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Lung cancer is one of the deadliest types of cancers and genetic and epigenetic alterations play major roles in its development. Chromodomain (CHD) protein family acts in chromatin organization, regulation of transcription and also genomic stability and cancer prevention. Although CHD5, a member of this family was shown to contribute to major cellular events and functions as a tumor suppressor gene in various types of cancer, it is not clear whether CHD5 plays a role in lung carcinogenesis. The aim of this study was to investigate the possible role of CHD5 in progression of non-small cell lung cancer (NSCLC). Expression levels of CHD5 gene in 59 tumor and corresponding non-cancerous lung tissue samples were analyzed by qRT-PCR and the methylation status of the promoter region was investigated by methylation specific PCR (MS-PCR). The Akt phosphorylation levels were investigated by Western Blot (WB). CHD5 was down-regulated in 17 (39.5%) and up-regulated in 24 (55.8%) of tumor specimens. Even though the promoter of CHD5 was hypermethylated in 8 patients, it was not found associated with CHD5 gene expression (p=0.08). Akt phosphorylation was increased in 14 (53.8%) and decreased in 12 (46.2%) of the samples but no significant association was found between p-Akt phosphorylation and CHD5 expression (p=0.67). We suggest that CHD5 may act as a tumor suppressor gene in NSCLC.

Background: Lung cancer is the leading cause of cancer deaths. The main risk factor is smoking but the risk is also associated with various genetic and epigenetic components in addition to environmental factors. Increases in the gene copy numbers due to chromosomal amplifications constitute a common mechanism for oncogene activation. A gene-dense region on chromosome 11q13 which harbors four core regions that are frequently amplified, has been associated with various types of cancer. The important cell cycle regulatory protein cyclin D1 (CCND1) is an essential driver of the first core region of the Chr11q13 amplicon. Deregulation of CCND1 has been associated with different kinds of human malignancies including lung cancer. The EMSY (c11orf30) gene has been proposed as the possible driver of the fourth core of the 11q13 amplicon and its amplification has been associated with breast and ovarian cancers. There is no report in the literature investigating the EMSY gene in lung cancer. Methods: In this study, expression levels of the EMSY and CCND1 genes were investigated in 85 patients with non small cell lung cancer by Real Time PCR. Results: Expression of the EMSY and CCND1 genes were increased in 56 (65.8%) and 50 (58.8%) of the patients, respectively. Both genes showed a higher expression in the tumors when compared to normal tissues. A strong correlation was present between the expression rates of both genes (p

Despite therapeutic advances, lung cancer remains one of the most common causes of cancer‐related deaths worldwide. The ZNF703 gene has been identified as the driver of the 8p11‐12 region and its amplification or overexpression has been associated with several types of cancers. It has also been shown that ZNF703 overexpression can activate the Akt/mTOR signaling pathway. The aim of our study was to investigate the role of the ZNF703 gene in association with Akt/mTOR activation in non‐small cell lung cancer (NSCLC). Expression levels in tumors and matched noncancerous tissue samples from 47 patients were analyzed by qRT‐PCR and the Akt phosphorylation levels were investigated by Western blotting. Our results show that ZNF703 is up‐regulated in 63.4% of NSCLC tumor samples. Althogh the correlation did not reach a statistically significant level Akt phosphorylation was increased in tumor tissues expressing high levels of ZNF703. The role of the ZNF703 gene has not been investigated in NSCLC. Our data show that ZNF703 may contribute to tumor development in NSCLC by activating the Akt/mTOR pathway.

