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ABSTRACT: The aetiology of head and neck cancer (HNC) has been shown to be associated with genetic and certain environmental factors that produce DNA damage. Base excision repair (BER) genes are responsible for repair of DNA damage caused by reactive oxygen species and other electrophiles and therefore are good candidate susceptibility genes for HNC. Apurinic/apyrimidinic endonuclease-1 (APEX1) proteins have important functions in the BER pathway. In this case-control study, all exons of the APEX1 gene and its exon/intron boundaries were amplified in 300 HNC cases and 300 matched healthy controls and then analysed by single-stranded conformational polymorphism. Amplified products showing altered mobility patterns were sequenced and analysed. To confirm our observations, we examined APEX1 expression at mRNA level on 50 head and neck squamous cell carcinoma (HNSCC) and 50 normal control samples by quantitative real-time polymerase chain reaction. At germ line level, three novel mutations (13T > G, Ser129Arg and Val131Gly) of APEX1 were observed. The homozygous and heterozygous genotypes of APEX1 13T > G, Ser129Arg and Val131Gly appear to be significantly involved in the development of HNC. In the case of expressional level, APEX1 mRNA expression was positively correlated with tumour size, clinical stage and positive lymph node metastasis. Statistical analysis showed a significantly higher APEX1 mRNA level in HNC tumour tissue than in control samples. Our study demonstrated that APEX1 mutations and deregulation of APEX1 are associated with increased risk of HNC in the Pakistani population.
Mutagenesis 03/2013; 28(2):213-8. · 3.18 Impact Factor
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ABSTRACT: Cytoskeletal rearrangement occurs in variety of cellular processes and involves a wide spectrum of proteins. Gelsolin super family proteins control actin organization by severing and capping filament ends and nucleating actin assembly. Gelsolin is the founding member of this family and plays important role in pathogenesis of human neoplasia. This study aimed to investigate the germline mutations and expressional profile of Gelsolin in human breast cancer tissues. For germ line screening PCR-SSCP technique was used while expression was analyzed through quantitative real time PCR. Different types of mutations were observed in Gelsolin coding regions on exons 4, 10, 11, 14 and 15. These mutations include 3 missense nonsynonymous substitution mutations, 2 deletions, 1 insertion and 1 synonymous substitution mutation. Gelsolin transcript level was found significantly lower in breast tumor tissues compared to control samples (p=0.03). Low level of Gelsolin was found in metastatic patients (p=0.002) and patients who died from breast cancer (P=0.03) compared to disease free patients at final follow up. This study shows that level of Gelsolin is down regulated in breast cancer tissues and is linked with metastasis development and death in patients. It is concluded that genetic changes in coding regions of Gelsolin can potentially contribute to genetic instability. These genetic variations and expressional correlation with patient survival may prove to be of significant importance.
Disease markers 01/2013; · 1.64 Impact Factor
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ABSTRACT: Cyclin D1 plays a key role in cell cycle control, particularly in the transition from G1 to S phase, regulated by cyclin-dependent kinases. The objective of the present study was to screen the cyclin D1 gene (CCND1) for polymorphisms in patients with head and neck cancer (HNC). Genomic DNA was isolated from blood samples of 380 HNC patients and 350 controls. In a hospital-based case-control study using the PCR-SSCP technique we found 3 novel germline mutations: g3578C>A, g3475G>C and g3383delA. The commonly reported guanine to adenine polymorphisms in exon 4 g7656G>A (rs9344) and g10861C>A (rs7177) in 3'UTR of CCND1 were also observed. The calculated frequencies of the g7656G>A (rs9344) polymorphism in GG, GA and AA genotypes were 27.3%, 38.6%, and 33.9% in HNC cases, and 44.2%, 29.4%, and 26.2% in normal healthy controls, respectively. Adjusted by age (in years), sex and smoking status, multivariate logistic regression analysis showed that the AA and GA genotypes were associated with a significantly increased risk (OR 1.34, 95% CI 1.03-1.64, p=0.028) for HNC. The CCND1 AA genotype variant was associated with an increased risk in individuals who were <40 years old (OR 1.45, 95% CI 1.02-2.08, p=0.04). In conclusion, it is suggested that the CCND1 G/A polymorphism is associated with the early onset of HNC and may contribute to HNC susceptibility in a Pakistani population.
