Research ArticleInvestigation of NF-120581B1 and NF-120581BIA Gene Polymorphism inNon-Small Cell Lung Cancer

Y M Oltulu1 E Coskunpinar1 G Ozkan1 E Aynaci2 P Yildiz3 T Isbir4 and I Yaylim1

1 Department of Molecular Medicine Institute of Experimental Medicine Research Istanbul University DETAEPO Box 7 34390 CapaIstanbul Turkey

2Department of Chest Diseases Faculty of Medicine Medipol University Istanbul Turkey3Third Clinics Yedikule Chest Diseases andThoracic Surgery Training Research Hospital Istanbul Turkey4Department of Medical Biology School of Medicine Yeditepe University Istanbul Turkey

Correspondence should be addressed to I Yaylim ilhanyaylimgmailcom

Received 20 October 2013 Revised 13 December 2013 Accepted 9 January 2014 Published 23 February 2014

Academic Editor Sherven Sharma

Copyright copy 2014 Y M Oltulu et al This is an open access article distributed under the Creative Commons Attribution Licensewhich permits unrestricted use distribution and reproduction in any medium provided the original work is properly cited

Lung cancer is a complex multifactorial disease which is the leading cause of cancer death in both men and women NF-120581B is atranscription factor which is known to affect the expression of more than 150 genes related to inflammation lymphocyte activationcell proliferation differentiation and apoptosis as well as contributing to cell apoptosis and survival However NF-120581BIA (I120581B120572) isthe inhibitor of the transcription factor The -94insdelATTG polymorphism of the NF-120581B1 gene promoter region which causes afunctional effect and NF-120581BIA 31015840UTR ArarrG polymorphism has been shown to be related to various inflammatory diseases andcancer Ninety-five NSCLC patients and 99 healthy controls were included in study The NF-120581B1 -94insdelATTG and NF-120581BIA31015840UTR ArarrG polymorphism have been studied by using PCR-RFLP method It was found that the NF-120581B1 -94insdelATTG DDgenotype and D allele frequencies were higher in patients than healthy controls and the presence of the DD genotype has a 35-foldincreased risk of the disease (P 0014)This study is the first to investigate the NF-120581B1 -94insdelATTG andNF-120581BIA 31015840UTRArarrGpolymorphism together in the Turkish population According to the results the NF-120581B1 -94insdel ATTG promoter polymorphismmay have a role in lung carcinogenesis and prognosis

1 Introduction

Non-small cell lung cancer (NSCLC) which includes squa-mous cell carcinoma adenocarcinoma and large-cell carci-noma is the most common lung cancer possessing approxi-mately 80ndash85 rates in the prevalence of lung cancer case [1]Despite all advances in the current treatments including sur-gical resection chemotherapy and radiation therapy alone orin combination the disease is rarely curable prognosis andremains poor [2] As all of these facts are considered recentresearches are tending to understand molecular biologicaland genetic factors and to find other prognostic factorsproviding long-term survival and intending new targetedtherapies [3 4] Lung cancer cells manage to escape from thesignal transduction pathways to facilitate their own survivaland proliferation by usingmultiplemechanisms [5] Carcino-gens and inflammatory cytokines contributing substantially

to cancer development are involved in activation of commoncell survival signaling pathways The one of this cell survivalsignal is nuclear factor-kappaB (NF-120581B) which is involved inmultiple steps in carcinogenesis and in cancer cellrsquos resistanceto chemo- and radiotherapy [6] Recently many studies withanimal models and cell culture systems indicate the interplaybetween NF-120581B and lung carcinogenesis which emphasizesthe importance of targeting the NF-120581B signaling pathway forlung cancer treatment and chemoprevention [7] NF-120581B anuclear transcription factor [8] was first identified in 1986by Sen and Baltimore [9] It was initially observed to be atranscription factor binding to the intronic enhancer of thekappa light chain gene (the 120581B site) in B cells but it waslater shown to be present in every cell type [10] AfterwardsNF-120581B emerged as a major regulator of more than 200genes involved in diverse process such as cell survival and

Hindawi Publishing CorporationBioMed Research InternationalVolume 2014 Article ID 530381 6 pageshttpdxdoiorg1011552014530381

2 BioMed Research International

cell adhesion inflammation differentiation and growth [11]NF-120581B is activated by phosphorylation of I120581B120572 which iscatalyzed by an I120581B120572 Kinase (IKK) complex consisting ofIkK-120572 IkK-120573 IkK-120574 (also called NEMO) and other proteinsyet to be identified [12 13] After activation of NF-120581B I120581B120572is degraded and p50ndashp65 heterodimer is translocated to thenucleus binds to the DNA (at the promoter region) andactivates gene [14 15] Currently studies show that activationof the transcription factor nuclear factor (NF) 120581B is a novelmechanism of chemoresistance in NSCLC and other tumors[16 17] NF-120581B1 is inhibited by IkB proteins (eg NF-120581BIA)[18] Phosphorylation of serine residues on the I-kappa-Bproteins by kinases andmarks them for degradation therebyallowing activation of the NF-120581B complex [19] As the firstpotential functional NF-120581B1 genetic variation was identified94 insertiondeletion ATTG located between two receivedkey promoter regulatory elements in the NF-120581B1 gene ATTGdeletion causes the loss of binding to nuclear proteinswhich leads to reduced promoter activity [20] The NF-120581B -94insdelATTG polymorphism of the NF-120581B1 gene promoterregion which causes a functional effect and NF-120581BIA 31015840UTRArarrG polymorphism have been shown to be associated withvarious inflammatory diseases and cancers [21] The aim ofthis study is firstly to investigate the NF-120581B1 -94insdelATTGand NF-120581BIA 31015840UTR ArarrG polymorphism together in theTurkish population

2 Materials and Methods

21 Study Groups Ninety-five primary non-small cell lungcancer (NSCLC) patients and 99 healthy individuals wereincluded in the study NSCLC patients were recruited fromthe Yedikule Chest Diseases and Thoracic Surgery TrainingResearch Hospital Istanbul The diagnosis of NSCLC wasmade by the pathologist based on histopathological exam-ination In NSCLC group all subjects were diagnosed andconfirmed with histopathological examination They wereall newly diagnosed without a history of prior radiotherapyandor chemotherapy Exclusion criteria included primaryextra pulmonary malignancy small cell lung cancer a his-tory of malignant disease and withdrawal of consent andpatient aged less than 18 Pathological staging informationon all NSCLC cases was confirmed by manual review ofthe pathology reports and clinical charts Nodal status wascategorized as no regional lymph nodes affected (N0) or atleast one nodal metastasis The mean ages of the patientsand controls were 612 plusmn 983 years and 5749 plusmn 1083 yearsrespectively The percentage of females was 63 for patientsand 323 for controls and percentage ofmales was 937 forpatients and 677 for controls 99 healthy subjects withoutany malignancy were selected for the control group that iscomprised only of individuals with a negative family historyof cancer The patient and control groups were matchedfor age All participants signed an informed consent beforeenrollment and Institutional Ethical committee approval wasobtained for the study

331bp242bp 285 bp

240bp

II DD II DD ID II II II DD II1 2 3 4 5 6 7 8 9 10 11

Figure 1 Representative genotypes of NF-120581B1 -94insdelATTGpolymorphism Lane 1 marker DNA ladder Lanes 2 4 7 8 9and 11 insins (ATTG2ATTG2) genotypes Lanes 3 5 and 10deldel (ATTG1ATTG1) genotypes Lane 6 heterozygous delins(ATTG1ATTG2) genotypes

1 2

GG3

AA4

GG5

AG6

AG

400bp300bp

100 bp

424bp316bp

108 bp

Figure 2 Representative genotypes of NF-120581BIA 31015840UTR ArarrGpolymorphism Lane 1 marker DNA ladder Lanes 2 and 4 GGmutant genotypes Lane 3 AA wild-type homozygous genotypesLanes 5 and 6 AG heterozygous genotypes

22 Polymorphism Analysis Blood samples from all studyparticipants were collected in EDTA-containing tubesGenomic DNA was extracted from peripheral wholeblood according to kit protocol (High Pure PCR TemplatePreparation Kit REF 11796828001 (Roche DiagnosticsGmbH Mannheim Germany) Genotyping was performedby polymerase chain reaction (PCR) and restriction fragmentlength polymorphism (RFLP) the procedures of PCR-RFLPare given in Table 1 Two separate PCR reactions wereused to detect the two types of polymorphism in NF-120581Bgene namely NF-120581B1 -94insdelATTG polymorphism andNF-120581BIA 31015840UTR ArarrG polymorphism The appropriateprimers were used to amplify the corresponding gene of thesubjects by PCR and the reaction products were digestedby using the appropriate enzyme and incubated at 37∘Covernight The digested products were analyzed on 3agarose gel stained with ethidium bromide and examinedunder transillumination (Figures 1 and 2) Each gel was readby two observers unaware of the subjectrsquos status In order toverify our PCR-RFLP results we repeated PCR-RFLP stage 2times for each of selected subject The expected results afterrestriction for each gene were also given in Table 1

23 Statistical Analysis Statistical analyses were performedusing the SPSS software package (revision 130 SPSS Inc

BioMed Research International 3

Table 1 PCR and RFLP procedures and products of NF-120581B1 -94insdelATTG and NF-120581BIA 31015840UTR ArarrG

Primers (forward and reverse) PCR product Restriction enzyme Restriction products

NF-120581B151015840-TGGGCACAAGTCGTTTATGA-31015840

285 bp Van91l (PflMI)ATTG2ATTG2 (insins) 240 bp 45 bp

51015840-CTGGAGCCGGTAGGGAAG-31015840 ATTG1ATTG1 (deldel) 281ATTG2ATTG1 (insdel) 281 240 45

NF-120581BIA51015840-GGCTGAAAGAACATGGACTTG-31015840

424 bp HaeIIIAA wild type 424

51015840-GTACACCATTTACAGGAGGG-31015840 AG heterozygous 316 108GG mutant 424 316 108

Table 2 Distribution of NF-120581B1 and NF-120581BIA genotypes and allele in NSCLC patients and controls

Genotypesalleles Controls 119899 () Patients 119899 () OR (95 CI) 119875 valueNF-120581B1

II 46 (4647) 35 (3684) ReferenceID 47 (4747) 44 (4632) 123 (067ndash225) 0500DD 6 (606) 16 (1684) 350 (124ndash987) 0014ID + DD 53 (5353) 60 (6316) 149 (084ndash264) 0174I allele 139 (702) 114 (60) ReferenceD allele 59 (298) 76 (40) 157 (103ndash239) 0035

NF-120581BIAAA 21 (2121) 17 (179) ReferenceAG 45 (4546) 45 (4736) 124 (058ndash265) 0587GG 33 (3333) 33 (3474) 124 (055ndash275) 0605AG + GG 78 (7879) 78 (821) 124 (061ndash252) 0561A allele 87 (4394) 79 (4158) ReferenceG allele 111 (5606) 111 (5842) 110 (074ndash165) 0639

OR Odds ratio CI confidence interval

Chicago IL USA) Data were expressed as means plusmn SDDifferences in the distribution of NF-120581B1 -94insdelATTGand NF-120581BIA 31015840UTR ArarrG genotypes or alleles betweencases and controls were tested using the Chi-square statisticrespectively (Table 2) Differences in characteristics betweenNSCLC patients and controls were assessed with Fisherrsquosexact test as well as disparities in genotype and allelefrequencies Relative risk at 95 confidence intervals (CI)was calculated as the odds ratio (OR) Values 119875 lt 005 wereconsidered statistically significant A multivariate logisticregression model was performed to investigate possibleeffects of genotypes and alleles after adjustment for age

3 Results

In this study we examined 194 volunteers 95 NSCLC (89males 6 females) patients and 99 healthy people (67 males32 females) detecting any chronic disease or any evidence ofmalignancy Distribution of NF-120581B1 and NF-120581BIA genotypesaccording to clinic features in NSCLC patients is shownin Table 3 The distribution of the NF-120581B1 -94insdelATTGgenotypes in control and NSCLC patients was found tobe significantly different (119875 0048) It was evaluated thatindividuals carrying DD genotype had 35-fold increased riskfor NSCLC (119875 0014 1205942 5605 OR 350 95 CI 124ndash987)No statistically significant differences between groups were

observed when the NF-120581BIA 31015840UTR ArarrG genotypes distri-butions were compared (119875 0844) A significant correlationbetween genotype combinations of NF-120581B1 and NF-120581BIA(DDAG genotype) and NSCLC risk was found compared toall other combinations (119875 0025 OR 5035 95 CI 1067ndash2414) and DDAG genotype had increased risk for NSCLCThe prevalence of IIAA genotype combinations versus toall other combinations was 53 in patients and 121 inthe control group but there are no statistically significantdifferences between groups (119875 0091 OR 0403 95 CI0136ndash1191) The results of multivariate logistic regressionanalysis are presented in Table 4 Gender age (lt57ge57)and NF-120581B1 DD genotype were associated with NSCLC inunivariate analysis and additionally these were associatedwith this disease in multivariate logistic regression analysis