The aim of this study was to investigate the relationships between the maternal levels of oxidized LDL (ox-LDL), lipid peroxidation marker malondialdehyde (MDA) and LOX-1 3'UTR188C/T and K167N single nucleotide polymorphisms in pregnant Turkish women with gestational diabetes mellitus (GDM).116 pregnant women with GDM and 120 healthy pregnant women from the same geographic region were included in the study. Polymerase chain reaction-based restriction analysis was used to identify 3'UTR188C/T and K167N polymorphisms of the LOX-1 gene. Plasma ox-LDL and MDA levels were determined by enzyme-linked immunosorbent assay and spectrophotometric method in all study subjects, respectively.Our results indicated that the distribution of the LOX-1 3'UTR188C/T and K167N genotypes and alleles did not differ significantly among subjects with or without GDM (p0.05). TT and NN genotype carriers are associated with some glucose metabolism parameters (p0.05). There were no significant differences among plasma ox-LDL and MDA levels with regard to LOX-1 3'UTR188C/T and K167N polymorphisms in GDM group and control subjects (p0.05). According to the combined genotype analysis of LOX-1 3'UTR 188 TT and K167N NN polymorphisms, plasma MDA and ox-LDL levels were significantly different between women with GDM and healthy subjects either with or without combined TT/NN genotype carriers (p0.001).According to our results, ox-LDL and MDA levels were increased in GDM pregnant women and healthy pregnant women either with or without combined TT/NN genotype carriers, for our Turkish sample, these genotype carriers appear to be related with increased oxidative stress in patients with GDM.

Chromosomal alterations are frequent events in lung carcinogenesis and usually display regions of focal amplification containing several overexpressed oncogenes. Although gains and losses of chromosomal loci have been reported copy number changes of the individual genes have not been analyzed in lung cancer. In this study 22 genes were analyzed by MLPA in tumors and matched normal tissue samples from 82 patients with non-small cell lung cancer. Gene amplifications were observed in 84% of the samples. Chromosome 8 was found to harbor the most frequent copy number alterations. The most frequently amplified genes were ZNF703, PRDM14 and MYC on chromosome 8 and the BIRC5 gene on chromosome 17. The frequency of deletions were much lower and the most frequently deleted gene was ADAM9. Amplification of the ZNF703, PRDM14 and MYC genes were highly correlated suggesting that the genes displaying high copy number changes on chromosome 8 collaborate during lung carcinogenesis.

Introduction Chronic obstructive pulmonary disease (COPD) is a common respiratory condition characterized by persistent airflow limitation and is associated with an enhanced chronic inflammatory response in the airways and the lung to noxious particles or gases. Interleukin-1 beta (IL-1β) is a major pro-inflammatory cytokine expressed by many cells such as macrophages, neutrophils and monocytes and functions in cellular activities such as proliferation, differentiation and apoptosis. Recent studies demonstrate controversial results about the relationship between IL-1β and COPD. The aim of this study is to investigate the association between IL-1β -511 (rs 16944) and +3954 (rs 1143634) gene polymorphisms and COPD in Turkish population. Materials and Methods A total of 152 subjects were recruited in the study and divided into three groups: 72 COPD patients, 41 healthy smokers and 39 never-smokers. PCR-RFLP method was used to determine the allele frequencies, genotype and haplotype distributions. Result We did not find any significant difference between the gene polymorphisms and COPD by means of genotype frequencies, haplotype association, stage, gender or smoking status (p< 0.05). Conclusions Our results do not show any evidence of association between COPD and IL-1β -511 and +3954 gene polymorphisms in Turkish population.

Radiotherapy (RT) may result in platelet activation and thrombosis development. To the best of our knowledge, the potential effect of volumetric-modulated arc therapy (VMAT), a novel radiotherapy technique, on platelet function and microRNA (miRNA/miR) expression has not been previously investigated. The present study aimed to determine the effect of VMAT on the alterations in platelet function parameters and miRNA expression levels. A total of 25 patients with prostate cancer and 25 healthy subjects were included in the present study. Blood samples were collected from the patient group on the day prior to RT (pre-RT), the day RT was completed (post-RT day 0), and 40 days following the end of therapy (post-RT day 40). Platelet count, mean platelet volume (MPV) value, platelet aggregation, plasma P-selectin, thrombospondin-1, platelet factor 4, plasma miR-223 and miR-126 expression levels were measured. A significant decrease in platelet count in the post-RT day 0 group was measured in comparison with the pre-RT and the post-RT day 40 groups. Pre-RT MPV values were higher than those of the post-RT day 0 and the post-RT day 40 groups. No significant differences were observed in the levels of platelet activation markers or miR-223 and miR-126 expression levels between the RT groups. Although RT may result in a reduction in platelet and MPV counts, the results of the present study indicate that platelet activation markers are not affected by VMAT. Therefore, it is possible that no platelet activation occurs during VMAT, owing to the conformal dose distributions, improved target volume coverage and the sparing of normal tissues from undesired radiation.