The International journal of biological markers 11/2012; · 1.48 Impact Factor
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ABSTRACT: DNA repair gene polymorphisms have been implicated as susceptibility factors in cancer development. It is possible that DNA repair polymorphisms may also influence the risk of gene mutation. Polymorphisms in the DNA repair gene XRCC1 have been indicated to have a contributive role in DNA adduct formation and an increased risk of cancer development. 300 head and neck cancer patients and 150 controls were included in this study. PCR-single-strand conformation polymorphism and DNA sequencing were used to analyze the whole exonic region of XRCC1 in head and neck cancer patients. Sequence analysis revealed two missense and two silent mutations in our study. Frequency of silent mutations; Pro206Pro (rs915927) and Gln632Gln (rs3547) was calculated as 0.16 (16 %) and 0.30 (30 %) respectively. Whereas, the frequency of missense mutations; Arg399Gln (rs25487) and Tyr576Asn (rs2307177) was calculated as 0.27 (27 %) and 0.28 (28 %) respectively. In our study, incidence of these mutations was found higher in larynx cancer (p < 0.005) as compared to oral cavity and pharynx cancer. Our finding suggests that the polymorphic XRCC1 gene may contribute to risk of developing head and neck cancer. To our knowledge, this is the first report that XRCC1 is associated with increased risk of head and neck cancer in a Pakistani population.
Pathology & Oncology Research 09/2012; · 1.37 Impact Factor
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ABSTRACT: Cyclin-dependent kinase 4 (CDK4) together with its regulatory subunit cyclin D1, governs cell cycle progression through G1 phase. Cyclin-dependent kinase inhibitors, including p16INK4A in turn regulate CDK4. In particular, deregulation of the p16/CDK4/cyclin D1 complex has been established in a variety of human tumors including gliomas, sarcomas, melanoma, breast and colorectal cancer. However, changes in CDK4 have rarely been observed.
In this study we used a combination of PCR-SSCP and direct sequencing for mutational screening of CDK4. DNA was isolated from peripheral blood leukocyte of patients with squamous cell carcinoma of head and neck, for screening germline mutations in coding regions of CDK4.
Variations observed in exon 2 and 5 were three missense mutations, g5051G > C (Ser52Thr), g5095G > C (Glu67Gln), g5906C > A, g5907C > G (Pro194Ser) and novel frame shift mutations g7321_23delTGA, g7121_7122insG, g7143delG in exon 7 and 3'UTR respectively.
In conclusion, two novel mutations were found in N terminal domain which indicates that CDK4 mutation may play a major role in the development and progression of squamous cell carcinoma of head and neck.
Hereditary Cancer in Clinical Practice 08/2012; 10(1):11. · 1.68 Impact Factor
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ABSTRACT: OGG1 (The human 8-oxoguanine glycosylase 1) is the primary enzyme in BER (base excision repair) pathway, responsible for the excision of 7, 8-dihydro-8-oxoguanine (8-oxoG), a mutagenic base byproduct that occurs as a result of exposure to reactive oxygen species. OGG1 gene is highly polymorphic among humans and is mutated in cancer cells. In this case control study, all exons of OGG1 gene and its exon/intron boundaries were amplified in 210 laryngeal cancer cases and 210 matched controls and then analyzed by single stranded conformational polymorphism. Amplified products showing altered mobility patterns were sequenced and analyzed. Two silent (Gln718Gln, His699-700His) and three missense (Ala597, Thr608-610Pro and Glu707Lys) mutations were observed in exon 2. In addition to this one missense mutation (1578G > A) was also observed in 3'UTR region. We found a significant association between OGG1 mutations and laryngeal cancer and observed that His699-His700, silent mutation exhibited an enhanced risk of ~9.0 folds (OR = 9.07, 95 % CI = 4.73-17.39) and 1578G > A, missense mutation ~0.4 folds (OR = 0.37, 95 % CI = 0.15-0.90). Furthermore, a positive association of OGG1 mutations with smoking was observed in laryngeal cancer cases when compared to controls. Heavy smokers have higher incidence of OGG1 mutations when compared to light smokers in present study. Our results demonstrate that OGG1 mutations are associated with an increased risk of laryngeal cancer. OGG1 mutations were found to accumulate more of 8-OHdG in smokers, which may serve as a biomarker for early diagnosis of laryngeal cancer.