4 Discussion

A functional polymorphism in the NF-120581B1 gene promoterregion (-94insdelATTG) has been identified and associatedwith both chronic inflammatory diseases and malignantdiseases [22] NF-120581B is inactivated in the cytoplasm byI120581B120572 120573 or 120574 and the most common protein of this familyis the NF-120581B inhibitor 120572 (NF-120581BIA) [23] -94insdelATTGpolymorphism has evidence from two independent func-tional assays in vitro promoter activity and differential anunidentified nuclear protein binding that the specific allele

4 BioMed Research International

Table 3 Distribution of NF-120581B1 and NF-120581BIA genotypes with clinic features in NSCLC patients

NF-120581B1 NF-120581BIAII ID DD

119875 value AA AG GG119875 value

119899 () 119899 () 119899 () 119899 () 119899 () 119899 ()Sex

Men 33 (3710) 41 (4610) 15 (1680) 0980 15 (1690) 43 (4830) 31 (3480) 0570Women 2 (3330) 3 (5000) 1 (1670) 2 (3330) 2 (3330) 2 (3330)

Agelt57 10 (294) 17 (50) 7 (206) 0496 5 (147) 19 (559) 10 (294) 0567ge57 27 (415) 27 (415) 11 (169) 12 (185) 29 (446) 24 (369)

Smoking (boxyear)lt50 25 (4810) 24 (4620) 3 (580) 002 10 (1920) 25 (4810) 17 (3270) 0876ge50 10 (2330) 20 (4650) 13 (3020) 7 (1630) 20 (4650) 16 (3720)

Alcohol consumptionNo 17 (3150) 28 (5190) 9 (1670) 0405 6 (1110) 31 (5740) 17 (3150) 0044Yes 18 (4390) 16 (3900) 7 (1710) 11 (2680) 14 (3410) 16 (3900)

HistopathologySquamous 16 (4570) 16 (4570) 3 (860) 0179 7 (2000) 20 (5710) 8 (2290) 0173Nonsquamous 19 (3170) 28 (4670) 13 (2170) 10 (1670) 25 (4170) 25 (4170)

Total proteinlt6 gL 2 (5000) 0 (0) 2 (5000) 0087 0 (0) 3 (7500) 1 (2500) 0452ge6 gL 31 (3830) 38 (4690) 12 (1480) 16 (1980) 37 (4570) 28 (3460)

Albuminlt3 gL 6 (5450) 1 (910) 4 (3640) 0019 1 (910) 6 (5450) 4 (3640) 0635ge3 gL 26 (3470) 39 (5200) 10 (1330) 16 (2130) 35 (4670) 24 (3200)

Calciumlt10mgDl 30 (4000) 30 (4000) 15 (2000) 0001 16 (2130) 35 (4670) 24 (3200) 0506ge10mgdL 0 (0) 11 (100) 0 (0) 1 (910) 7 (6360) 3 (2730)

LDHlt250UL 22 (3610) 28 (4590) 11 (1800) 0644 13 (2130) 31 (5080) 17 (2790) 0249ge250UL 10 (4760) 8 (3810) 3 (1430) 3 (1430) 8 (3810) 10 (4760)

Table 4 The results of multivariate logistic regression

Covariates 119875 value Exp (B) 95 CI for Exp (B)Gender lt0001 7866 2915ndash21231Age (lt57ge57) lt0001 5074 2633ndash9776NF-120581B1 DD genotype 0035 3167 1086ndash9234

inherited likely has functional consequences [24] NF-120581BIA31015840UTR ArarrG polymorphismmay affect mRNA stability andtranslational efficacy or conduces to differential nuclear RNAprocessing or export also cannot be completely excludedMany studies have been conducted to investigate a possibleassociation between NF-120581B1 -94insdelATTG and NF-120581BIA31015840UTR ArarrG polymorphism and both inflammatory dis-eases and various cancer types [25] However no data areavailable in the English literature to report the associationwith NSCLC to date Our study is the initial report on thesetwo forms of polymorphism (both NF-120581B1 -94insdelATTGand NF-120581BIA 31015840UTR ArarrG) studied together in NSCLCpatients to our knowledge The genotypic combinations ofNF-120581B1 and NF-120581B2 polymorphism have been shown to be

associated with the development of common inflammatorydiseases including ulcerative colitis (UC) Crohnrsquos diseaseand Type I diabetes as well as susceptibility of severalcancers such as oral squamous cell carcinoma and colorectalcancer [26] It can be concluded that previous studies haveconflicting results [27] Oliver et al suggest that the NF-120581B1-94insdelATTG gene variation previously associated withUC susceptibility in North Americans does not influenceeither susceptibility or phenotype of UC in the Spanishpopulation [28] In this study we performed a risk associationbetween the NF-120581B1 -94insdelATTG promoter polymor-phism and NSCLC The -94insdelATTG polymorphism hasbeen shown as a first potential functional NF-120581B1 polymor-phism by Karban et al Nuclear proteins from normal humancolon tissue showed significant binding to -94insATTGbut not to -94delATTG containing oligonucleotides NF-120581B1 promoterexon 1 luciferase reporter plasmid constructscontaining the -94delATTG allele and transfected into eitherHeLa or HT-29 cell lines showed low promoter activity morethan comparable constructs containing the -94insATTGallele Therefore it is known that D allele promoter activ-ity is low and I allele promoter activity is high Previous

BioMed Research International 5

studies have suggested that D allele may result in decreasedNF-120581B1 message and hence decreased p50p105 NF-120581B pro-tein production leads to increased inflammatory responseOtherwise a potential explanation of decreased NF-120581B1 Dallele gene expression may be the resulting decreases inNF-120581B p50p65 heterodimers that are major mediators ofinflammation [29]

Defects in components that regulate NF-120581B release fromI120581B120572 result in constitutive or decreased NF-120581B activationThese components may be any of the kinases phosphatasesor other signal transducers normally involved in NF-120581B-activation pathways [30] Sonenshein suggests that alterationsof NF-120581B1 expression play an important role in the protec-tion of cells from apoptosis [31] NF-120581B1 activity has beenobserved in various types of cancer as well as colorectalcancer and breast cancer to contribute to tumor angiogenesisinvasion and progression [32] Therefore the variants ofthe NF-120581B1 gene could be expected to have an effect oncell death and thus carcinogenesis NF-120581B1 31015840UTR ArarrGpolymorphism has functional effects on expression of theNF-120581BIA gene and altered NF-120581B transcription [33] Thereare many studies with different results on NF-120581BIA 31015840UTRpolymorphism in the literature [34] Our results suggestedthat NF-120581BIA polymorphism has no effect on risk of NSCLCIn conclusion we here clearly demonstrated that NF-120581B1 -94insdelATTG promoter polymorphism and the presenceof the DD genotype might have a risk factor for NSCLCpathogenesis in our ethnic population Larger trials thatincluded different ethnic groups are necessary to defineobjectively the correlation between NF-120581B1 -94insdelATTGpromoter and development of NSCLC as well as prognosis ofdisease

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

The present work was supported by a Grant from theScientific Research Projects Coordination Unit of IstanbulUniversity (Project no 10537) The authors would like tothank MSc Allison P Eronat BSc Nesibe Selma Guler andPhDAylaKarimova for their understanding and suggestionsin English grammar of our article and the Editor and anony-mous reviewers for their valuable comments and suggestionswhich were helpful in improving the paper

References

[1] A Jemal R Siegel E Ward et al ldquoCancer statistics 2006rdquo CACancer Journal for Clinicians vol 56 no 2 pp 106ndash130 2006

[2] S Sun J H Schiller M Spinola and J D Minna ldquoNewmolecularly targeted therapies for lung cancerrdquo The Journal ofClinical Investigation vol 117 no 10 pp 2740ndash2750 2007

[3] M Fleischhacker T Beinert and K Possinger ldquoMoleculargenetic characteristics of lung cancermdashuseful as ldquorealrdquo tumormarkersrdquo Lung Cancer vol 25 no 1 pp 7ndash24 1999

[4] X Tang D Liu S Shishodia et al ldquoNuclear factor-120581B (NF-120581B) is frequently expressed in lung cancer and preneoplasticlesionsrdquo Cancer vol 107 no 11 pp 2637ndash2646 2006

[5] R Doll and A B Hill ldquoThe mortality of doctors in relationto their smoking habits a preliminary reportrdquo British MedicalJournal vol 1 no 4877 pp 1451ndash1455 1954

[6] M Koti R J Gooding P Nuin et al ldquoIdentification ofthe IGF1PI3KNF120581BERK gene signalling networks associatedwith chemotherapy resistance and treatment response in high-grade serous epithelial ovarian cancerrdquo BMC Cancer vol 13article 549 2013

[7] W Chen Z Li L Bai and Y Lin ldquoNF-120581B in lung cancer acarcinogenesis mediator and a prevention and therapy targetrdquoFrontiers in Bioscience vol 16 no 3 pp 1172ndash1185 2011

[8] P H Salim M Jobim M Bredemeier et al ldquoInterleukin-10 gene promoter and NFKB1 promoter insertiondeletionpolymorphisms in systemic sclerosisrdquo Scandinavian Journal ofImmunology vol 77 no 2 pp 162ndash168 2013

[9] R Sen and D Baltimore ldquoMultiple nuclear factors interact withthe immunoglobulin enhancer sequencesrdquo Cell vol 46 no 5pp 705ndash716 1986

[10] G Sethi B Sung and B B Aggarwal ldquoNuclear factor-120581Bactivation from bench to bedsiderdquo Experimental Biology andMedicine vol 233 no 1 pp 21ndash31 2008

[11] M S Hayden and S Ghosh ldquoSignaling to NF-120581Brdquo Genes ampDevelopment vol 18 no 18 pp 2195ndash2224 2004

[12] M Karin ldquoThe beginning of the end I120581B kinase (IKK) and NF-120581B activationrdquoThe Journal of Biological Chemistry vol 274 no39 pp 27339ndash27342 1999

[13] S Khan Z Lopez-Dee R Kumar and J Ling ldquoActivationof NFkB is a novel mechanism of pro-survival activity ofglucocorticoids in breast cancer cellsrdquo Cancer Letters vol 337no 1 pp 90ndash95 2013

[14] A C Bharti and B B Aggarwal ldquoNuclear factor-120581B and cancerits role in prevention and therapyrdquo Biochemical Pharmacologyvol 64 no 5-6 pp 883ndash888 2002

[15] D D Datta A Datta S Bhattacharjya and S RoychoudhuryldquoNF-120581B mediated transcriptional repression of acid modifyinghormone gastrinrdquo PLoS ONE vol 8 no 8 Article ID e734092013

[16] D R Jones R M Broad L V Madrid A S Baldwin Jr andMW Mayo ldquoInhibition of NF-120581B sensitizes non-small cell lungcancer cells to chemotherapy-induced apoptosisrdquoTheAnnals ofThoracic Surgery vol 70 no 3 pp 930ndash936 2000

[17] C-Y Wang J C Cusack Jr R Liu and A S Baldwin JrldquoControl of inducible chemoresistance enhanced anti-tumortherapy through increased apoptosis by inhibition of NF-120581BrdquoNature Medicine vol 5 no 4 pp 412ndash417 1999

[18] M Adamzik S Schafer U H Frey et al ldquoTheNFKB1 promoterpolymorphism (minus94insdelATTG) alters nuclear translocationof NF-120581B1 in monocytes after lipopolysaccharide stimulationand is associated with increased mortality in sepsisrdquo Anesthe-siology vol 118 no 1 pp 123ndash133 2013

[19] H Nakshatri P Bhat-Nakshatri D A Martin R J Goulet Jrand G W Sledge Jr ldquoConstitutive activation of NF-120581B duringprogression of breast cancer to hormone-independent growthrdquoMolecular andCellular Biology vol 17 no 7 pp 3629ndash3639 1997

[20] D M Hegazy D A OrsquoReilly B M Yang A D Hodgkinson BA Millward and A G Demaine ldquoNF120581B polymorphisms andsusceptibility to type 1 diabetesrdquo Genes amp Immunity vol 2 no6 pp 304ndash308 2001

6 BioMed Research International

[21] A F Gazdar ldquoDNA repair and survival in lung cancermdashthe twofaces of Janusrdquo The New England Journal of Medicine vol 356no 8 pp 771ndash773 2007

[22] J Vangsted T W Klausen N Abildgaard et al ldquoSinglenucleotide polymorphisms in the promoter region of the IL1Bgene influence outcome in multiple myeloma patients treatedwith high-dose chemotherapy independently of relapse treat-ment with thalidomide and bortezomibrdquoAnnals of Hematologyvol 90 no 10 pp 1173ndash1181 2011