Background: CT120 is a universally expressed protein with seven transmembrane domains. It functions in cell proliferation, survival and apoptosis by activating Raf/MEK/ERK and PI3K/Akt signaling pathways. Evidence suggests that CT120 plays important roles in lung carcinogenesis and oncogenic pathway activation. c-Myc is an important transcription factor modulating cell progression, apoptosis and cellular transformation. Previous studies have shown that MYC gene is amplified in many types of cancer including head and neck squamous cell carcinoma (HNSCC). Myc can regulate expression of many genes by binding to E-boxes. The aim of this study was to investigate the relationship between c-Myc protein and CT120 gene. Methods: Tumor and normal tissue samples from 50 patients with HNSCC were investigated with chromatin immunoprecipitation assay (ChIP), Illumina MiSeq, bisulphite sequencing and qRT-PCR. Results: c-Myc binds to all E-boxes except E-box 5 on CT120 promoter. The CpG dinucleotides were found to be partially methylated in all tumor and normal tissue samples. Bisulphite sequencing showed a 10% down-regulation in the methylation levels of the tumor tissues. CT120 gene was hypomethylated and up-regulated in 56% of the tumor tissue samples. Expression of c-Myc was significantly higher in tumor tissues than in non-cancerous tissue samples. MYC was overexpressed in 68% of the tumor tissue samples compared to normal tissues. The mean MYC levels were 2.42-fold higher in the tumor tissue samples. In 48% of the tumor tissues, MYC and CT120A mRNA were up- or down-regulated simultaneously (p

Background Graves Disease (GD) is an autoimmune disorder affected by an interaction of multiple genes such as Nuclear Factor-κB (NF-κB), Nuclear Factor-κB Inhibitor (NF-κBIA), Poly (ADP-ribose) polymerase-1 (PARP-1) and cytokines like Interleukin-1β (IL-1β), Interleukin-6 (IL-6) and Tumor Necrosis Factor-α (TNF-α) and mostly accompanied by an ocular disorder, Graves Ophthalmopathy (GO). We hypothesize that there is a relationship between GD, GO, polymorphisms of inflammatory related genes and their association with cytokines, which may play important roles in autoimmune and inflammatory processes. Subjects and methods To confirm our hypothesis, we studied the polymorphisms and cytokine levels of 120 patients with GD and GO using PCR-RFLP and ELISA methods, respectively. Results We found that patients with GG genotype and carriers of G allele of PARP-1 G1672A polymorphism are at risk in the group having GD (p = 0.0007) while having GA genotype may be protective against the disease. PARP-1 C410T polymorphism was found to be associated with GO by increasing the risk by 1.7 times (p = 0.004). Another risk factor for development of GO was the polymorphism of del/ins of NFkB1 gene (p = 0.032) that increases the risk by 39%. Levels of cytokines were also elevated in patients with GD, but no association was found between levels of cytokines and the development of GO as there was no change in levels of cytokines. Conclusions We suggest that, PARP-1 and NFkB1 gene polymorphisms may be risk factors for developing Graves Disease and Ophthalmopathy.

Lung cancer, a major health problem affecting the epithelial lining of the lower respiratory tract, is considered to be one of the deadliest types of cancer in males and females and it is well-known that smoking is the chief cause of lung cancer. In addition to smoking and environmental factors, genetic susceptibility may also contribute to the development of lung cancer. Previous studies have shown that certain non-small cell lung cancer (NSCLC) patients harbor gain-of-function mutations in the epidermal growth factor receptor gene (EGFR). Phosphorylated EGFR triggers the activation of intracellular signal transduction pathways, including the RAS-MAPK, PI3K-Akt and STAT pathways. However, K-Ras gene point mutations in codons 12, 13 or 61 cause the inactivation of GTPase activity which results in overstimulation of cellular growth and gives rise to neoplastic development. Our aim was to investigate the presence and association of EGFR and K-Ras mutations in 50 primary NSCLC patients with a smoking history by using real-time PCR and sequencing. EGFR mutations were detected in four patients (8%). Two of these mutations were L858R mutations and the remaining two were deletion mutations spanning between codons 746 and 750. The L858R mutation was significantly associated with smoking status (P=0.003). K-Ras codon 12 and 61 mutations were also observed in four patients. However, no association was observed between K-Ras mutations and the tumor staging, gender, histology and smoking status of the patients.

