sRAGE and Plt in the total, hypertensive and no systemic disease control subgroups. Multivariate logistic regression analysis confirmed that the -374A allele (p < 0.001), hyperlipidemia (p < 0.05), and high sOLR1 level (p < 0.05) are risk factors for CAE. ROC curve analysis shows that RAGE -374A allele has AUC of 0.713 (sensitivity: 83.7 %, specificity: 59.0 %), which is higher than HLD (sensitivity: 59.2 %, specificity: 69.0 %), HT (sensitivity: 62.4 %, specificity: 61.1 %) and high sOLR1 level (≥0.67 ng/ml)) (sensitivity: 59.8 %, specificity: 58.5 %). CONCLUSION: Beside the demonstration of the relationship between -374A allele and increased risk of CAE for the first time, our results indicate that antihypertensive and antidiabetic treatment in CAE patients causes an increase in sRAGE levels. The lack of an association between the expected -374A allele and low sRAGE levels in total CAE group was attributed to the high proportion of hypertensive patients and hence to antihypertensive treatment. Moreover, the RAGE -374A allele is associated with younger age at CAE and higher Plt, suggesting that -374A may also be associated with platelet activation, which plays a role in the pathogenesis of CAE. However, our data need to be confirmed in a large study for definitive conclusions.

Unusual effects of PCSK9 E670G (rs505151) variation in patients with in-stent restenosis: Variable effects on restenosis risk according to concomitant chronic conditions.

Recent reports showing that neo-atherosclerosis formation in stented coronary artery is characterized by the accumulation of lipid-laden macrophages within the neointima has strengthened the possibility that elevated low-density lipoprotein (LDL)-cholesterol may be a risk factor for in-stent restenosis (ISR). Protein Convertase Subtilisin/Kexin-9 (PCSK9) protein plays an important role in cholesterol metabolism by degrading of LDL receptors. The gain-of-function E670G (rs505151) mutation of the PCSK9 gene is a well-known genetic risk factor for hypercholesterolemia. This study evaluated for the first time the association of the E670G variation with the serum lipids, PCSK9 levels and concomitant diseases on the ISR risk. The study included 109 ISR, and 82 Non-ISR patients, based on the results of coronary angiography. Genotypes were determined using the real-time PCR and serum PCSK9 levels were measured by ELISA technique. The rare G allele of PCSK9 E670G (p < 0.05), hyperlipidemia (HL) (p < 0.001), and type 2 diabetes (T2DM) (p < 0.01) were associated with increased risk for ISR. In hyperlipidemic conditions, the E670G-G allele was associated with hypercholesterolemia and a higher risk of ISR (p < 0.001), while the E670G-AA genotype has been associated with a high prevalence of T2DM and hypertension. In addition, diabetic ISRs had higher serum PCSK9 levels (p < 0.05) and the E670G-AA genotype was associated with increased levels of diabetes markers. Our results indicated that the unusual effects of both G allele and AA genotype of the PCSK9 E670G variation may be involved in the risk of ISR in association with concomitant metabolic diseases.

This study evaluated the association of the Protein Convertase Subtilisin/Kexin-9 (PCSK9) E670G mutation with the serum lipids, PCSK9 levels and concomitant diseases on the in-stent restenosis (ISR) risk. The E670G-G allele, hyperlipidemia, and type 2 diabetes (T2DM) were found risk factors for ISR. In hyperlipidemic conditions, the E670G-G allele was associated with hypercholesterolemia and a higher risk of ISR, while the E670G-AA genotype has been associated with a high prevalence of T2DM and hypertension. Our results indicated that the unusual effects of both genotypes of the E670G that may be involved in the ISR risk in association with concomitant diseases.

A New Promising Pathway in Aggressive Prostate Cancer: Treg/mir-let8c/lin28b.