[23] S Song D Chen J Lu et al ldquoNF120581B1 and NF120581BIA polymor-phisms are associatedwith increased risk for sporadic colorectalcancer in a southern Chinese populationrdquo PLoS ONE vol 6 no6 Article ID e21726 2011

[24] M Liang X Xu Y Gong Y Tang and L Lin ldquoRisk associationbetween theNF-120581B1minus94insdelATTGpromoter polymorphismand inflammatory bowel diseases a meta-analysisrdquo DigestiveDiseases and Sciences vol 57 no 9 pp 2304ndash2309 2012

[25] C W Cheng J L Su C W Lin et al ldquoEffects of NFKB1andNFKBIA gene polymorphisms on hepatocellular carcinomasusceptibility and clinicopathological featuresrdquo PLoS ONE vol8 no 2 Article ID e56130 2013

[26] D Vu E Tellez-Corrales P Sakharkar et al ldquoImpact of NF-120581B gene polymorphism on allograft outcome in Hispanic renaltransplant recipientsrdquo Transplant Immunology vol 28 no 1 pp18ndash23 2013

[27] Y-F Zou F-L Yuan X-L Feng et al ldquoAssociation betweenNFKB1 minus94insdelATTG promoter polymorphism and cancerrisk a meta-analysisrdquoCancer Investigation vol 29 no 1 pp 78ndash85 2011

[28] J Oliver M Gomez-Garcıa L Paco et al ldquoA functionalpolymorphism of the NFKB1 promoter is not associated withulcerative colitis in a Spanish populationrdquo Inflammatory BowelDiseases vol 11 no 6 pp 576ndash579 2005

[29] A S Karban T Okazaki C I M Panhuysen et al ldquoFunctionalannotation of a novel NFKB1 promoter polymorphism thatincreases risk for ulcerative colitisrdquoHuman Molecular Geneticsvol 13 no 1 pp 35ndash45 2004

[30] R C Bargou F Emmerich D Krappmann et al ldquoConstitutivenuclear factor-120581B-RelA activation is required for proliferationand survival of Hodgkinrsquos disease tumor cellsrdquo The Journal ofClinical Investigation vol 100 no 12 pp 2961ndash2969 1997

[31] G E Sonenshein ldquoRelNF-120581B transcription factors and thecontrol of apoptosisrdquo Seminars in Cancer Biology vol 8 no 2pp 113ndash119 1997

[32] X Dolcet D Llobet J Pallares and X Matias-Guiu ldquoNF-120581Bin development and progression of human cancerrdquo VirchowsArchiv vol 446 no 5 pp 475ndash482 2005

[33] D Glavac M Ravnik-Glavac S J OrsquoBrien and M DeanldquoPolymorphisms in the 31015840 untranslated region of the I120581BMAD-3 (NFKBI) gene located on chromosome 14rdquo Human Geneticsvol 93 no 6 pp 694ndash696 1994

[34] J Gao D Pfeifer L-J He et al ldquoAssociation of NFKBIApolymorphism with colorectal cancer risk and prognosis inSwedish and Chinese populationsrdquo Scandinavian Journal ofGastroenterology vol 42 no 3 pp 345ndash350 2007

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2 BioMed Research International

cell adhesion inflammation differentiation and growth [11]NF-120581B is activated by phosphorylation of I120581B120572 which iscatalyzed by an I120581B120572 Kinase (IKK) complex consisting ofIkK-120572 IkK-120573 IkK-120574 (also called NEMO) and other proteinsyet to be identified [12 13] After activation of NF-120581B I120581B120572is degraded and p50ndashp65 heterodimer is translocated to thenucleus binds to the DNA (at the promoter region) andactivates gene [14 15] Currently studies show that activationof the transcription factor nuclear factor (NF) 120581B is a novelmechanism of chemoresistance in NSCLC and other tumors[16 17] NF-120581B1 is inhibited by IkB proteins (eg NF-120581BIA)[18] Phosphorylation of serine residues on the I-kappa-Bproteins by kinases andmarks them for degradation therebyallowing activation of the NF-120581B complex [19] As the firstpotential functional NF-120581B1 genetic variation was identified94 insertiondeletion ATTG located between two receivedkey promoter regulatory elements in the NF-120581B1 gene ATTGdeletion causes the loss of binding to nuclear proteinswhich leads to reduced promoter activity [20] The NF-120581B -94insdelATTG polymorphism of the NF-120581B1 gene promoterregion which causes a functional effect and NF-120581BIA 31015840UTRArarrG polymorphism have been shown to be associated withvarious inflammatory diseases and cancers [21] The aim ofthis study is firstly to investigate the NF-120581B1 -94insdelATTGand NF-120581BIA 31015840UTR ArarrG polymorphism together in theTurkish population

2 Materials and Methods

21 Study Groups Ninety-five primary non-small cell lungcancer (NSCLC) patients and 99 healthy individuals wereincluded in the study NSCLC patients were recruited fromthe Yedikule Chest Diseases and Thoracic Surgery TrainingResearch Hospital Istanbul The diagnosis of NSCLC wasmade by the pathologist based on histopathological exam-ination In NSCLC group all subjects were diagnosed andconfirmed with histopathological examination They wereall newly diagnosed without a history of prior radiotherapyandor chemotherapy Exclusion criteria included primaryextra pulmonary malignancy small cell lung cancer a his-tory of malignant disease and withdrawal of consent andpatient aged less than 18 Pathological staging informationon all NSCLC cases was confirmed by manual review ofthe pathology reports and clinical charts Nodal status wascategorized as no regional lymph nodes affected (N0) or atleast one nodal metastasis The mean ages of the patientsand controls were 612 plusmn 983 years and 5749 plusmn 1083 yearsrespectively The percentage of females was 63 for patientsand 323 for controls and percentage ofmales was 937 forpatients and 677 for controls 99 healthy subjects withoutany malignancy were selected for the control group that iscomprised only of individuals with a negative family historyof cancer The patient and control groups were matchedfor age All participants signed an informed consent beforeenrollment and Institutional Ethical committee approval wasobtained for the study

331bp242bp 285 bp

240bp

II DD II DD ID II II II DD II1 2 3 4 5 6 7 8 9 10 11

Figure 1 Representative genotypes of NF-120581B1 -94insdelATTGpolymorphism Lane 1 marker DNA ladder Lanes 2 4 7 8 9and 11 insins (ATTG2ATTG2) genotypes Lanes 3 5 and 10deldel (ATTG1ATTG1) genotypes Lane 6 heterozygous delins(ATTG1ATTG2) genotypes

1 2

GG3

AA4

GG5

AG6

AG

400bp300bp

100 bp

424bp316bp

108 bp

Figure 2 Representative genotypes of NF-120581BIA 31015840UTR ArarrGpolymorphism Lane 1 marker DNA ladder Lanes 2 and 4 GGmutant genotypes Lane 3 AA wild-type homozygous genotypesLanes 5 and 6 AG heterozygous genotypes

22 Polymorphism Analysis Blood samples from all studyparticipants were collected in EDTA-containing tubesGenomic DNA was extracted from peripheral wholeblood according to kit protocol (High Pure PCR TemplatePreparation Kit REF 11796828001 (Roche DiagnosticsGmbH Mannheim Germany) Genotyping was performedby polymerase chain reaction (PCR) and restriction fragmentlength polymorphism (RFLP) the procedures of PCR-RFLPare given in Table 1 Two separate PCR reactions wereused to detect the two types of polymorphism in NF-120581Bgene namely NF-120581B1 -94insdelATTG polymorphism andNF-120581BIA 31015840UTR ArarrG polymorphism The appropriateprimers were used to amplify the corresponding gene of thesubjects by PCR and the reaction products were digestedby using the appropriate enzyme and incubated at 37∘Covernight The digested products were analyzed on 3agarose gel stained with ethidium bromide and examinedunder transillumination (Figures 1 and 2) Each gel was readby two observers unaware of the subjectrsquos status In order toverify our PCR-RFLP results we repeated PCR-RFLP stage 2times for each of selected subject The expected results afterrestriction for each gene were also given in Table 1

23 Statistical Analysis Statistical analyses were performedusing the SPSS software package (revision 130 SPSS Inc

BioMed Research International 3

Table 1 PCR and RFLP procedures and products of NF-120581B1 -94insdelATTG and NF-120581BIA 31015840UTR ArarrG

Primers (forward and reverse) PCR product Restriction enzyme Restriction products

NF-120581B151015840-TGGGCACAAGTCGTTTATGA-31015840

285 bp Van91l (PflMI)ATTG2ATTG2 (insins) 240 bp 45 bp

51015840-CTGGAGCCGGTAGGGAAG-31015840 ATTG1ATTG1 (deldel) 281ATTG2ATTG1 (insdel) 281 240 45

NF-120581BIA51015840-GGCTGAAAGAACATGGACTTG-31015840

424 bp HaeIIIAA wild type 424

51015840-GTACACCATTTACAGGAGGG-31015840 AG heterozygous 316 108GG mutant 424 316 108

Table 2 Distribution of NF-120581B1 and NF-120581BIA genotypes and allele in NSCLC patients and controls

Genotypesalleles Controls 119899 () Patients 119899 () OR (95 CI) 119875 valueNF-120581B1

II 46 (4647) 35 (3684) ReferenceID 47 (4747) 44 (4632) 123 (067ndash225) 0500DD 6 (606) 16 (1684) 350 (124ndash987) 0014ID + DD 53 (5353) 60 (6316) 149 (084ndash264) 0174I allele 139 (702) 114 (60) ReferenceD allele 59 (298) 76 (40) 157 (103ndash239) 0035

NF-120581BIAAA 21 (2121) 17 (179) ReferenceAG 45 (4546) 45 (4736) 124 (058ndash265) 0587GG 33 (3333) 33 (3474) 124 (055ndash275) 0605AG + GG 78 (7879) 78 (821) 124 (061ndash252) 0561A allele 87 (4394) 79 (4158) ReferenceG allele 111 (5606) 111 (5842) 110 (074ndash165) 0639

OR Odds ratio CI confidence interval

Chicago IL USA) Data were expressed as means plusmn SDDifferences in the distribution of NF-120581B1 -94insdelATTGand NF-120581BIA 31015840UTR ArarrG genotypes or alleles betweencases and controls were tested using the Chi-square statisticrespectively (Table 2) Differences in characteristics betweenNSCLC patients and controls were assessed with Fisherrsquosexact test as well as disparities in genotype and allelefrequencies Relative risk at 95 confidence intervals (CI)was calculated as the odds ratio (OR) Values 119875 lt 005 wereconsidered statistically significant A multivariate logisticregression model was performed to investigate possibleeffects of genotypes and alleles after adjustment for age

3 Results

In this study we examined 194 volunteers 95 NSCLC (89males 6 females) patients and 99 healthy people (67 males32 females) detecting any chronic disease or any evidence ofmalignancy Distribution of NF-120581B1 and NF-120581BIA genotypesaccording to clinic features in NSCLC patients is shownin Table 3 The distribution of the NF-120581B1 -94insdelATTGgenotypes in control and NSCLC patients was found tobe significantly different (119875 0048) It was evaluated thatindividuals carrying DD genotype had 35-fold increased riskfor NSCLC (119875 0014 1205942 5605 OR 350 95 CI 124ndash987)No statistically significant differences between groups were

observed when the NF-120581BIA 31015840UTR ArarrG genotypes distri-butions were compared (119875 0844) A significant correlationbetween genotype combinations of NF-120581B1 and NF-120581BIA(DDAG genotype) and NSCLC risk was found compared toall other combinations (119875 0025 OR 5035 95 CI 1067ndash2414) and DDAG genotype had increased risk for NSCLCThe prevalence of IIAA genotype combinations versus toall other combinations was 53 in patients and 121 inthe control group but there are no statistically significantdifferences between groups (119875 0091 OR 0403 95 CI0136ndash1191) The results of multivariate logistic regressionanalysis are presented in Table 4 Gender age (lt57ge57)and NF-120581B1 DD genotype were associated with NSCLC inunivariate analysis and additionally these were associatedwith this disease in multivariate logistic regression analysis

4 Discussion

A functional polymorphism in the NF-120581B1 gene promoterregion (-94insdelATTG) has been identified and associatedwith both chronic inflammatory diseases and malignantdiseases [22] NF-120581B is inactivated in the cytoplasm byI120581B120572 120573 or 120574 and the most common protein of this familyis the NF-120581B inhibitor 120572 (NF-120581BIA) [23] -94insdelATTGpolymorphism has evidence from two independent func-tional assays in vitro promoter activity and differential anunidentified nuclear protein binding that the specific allele

4 BioMed Research International

Table 3 Distribution of NF-120581B1 and NF-120581BIA genotypes with clinic features in NSCLC patients