Lung cancer is the most common cause of cancer-related death worldwide and, like many other cancers, is affected by different genetic, epigenetic, and environmental factors. The WW domain-containing oxidoreductase (WWOX) gene is a tumor-suppressor gene located on chromosome 16q23.3-24.1, and it has been shown that it loses its function due to alterations in genetic and epigenetic mechanisms. The aim of this study is to investigate the relationship between lung cancer and WWOX gene. Tumor tissue samples, corresponding normal tissues, and blood samples obtained from 50 lung cancer patients were involved in the study. We analyzed methylation profile by methylation-specific PCR and mutations and polymorphisms by DNA sequencing. Methylation analysis showed that promoter hypermethylation was present in 38 of 50 (76%) patients. In addition, promoter region of WWOX gene of younger patients was more frequently methylated than older patients. Using DNA sequencing, we found four genetic alterations in WWOX gene. Two of them were germline mutations (Exon 4 and 7), and two of them were polymorphic (Exon 6 and 8). We found a new mutation in exon 7 (Arg-254--Cys) which has not been described previously. The changes in the short-chain dehydrogenase domain of the protein caused by the genetic alterations may affect the function of the gene. We conclude that hypermethylation of WWOX gene promoter region and mutations in the gene might be related to lung carcinogenesis.

About 10% of patients with severe hemophilia exhibit a milder clinical phenotype with less frequent bleeds. Among many other factors, coinheritance of prothrombotic mutations have been proposed to act as modulators of clinical severity in severe hemophilia. We conducted a study to evaluate the impact of 3 prothrombotic mutations (factor V Leiden, factor II, and methylenetetrahydrofolate reductase mutations) on clinical phenotype of patients with severe hemophilia in our institution. For this purpose we compared the average annual factor concentrate consumption between carriers and noncarriers of prothrombotic mutations. A total of 38 hemophilia A and B patients with factor levels less than 1 were recruited between October 2006 and October 2007. Prothrombotic mutations were detected in venous blood using polymerase chain reaction amplification technique. Eighteen patients (47%) carried no prothrombotic mutations. The remaining 20 patients (53%) were found to be carriers of either 1 or 2 mutations. Median age in both carrier and the non-carrier groups was 27 years. None of the patients in either group gave a history of thromboembolic event. Median annual factor concentrate consumptions in carriers and noncarriers were 610 ± 530 units/kg and 770 ± 670 units/kg, respectively ( P = .203). Our results demonstrated no significant difference in annual factor concentrate consumption between carriers and noncarriers of prothrombotic mutations. Considering that average annual factor consumption is a surrogate indicator of clinical phenotype, we concluded that coinheritance of prothrombotic mutations was not associated with occurrence of different clinical phenotypes in severe hemophilia.

Mesial temporal lobe epilepsy with hippocampal sclerosis (MTLE-HS) is one of the most common medically intractable epilepsy syndromes and the pathogenesis of HS remains highly obscure. Recent studies demonstrated controversial results about the relationship between interleukin (IL) gene polymorphism and epilepsy in different ethnic groups. This correlation was investigated in Turkish patients with MTLE-HS. The allele distribution of IL-1alpha and IL-1beta in 47 patients of Turkish ancestry was determined and compared with 99 ethnically matched control subjects. Analysis of genotype frequencies between patients and controls showed no statistically significant difference (p0.05). Our data suggest that IL-1alpha and IL-1beta gene polymorphisms do not act as a strong susceptibility factor for MTLE-HS in individuals of Turkish ancestry.