PURPOSE: The progress of prostate cancer entails complex contemporaneous tumor developmental events in diverse stages that they are still yet to be clarified. miRNAs might accompany to balance between regulatory and cytotoxic T cells in tumors. Here, we investigated miRNAs and Regulatory T cell (Treg) marker FOXP3 expressions within prostate cancer spectrum. METHODS: Thirty-eight prostate cancer patients enrolled within two groups to the study as having Gleason Score ≤ 7 (Group-1) and ≥ 8 (Group-2) that compared to 19 benign prostate hyperplasia controls. Twelve miRNAs expressions were analyzed by real time PCR from paraffin-embedded prostate tissue samples. Correlations between serum PSA levels, immunohistochemical staining of CD3, CD4, FOXP3 and miRNA expressions were analyzed. RESULTS: In our study, hsa-let7c-3p significantly 1,52 (p=0.018) and 1,84 (p=0.0095) fold down- regulated whereas, miR-141-3p was significantly 2,36 (p=0.0006) and 2,24 (p=0.001) fold upregulated in the prostate cancer patients compared to benign prostate hyperplasia in group 1 and 2, respectively. Only CD4 (p=0.004) and PSA (p<0.001) have statistically significant differences among groups when compared to benign prostate hyperplasia. miR-143-p, miR-221-3p, hsa-let7c-3p and miR-17-3p expressions were significantly correlated with regulatory T cell marker FOXP3 expression. CONCLUSIONS: For the first time, we reported significantly altered expression levels of miRNAs (miR-let7c, miR221, miR-146a, miR-141, miR-143, miR17) and correlations between Treg marker FOXP3 in the aggressive prostate cancer patients suggesting that prostate cancer progression might be under the regulation of crosstalk between Tregs and miRNAs.

The comparison of serum TGF-beta levels and associated polymorphisms in patients with coronary artery ectasia and normal coronary artery.

BACKGROUND: Coronary artery ectasia (CAE) is described as the enlargement of a coronary artery segment by 1.5 times or more, which is generally associated with the atherosclerotic process. Atherosclerotic changes lead to arterial remodeling result in CAE. In our study, we measured serum transforming growth factor (TGF)-ß1 levels, which have a protective role against atherosclerosis. Further, we aimed to assess the TGF-ß1 gene variants rs1800469 (-509C>T, c.-1347C>T) and rs1800470 (c.+29T>C, p.Pro10Leu, rs1982073), which might have an effect on TGF production. Overall, 2877 patients were screened including 56 patients with CAE and 44 patients with normal coronary arteries who were included in the study. Serum TGF-ß1 levels were measured using ELISA and compared between two groups. Additionally, TGF-ß1 rs1800469 and rs1800470 gene variations were determined using TaqMan® SNP Genotyping Assays. RESULTS: Serum TGF-ß1 levels were significantly lower in patients with CAE than in controls (p=0.012). However, there was no difference in terms of the genotype and allele distributions of TGF-ß1 rs1800469 and rs1800470 polymorphisms. Serum TGF-ß1 levels were higher in individuals carrying the TGF-ß1 rs1800470 G allele (GG+AG) than in individuals with normal homozygous AA genotype in the CAE group (p=0.012). CONCLUSION: Our findings suggest that lower serum TGF-ß1 levels are associated with an increased risk for CAE development and that TGF-ß1 polymorphisms exert a protective effect. Furthermore, TGF-ß1 rs1800470 G allele carriers were shown to have higher TGF-ß1 levels in the CAE group. This suggests that having the G allele in the TGF-ß1 rs1800470 polymorphism could prevent CAE development.

Ear atresia: Is there a role of apoptosis-regulating miRNAs?

OBJECTIVE: The molecular events underlying ear development involve numerous regulatory molecules; however, the role of microRNAs (miRNAs) has not been explored in patients with ear atresia. Here, we aimed to investigate the expressions of 20-22 nucleotide noncoding RNAs. METHODS: We selected 12 miRNAs that function to control post-transcriptional gene expression in different pathways, including apoptosis, angiogenesis, and chondrogenesis. The altered miRNA expressions were analyzed by real-time PCR from serum samples of 7 patients with ear atresia and 8 controls. RESULTS: We found that the expression of apoptosis-regulating miRNAs was significantly downregulated in patients with ear atresia. TThe expressions of miR126, miR146a, miR222, and miR21 were significantly decreased by 76.2-(p=0.041), 61.8-(p=0.000), 30.5-(p=0.009), and 71.21-fold (p=0.042), respectively, compared with controls. CONCLUSION: Abnormal ear development in ear atresia patients, could possibly be due to the reduced expression of apoptosis regulating miRNAs. Changes in the regulation of tumor protein p53 (TP53), p53 upregulated modulator of apoptosis (PUMA), Fas cell surface death receptor (FAS), FAS ligand (FasL), and phosphatase and tensin homolog (PTEN) directly or within the apoptosis-related cascades may play important roles during development, particularly in the external ear. This is the first report to present the possible association between apoptosis-regulating miRNAs and ear atresia/microtia.