NF-120581B1 NF-120581BIAII ID DD

119875 value AA AG GG119875 value

119899 () 119899 () 119899 () 119899 () 119899 () 119899 ()Sex

Men 33 (3710) 41 (4610) 15 (1680) 0980 15 (1690) 43 (4830) 31 (3480) 0570Women 2 (3330) 3 (5000) 1 (1670) 2 (3330) 2 (3330) 2 (3330)

Agelt57 10 (294) 17 (50) 7 (206) 0496 5 (147) 19 (559) 10 (294) 0567ge57 27 (415) 27 (415) 11 (169) 12 (185) 29 (446) 24 (369)

Smoking (boxyear)lt50 25 (4810) 24 (4620) 3 (580) 002 10 (1920) 25 (4810) 17 (3270) 0876ge50 10 (2330) 20 (4650) 13 (3020) 7 (1630) 20 (4650) 16 (3720)

Alcohol consumptionNo 17 (3150) 28 (5190) 9 (1670) 0405 6 (1110) 31 (5740) 17 (3150) 0044Yes 18 (4390) 16 (3900) 7 (1710) 11 (2680) 14 (3410) 16 (3900)

HistopathologySquamous 16 (4570) 16 (4570) 3 (860) 0179 7 (2000) 20 (5710) 8 (2290) 0173Nonsquamous 19 (3170) 28 (4670) 13 (2170) 10 (1670) 25 (4170) 25 (4170)

Total proteinlt6 gL 2 (5000) 0 (0) 2 (5000) 0087 0 (0) 3 (7500) 1 (2500) 0452ge6 gL 31 (3830) 38 (4690) 12 (1480) 16 (1980) 37 (4570) 28 (3460)

Albuminlt3 gL 6 (5450) 1 (910) 4 (3640) 0019 1 (910) 6 (5450) 4 (3640) 0635ge3 gL 26 (3470) 39 (5200) 10 (1330) 16 (2130) 35 (4670) 24 (3200)

Calciumlt10mgDl 30 (4000) 30 (4000) 15 (2000) 0001 16 (2130) 35 (4670) 24 (3200) 0506ge10mgdL 0 (0) 11 (100) 0 (0) 1 (910) 7 (6360) 3 (2730)

LDHlt250UL 22 (3610) 28 (4590) 11 (1800) 0644 13 (2130) 31 (5080) 17 (2790) 0249ge250UL 10 (4760) 8 (3810) 3 (1430) 3 (1430) 8 (3810) 10 (4760)

Table 4 The results of multivariate logistic regression

Covariates 119875 value Exp (B) 95 CI for Exp (B)Gender lt0001 7866 2915ndash21231Age (lt57ge57) lt0001 5074 2633ndash9776NF-120581B1 DD genotype 0035 3167 1086ndash9234

inherited likely has functional consequences [24] NF-120581BIA31015840UTR ArarrG polymorphismmay affect mRNA stability andtranslational efficacy or conduces to differential nuclear RNAprocessing or export also cannot be completely excludedMany studies have been conducted to investigate a possibleassociation between NF-120581B1 -94insdelATTG and NF-120581BIA31015840UTR ArarrG polymorphism and both inflammatory dis-eases and various cancer types [25] However no data areavailable in the English literature to report the associationwith NSCLC to date Our study is the initial report on thesetwo forms of polymorphism (both NF-120581B1 -94insdelATTGand NF-120581BIA 31015840UTR ArarrG) studied together in NSCLCpatients to our knowledge The genotypic combinations ofNF-120581B1 and NF-120581B2 polymorphism have been shown to be

associated with the development of common inflammatorydiseases including ulcerative colitis (UC) Crohnrsquos diseaseand Type I diabetes as well as susceptibility of severalcancers such as oral squamous cell carcinoma and colorectalcancer [26] It can be concluded that previous studies haveconflicting results [27] Oliver et al suggest that the NF-120581B1-94insdelATTG gene variation previously associated withUC susceptibility in North Americans does not influenceeither susceptibility or phenotype of UC in the Spanishpopulation [28] In this study we performed a risk associationbetween the NF-120581B1 -94insdelATTG promoter polymor-phism and NSCLC The -94insdelATTG polymorphism hasbeen shown as a first potential functional NF-120581B1 polymor-phism by Karban et al Nuclear proteins from normal humancolon tissue showed significant binding to -94insATTGbut not to -94delATTG containing oligonucleotides NF-120581B1 promoterexon 1 luciferase reporter plasmid constructscontaining the -94delATTG allele and transfected into eitherHeLa or HT-29 cell lines showed low promoter activity morethan comparable constructs containing the -94insATTGallele Therefore it is known that D allele promoter activ-ity is low and I allele promoter activity is high Previous

BioMed Research International 5

studies have suggested that D allele may result in decreasedNF-120581B1 message and hence decreased p50p105 NF-120581B pro-tein production leads to increased inflammatory responseOtherwise a potential explanation of decreased NF-120581B1 Dallele gene expression may be the resulting decreases inNF-120581B p50p65 heterodimers that are major mediators ofinflammation [29]

Defects in components that regulate NF-120581B release fromI120581B120572 result in constitutive or decreased NF-120581B activationThese components may be any of the kinases phosphatasesor other signal transducers normally involved in NF-120581B-activation pathways [30] Sonenshein suggests that alterationsof NF-120581B1 expression play an important role in the protec-tion of cells from apoptosis [31] NF-120581B1 activity has beenobserved in various types of cancer as well as colorectalcancer and breast cancer to contribute to tumor angiogenesisinvasion and progression [32] Therefore the variants ofthe NF-120581B1 gene could be expected to have an effect oncell death and thus carcinogenesis NF-120581B1 31015840UTR ArarrGpolymorphism has functional effects on expression of theNF-120581BIA gene and altered NF-120581B transcription [33] Thereare many studies with different results on NF-120581BIA 31015840UTRpolymorphism in the literature [34] Our results suggestedthat NF-120581BIA polymorphism has no effect on risk of NSCLCIn conclusion we here clearly demonstrated that NF-120581B1 -94insdelATTG promoter polymorphism and the presenceof the DD genotype might have a risk factor for NSCLCpathogenesis in our ethnic population Larger trials thatincluded different ethnic groups are necessary to defineobjectively the correlation between NF-120581B1 -94insdelATTGpromoter and development of NSCLC as well as prognosis ofdisease

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

The present work was supported by a Grant from theScientific Research Projects Coordination Unit of IstanbulUniversity (Project no 10537) The authors would like tothank MSc Allison P Eronat BSc Nesibe Selma Guler andPhDAylaKarimova for their understanding and suggestionsin English grammar of our article and the Editor and anony-mous reviewers for their valuable comments and suggestionswhich were helpful in improving the paper

References

[1] A Jemal R Siegel E Ward et al ldquoCancer statistics 2006rdquo CACancer Journal for Clinicians vol 56 no 2 pp 106ndash130 2006

[2] S Sun J H Schiller M Spinola and J D Minna ldquoNewmolecularly targeted therapies for lung cancerrdquo The Journal ofClinical Investigation vol 117 no 10 pp 2740ndash2750 2007

[3] M Fleischhacker T Beinert and K Possinger ldquoMoleculargenetic characteristics of lung cancermdashuseful as ldquorealrdquo tumormarkersrdquo Lung Cancer vol 25 no 1 pp 7ndash24 1999

[4] X Tang D Liu S Shishodia et al ldquoNuclear factor-120581B (NF-120581B) is frequently expressed in lung cancer and preneoplasticlesionsrdquo Cancer vol 107 no 11 pp 2637ndash2646 2006

[5] R Doll and A B Hill ldquoThe mortality of doctors in relationto their smoking habits a preliminary reportrdquo British MedicalJournal vol 1 no 4877 pp 1451ndash1455 1954

[6] M Koti R J Gooding P Nuin et al ldquoIdentification ofthe IGF1PI3KNF120581BERK gene signalling networks associatedwith chemotherapy resistance and treatment response in high-grade serous epithelial ovarian cancerrdquo BMC Cancer vol 13article 549 2013

[7] W Chen Z Li L Bai and Y Lin ldquoNF-120581B in lung cancer acarcinogenesis mediator and a prevention and therapy targetrdquoFrontiers in Bioscience vol 16 no 3 pp 1172ndash1185 2011

[8] P H Salim M Jobim M Bredemeier et al ldquoInterleukin-10 gene promoter and NFKB1 promoter insertiondeletionpolymorphisms in systemic sclerosisrdquo Scandinavian Journal ofImmunology vol 77 no 2 pp 162ndash168 2013

[9] R Sen and D Baltimore ldquoMultiple nuclear factors interact withthe immunoglobulin enhancer sequencesrdquo Cell vol 46 no 5pp 705ndash716 1986

[10] G Sethi B Sung and B B Aggarwal ldquoNuclear factor-120581Bactivation from bench to bedsiderdquo Experimental Biology andMedicine vol 233 no 1 pp 21ndash31 2008

[11] M S Hayden and S Ghosh ldquoSignaling to NF-120581Brdquo Genes ampDevelopment vol 18 no 18 pp 2195ndash2224 2004

[12] M Karin ldquoThe beginning of the end I120581B kinase (IKK) and NF-120581B activationrdquoThe Journal of Biological Chemistry vol 274 no39 pp 27339ndash27342 1999

[13] S Khan Z Lopez-Dee R Kumar and J Ling ldquoActivationof NFkB is a novel mechanism of pro-survival activity ofglucocorticoids in breast cancer cellsrdquo Cancer Letters vol 337no 1 pp 90ndash95 2013

[14] A C Bharti and B B Aggarwal ldquoNuclear factor-120581B and cancerits role in prevention and therapyrdquo Biochemical Pharmacologyvol 64 no 5-6 pp 883ndash888 2002

[15] D D Datta A Datta S Bhattacharjya and S RoychoudhuryldquoNF-120581B mediated transcriptional repression of acid modifyinghormone gastrinrdquo PLoS ONE vol 8 no 8 Article ID e734092013

[16] D R Jones R M Broad L V Madrid A S Baldwin Jr andMW Mayo ldquoInhibition of NF-120581B sensitizes non-small cell lungcancer cells to chemotherapy-induced apoptosisrdquoTheAnnals ofThoracic Surgery vol 70 no 3 pp 930ndash936 2000

[17] C-Y Wang J C Cusack Jr R Liu and A S Baldwin JrldquoControl of inducible chemoresistance enhanced anti-tumortherapy through increased apoptosis by inhibition of NF-120581BrdquoNature Medicine vol 5 no 4 pp 412ndash417 1999

[18] M Adamzik S Schafer U H Frey et al ldquoTheNFKB1 promoterpolymorphism (minus94insdelATTG) alters nuclear translocationof NF-120581B1 in monocytes after lipopolysaccharide stimulationand is associated with increased mortality in sepsisrdquo Anesthe-siology vol 118 no 1 pp 123ndash133 2013

[19] H Nakshatri P Bhat-Nakshatri D A Martin R J Goulet Jrand G W Sledge Jr ldquoConstitutive activation of NF-120581B duringprogression of breast cancer to hormone-independent growthrdquoMolecular andCellular Biology vol 17 no 7 pp 3629ndash3639 1997

[20] D M Hegazy D A OrsquoReilly B M Yang A D Hodgkinson BA Millward and A G Demaine ldquoNF120581B polymorphisms andsusceptibility to type 1 diabetesrdquo Genes amp Immunity vol 2 no6 pp 304ndash308 2001

6 BioMed Research International

[21] A F Gazdar ldquoDNA repair and survival in lung cancermdashthe twofaces of Janusrdquo The New England Journal of Medicine vol 356no 8 pp 771ndash773 2007

[22] J Vangsted T W Klausen N Abildgaard et al ldquoSinglenucleotide polymorphisms in the promoter region of the IL1Bgene influence outcome in multiple myeloma patients treatedwith high-dose chemotherapy independently of relapse treat-ment with thalidomide and bortezomibrdquoAnnals of Hematologyvol 90 no 10 pp 1173ndash1181 2011

[23] S Song D Chen J Lu et al ldquoNF120581B1 and NF120581BIA polymor-phisms are associatedwith increased risk for sporadic colorectalcancer in a southern Chinese populationrdquo PLoS ONE vol 6 no6 Article ID e21726 2011

[24] M Liang X Xu Y Gong Y Tang and L Lin ldquoRisk associationbetween theNF-120581B1minus94insdelATTGpromoter polymorphismand inflammatory bowel diseases a meta-analysisrdquo DigestiveDiseases and Sciences vol 57 no 9 pp 2304ndash2309 2012

[25] C W Cheng J L Su C W Lin et al ldquoEffects of NFKB1andNFKBIA gene polymorphisms on hepatocellular carcinomasusceptibility and clinicopathological featuresrdquo PLoS ONE vol8 no 2 Article ID e56130 2013