Recent studies focusing on the genetic influences on outcome after head injury (HI) have suggested that different alleles of certain genes are associated with different outcomes. Interleukin-1 beta (IL-1beta) gene, especially beta2 polymorphism, is frequently observed in Alzheimer's disease, a remarkable degenerative state in which HI is among the known risk factors. Therefore, the aim of this paper was to search for the possible association between the outcome and IL-1beta gene polymorphism in human HI.The study group was composed of the 69 patients admitted to the neurosurgery department after HI. The severity of the initial injury was evaluated by means of the Glasgow Coma Scale and outcome six months later was assessed by means of the Glasgow Outcome Scale. IL-1beta genotypes were determined from blood samples by standard methods.Fourteen of 25 (56%) patients with IL-1beta +3953 allele 2 had an unfavourable outcome (dead, vegetative state or severe disability) compared with eight of 44 (18.1%) patients without IL-1beta +3953 (p = 0.0004). Similarly, 20 of 28 (71.4%) patients with IL-1beta -511 allele 2 had an unfavourable outcome compared with two of 41 (4.8%) patients without IL-1beta -511 (p = 0.005). Patients who had a composite of IL-1beta 2/2 or 1/2 genotype from both -511 and +3953 region of the chromosome 2 were more prone to have bad prognosis.Results of our study demonstrated that there might be a significant association between IL-1beta gene polymorphism and outcome after HI, supporting the hypothesis of a genetically determined influence.

HNSCC is the sixth most common form of cancer and represents the third common cause of cancer-related deaths worldwide. Therefore, there is a need to understand the molecular mechanisms involved in the pathogenesis of HNSCC. The c-myc protein belongs to the myc family of transcription factors and plays a fundamental role in cell cycle progression, apoptosis and cellular transformation. In the human genome, the MYC gene is located on chromosome 8 and its protein product regulates expression of 15% of all genes through binding to the Enhancer box (E-box) sequences. An E-box is a DNA sequence found in the promoter regions of eukaryotic genes that acts as a protein binding site to regulate gene expression. The CANNTG consensus sequence of the E-box is known as the canonical E-boxes. There are several other non-canonical E-box motifs such as the GATGTG, CATGCG, CACGCG, CACGAG, CGCGAG and CAACGTG sequences. Amplification of the MYC gene is observed in many different types of cancers including HNSCC. CT120 is a novel gene which encodes a human trans-membrane plasma protein. It has been shown that up-regulation of CT120 is associated with lung and head and neck carcinogenesis. In this study we investigated the promoter region of the CT120 gene and identified 6 E-boxes. To test whether c-myc binds to these E-boxes we performed chromatin immunoprecipitation (ChIP) assays. We demonstrated that c-myc binds strongly to the E-boxes 2, 3 and 4. It is well known that the expression rates of the genes are under the control of epigenetic modifications. Methylation of the CpG islands which are found in the promoter regions is one of the most studied epigenetic modifications. We identified two putative CpG islands in the promoter of the CT120 gene. To investigate the possible epigenetic regulation of CT120 and the effect of the promoter methylation to the binding of c-myc, we also investigated the methylation levels of the two islands in the CT120 promoter by MS-PCR. We observed a slight decrease (3.1%) in the methylation level of the first CpG-rich region. In 56% (28/50) of the tumor tissues methylation was decreased compared to the non-cancerous tissues. However, there was no statistically significant correlation between the promoter methylation and CT120 mRNA level. Depending on these data we investigated the expression levels of the c-myc and CT120 in the 50 HNSCC and normal tissue samples by qRT-PCR and observed a direct correlation between the overexpression of the c-myc and CT120 genes. Overexpression of CT120 and c-myc were observed in 29 (58%) and 34 (68%) of the tumors, respectively, compared to non-cancerous tissues. A total of 24 (48%) patients out of 50 showed a concurrent pattern of either up- or down regulation. Our results indicate that the CT120 gene is a target of c-myc and that both proteins may participate in the progression of HNSCC. Citation Format: Onur Baykara, Elif Baltaci, Betul Seyhan, Nejat Dalay, Nur Buyru. Regulation of CT120 gene as a new target of c-myc in head and neck squamous cell carcinoma (HNSCC). [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1132.