Familial Cancer 07/2012; · 1.30 Impact Factor
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ABSTRACT: Retinoblastoma gene (Rb1) is a tumor suppressor gene, which plays a pivotal role in cell cycle regulation, promoting G1/S arrest and growth restriction through inhibition of the E2F transcription factor. Abnormalities in the genes involved in cell cycle, including Rb1, have been reported in head and neck cancer (HNC) patients. Studies regarding Rb1 have been observed in different world populations but data is missing for Pakistani population. This study was aimed to analyze the genetic aberrations of Rb1 and their association with the development of HNC in Pakistani population. Genomic DNA was isolated from blood samples of 300 HNC patients and 270 controls. Salient coding region of gene was amplified by using Polymerase Chain Reaction (PCR). PCR conditions were optimized for each exon separately. Amplified products were analyzed for mutational screening using Single strand confirmation polymorphism (SSCP) technique followed by sequence analysis. Sequence analysis revealed five missense mutations g77082G>C, g77083G>A, g170220A>T, g170221G>C, g170228T>A, two frameshift mutations, two stop codon and two intronic substitutions in this study. The overall frequency of these mutations was 0.71. Frequency of nonsense mutations; Lys462stop (Novel) and Ser834stop (CM952105) were 0.15 and 0.14 respectively. We also report here novel missense mutations, frameshift mutation and a stop codon Lys462stop in HNC patients of Pakistani origin.This study suggests that the Rb1 germline mutations may contribute to genetic susceptibility for HNC. To our knowledge, this is the first report that Rb1 gene may be associated with risk of cancer in Pakistani population.
Molecular Biology Reports 06/2012; 39(10):9573-81. · 2.93 Impact Factor
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ABSTRACT: The PTEN gene, a candidate tumor suppressor, is one of the more commonly inactivated and extensively studied genes in cancer. However, few data are available about the role of germ line mutations of this gene in sporadic breast cancer cases. The purpose of this study was to determine extent of involvement of this gene in breast cancer in Pakistan. To test the hypothesis that genetic variations of PTEN play a role in the etiology of breast cancer, a population based case-control study was conducted in 350 breast cancer patients along 400 healthy controls. After extracting DNA from blood, the whole coding sequence of PTEN along with intron/exon boundaries was genotyped by polymerase chain reaction-single stranded conformational polymorphism. Sequencing analysis revealed nineteen different types of mutations in different regions of PTEN (in exon 2, 4, 5, 6, 7 and splicing sites of intron 2 and 4 and also in the 3' UTR region), including 3 silent, 8 missense, 2 frame shift and 6 splice site variations. Among the observed variations in this study, three missense mutations have already been reported i.e. 319G>A (Asp106Asn), 389G>A (Arg129Gln) and 482G>A (Arg160Lys) in different populations. The present results suggest that a wide range of germline PTEN mutations may play a role in the pathogenesis of breast cancer.
Asian Pacific journal of cancer prevention: APJCP 01/2011; 12(10):2773-8. · 0.66 Impact Factor
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ABSTRACT: In Pakistani culture tobacco use is very high and a well known risk factor for developing head and neck cancer (HNC), tobacco smoke containing high quantities of chemical carcinogens such as aromatic amines and reactive oxygen species. OGG1 is the primary enzyme in the base excision repair (BER) pathway, responsible for the excision of 7, 8-dihydro-8-oxoguanine, a mutagenic base byproduct that occurs as a result of exposure to reactive oxygen species. Groups of 300 already diagnosed HNC patients along with normal controls were included in this study. PCR-single-strand conformation polymorphism and DNA sequencing were used to analyze the whole coding region of OGG1 gene. Sequence analysis revealed eight novel mutations (six missense and two frame shift mutations). Frequencies of missense mutations, Asp267Asn, Ser279Gly and Ile253Phe were 0.12, 0.13 and 0.06 respectively. Frequencies of other missense mutations, 1578A> T, 1582C> T and Ala399Glu (1542C> A) were 0.13, 0.13 and 0.16, whereas values for the frame shift mutations 1582insG and 1543-1544delCT were 0.13 and 0.16. In our study, incidence of these mutations was found higher in oral cancers (p<0.002) and in smokers (p<0.002) when compared with other sites of HNC and nonsmokers, respectively. Our finding suggests that these germline mutations in OGG1 gene contribute to risk of developing HNC.
Asian Pacific journal of cancer prevention: APJCP 01/2011; 12(10):2779-83. · 0.66 Impact Factor