[26] D Vu E Tellez-Corrales P Sakharkar et al ldquoImpact of NF-120581B gene polymorphism on allograft outcome in Hispanic renaltransplant recipientsrdquo Transplant Immunology vol 28 no 1 pp18ndash23 2013

[27] Y-F Zou F-L Yuan X-L Feng et al ldquoAssociation betweenNFKB1 minus94insdelATTG promoter polymorphism and cancerrisk a meta-analysisrdquoCancer Investigation vol 29 no 1 pp 78ndash85 2011

[28] J Oliver M Gomez-Garcıa L Paco et al ldquoA functionalpolymorphism of the NFKB1 promoter is not associated withulcerative colitis in a Spanish populationrdquo Inflammatory BowelDiseases vol 11 no 6 pp 576ndash579 2005

[29] A S Karban T Okazaki C I M Panhuysen et al ldquoFunctionalannotation of a novel NFKB1 promoter polymorphism thatincreases risk for ulcerative colitisrdquoHuman Molecular Geneticsvol 13 no 1 pp 35ndash45 2004

[30] R C Bargou F Emmerich D Krappmann et al ldquoConstitutivenuclear factor-120581B-RelA activation is required for proliferationand survival of Hodgkinrsquos disease tumor cellsrdquo The Journal ofClinical Investigation vol 100 no 12 pp 2961ndash2969 1997

[31] G E Sonenshein ldquoRelNF-120581B transcription factors and thecontrol of apoptosisrdquo Seminars in Cancer Biology vol 8 no 2pp 113ndash119 1997

[32] X Dolcet D Llobet J Pallares and X Matias-Guiu ldquoNF-120581Bin development and progression of human cancerrdquo VirchowsArchiv vol 446 no 5 pp 475ndash482 2005

[33] D Glavac M Ravnik-Glavac S J OrsquoBrien and M DeanldquoPolymorphisms in the 31015840 untranslated region of the I120581BMAD-3 (NFKBI) gene located on chromosome 14rdquo Human Geneticsvol 93 no 6 pp 694ndash696 1994

[34] J Gao D Pfeifer L-J He et al ldquoAssociation of NFKBIApolymorphism with colorectal cancer risk and prognosis inSwedish and Chinese populationsrdquo Scandinavian Journal ofGastroenterology vol 42 no 3 pp 345ndash350 2007

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

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Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

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Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

BioMed Research International 3

Table 1 PCR and RFLP procedures and products of NF-120581B1 -94insdelATTG and NF-120581BIA 31015840UTR ArarrG

Primers (forward and reverse) PCR product Restriction enzyme Restriction products

NF-120581B151015840-TGGGCACAAGTCGTTTATGA-31015840

285 bp Van91l (PflMI)ATTG2ATTG2 (insins) 240 bp 45 bp

51015840-CTGGAGCCGGTAGGGAAG-31015840 ATTG1ATTG1 (deldel) 281ATTG2ATTG1 (insdel) 281 240 45

NF-120581BIA51015840-GGCTGAAAGAACATGGACTTG-31015840

424 bp HaeIIIAA wild type 424

51015840-GTACACCATTTACAGGAGGG-31015840 AG heterozygous 316 108GG mutant 424 316 108

Table 2 Distribution of NF-120581B1 and NF-120581BIA genotypes and allele in NSCLC patients and controls

Genotypesalleles Controls 119899 () Patients 119899 () OR (95 CI) 119875 valueNF-120581B1

II 46 (4647) 35 (3684) ReferenceID 47 (4747) 44 (4632) 123 (067ndash225) 0500DD 6 (606) 16 (1684) 350 (124ndash987) 0014ID + DD 53 (5353) 60 (6316) 149 (084ndash264) 0174I allele 139 (702) 114 (60) ReferenceD allele 59 (298) 76 (40) 157 (103ndash239) 0035

NF-120581BIAAA 21 (2121) 17 (179) ReferenceAG 45 (4546) 45 (4736) 124 (058ndash265) 0587GG 33 (3333) 33 (3474) 124 (055ndash275) 0605AG + GG 78 (7879) 78 (821) 124 (061ndash252) 0561A allele 87 (4394) 79 (4158) ReferenceG allele 111 (5606) 111 (5842) 110 (074ndash165) 0639

OR Odds ratio CI confidence interval

Chicago IL USA) Data were expressed as means plusmn SDDifferences in the distribution of NF-120581B1 -94insdelATTGand NF-120581BIA 31015840UTR ArarrG genotypes or alleles betweencases and controls were tested using the Chi-square statisticrespectively (Table 2) Differences in characteristics betweenNSCLC patients and controls were assessed with Fisherrsquosexact test as well as disparities in genotype and allelefrequencies Relative risk at 95 confidence intervals (CI)was calculated as the odds ratio (OR) Values 119875 lt 005 wereconsidered statistically significant A multivariate logisticregression model was performed to investigate possibleeffects of genotypes and alleles after adjustment for age

3 Results

In this study we examined 194 volunteers 95 NSCLC (89males 6 females) patients and 99 healthy people (67 males32 females) detecting any chronic disease or any evidence ofmalignancy Distribution of NF-120581B1 and NF-120581BIA genotypesaccording to clinic features in NSCLC patients is shownin Table 3 The distribution of the NF-120581B1 -94insdelATTGgenotypes in control and NSCLC patients was found tobe significantly different (119875 0048) It was evaluated thatindividuals carrying DD genotype had 35-fold increased riskfor NSCLC (119875 0014 1205942 5605 OR 350 95 CI 124ndash987)No statistically significant differences between groups were

observed when the NF-120581BIA 31015840UTR ArarrG genotypes distri-butions were compared (119875 0844) A significant correlationbetween genotype combinations of NF-120581B1 and NF-120581BIA(DDAG genotype) and NSCLC risk was found compared toall other combinations (119875 0025 OR 5035 95 CI 1067ndash2414) and DDAG genotype had increased risk for NSCLCThe prevalence of IIAA genotype combinations versus toall other combinations was 53 in patients and 121 inthe control group but there are no statistically significantdifferences between groups (119875 0091 OR 0403 95 CI0136ndash1191) The results of multivariate logistic regressionanalysis are presented in Table 4 Gender age (lt57ge57)and NF-120581B1 DD genotype were associated with NSCLC inunivariate analysis and additionally these were associatedwith this disease in multivariate logistic regression analysis

4 Discussion

A functional polymorphism in the NF-120581B1 gene promoterregion (-94insdelATTG) has been identified and associatedwith both chronic inflammatory diseases and malignantdiseases [22] NF-120581B is inactivated in the cytoplasm byI120581B120572 120573 or 120574 and the most common protein of this familyis the NF-120581B inhibitor 120572 (NF-120581BIA) [23] -94insdelATTGpolymorphism has evidence from two independent func-tional assays in vitro promoter activity and differential anunidentified nuclear protein binding that the specific allele

4 BioMed Research International

Table 3 Distribution of NF-120581B1 and NF-120581BIA genotypes with clinic features in NSCLC patients

NF-120581B1 NF-120581BIAII ID DD

119875 value AA AG GG119875 value

119899 () 119899 () 119899 () 119899 () 119899 () 119899 ()Sex

Men 33 (3710) 41 (4610) 15 (1680) 0980 15 (1690) 43 (4830) 31 (3480) 0570Women 2 (3330) 3 (5000) 1 (1670) 2 (3330) 2 (3330) 2 (3330)

Agelt57 10 (294) 17 (50) 7 (206) 0496 5 (147) 19 (559) 10 (294) 0567ge57 27 (415) 27 (415) 11 (169) 12 (185) 29 (446) 24 (369)

Smoking (boxyear)lt50 25 (4810) 24 (4620) 3 (580) 002 10 (1920) 25 (4810) 17 (3270) 0876ge50 10 (2330) 20 (4650) 13 (3020) 7 (1630) 20 (4650) 16 (3720)

Alcohol consumptionNo 17 (3150) 28 (5190) 9 (1670) 0405 6 (1110) 31 (5740) 17 (3150) 0044Yes 18 (4390) 16 (3900) 7 (1710) 11 (2680) 14 (3410) 16 (3900)

HistopathologySquamous 16 (4570) 16 (4570) 3 (860) 0179 7 (2000) 20 (5710) 8 (2290) 0173Nonsquamous 19 (3170) 28 (4670) 13 (2170) 10 (1670) 25 (4170) 25 (4170)

Total proteinlt6 gL 2 (5000) 0 (0) 2 (5000) 0087 0 (0) 3 (7500) 1 (2500) 0452ge6 gL 31 (3830) 38 (4690) 12 (1480) 16 (1980) 37 (4570) 28 (3460)

Albuminlt3 gL 6 (5450) 1 (910) 4 (3640) 0019 1 (910) 6 (5450) 4 (3640) 0635ge3 gL 26 (3470) 39 (5200) 10 (1330) 16 (2130) 35 (4670) 24 (3200)

Calciumlt10mgDl 30 (4000) 30 (4000) 15 (2000) 0001 16 (2130) 35 (4670) 24 (3200) 0506ge10mgdL 0 (0) 11 (100) 0 (0) 1 (910) 7 (6360) 3 (2730)

LDHlt250UL 22 (3610) 28 (4590) 11 (1800) 0644 13 (2130) 31 (5080) 17 (2790) 0249ge250UL 10 (4760) 8 (3810) 3 (1430) 3 (1430) 8 (3810) 10 (4760)

Table 4 The results of multivariate logistic regression

Covariates 119875 value Exp (B) 95 CI for Exp (B)Gender lt0001 7866 2915ndash21231Age (lt57ge57) lt0001 5074 2633ndash9776NF-120581B1 DD genotype 0035 3167 1086ndash9234

inherited likely has functional consequences [24] NF-120581BIA31015840UTR ArarrG polymorphismmay affect mRNA stability andtranslational efficacy or conduces to differential nuclear RNAprocessing or export also cannot be completely excludedMany studies have been conducted to investigate a possibleassociation between NF-120581B1 -94insdelATTG and NF-120581BIA31015840UTR ArarrG polymorphism and both inflammatory dis-eases and various cancer types [25] However no data areavailable in the English literature to report the associationwith NSCLC to date Our study is the initial report on thesetwo forms of polymorphism (both NF-120581B1 -94insdelATTGand NF-120581BIA 31015840UTR ArarrG) studied together in NSCLCpatients to our knowledge The genotypic combinations ofNF-120581B1 and NF-120581B2 polymorphism have been shown to be

associated with the development of common inflammatorydiseases including ulcerative colitis (UC) Crohnrsquos diseaseand Type I diabetes as well as susceptibility of severalcancers such as oral squamous cell carcinoma and colorectalcancer [26] It can be concluded that previous studies haveconflicting results [27] Oliver et al suggest that the NF-120581B1-94insdelATTG gene variation previously associated withUC susceptibility in North Americans does not influenceeither susceptibility or phenotype of UC in the Spanishpopulation [28] In this study we performed a risk associationbetween the NF-120581B1 -94insdelATTG promoter polymor-phism and NSCLC The -94insdelATTG polymorphism hasbeen shown as a first potential functional NF-120581B1 polymor-phism by Karban et al Nuclear proteins from normal humancolon tissue showed significant binding to -94insATTGbut not to -94delATTG containing oligonucleotides NF-120581B1 promoterexon 1 luciferase reporter plasmid constructscontaining the -94delATTG allele and transfected into eitherHeLa or HT-29 cell lines showed low promoter activity morethan comparable constructs containing the -94insATTGallele Therefore it is known that D allele promoter activ-ity is low and I allele promoter activity is high Previous

BioMed Research International 5

studies have suggested that D allele may result in decreasedNF-120581B1 message and hence decreased p50p105 NF-120581B pro-tein production leads to increased inflammatory responseOtherwise a potential explanation of decreased NF-120581B1 Dallele gene expression may be the resulting decreases inNF-120581B p50p65 heterodimers that are major mediators ofinflammation [29]

Defects in components that regulate NF-120581B release fromI120581B120572 result in constitutive or decreased NF-120581B activationThese components may be any of the kinases phosphatasesor other signal transducers normally involved in NF-120581B-activation pathways [30] Sonenshein suggests that alterationsof NF-120581B1 expression play an important role in the protec-tion of cells from apoptosis [31] NF-120581B1 activity has beenobserved in various types of cancer as well as colorectalcancer and breast cancer to contribute to tumor angiogenesisinvasion and progression [32] Therefore the variants ofthe NF-120581B1 gene could be expected to have an effect oncell death and thus carcinogenesis NF-120581B1 31015840UTR ArarrGpolymorphism has functional effects on expression of theNF-120581BIA gene and altered NF-120581B transcription [33] Thereare many studies with different results on NF-120581BIA 31015840UTRpolymorphism in the literature [34] Our results suggestedthat NF-120581BIA polymorphism has no effect on risk of NSCLCIn conclusion we here clearly demonstrated that NF-120581B1 -94insdelATTG promoter polymorphism and the presenceof the DD genotype might have a risk factor for NSCLCpathogenesis in our ethnic population Larger trials thatincluded different ethnic groups are necessary to defineobjectively the correlation between NF-120581B1 -94insdelATTGpromoter and development of NSCLC as well as prognosis ofdisease