Like most other epithelial malignancies, NSCLC is characterized by a complex karyotypic pattern with multiple, non-randomly gains and deletions of chromosomal segments. Therefore, several investigators have analyzed NSCLC tumors using conventional comperative genomic hybridization (CGH) or fluorescence in situ hybridization (FISH) techniques to identify the chromosomal aberrations. The 8p11-12 region is one of the most frequently rearranged chromosomal regions in the human genome. The region is prone to chromosomal breaks which makes it a candidate region for cancer research. It can contribute to oncogenesis via inactivation of one or multiple potential tumor suppressor genes and/or amplification and overexpression of candidate oncogenes. Four different amplicons have been identified in the 8p11-12 region. The ZNF703, PROSC, BRF2 and RAB11F1P1 genes reside on the telomeric amplicon A1 and are considered as potential drivers of this amplicon. ZNF703 is one of the candidate oncogenes and its amplification and/or overexpression has been associated with luminal type breast cancer. ZNF703 belongs to the NET family zinc-finger transcription factors and its function depends on the cellular context. More recently, it has been shown that ZNF703 overexpression can activate the Akt/mTOR signaling pathway in breast cancer cell lines. The Akt1 protein can be activated by different pathways and phosphorylation at theSer473 residue is an indicator of activation. In this study we analyzed the copy numbers and expression levels in association with Akt1 phosphorylation levels by MLPA, qRT-PCR and Western blotting, respectively. ZNF703 amplification was detected in 46.3% of the tumor tissue samples compared to adjacent non-cancerous tissues. When we analyzed the mRNA expression levels of the ZNF703 gene in the tumor samples we observed overexpression in 28 (59.6%) of the samples. We also investigated phosphorylation of the Akt protein in 43 tumor samples in association with ZNF703 expression. An increase in Akt phosphorylation was observed in 51.1% of the tumor tissue samples. When we correlated Akt1 phosphorylation with CNV and ZNF703 mRNA expression levels, the increase of p-Akt1 was more frequent in the samples displaying ZNF703 gene amplification and overexpression. However, this positive association was not statistically significant (p = 0.07). On the other hand, when association of ZNF703 expression and Akt1 phoshorylation levels with clinicopathological parameters were analyzed, higher ZNF703 mRNA levels were observed in SCC tumors compared to adenocarcinomas (p = 0.05). Our results indicate that ZNF703 may contribute to the development of SSCs by activating the Akt signaling pathway. Citation Format: NUR BUYRU, Burak Bakir, Onur Baykara, Kamil Kaynak, Nejat Dalay. Overexpression of ZNF703 may activate Akt1 in NSCLC. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3929. doi:10.1158/1538-7445.AM2015-3929

Lung cancer is a result of uncontrolled cell growth in lung tissue and is the most common type of cancer leading to death in men and second after breast cancer in women. Even though smoking is the main reason of lung cancer, genetic and environmental factors play an important role in the onset and prognosis of the disease. The WW domain-containing oxidoreductase (WWOX) gene is a putative tumor suppressor gene located on chromosome 16q23.3-24.1 and is composed of 9 exons. Several studies have reported the role of WWOX gene in different types of carcinomas including lung, breast, ovarian, stomach and nasopharyngeal cancer. The 46kDa WWOX protein has two WW (tryptophan) domains which can recognize and bind the proline-rich region (PPXY) of other proteins. Its main function is to inhibit tumor cell growth. WWOX also plays an important role in wnt/beta-catenin signaling pathway. The lost-on-transformation 1 (LOT1/ZAC1/PLAGL1) gene is another putative tumor suppressor gene located on chromosome 6q24-25, a common site known for loss of heterozygosity in many tumors. Deletions of the 6q region are observed in different types of cancers including breast, prostate, small cell lung cancer and mesotheliomas. The LOT1 gene codes for a 55 kDa protein with seven C2H2 type zinc finger motifs located in amino terminal half of the protein. This zinc-finger nuclear transcription factor interacts with other proteins and transcription factors exerting antiproliferative effects in cancer cells. The aim of this study was to investigate the expression levels of the WWOX and LOT1 genes by RT-PCR in tumor tissue samples from 39 patients with non small cell lung cancer (NSCLC). We observed down-regulation of WWOX and LOT1 expression in 82.1% and 48.7% of the patients, respectively. Expression levels of both genes were down-regulated simultaneously in 18 (46.1%) of the patients. When we analyzed the association between gene expression and age, sex, smoking, histology and stage, we found that WWOX expression was decreased in 28 of 31 (90.3%) patients older than 50 years (p = 0.02). Lower expression levels of both genes supports the hypothesis that WWOX and LOT1 act as tumor suppressors in NSCLC and that both genes may have a common effect on tumor progression which warrants further studies to analyze a possible cooperation between the two proteins. Citation Format: Onur Baykara, Seda Ekizoglu Eratak, Kamil Kaynak, Hikmet Koseoglu, Nur Buyru. WWOX and LOT1 genes are down-regulated in non-small cell lung cancer (NSCLC). [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4952. doi:10.1158/1538-7445.AM2015-4952

