• Asian Pacific Journal of Cancer Prevention
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• v.15 no.20
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• pp.8595-8601
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• 2014

Background: miR-200a expression is frequently altered in numerous cancers. The aim of the present study was to determine the role of microRNA-200a in advanced ovarian carcinomas. Materials and Methods: We measured miR-200a expression in 72 matched normal ovarian tissues and advanced ovarian carcinomas, and also two ovarian carcinoma cell lines (SKOV3 and SKOV3.ip1 - the latter being more invasive and metastatic than the parental SKOV3) by stem-loop real-time RT-PCR based on TaqMan microRNA assay using U6 as a reference. Levels of miR-200a expression were compared by disease stage, tumor grade, histology, and lymph node involvement. To evaluate the role of microRNA-200a, cell proliferation and invasion of SKOV-3 and SKOV-3.ip1 were analyzed with miR-200a inhibitor/mimic transfected cells. Results: Of 72 paired samples, 65 cancer tissues overexpressed microRNA-200a greater than two fold in comparison with matched normal epithelium. Specifically, patients with lymph node metastasis showed significant elevation. The level correlated with clinicopathological features, including high tumor grade, late disease stage, most notably with lymph node metastasis, but not with tumor histology. In addition, SKOV-3.ip1 cells also overexpressed miR-200a compared with SKOV-3, and miR-200a inhibitor transfected SKOV-3.ip1 cells showed significant reduction in cellular proliferation and invasion, while a miR-200a mimic stimulated the opposite behavior. Conclusions: We provide definitive evidence that miR-200a is up-regulated in a significant proportion of advanced ovarian carcinomas, and that elevated miR-200a expression facilitates tumor progression. Our findings support the notion that miR-200a is an onco-microRNA for ovarian cancer, and elevation is a useful potential diagnostic indicator. This study also provides a solid basis for further functional analysis of miR-200a in advanced ovarian cancer.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.9
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• pp.5117-5122
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• 2013

Purpose: To explore the association between PIK3CA and AKT single nucleotide polymorphisms(SNP) and osteosarcoma susceptibility. Methods: TaqMan polymerase chain reaction(PCR) was used to detect the genotypes of SNPs (rs7646409, rs6973569 and rs9866361) in peripheral blood samples from 59 patients with osteosarcoma and from 63 healthy controls. Unconditional logistic regression was used to analyze the correlation between SNPs and osteosarcoma risk. Results: No statistically significant difference was found between osteosarcoma patients and healthy controls in the genotype of AKT rs6973569 (P=0.7). However, after stratified analysis, the genotype AA of AKT rs6973569 carried a higher risk of osteosarcoma metastasis (OR:2.94, 95%CL:1.00-8.59); the difference of rs7646409 genotype distributions between the case and control groups was statistically significant (P=0.032). Taking genotype TT as a reference, the risk of osteosarcoma increased three fold in patients with genotype CC (OR:3.47, 95%CL:1.26-9.56). A statistically significant difference was found between the alleles C and T (P=0.005). Further analysis showed that the risk factor was more pronounced in male patients with Enneking's stage IIB and osteoblastic osteosarcoma. PIK3CA rs9866361 did not fit Hardy-Weinberg equilibrium (P<0.05). Conclusions: Genotype CC in locus PIK3CA rs7646409 may increase the risk of osteosarcoma in the Chinese population.

• Korean Journal of Head & Neck Oncology
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• v.25 no.1
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• pp.28-32
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• 2009

Background : The most frequently reported risk factors for head and neck suamous cell carcinoma are smoking and alcohol. But in a recent overview, human papilloma virus(HPV) infection was revealed the important carcinogenic factor in oropharyngeal cancer. We aimed to clarify whether HPV directly effects on the oncogenesis and biologic behavior of hean and neck squamous cell carcinoma by comparison with infection prevalence, and physical status of virus. Material and Method : We used HPV genotyping DNA chip(Biocore, Korea, Seoul) arrayed by multiple oligonucleotide probes of L1 sequence of 26 types of HPV and HPV genotypes are identified by fluorescence scanner. The copy numbers of HPV E2 and E6 open reading frames(ORF) were assessed using a TaqMan-based 5'-exonuclease quantitative real-time PCR assay. The ratio of E2 to E6 copy numbers was calculated to determine the physical status of HPV-16 viral gene. Results : We observed a significant difference in HPV prevalence between tonsillar cancer group and control group(73.1% vs. 11.6%), and most of the HPVs were type 16(87.2%) and integrated(94.1%) state. In terms of oral tongue cancer, we demonstrate that 30.5% has integrated HPV-16 in cancer tissue. But Glottic cancer only 1% is related to HPV-16 integration. Conclusion : This study revealed significant relationship of HPV prevalence with oropharyngeal and oral tongue squamous cell carcinoma. Most of HPV were 16 type and integrated or mixed, HPV-16 integration could be directly related to the carcinogenesis.