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

The present work was supported by a Grant from theScientific Research Projects Coordination Unit of IstanbulUniversity (Project no 10537) The authors would like tothank MSc Allison P Eronat BSc Nesibe Selma Guler andPhDAylaKarimova for their understanding and suggestionsin English grammar of our article and the Editor and anony-mous reviewers for their valuable comments and suggestionswhich were helpful in improving the paper

References

[1] A Jemal R Siegel E Ward et al ldquoCancer statistics 2006rdquo CACancer Journal for Clinicians vol 56 no 2 pp 106ndash130 2006

[2] S Sun J H Schiller M Spinola and J D Minna ldquoNewmolecularly targeted therapies for lung cancerrdquo The Journal ofClinical Investigation vol 117 no 10 pp 2740ndash2750 2007

[3] M Fleischhacker T Beinert and K Possinger ldquoMoleculargenetic characteristics of lung cancermdashuseful as ldquorealrdquo tumormarkersrdquo Lung Cancer vol 25 no 1 pp 7ndash24 1999

[4] X Tang D Liu S Shishodia et al ldquoNuclear factor-120581B (NF-120581B) is frequently expressed in lung cancer and preneoplasticlesionsrdquo Cancer vol 107 no 11 pp 2637ndash2646 2006

[5] R Doll and A B Hill ldquoThe mortality of doctors in relationto their smoking habits a preliminary reportrdquo British MedicalJournal vol 1 no 4877 pp 1451ndash1455 1954

[6] M Koti R J Gooding P Nuin et al ldquoIdentification ofthe IGF1PI3KNF120581BERK gene signalling networks associatedwith chemotherapy resistance and treatment response in high-grade serous epithelial ovarian cancerrdquo BMC Cancer vol 13article 549 2013

[7] W Chen Z Li L Bai and Y Lin ldquoNF-120581B in lung cancer acarcinogenesis mediator and a prevention and therapy targetrdquoFrontiers in Bioscience vol 16 no 3 pp 1172ndash1185 2011

[8] P H Salim M Jobim M Bredemeier et al ldquoInterleukin-10 gene promoter and NFKB1 promoter insertiondeletionpolymorphisms in systemic sclerosisrdquo Scandinavian Journal ofImmunology vol 77 no 2 pp 162ndash168 2013

[9] R Sen and D Baltimore ldquoMultiple nuclear factors interact withthe immunoglobulin enhancer sequencesrdquo Cell vol 46 no 5pp 705ndash716 1986

[10] G Sethi B Sung and B B Aggarwal ldquoNuclear factor-120581Bactivation from bench to bedsiderdquo Experimental Biology andMedicine vol 233 no 1 pp 21ndash31 2008

[11] M S Hayden and S Ghosh ldquoSignaling to NF-120581Brdquo Genes ampDevelopment vol 18 no 18 pp 2195ndash2224 2004

[12] M Karin ldquoThe beginning of the end I120581B kinase (IKK) and NF-120581B activationrdquoThe Journal of Biological Chemistry vol 274 no39 pp 27339ndash27342 1999

[13] S Khan Z Lopez-Dee R Kumar and J Ling ldquoActivationof NFkB is a novel mechanism of pro-survival activity ofglucocorticoids in breast cancer cellsrdquo Cancer Letters vol 337no 1 pp 90ndash95 2013

[14] A C Bharti and B B Aggarwal ldquoNuclear factor-120581B and cancerits role in prevention and therapyrdquo Biochemical Pharmacologyvol 64 no 5-6 pp 883ndash888 2002

[15] D D Datta A Datta S Bhattacharjya and S RoychoudhuryldquoNF-120581B mediated transcriptional repression of acid modifyinghormone gastrinrdquo PLoS ONE vol 8 no 8 Article ID e734092013

[16] D R Jones R M Broad L V Madrid A S Baldwin Jr andMW Mayo ldquoInhibition of NF-120581B sensitizes non-small cell lungcancer cells to chemotherapy-induced apoptosisrdquoTheAnnals ofThoracic Surgery vol 70 no 3 pp 930ndash936 2000

[17] C-Y Wang J C Cusack Jr R Liu and A S Baldwin JrldquoControl of inducible chemoresistance enhanced anti-tumortherapy through increased apoptosis by inhibition of NF-120581BrdquoNature Medicine vol 5 no 4 pp 412ndash417 1999

[18] M Adamzik S Schafer U H Frey et al ldquoTheNFKB1 promoterpolymorphism (minus94insdelATTG) alters nuclear translocationof NF-120581B1 in monocytes after lipopolysaccharide stimulationand is associated with increased mortality in sepsisrdquo Anesthe-siology vol 118 no 1 pp 123ndash133 2013

[19] H Nakshatri P Bhat-Nakshatri D A Martin R J Goulet Jrand G W Sledge Jr ldquoConstitutive activation of NF-120581B duringprogression of breast cancer to hormone-independent growthrdquoMolecular andCellular Biology vol 17 no 7 pp 3629ndash3639 1997

[20] D M Hegazy D A OrsquoReilly B M Yang A D Hodgkinson BA Millward and A G Demaine ldquoNF120581B polymorphisms andsusceptibility to type 1 diabetesrdquo Genes amp Immunity vol 2 no6 pp 304ndash308 2001

6 BioMed Research International

[21] A F Gazdar ldquoDNA repair and survival in lung cancermdashthe twofaces of Janusrdquo The New England Journal of Medicine vol 356no 8 pp 771ndash773 2007

[22] J Vangsted T W Klausen N Abildgaard et al ldquoSinglenucleotide polymorphisms in the promoter region of the IL1Bgene influence outcome in multiple myeloma patients treatedwith high-dose chemotherapy independently of relapse treat-ment with thalidomide and bortezomibrdquoAnnals of Hematologyvol 90 no 10 pp 1173ndash1181 2011

[23] S Song D Chen J Lu et al ldquoNF120581B1 and NF120581BIA polymor-phisms are associatedwith increased risk for sporadic colorectalcancer in a southern Chinese populationrdquo PLoS ONE vol 6 no6 Article ID e21726 2011

[24] M Liang X Xu Y Gong Y Tang and L Lin ldquoRisk associationbetween theNF-120581B1minus94insdelATTGpromoter polymorphismand inflammatory bowel diseases a meta-analysisrdquo DigestiveDiseases and Sciences vol 57 no 9 pp 2304ndash2309 2012

[25] C W Cheng J L Su C W Lin et al ldquoEffects of NFKB1andNFKBIA gene polymorphisms on hepatocellular carcinomasusceptibility and clinicopathological featuresrdquo PLoS ONE vol8 no 2 Article ID e56130 2013

[26] D Vu E Tellez-Corrales P Sakharkar et al ldquoImpact of NF-120581B gene polymorphism on allograft outcome in Hispanic renaltransplant recipientsrdquo Transplant Immunology vol 28 no 1 pp18ndash23 2013

[27] Y-F Zou F-L Yuan X-L Feng et al ldquoAssociation betweenNFKB1 minus94insdelATTG promoter polymorphism and cancerrisk a meta-analysisrdquoCancer Investigation vol 29 no 1 pp 78ndash85 2011

[28] J Oliver M Gomez-Garcıa L Paco et al ldquoA functionalpolymorphism of the NFKB1 promoter is not associated withulcerative colitis in a Spanish populationrdquo Inflammatory BowelDiseases vol 11 no 6 pp 576ndash579 2005

[29] A S Karban T Okazaki C I M Panhuysen et al ldquoFunctionalannotation of a novel NFKB1 promoter polymorphism thatincreases risk for ulcerative colitisrdquoHuman Molecular Geneticsvol 13 no 1 pp 35ndash45 2004

[30] R C Bargou F Emmerich D Krappmann et al ldquoConstitutivenuclear factor-120581B-RelA activation is required for proliferationand survival of Hodgkinrsquos disease tumor cellsrdquo The Journal ofClinical Investigation vol 100 no 12 pp 2961ndash2969 1997

[31] G E Sonenshein ldquoRelNF-120581B transcription factors and thecontrol of apoptosisrdquo Seminars in Cancer Biology vol 8 no 2pp 113ndash119 1997

[32] X Dolcet D Llobet J Pallares and X Matias-Guiu ldquoNF-120581Bin development and progression of human cancerrdquo VirchowsArchiv vol 446 no 5 pp 475ndash482 2005

[33] D Glavac M Ravnik-Glavac S J OrsquoBrien and M DeanldquoPolymorphisms in the 31015840 untranslated region of the I120581BMAD-3 (NFKBI) gene located on chromosome 14rdquo Human Geneticsvol 93 no 6 pp 694ndash696 1994

[34] J Gao D Pfeifer L-J He et al ldquoAssociation of NFKBIApolymorphism with colorectal cancer risk and prognosis inSwedish and Chinese populationsrdquo Scandinavian Journal ofGastroenterology vol 42 no 3 pp 345ndash350 2007

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

4 BioMed Research International

Table 3 Distribution of NF-120581B1 and NF-120581BIA genotypes with clinic features in NSCLC patients

NF-120581B1 NF-120581BIAII ID DD

119875 value AA AG GG119875 value

119899 () 119899 () 119899 () 119899 () 119899 () 119899 ()Sex

Men 33 (3710) 41 (4610) 15 (1680) 0980 15 (1690) 43 (4830) 31 (3480) 0570Women 2 (3330) 3 (5000) 1 (1670) 2 (3330) 2 (3330) 2 (3330)

Agelt57 10 (294) 17 (50) 7 (206) 0496 5 (147) 19 (559) 10 (294) 0567ge57 27 (415) 27 (415) 11 (169) 12 (185) 29 (446) 24 (369)

Smoking (boxyear)lt50 25 (4810) 24 (4620) 3 (580) 002 10 (1920) 25 (4810) 17 (3270) 0876ge50 10 (2330) 20 (4650) 13 (3020) 7 (1630) 20 (4650) 16 (3720)

Alcohol consumptionNo 17 (3150) 28 (5190) 9 (1670) 0405 6 (1110) 31 (5740) 17 (3150) 0044Yes 18 (4390) 16 (3900) 7 (1710) 11 (2680) 14 (3410) 16 (3900)

HistopathologySquamous 16 (4570) 16 (4570) 3 (860) 0179 7 (2000) 20 (5710) 8 (2290) 0173Nonsquamous 19 (3170) 28 (4670) 13 (2170) 10 (1670) 25 (4170) 25 (4170)

Total proteinlt6 gL 2 (5000) 0 (0) 2 (5000) 0087 0 (0) 3 (7500) 1 (2500) 0452ge6 gL 31 (3830) 38 (4690) 12 (1480) 16 (1980) 37 (4570) 28 (3460)

Albuminlt3 gL 6 (5450) 1 (910) 4 (3640) 0019 1 (910) 6 (5450) 4 (3640) 0635ge3 gL 26 (3470) 39 (5200) 10 (1330) 16 (2130) 35 (4670) 24 (3200)

Calciumlt10mgDl 30 (4000) 30 (4000) 15 (2000) 0001 16 (2130) 35 (4670) 24 (3200) 0506ge10mgdL 0 (0) 11 (100) 0 (0) 1 (910) 7 (6360) 3 (2730)

LDHlt250UL 22 (3610) 28 (4590) 11 (1800) 0644 13 (2130) 31 (5080) 17 (2790) 0249ge250UL 10 (4760) 8 (3810) 3 (1430) 3 (1430) 8 (3810) 10 (4760)

Table 4 The results of multivariate logistic regression

Covariates 119875 value Exp (B) 95 CI for Exp (B)Gender lt0001 7866 2915ndash21231Age (lt57ge57) lt0001 5074 2633ndash9776NF-120581B1 DD genotype 0035 3167 1086ndash9234

inherited likely has functional consequences [24] NF-120581BIA31015840UTR ArarrG polymorphismmay affect mRNA stability andtranslational efficacy or conduces to differential nuclear RNAprocessing or export also cannot be completely excludedMany studies have been conducted to investigate a possibleassociation between NF-120581B1 -94insdelATTG and NF-120581BIA31015840UTR ArarrG polymorphism and both inflammatory dis-eases and various cancer types [25] However no data areavailable in the English literature to report the associationwith NSCLC to date Our study is the initial report on thesetwo forms of polymorphism (both NF-120581B1 -94insdelATTGand NF-120581BIA 31015840UTR ArarrG) studied together in NSCLCpatients to our knowledge The genotypic combinations ofNF-120581B1 and NF-120581B2 polymorphism have been shown to be

associated with the development of common inflammatorydiseases including ulcerative colitis (UC) Crohnrsquos diseaseand Type I diabetes as well as susceptibility of severalcancers such as oral squamous cell carcinoma and colorectalcancer [26] It can be concluded that previous studies haveconflicting results [27] Oliver et al suggest that the NF-120581B1-94insdelATTG gene variation previously associated withUC susceptibility in North Americans does not influenceeither susceptibility or phenotype of UC in the Spanishpopulation [28] In this study we performed a risk associationbetween the NF-120581B1 -94insdelATTG promoter polymor-phism and NSCLC The -94insdelATTG polymorphism hasbeen shown as a first potential functional NF-120581B1 polymor-phism by Karban et al Nuclear proteins from normal humancolon tissue showed significant binding to -94insATTGbut not to -94delATTG containing oligonucleotides NF-120581B1 promoterexon 1 luciferase reporter plasmid constructscontaining the -94delATTG allele and transfected into eitherHeLa or HT-29 cell lines showed low promoter activity morethan comparable constructs containing the -94insATTGallele Therefore it is known that D allele promoter activ-ity is low and I allele promoter activity is high Previous