Lung cancer is the main cause of cancer-related death worldwide and genetic, epigenetic and environmental factors affect the outcome of the disease. The WW domain-containing oxidoreductase (WWOX) gene is a putative tumor suppressor gene located on chromosome 16q23.3-24.1. It is composed of 9 exons and codes for a 414 amino acid protein. The WWOX gene has been previously reported to lose its tumor suppressor effect in several types of cancer including lung, ovarian, breast, stomach, uroepithelial and nasopharyngeal carcinomas. Docking Protein 1 (DOK1) is another putative tumor suppressor gene located on chromosome 2p13 and is known to be down-regulated and hypermethylated in various types of cancer including lung, breast or head and neck cancers and Burkitt lymphoma. WWOX activates MEK2 (MAP2K2-mitogen activated protein kinase kinase 2) which plays a crucial role in mitogen growth factor signal transduction. This activation increases ERK signaling in the MAPK/ERK pathway leading to transcription. The WWOX protein has two WW (tryptophan) domains which recognize the proline-rich region of other proteins. The DOK1 protein has a proline-rich region on its COOH terminal which may interact with the WWOX protein. DOK1 activates the RAS-GAP (GTPase activating proteins) signaling pathway leading to transcription when it is phosphorylated. Both WWOX and DOK1 also play important roles in the wnt/beta-catenin signaling pathway. The aim of this study was to investigate the relationship between WWOX and DOK genes by analyzing the expression levels of WWOX and DOK1 using RT-PCR in tumor tissue samples obtained from patients with non small cell lung cancer (NSCLC). When we analyzed expression levels by Spearman's Rank Correlation Coefficient Test, we observed down-regulation of the WWOX and DOK1 expression by 92.3% and 69.4% of the patients, respectively. Expression levels of WWOX and DOK both decreased simultaneously in 66.7% of the patients. We found a significant positive correlation between the expression levels of the WWOX and DOK1 genes when analyzed using the Wilcoxon Signed Ranks Test (p=0.033). Citation Format: Onur Baykara, Seda Ekizoglu, Esin Tuna, Ahmet Demirkaya, Kamil Kaynak, Hikmet Kose, Nur Buyru. Simultaneous down-regulation of WWOX and DOK1 genes in non-small cell lung cancer (NSCLC) patients. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1397. doi:10.1158/1538-7445.AM2014-1397

Lung cancer, which involves malignant proliferation of the epithelial lining of the lower respiratory tract is the most common cause of cancer death in men and is second only to breast cancer in women. Smoking is considered to be one of the principal causes of lung cancer. However, as only a subgroup of smokers ever develop the disease, it has been suggested that genetic susceptibility may contribute to the risk. Recent studies have shown that some non-small cell lung cancer (NSCLC) patients harbor gain-of-function mutations in the epidermal growth factor receptor gene (EGFR). Phosphorylated EGFR triggers the activation of intracellular signal transduction pathways such as RAS-MAPK, PI3K-Akt and STAT. On the other hand, point mutations in codons 12, 13 or 61 of the K-Ras gene inactivate the GTPase activity of the K-Ras resulting in continuous stimulation of autonomous growth and contribute to neoplastic development. The aim of this study was to investigate the presence and relationship of EGFR and K-Ras mutations with smoking history in a series of 50 primary NSCLC tumors. DNA was isolated by digestion of tissue samples using a DNA isolation kit. EGFR and K-ras mutations were investigated by Real Time-PCR assay. To confirm the mutations exons 18-21 of the EGFR gene and the whole K-ras gene were also analyzed by cycle sequencing in the samples found positive by the real-time PCR assay. In four patients (8%) EGFR mutations were detected. Two of these mutations were L858R and the remaining two were deletion mutations spanning codons from 746 to 750. The L858R mutation was significantly associated with smoking status (p=0.003). K-ras codon 12 and 61 mutations were also observed in 4 patients. However, no association was observed between K-ras mutations and tumor staging, sex, histology and smoking status of the patients. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1126. doi:10.1158/1538-7445.AM2011-1126