• Journal of Plant Biotechnology
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• v.44 no.1
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• pp.89-96
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• 2017

The CRISPR/Cas9 is a core technology that can result in a paradigm for breeding new varieties. This study describes in detail the sgRNA design, vector construction, and the development of a transgenic plant and its molecular analysis, and demonstrates how gene editing technology through the CRISPR/Cas9 system can be applied easily and accurately. CRISPR/Cas9 facilitates targeted gene editing through RNA-guided DNA cleavage, followed by cellular DNA repair mechanisms that introduce sequence changes at the site of cleavage. It also allows the generation of heritable-targeted gene mutations and corrections. Here, we present detailed procedures involved in the CRISPR/Cas9 system to acquire faster, easier and more cost-efficient gene edited transgenic rice. The protocol described here establishes the strategies and steps for the selection of targets, design of sgRNA, vector construction, and analysis of the transgenic lines. The same principles can be used to customize the versatile CRISPR/Cas9 system, for application to other plant species.

• Journal of Microbiology
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• v.44 no.6
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• pp.660-664
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• 2006

The relationship between H. pylori clarithromycin resistance and genetic pattern distribution has been differently explained from different geographic areas. Therefore, we aimed to assess the clarithromycin resistance rate, to evaluate the bacterial genetic pattern, and to search for a possible association between clarithromycin resistance and cagA or vacA genes. This prospective study enrolled 62 consecutive H. pylori infected patients. The infection was established by histology and rapid urease test. Clarithromycin resistance, cagA and vacA status, including s/m subtypes, were assessed on paraffin-embedded antral biopsy specimens by TaqMan real time polymerase chain reaction (PCR). Primary clarithromycin resistance was detected in 24.1 % of cases. The prevalence of cagA was 69.3%, and a single vacA mosaicism was observed in 95.1 % cases. In detail, the s1m1 was observed in 23 (38.9%) patients, the s1m2 in 22 (37.2%), and the s2m2 in 14 (23.7%), whereas the s2m1 combination was never found. The prevalence of cagA and the vacA alleles distribution did not significantly differ between susceptible and resistant strains. Primary clarithromycin resistance is high in our area. The s1m1 and s1m2 are the most frequent vacA mosaicisms. There is no a relationship between clarithromycin resistance and bacterial genotypic pattern and/or cagA positivity.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.11
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• pp.4437-4441
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• 2014

SMAD7 has been identified as a functional candidate gene for colorectal cancer (CRC). SMAD7 protein is a known antagonist of the transforming growth factor beta ($TGF-{\beta}$) signaling pathway which is involved in tumorigenesis. Polymorphisms in SMAD7 may thus alter cancer risk. The aim of this study was to investigate the influence of a SMAD7 gene polymorphism (rs2337107) on risk of CRC and clinicopathological features in an Iranian population. In total, 210 subjects including 105 patients with colorectal cancer and 105 healthy controls were recruited in our study. All samples were genotyped by TaqMan assay via an ABI 7500 Real Time PCR System (Applied Biosystems) with DNA from peripheral blood. The polymorphism was statistically analyzed to investigate the relationship with the risk of colorectal cancer and clinicopathological properties. Logistic regression analysis revealed that there was no significant association between rs2337107and the risk of colorectal cancer. In addition, no significant association between genotypes and clinicopathological features was observed (p value>0.05). Although there was not any association between genotypes and disorder, CT was the most common genotype in this population. This genotype prevalence was also higher in the patients with well grade (54.9%) and colon (72.0%) tumors. Our results provide the first evidence that this polymorphism is not a potential contributor to the risk of colorectal cancer and clinicopathological features in an Iranian population, and suggests the need of a large-scale case-control study to validate our results.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.1
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• pp.369-374
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• 2014

Background: Colorectal cancer (CRC) is the fourth most common cause of cancer death in the world. Genetic variants in 8q24.21 including rs10505477 and rs6983267 have been hypothesized to be involved in susceptibility to CRC. This study aims to investigate the possible association between these loci and their haplotypes with CRC risk in Iranian population. Materials and Methods: Subjects were recruited from two hospitals in Tehran. The rs10505477 and rs6983267 polymorphisms were genotyped by TaqMan real time PCR using subject genomic DNA, extracted either from formalin-fixed, paraffin-embedded tissue of patients or from blood of the controls by standard methods. Results: A total of 715 subjects (380 CRC patients and 335 matched controls) were genotyped in this study. Allele and genotype analysis of the rs10505477 and rs6983267 polymorphisms by gender, age at diagnosis, tumor location, tumor grade, and tumor node metastasis (TNM) showed no significant association with CRC risk. There was a significant relationship between GG haplotype and susceptibility to age at diagnosis for both <60 and ${\geq}60$ (p=0.0005 and p=0.000004, respectively) and between GT and CRC in the age at diagnosis ${\geq}60$ (Table 3: p=0.031). The GG haplotype was less frequent in CRC patients with the age at diagnosis <60, but was more common in subjects with the age at diagnosis ${\geq}60$. Conclusions: Results of this study suggests that the rs6983267 and rs10505477 polymorphisms alone may not be relevant to CRC risk, but their GG haplotype plays a notable role in age at diagnosis of CRC in the Iranian population.

• Asian Pacific Journal of Cancer Prevention
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• v.16 no.12
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• pp.5069-5073
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• 2015

The single nucleotide polymorphism (SNP) rs1053004 in Signal transducer and activator of transcription 3 (STAT3) was recently reported to be associated with chronic hepatitis B (CHB)-related hepatocellular carcinoma (HCC) in a Chinese cohort. This study was aimed at investigating whether the SNP might also contribute to HCC susceptibility in the Thai population. Study subjects were enrolled and divided into 3 groups including CHB-related HCC (n=211), CHB without HCC (n=233) and healthy controls (n=206). The SNP was genotyped using allelic discrimination assays based on TaqMan real-time PCR. Data analysis revealed that the distribution of different genotypes was in Hardy-Weinberg equilibrium (P>0.05). The frequencies of allele T (major allele) in HCC patients, CHB patients and healthy controls were 51.4%, 58.6% and 61.4%, respectively, whereas the frequencies of C allele (minor allele) were 48.6%, 41.4% and 38.6%. The C allele frequency was higher in HCC when compared with CHB patients (odds ratio (OR)=1.34, 95% confidence interval (CI)=1.02-1.74, P=0.032). The genotype of SNP rs1053004 (CC versus TT+TC) was significantly associated with an increased risk when compared with CHB patients (OR=1.83, 95% CI=1.13-2.99, P=0.015). In addition, we observed a similar trend of association when comparing HCC patients with healthy controls (OR=1.77, 95% CI=1.07-2.93, P=0.025) and all controls (OR=1.81, 95% CI=1.19-2.74, P=0.005). These findings suggest that the SNP rs1053004 in STAT3 might contribute to HCC susceptibility and could be used as a genetic marker for HCC in the Thai population.

• Asian Pacific Journal of Cancer Prevention
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• v.17 no.4
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• pp.1801-1810
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• 2016

Recent discovery showing the presence of microRNAs (miRNAs) in the circulation sparked interest in their use as potential biomarkers. Our previous studies showed the diagnostic potential of miR-451 as a serological marker for inflammatory breast cancer (IBC), miR-337-5p and miR-30b for non-inflammatory breast cancer (non-IBC). The aim of this study is to investigate the prognostic values of circulating miRNAs by comparing the amounts of 12 circulating miRNAs in the serum of IBC and non-IBC from Tunisian breast cancer patients, and by determinating whether correlated pairs of miRNAs could provide useful information in the diagnosis of IBC and non-IBC patients. TaqMan qPCR was performed to detect circulating expression of miRNAs in serum of 20 IBC, 20 non-IBC and 20 healthy controls. Nonparametric rank Spearman rho correlation coefficient was used to examine the prognostic value of miRNAs and to assess the correlation profile between miRNAs expression. Further, a large number of miRNAs were highly correlated (rho>0.5) in both patients groups and controls. Also, the correlations profiles were different between IBC, non-IBC and healthy controls indicating important changes in molecular pathways in cancer cells. Our results showed that miR-335 was significantly overexpressed in premenopausal non-IBC patients; miR-24 was significantly overexpressed in non-IBC postmenopausal patients. Patients with previous parity had higher serum of miR-342-5p levels than those without. Furthermore, patients with HER2+ IBC present lower serum levels of miR-15a than patients with HER2-disease. Together, these results underline the potential of miRNAs to function as diagnostic and prognostic markers for IBC and non-IBC, with links to the menopausal state, Her2 status and parity.

• Asian Pacific Journal of Cancer Prevention
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• v.17 no.4
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• pp.2019-2025
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• 2016

Hepatocellular carcinoma (HCC) is major health problem with high mortality rates, especially in patients with hepatitis B virus (HBV) infection. Telomerase function is one of common mechanisms affecting genome stability and cancer development. Recent studies demonstrated that genetic polymorphisms of telomerase associated genes such as telomerase associated protein 1 (TEP1) rs1713449 and PIN2/TERF1-interacting telomerase inhibitor 1 (PINX1) rs1469557 may be associated with risk of HCC and other cancers. In this study, 325 patients with HCC and 539 non-HCC groups [193 healthy controls, 80 patients with HBV-related liver cirrhosis (LC) and 266 patients with HBV-related chronic hepatitis (CH)] were enrolled to explore genetic polymorphisms of both SNPs using the allelic discrimination method based on MGB probe TaqMan real time PCR. We demonstrated that all genotypes of both genes were in Hardy-Wienberg equilibrium (P>0.05). Moreover, there was no significant association between rs1713449 genotypes and HCC risk, HCC progression and overall survival (P>0.05). Interestingly, we observed positive association of rs1469557 with risk of HCC when compared with the LC group under dominant (CC versus CT+TT, OR=1.89, 95% CI= 1.06-3.40, P=0.031) and allelic (C versus T alleles, OR=1.75, 95% CI=1.04-2.94, P=0.033) models, respectively. Moreover, overall survival of HCC patients with CC genotype of rs1469557 was significantly higher than non-CC genotype (Log-rank P=0.015). These findings suggest that PINX1 rs1469557 but not TEP1 rs1469557 might play a role in HCC progression in Thai patients with LC and be used as the prognosis marker to predict overall survival in HCC patients.