BioMed Research International 5

studies have suggested that D allele may result in decreasedNF-120581B1 message and hence decreased p50p105 NF-120581B pro-tein production leads to increased inflammatory responseOtherwise a potential explanation of decreased NF-120581B1 Dallele gene expression may be the resulting decreases inNF-120581B p50p65 heterodimers that are major mediators ofinflammation [29]

Defects in components that regulate NF-120581B release fromI120581B120572 result in constitutive or decreased NF-120581B activationThese components may be any of the kinases phosphatasesor other signal transducers normally involved in NF-120581B-activation pathways [30] Sonenshein suggests that alterationsof NF-120581B1 expression play an important role in the protec-tion of cells from apoptosis [31] NF-120581B1 activity has beenobserved in various types of cancer as well as colorectalcancer and breast cancer to contribute to tumor angiogenesisinvasion and progression [32] Therefore the variants ofthe NF-120581B1 gene could be expected to have an effect oncell death and thus carcinogenesis NF-120581B1 31015840UTR ArarrGpolymorphism has functional effects on expression of theNF-120581BIA gene and altered NF-120581B transcription [33] Thereare many studies with different results on NF-120581BIA 31015840UTRpolymorphism in the literature [34] Our results suggestedthat NF-120581BIA polymorphism has no effect on risk of NSCLCIn conclusion we here clearly demonstrated that NF-120581B1 -94insdelATTG promoter polymorphism and the presenceof the DD genotype might have a risk factor for NSCLCpathogenesis in our ethnic population Larger trials thatincluded different ethnic groups are necessary to defineobjectively the correlation between NF-120581B1 -94insdelATTGpromoter and development of NSCLC as well as prognosis ofdisease

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

The present work was supported by a Grant from theScientific Research Projects Coordination Unit of IstanbulUniversity (Project no 10537) The authors would like tothank MSc Allison P Eronat BSc Nesibe Selma Guler andPhDAylaKarimova for their understanding and suggestionsin English grammar of our article and the Editor and anony-mous reviewers for their valuable comments and suggestionswhich were helpful in improving the paper

References

[1] A Jemal R Siegel E Ward et al ldquoCancer statistics 2006rdquo CACancer Journal for Clinicians vol 56 no 2 pp 106ndash130 2006

[2] S Sun J H Schiller M Spinola and J D Minna ldquoNewmolecularly targeted therapies for lung cancerrdquo The Journal ofClinical Investigation vol 117 no 10 pp 2740ndash2750 2007

[3] M Fleischhacker T Beinert and K Possinger ldquoMoleculargenetic characteristics of lung cancermdashuseful as ldquorealrdquo tumormarkersrdquo Lung Cancer vol 25 no 1 pp 7ndash24 1999

[4] X Tang D Liu S Shishodia et al ldquoNuclear factor-120581B (NF-120581B) is frequently expressed in lung cancer and preneoplasticlesionsrdquo Cancer vol 107 no 11 pp 2637ndash2646 2006

[5] R Doll and A B Hill ldquoThe mortality of doctors in relationto their smoking habits a preliminary reportrdquo British MedicalJournal vol 1 no 4877 pp 1451ndash1455 1954

[6] M Koti R J Gooding P Nuin et al ldquoIdentification ofthe IGF1PI3KNF120581BERK gene signalling networks associatedwith chemotherapy resistance and treatment response in high-grade serous epithelial ovarian cancerrdquo BMC Cancer vol 13article 549 2013

[7] W Chen Z Li L Bai and Y Lin ldquoNF-120581B in lung cancer acarcinogenesis mediator and a prevention and therapy targetrdquoFrontiers in Bioscience vol 16 no 3 pp 1172ndash1185 2011

[8] P H Salim M Jobim M Bredemeier et al ldquoInterleukin-10 gene promoter and NFKB1 promoter insertiondeletionpolymorphisms in systemic sclerosisrdquo Scandinavian Journal ofImmunology vol 77 no 2 pp 162ndash168 2013

[9] R Sen and D Baltimore ldquoMultiple nuclear factors interact withthe immunoglobulin enhancer sequencesrdquo Cell vol 46 no 5pp 705ndash716 1986

[10] G Sethi B Sung and B B Aggarwal ldquoNuclear factor-120581Bactivation from bench to bedsiderdquo Experimental Biology andMedicine vol 233 no 1 pp 21ndash31 2008

[11] M S Hayden and S Ghosh ldquoSignaling to NF-120581Brdquo Genes ampDevelopment vol 18 no 18 pp 2195ndash2224 2004

[12] M Karin ldquoThe beginning of the end I120581B kinase (IKK) and NF-120581B activationrdquoThe Journal of Biological Chemistry vol 274 no39 pp 27339ndash27342 1999

[13] S Khan Z Lopez-Dee R Kumar and J Ling ldquoActivationof NFkB is a novel mechanism of pro-survival activity ofglucocorticoids in breast cancer cellsrdquo Cancer Letters vol 337no 1 pp 90ndash95 2013

[14] A C Bharti and B B Aggarwal ldquoNuclear factor-120581B and cancerits role in prevention and therapyrdquo Biochemical Pharmacologyvol 64 no 5-6 pp 883ndash888 2002

[15] D D Datta A Datta S Bhattacharjya and S RoychoudhuryldquoNF-120581B mediated transcriptional repression of acid modifyinghormone gastrinrdquo PLoS ONE vol 8 no 8 Article ID e734092013

[16] D R Jones R M Broad L V Madrid A S Baldwin Jr andMW Mayo ldquoInhibition of NF-120581B sensitizes non-small cell lungcancer cells to chemotherapy-induced apoptosisrdquoTheAnnals ofThoracic Surgery vol 70 no 3 pp 930ndash936 2000

[17] C-Y Wang J C Cusack Jr R Liu and A S Baldwin JrldquoControl of inducible chemoresistance enhanced anti-tumortherapy through increased apoptosis by inhibition of NF-120581BrdquoNature Medicine vol 5 no 4 pp 412ndash417 1999

[18] M Adamzik S Schafer U H Frey et al ldquoTheNFKB1 promoterpolymorphism (minus94insdelATTG) alters nuclear translocationof NF-120581B1 in monocytes after lipopolysaccharide stimulationand is associated with increased mortality in sepsisrdquo Anesthe-siology vol 118 no 1 pp 123ndash133 2013

[19] H Nakshatri P Bhat-Nakshatri D A Martin R J Goulet Jrand G W Sledge Jr ldquoConstitutive activation of NF-120581B duringprogression of breast cancer to hormone-independent growthrdquoMolecular andCellular Biology vol 17 no 7 pp 3629ndash3639 1997

[20] D M Hegazy D A OrsquoReilly B M Yang A D Hodgkinson BA Millward and A G Demaine ldquoNF120581B polymorphisms andsusceptibility to type 1 diabetesrdquo Genes amp Immunity vol 2 no6 pp 304ndash308 2001

6 BioMed Research International

[21] A F Gazdar ldquoDNA repair and survival in lung cancermdashthe twofaces of Janusrdquo The New England Journal of Medicine vol 356no 8 pp 771ndash773 2007

[22] J Vangsted T W Klausen N Abildgaard et al ldquoSinglenucleotide polymorphisms in the promoter region of the IL1Bgene influence outcome in multiple myeloma patients treatedwith high-dose chemotherapy independently of relapse treat-ment with thalidomide and bortezomibrdquoAnnals of Hematologyvol 90 no 10 pp 1173ndash1181 2011

[23] S Song D Chen J Lu et al ldquoNF120581B1 and NF120581BIA polymor-phisms are associatedwith increased risk for sporadic colorectalcancer in a southern Chinese populationrdquo PLoS ONE vol 6 no6 Article ID e21726 2011

[24] M Liang X Xu Y Gong Y Tang and L Lin ldquoRisk associationbetween theNF-120581B1minus94insdelATTGpromoter polymorphismand inflammatory bowel diseases a meta-analysisrdquo DigestiveDiseases and Sciences vol 57 no 9 pp 2304ndash2309 2012

[25] C W Cheng J L Su C W Lin et al ldquoEffects of NFKB1andNFKBIA gene polymorphisms on hepatocellular carcinomasusceptibility and clinicopathological featuresrdquo PLoS ONE vol8 no 2 Article ID e56130 2013

[26] D Vu E Tellez-Corrales P Sakharkar et al ldquoImpact of NF-120581B gene polymorphism on allograft outcome in Hispanic renaltransplant recipientsrdquo Transplant Immunology vol 28 no 1 pp18ndash23 2013

[27] Y-F Zou F-L Yuan X-L Feng et al ldquoAssociation betweenNFKB1 minus94insdelATTG promoter polymorphism and cancerrisk a meta-analysisrdquoCancer Investigation vol 29 no 1 pp 78ndash85 2011

[28] J Oliver M Gomez-Garcıa L Paco et al ldquoA functionalpolymorphism of the NFKB1 promoter is not associated withulcerative colitis in a Spanish populationrdquo Inflammatory BowelDiseases vol 11 no 6 pp 576ndash579 2005

[29] A S Karban T Okazaki C I M Panhuysen et al ldquoFunctionalannotation of a novel NFKB1 promoter polymorphism thatincreases risk for ulcerative colitisrdquoHuman Molecular Geneticsvol 13 no 1 pp 35ndash45 2004

[30] R C Bargou F Emmerich D Krappmann et al ldquoConstitutivenuclear factor-120581B-RelA activation is required for proliferationand survival of Hodgkinrsquos disease tumor cellsrdquo The Journal ofClinical Investigation vol 100 no 12 pp 2961ndash2969 1997

[31] G E Sonenshein ldquoRelNF-120581B transcription factors and thecontrol of apoptosisrdquo Seminars in Cancer Biology vol 8 no 2pp 113ndash119 1997

[32] X Dolcet D Llobet J Pallares and X Matias-Guiu ldquoNF-120581Bin development and progression of human cancerrdquo VirchowsArchiv vol 446 no 5 pp 475ndash482 2005

[33] D Glavac M Ravnik-Glavac S J OrsquoBrien and M DeanldquoPolymorphisms in the 31015840 untranslated region of the I120581BMAD-3 (NFKBI) gene located on chromosome 14rdquo Human Geneticsvol 93 no 6 pp 694ndash696 1994

[34] J Gao D Pfeifer L-J He et al ldquoAssociation of NFKBIApolymorphism with colorectal cancer risk and prognosis inSwedish and Chinese populationsrdquo Scandinavian Journal ofGastroenterology vol 42 no 3 pp 345ndash350 2007

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

BioMed Research International 5

studies have suggested that D allele may result in decreasedNF-120581B1 message and hence decreased p50p105 NF-120581B pro-tein production leads to increased inflammatory responseOtherwise a potential explanation of decreased NF-120581B1 Dallele gene expression may be the resulting decreases inNF-120581B p50p65 heterodimers that are major mediators ofinflammation [29]

Defects in components that regulate NF-120581B release fromI120581B120572 result in constitutive or decreased NF-120581B activationThese components may be any of the kinases phosphatasesor other signal transducers normally involved in NF-120581B-activation pathways [30] Sonenshein suggests that alterationsof NF-120581B1 expression play an important role in the protec-tion of cells from apoptosis [31] NF-120581B1 activity has beenobserved in various types of cancer as well as colorectalcancer and breast cancer to contribute to tumor angiogenesisinvasion and progression [32] Therefore the variants ofthe NF-120581B1 gene could be expected to have an effect oncell death and thus carcinogenesis NF-120581B1 31015840UTR ArarrGpolymorphism has functional effects on expression of theNF-120581BIA gene and altered NF-120581B transcription [33] Thereare many studies with different results on NF-120581BIA 31015840UTRpolymorphism in the literature [34] Our results suggestedthat NF-120581BIA polymorphism has no effect on risk of NSCLCIn conclusion we here clearly demonstrated that NF-120581B1 -94insdelATTG promoter polymorphism and the presenceof the DD genotype might have a risk factor for NSCLCpathogenesis in our ethnic population Larger trials thatincluded different ethnic groups are necessary to defineobjectively the correlation between NF-120581B1 -94insdelATTGpromoter and development of NSCLC as well as prognosis ofdisease