Non-small cell lung cancer (NSCLC) is the leading cause of cancer mortality worldwide with poor differential diagnosis of the disease and with low response rates to standard chemotherapy. Individuals classified as Stage I by the present clinical-based scoring system may display survival characteristics ranging from 10 to 90 %. Identification of reliable biomarkers to detect NSCLC at early stages is therefore a subject of extensive research. Surface-enhanced laser-desorption-ionization time-of-flight mass spectrophotometry (SELDI-TOF-MS) combines chromatographic surfaces with solid-state TOF-MS for the high throughput analysis of proteins in diverse clinical samples such as serum, urine, cerebrospinal fluids and has overcome the limitations of earlier proteomic approaches. The method has been widely used for biomarker discovery. Therefore, in this study we used the SELDI-TOF protein Chip system to achieve a stratification based on the protein expression profile and to identify differentially expressed proteins in sera from 50 patients with NSCLC and 50 healthy controls. IMAC 30-Cu protein chips were used to fractionate the proteins according to their metal binding affinity. The arrays were analyzed using the Ciphergen Protein Chip Reader PB IIa. The spectra were normalized to the total ion current of the mass to charge between 3000 — 10 000 Da. Protein peak clustering and classification analysis were performed using the Express 3.0 Biomarker Pattern Analysis Software. Several proteins with masses ranging from 2000-20000 Da were differentially expressed between NSCLC patients and healthy controls. Among the peak clusters revealing differences we selected two specific clusters as prospective candidate biomarkers in sera of the patients. High expression of one peak and underexpression of the other with respect to the healthy control sera were evaluated in a blind test in terms of the ability to classify a separate group of patients and controls.

Lung cancer is the most common cause of cancer-related deaths worldwide and like many other cancers is affected by different genetic, epigenetic and environmental factors. The WWOX gene is a tumor supressor gene located on 16q23.3–24.1. It has been shown that it loses its function due to differences in genetic and epigenetic mechanisms. In this study we investigated whether the presence of WWOX gene mutations in lung cancer. The entire coding exons and flanking intronic sequences of the gene were analysed by PCR amplification from the genomic DNA and direct sequencing in 50 patients with non-small cell lung cancer (NSCLC). WWOX gene mutations were detected in five out of 50 primary tumors (10%). One of these mutations was an insertion in exon 4 at the third nucleotide of codon 120. This mutation was observed in 2 patients and leads to a frameshift creating a stop codon at codon 133. The second mutation was an insertion at position 2583 which also causes a shift in the reading frame. Finally, heterozygous C→G transitions on position 2807 at the first nucleotide of codon 282 (exon 8) were identified in 2 patients. This mutation affecting codon 282 leads to the substitution of alanine instead of proline. We also observed a previously defined G to A polymorphism at the nucleotide 2196 in homozygous and heterozygous form in 20 and 5 patients, respectively. Additionally we detected 12 different intronic substitutions. In addition to inactivating mutations target genes may also be silenced epigenetically. Therefore, besides mutation screening we also analyzed the methylation profile of the promoter region of the WWOX gene in tumor tissue and adjacent non-cancerous tissue samples by bisulphite modification and methylation-specific PCR. Promoter methylation was detected in 76% of the tumor tissue samples. While 20% of the samples displayed partial methylation 56% were completely methylated. In the adjacent normal tissue samples partial and complete methylation frequencies were 6% and 58%, respectively. Methylation frequency was significantly higher in the tumor tissue samples than in normal tissue samples (p=0.014). The high frequency of mutations and epigenetic modifications suggest that the WWOX gene may play a significant role in the etiology of NSCLC. Citation Information: Cancer Res 2009;69(23 Suppl):A8.