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

The present work was supported by a Grant from theScientific Research Projects Coordination Unit of IstanbulUniversity (Project no 10537) The authors would like tothank MSc Allison P Eronat BSc Nesibe Selma Guler andPhDAylaKarimova for their understanding and suggestionsin English grammar of our article and the Editor and anony-mous reviewers for their valuable comments and suggestionswhich were helpful in improving the paper

References

[1] A Jemal R Siegel E Ward et al ldquoCancer statistics 2006rdquo CACancer Journal for Clinicians vol 56 no 2 pp 106ndash130 2006

[2] S Sun J H Schiller M Spinola and J D Minna ldquoNewmolecularly targeted therapies for lung cancerrdquo The Journal ofClinical Investigation vol 117 no 10 pp 2740ndash2750 2007

[3] M Fleischhacker T Beinert and K Possinger ldquoMoleculargenetic characteristics of lung cancermdashuseful as ldquorealrdquo tumormarkersrdquo Lung Cancer vol 25 no 1 pp 7ndash24 1999

[4] X Tang D Liu S Shishodia et al ldquoNuclear factor-120581B (NF-120581B) is frequently expressed in lung cancer and preneoplasticlesionsrdquo Cancer vol 107 no 11 pp 2637ndash2646 2006

[5] R Doll and A B Hill ldquoThe mortality of doctors in relationto their smoking habits a preliminary reportrdquo British MedicalJournal vol 1 no 4877 pp 1451ndash1455 1954

[6] M Koti R J Gooding P Nuin et al ldquoIdentification ofthe IGF1PI3KNF120581BERK gene signalling networks associatedwith chemotherapy resistance and treatment response in high-grade serous epithelial ovarian cancerrdquo BMC Cancer vol 13article 549 2013

[7] W Chen Z Li L Bai and Y Lin ldquoNF-120581B in lung cancer acarcinogenesis mediator and a prevention and therapy targetrdquoFrontiers in Bioscience vol 16 no 3 pp 1172ndash1185 2011

[8] P H Salim M Jobim M Bredemeier et al ldquoInterleukin-10 gene promoter and NFKB1 promoter insertiondeletionpolymorphisms in systemic sclerosisrdquo Scandinavian Journal ofImmunology vol 77 no 2 pp 162ndash168 2013

[9] R Sen and D Baltimore ldquoMultiple nuclear factors interact withthe immunoglobulin enhancer sequencesrdquo Cell vol 46 no 5pp 705ndash716 1986

[10] G Sethi B Sung and B B Aggarwal ldquoNuclear factor-120581Bactivation from bench to bedsiderdquo Experimental Biology andMedicine vol 233 no 1 pp 21ndash31 2008

[11] M S Hayden and S Ghosh ldquoSignaling to NF-120581Brdquo Genes ampDevelopment vol 18 no 18 pp 2195ndash2224 2004

[12] M Karin ldquoThe beginning of the end I120581B kinase (IKK) and NF-120581B activationrdquoThe Journal of Biological Chemistry vol 274 no39 pp 27339ndash27342 1999

[13] S Khan Z Lopez-Dee R Kumar and J Ling ldquoActivationof NFkB is a novel mechanism of pro-survival activity ofglucocorticoids in breast cancer cellsrdquo Cancer Letters vol 337no 1 pp 90ndash95 2013

[14] A C Bharti and B B Aggarwal ldquoNuclear factor-120581B and cancerits role in prevention and therapyrdquo Biochemical Pharmacologyvol 64 no 5-6 pp 883ndash888 2002

[15] D D Datta A Datta S Bhattacharjya and S RoychoudhuryldquoNF-120581B mediated transcriptional repression of acid modifyinghormone gastrinrdquo PLoS ONE vol 8 no 8 Article ID e734092013

[16] D R Jones R M Broad L V Madrid A S Baldwin Jr andMW Mayo ldquoInhibition of NF-120581B sensitizes non-small cell lungcancer cells to chemotherapy-induced apoptosisrdquoTheAnnals ofThoracic Surgery vol 70 no 3 pp 930ndash936 2000

[17] C-Y Wang J C Cusack Jr R Liu and A S Baldwin JrldquoControl of inducible chemoresistance enhanced anti-tumortherapy through increased apoptosis by inhibition of NF-120581BrdquoNature Medicine vol 5 no 4 pp 412ndash417 1999

[18] M Adamzik S Schafer U H Frey et al ldquoTheNFKB1 promoterpolymorphism (minus94insdelATTG) alters nuclear translocationof NF-120581B1 in monocytes after lipopolysaccharide stimulationand is associated with increased mortality in sepsisrdquo Anesthe-siology vol 118 no 1 pp 123ndash133 2013

[19] H Nakshatri P Bhat-Nakshatri D A Martin R J Goulet Jrand G W Sledge Jr ldquoConstitutive activation of NF-120581B duringprogression of breast cancer to hormone-independent growthrdquoMolecular andCellular Biology vol 17 no 7 pp 3629ndash3639 1997

[20] D M Hegazy D A OrsquoReilly B M Yang A D Hodgkinson BA Millward and A G Demaine ldquoNF120581B polymorphisms andsusceptibility to type 1 diabetesrdquo Genes amp Immunity vol 2 no6 pp 304ndash308 2001

6 BioMed Research International

[21] A F Gazdar ldquoDNA repair and survival in lung cancermdashthe twofaces of Janusrdquo The New England Journal of Medicine vol 356no 8 pp 771ndash773 2007

[22] J Vangsted T W Klausen N Abildgaard et al ldquoSinglenucleotide polymorphisms in the promoter region of the IL1Bgene influence outcome in multiple myeloma patients treatedwith high-dose chemotherapy independently of relapse treat-ment with thalidomide and bortezomibrdquoAnnals of Hematologyvol 90 no 10 pp 1173ndash1181 2011

[23] S Song D Chen J Lu et al ldquoNF120581B1 and NF120581BIA polymor-phisms are associatedwith increased risk for sporadic colorectalcancer in a southern Chinese populationrdquo PLoS ONE vol 6 no6 Article ID e21726 2011

[24] M Liang X Xu Y Gong Y Tang and L Lin ldquoRisk associationbetween theNF-120581B1minus94insdelATTGpromoter polymorphismand inflammatory bowel diseases a meta-analysisrdquo DigestiveDiseases and Sciences vol 57 no 9 pp 2304ndash2309 2012

[25] C W Cheng J L Su C W Lin et al ldquoEffects of NFKB1andNFKBIA gene polymorphisms on hepatocellular carcinomasusceptibility and clinicopathological featuresrdquo PLoS ONE vol8 no 2 Article ID e56130 2013

[26] D Vu E Tellez-Corrales P Sakharkar et al ldquoImpact of NF-120581B gene polymorphism on allograft outcome in Hispanic renaltransplant recipientsrdquo Transplant Immunology vol 28 no 1 pp18ndash23 2013

[27] Y-F Zou F-L Yuan X-L Feng et al ldquoAssociation betweenNFKB1 minus94insdelATTG promoter polymorphism and cancerrisk a meta-analysisrdquoCancer Investigation vol 29 no 1 pp 78ndash85 2011

[28] J Oliver M Gomez-Garcıa L Paco et al ldquoA functionalpolymorphism of the NFKB1 promoter is not associated withulcerative colitis in a Spanish populationrdquo Inflammatory BowelDiseases vol 11 no 6 pp 576ndash579 2005

[29] A S Karban T Okazaki C I M Panhuysen et al ldquoFunctionalannotation of a novel NFKB1 promoter polymorphism thatincreases risk for ulcerative colitisrdquoHuman Molecular Geneticsvol 13 no 1 pp 35ndash45 2004

[30] R C Bargou F Emmerich D Krappmann et al ldquoConstitutivenuclear factor-120581B-RelA activation is required for proliferationand survival of Hodgkinrsquos disease tumor cellsrdquo The Journal ofClinical Investigation vol 100 no 12 pp 2961ndash2969 1997

[31] G E Sonenshein ldquoRelNF-120581B transcription factors and thecontrol of apoptosisrdquo Seminars in Cancer Biology vol 8 no 2pp 113ndash119 1997

[32] X Dolcet D Llobet J Pallares and X Matias-Guiu ldquoNF-120581Bin development and progression of human cancerrdquo VirchowsArchiv vol 446 no 5 pp 475ndash482 2005

[33] D Glavac M Ravnik-Glavac S J OrsquoBrien and M DeanldquoPolymorphisms in the 31015840 untranslated region of the I120581BMAD-3 (NFKBI) gene located on chromosome 14rdquo Human Geneticsvol 93 no 6 pp 694ndash696 1994

[34] J Gao D Pfeifer L-J He et al ldquoAssociation of NFKBIApolymorphism with colorectal cancer risk and prognosis inSwedish and Chinese populationsrdquo Scandinavian Journal ofGastroenterology vol 42 no 3 pp 345ndash350 2007

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

6 BioMed Research International

[21] A F Gazdar ldquoDNA repair and survival in lung cancermdashthe twofaces of Janusrdquo The New England Journal of Medicine vol 356no 8 pp 771ndash773 2007

[22] J Vangsted T W Klausen N Abildgaard et al ldquoSinglenucleotide polymorphisms in the promoter region of the IL1Bgene influence outcome in multiple myeloma patients treatedwith high-dose chemotherapy independently of relapse treat-ment with thalidomide and bortezomibrdquoAnnals of Hematologyvol 90 no 10 pp 1173ndash1181 2011

[23] S Song D Chen J Lu et al ldquoNF120581B1 and NF120581BIA polymor-phisms are associatedwith increased risk for sporadic colorectalcancer in a southern Chinese populationrdquo PLoS ONE vol 6 no6 Article ID e21726 2011

[24] M Liang X Xu Y Gong Y Tang and L Lin ldquoRisk associationbetween theNF-120581B1minus94insdelATTGpromoter polymorphismand inflammatory bowel diseases a meta-analysisrdquo DigestiveDiseases and Sciences vol 57 no 9 pp 2304ndash2309 2012

[25] C W Cheng J L Su C W Lin et al ldquoEffects of NFKB1andNFKBIA gene polymorphisms on hepatocellular carcinomasusceptibility and clinicopathological featuresrdquo PLoS ONE vol8 no 2 Article ID e56130 2013

[26] D Vu E Tellez-Corrales P Sakharkar et al ldquoImpact of NF-120581B gene polymorphism on allograft outcome in Hispanic renaltransplant recipientsrdquo Transplant Immunology vol 28 no 1 pp18ndash23 2013

[27] Y-F Zou F-L Yuan X-L Feng et al ldquoAssociation betweenNFKB1 minus94insdelATTG promoter polymorphism and cancerrisk a meta-analysisrdquoCancer Investigation vol 29 no 1 pp 78ndash85 2011

[28] J Oliver M Gomez-Garcıa L Paco et al ldquoA functionalpolymorphism of the NFKB1 promoter is not associated withulcerative colitis in a Spanish populationrdquo Inflammatory BowelDiseases vol 11 no 6 pp 576ndash579 2005

[29] A S Karban T Okazaki C I M Panhuysen et al ldquoFunctionalannotation of a novel NFKB1 promoter polymorphism thatincreases risk for ulcerative colitisrdquoHuman Molecular Geneticsvol 13 no 1 pp 35ndash45 2004

[30] R C Bargou F Emmerich D Krappmann et al ldquoConstitutivenuclear factor-120581B-RelA activation is required for proliferationand survival of Hodgkinrsquos disease tumor cellsrdquo The Journal ofClinical Investigation vol 100 no 12 pp 2961ndash2969 1997

[31] G E Sonenshein ldquoRelNF-120581B transcription factors and thecontrol of apoptosisrdquo Seminars in Cancer Biology vol 8 no 2pp 113ndash119 1997

[32] X Dolcet D Llobet J Pallares and X Matias-Guiu ldquoNF-120581Bin development and progression of human cancerrdquo VirchowsArchiv vol 446 no 5 pp 475ndash482 2005

[33] D Glavac M Ravnik-Glavac S J OrsquoBrien and M DeanldquoPolymorphisms in the 31015840 untranslated region of the I120581BMAD-3 (NFKBI) gene located on chromosome 14rdquo Human Geneticsvol 93 no 6 pp 694ndash696 1994

[34] J Gao D Pfeifer L-J He et al ldquoAssociation of NFKBIApolymorphism with colorectal cancer risk and prognosis inSwedish and Chinese populationsrdquo Scandinavian Journal ofGastroenterology vol 42 no 3 pp 345ndash350 2007

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology