Cancer Epidemiology 37 (2013) 625–628

UDP-glucuronosyltransferase 2B17 genotype and the risk of lung canceramong Austrian Caucasians

Michaela Gruber a, Trang Le a, Martin Filipits b, Andrea Gsur b, Christine Mannhalter c,Ulrich Jager a, Katrina Vanura a,*a Department of Medicine I, Division of Hematology and Hemostaseology, Comprehensive Cancer Center, Medical University of Vienna, Waehringer Guertel 18-20, 1090 Vienna, Austriab Department of Medicine I, Institute of Cancer Research, Medical University of Vienna, Borschkegasse 8a, 1090 Vienna, Austriac Department of Clinical Chemistry and Laboratory Medicine, Medical University of Vienna, Waehringer Guertel 18-20, 1090 Vienna, Austria

A R T I C L E I N F O

Article history:

Received 3 May 2013

Received in revised form 7 June 2013

Accepted 13 June 2013

Available online 11 July 2013

Keywords:

UGT2B17

Lung cancer

Copy number variation

Polymorphisms in carcinogen metabolism

Cancer risk

Cancer susceptibility genes

A B S T R A C T

Background: The enzyme uridine diphospho glucuronosyltansferase 2B17 (UGT2B17) glucuronidates

several endogenous and exogenous compounds, including carcinogens from tobacco smoke like 4-

(methylnitrosamino)-1-(3-pyridyl)-1-butanl (NNAL). UGT2B17 shows a remarkable copy number

variation (CNV) and an association between deletion genotype and increased risk of lung

adenocarcinoma in women has been previously reported. Methods: We investigated the UGT2B17

CNV by PCR in 453 Austrian lung cancer patients and in 449 healthy donors and analyzed the impact on

lung cancer susceptibility and outcome. Results: Copy numbers of UGT2B17 were 44.4% (+/+), 42.2% (+/�)

and 13.5% (�/�) in lung cancer patients and 43.0% (+/+), 46.3% (+/�) and 10.7% (�/�) among healthy

donors. The null genotype was not significantly more frequent among women with adenocarcinoma

compared to healthy women (p = 0.59). There was no association with overall survival (p = 0.622) and no

significant sex-associated (p = 0.423) or histology-related impact on development of lung cancer.

Conclusion: UGT2B17 deletion genotype was not associated with a significant risk for lung cancer

development or outcome in our Central European patient cohort. Our study indicates that UGT2B17 is

not a crucial factor in lung carcinogenesis among Caucasians and shows the importance of investigating

such markers in large cohorts from different populations.

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Cancer EpidemiologyThe International Journal of Cancer Epidemiology, Detection, and Prevention

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1. Introduction

Lung cancer is a leading cause of death in Western countries,smoking being the strongest established risk factor for thismalignancy. Still, fewer than 20% of smokers develop lung cancer,indicating that there may be unknown genetic componentsresponsible for this variable susceptibility [1]. Genetically inducedvariations in metabolic processes involved in detoxification oftobacco carcinogens might play an important role in the etiologyof lung cancer. UDP-glucuronosyltransferases (UGT) have beenreported to detoxify carcinogenic tobacco substances, likebenzo[a]pyrene (B[a]P) and 4-(methylnitrosamino)-1-(3-pyri-dyl)-1-butanol (NNAL) [2–4].

One important enzyme for glucuronidation of NNAL is Uridinediphospho glucuronosyltansferase 2B17 (UGT2B17) which alsoplays a major role in steroid hormone metabolism. The UGT2B17gene shows a copy number variation (CNV) and a deletion of bothcopies is associated with a reduced glucuronidation not only ofendogenous substrates but also NNAL [5–7]. In line with this

* Corresponding author. Tel.: +43 1 40400 73781; fax: +43 1 40400 73595.

E-mail address: katrina.vanura@meduniwien.ac.at (K. Vanura).

1877-7821/$ – see front matter � 2013 Elsevier Ltd. All rights reserved.

http://dx.doi.org/10.1016/j.canep.2013.06.004

genotype dependent functionality, an association between the nullgenotype and an increased risk of lung adenocarcinoma has beenreported in North American women although not in men [8]. Forprostate cancer risk, such a functional link, in particular the role ofthe null genotype, has not been proven conclusively [9–14]. TheUGT2B17 null genotype was not associated with an increased riskfor chronic lymphocytic leukemia (CLL) for either sex [15], it was,however, associated with a decreased risk for colorectal cancer inmen but not in women [16], and was shown to be a risk factor forbreast cancer [17].

To more precisely define the potential risk for lung cancer inpersons carrying the null genotype, in particular among women,we analyzed UGT2B17 copy number variation in 453 Austrianpatients diagnosed with lung cancer and 449 Austrian Caucasianhealthy controls.

2. Patients and methods

2.1. Study population

This retrospective study was carried out at the MedicalUniversity of Vienna, Austria. Cases were newly diagnosed,histologically confirmed lung cancer patients (N = 453). A short

Table 1Distribution of UGT2B17 genotype by sex and type of lung cancer. Median age at

diagnosis/recruitment was 65 and 66 years for lung cancer patients and healthy

donors, respectively. N, number of patients; NSCLC, non-small cell lung cancer;

(+/+), homozygous presence of UGT2B17; (+/�), heterozygous presence of

UGT2B17; (�/�), homozygous absence of UGT2B17.

All N cases (%) Male N cases (%) Female N cases (%)

Healthy donors (N = 449)

(+/+) 193 (43.0) 133 (43.3) 60 (42.3)

(+/�) 208 (46.3) 140 (45.6) 68 (47.9)

(�/�) 48 (10.7)a 34 (11.1) 14 (9.9)c

Lung cancer patients

All (N = 453)

(+/+) 201 (44.4) 143 (44.4) 58 (44.3)

(+/�) 191 (42.2) 133 (41.3) 58 (44.3)

(�/�) 61 (13.5)a 46 (14.3)b 15 (11.5)b

Adenocarcinoma (N = 222)

(+/+) + (+/�) 191 (86.0) 118 (85.5) 73 (86.9)

(�/�) 31 (14.0) 20 (14.5) 11 (13.1)c

Squamous cell carcinoma (N = 106)

(+/+) + (+/�) 92 (86.8) 77 (85.6) 15 (93.8)

(�/�) 14 (13.2) 13 (14.4) 1 (6.3)

Small cell lung cancer (N = 82)

(+/+) + (+/�) 72 (87.8) 51 (87.9) 21 (87.5)

(�/�) 10 (12.2) 7 (12.9) 3 (12.5)

NSCLC undiff. (N = 29)

(+/+) + (+/�) 25 (86.2) 20 (83.3) 5 (100.0)

(�/�) 4 (13.8) 4 (14.5) 0 (0.0)

Regular smoker � 1 year = yes (N = 391)

(+/+) + (+/�) 342 (87.5) 259 (86.3) 83 (91.2)

(�/�) 49 (12.5) 41 (13.7) 8 (8.8)

Regular smoker � 1 year = no (N = 46)

(+/+) + (+/�) 39 (84.8) 11 (84.6) 28 (84.4)

(�/�) 7 (15.2) 2 (15.4) 5 (15.2)

a (�/�) healthy donors vs. (�/�) lung cancer patients p = 0.201.b (�/�) male patients vs. (�/�) female patients p = 0.423.c (�/�) female healthy donors vs. (�/�) female adenocarcinoma patients p = 0.59.

M. Gruber et al. / Cancer Epidemiology 37 (2013) 625–628626

questionnaire about smoking habits was performed. The controlsubjects were unrelated Caucasian men and women (N = 449), whowere participants in a Viennese health care program, 45 years orolder, free of vascular disease and any other severe disease atthe time of sample collection between 1998 and 2000. Writteninformed consent was obtained from all study participantsaccording to the Institutional Review Board approved protocols(Ethical Approval at the Medical University of Vienna, EK-No. 025/2009).

2.2. Genotyping

DNA from healthy controls was isolated from frozen EDTAblood using the MagnaPure DNA isolation System1 (RocheDiagnostics GmbH, Vienna, Austria). DNA from lung cancerpatients was isolated from peripheral blood using QIAmp bloodkit (QIAGEN, Valencia, CA, USA). The UGT2B17 copy numbervariation was analyzed by conventional PCR as describedpreviously [18,19]. This PCR gave consistent and robust bands ofthe expected sizes. Due to a recent evaluation of primers used forUGT2B17 CNV detection which reported questionable specificityfor a number of primers used in various studies [14], PCR productsof a subset of samples were sequenced. All sequences were specificfor the UGT2B17 gene (data not shown).

2.3. Statistical analysis

Unconditional logistic regression models were used to assessthe associations between UGT2B17 genotype and risk for lungcancer. Adjustment variables were gender, smoking status (ever-smokers versus never-smokers), and histologic subtype of lungcancer (adenocarcinoma, squamous cell carcinoma, small cell lungcarcinoma and other subtypes). Calculations were done usingPASW Statistics version 17 (IBM Corporation, Armonk, NY, USA).

3. Results

Of the 453 lung cancer patients, 322 were male and 131 werefemale (71.1 and 28.9%, respectively), median age was 65 years.The controls comprised 307 men and 142 women (68.4 and 31.6%,respectively) with a median age of 66 years (see also Table 1).

The genotype-distribution of UGT2B17 was 44.4% (+/+), 42.2%(+/�) and 13.5% (�/�) in lung cancer patients (Table 1). This did notdiffer significantly from 43.3% (+/+), 46.3% (+/�) and 10.7% (�/�)among healthy donors and was similar to frequencies reported inother studies [5,10,14,19]. Among women with adenocarcinoma,the frequency of the UGT2B17 null genotype was 13.1% comparedto 9.9% in female healthy donors. This trend is in line withpreviously reported data [8], however, the difference was notstatistically significant in our cohort (p = 0.59; Odds Ratio 1.38, 95%CI = 0.6;3.2). Generally, the UGT2B17 (�/�) genotype was notsignificantly more frequent among lung cancer patients (13.5%)compared to healthy subjects (10.7%) (p = 0.201) and among malepatients (14.3%) compared to females (11.5%) (p = 0.423). Wefurthermore did not observe an impact of the UGT2B17 CNV on theonset or outcome of lung cancer in our Austrian Caucasianpopulation (p = 0.289 and p = 0.622, respectively; Fig. 1).

4. Discussion

Compared to the healthy population, we observed a higherfrequency of the UGT2B17 null genotype in lung cancer patients,both in male and female individuals. However, none of thedifferences detected reached statistical significance in our cohortand, in contrast to a previous report [8], did not translate into anincreased risk for lung cancer for either sex.

The frequency of the double deletion varies considerablybetween different populations occurring roughly in 10–15% ofCaucasians but in up to 90% among Asian populations [20]. Buteven within populations considerable differences have beenreported. Thus, ranges of UGT2B17 genotypes were 41–54% and33–64% (+/+), 33–48% and 32–52% (+/�), and 4–19% and 3–27% (�/�) in cancer patients and healthy individuals, respectively, ofCaucasian descent from different geographic regions [8–14,16,21–24]. This high variability of genotype frequencies may partlyexplain the ambiguous data regarding the risks for individuals todevelop prostate cancer [9–15], although Cai and colleaguesdemonstrated in a recent meta-analysis a consistent and highprostate cancer risk for men carrying the null genotype whencompared to men-based controls but not in studies usingpopulation as controls [25].

Considering the heterogeneity of genotype frequencies, onelimitation of our study is sample size. It may also explain thedifferences in patients of lung cancer observed by Gallagher et al.[8] and by us. This points to the necessity of conducting UGT2B17genotype analyses with regard to cancer risk in much largercohorts, not only from Europe and North America but most likelyalso from other geographic regions.

However, absence or presence of one or both copies of a genedoes not necessarily involve functional consequences. In particu-lar, UGT2B17 gene expression shows a high interindividualvariability [26] which could modulate the impact of genotype.Although UGT2B17 glucuronidation activities show a gene dosageeffect in most reports [6–8,23,27] and glucuronidation levels in

Fig. 1. Survival curves in relation to UGT2B17 copy number variation (CNV). (A) Cumulative hazard of developing lung cancer. (B) Overall survival among lung cancer patients.

(+/+), Homozygous presence of UGT2B17; (+/�), heterozygous presence of UGT2B17; and (�/�), homozygous absence of UGT2B17.

M. Gruber et al. / Cancer Epidemiology 37 (2013) 625–628 627

women are much lower compared to men [8,27], Grant andcolleagues reported that there was no link between glucuronida-tion levels and genotype in a cohort of African American men [24].Also, while people of Japanese origin have a high prevalence of theUGT2B17 null genotype, they even showed a significantly reducedlung cancer risk with cigarette consumption equal to Caucasians[28], suggesting that UGT2B17 CNV by itself is no pivotal risk factorfor lung cancer. Other members of the UGT family of glucur-onidating enzymes have been shown to significantly contribute toglucuronidation of NNAL, some of which also show geneticpolymorphisms that may influence rates of glucuronidation andsubsequently NNAL detoxification [3,7,29]. In addition, manyvariations in other important tobacco carcinogen detoxifyingenzymes, tumor promoters or tumor suppressor genes have beenreported, which might play a role [30]. Such variations could alsobe in linkage disequilibrium with the UGT2B17 deletion.

With current treatment strategies, we did not observe adifference in overall survival between various UGT2B17 CNVgroups in our cohort receiving heterogeneous therapy. We cannotexclude that UGT2B17 mediated drug metabolism could result indifferent treatment responses to specific drugs or regimens.

In conclusion, our data suggest that the UGT2B17 genotypealone is unlikely to have a crucial impact on lung cancer risk amongCentral European Caucasians. As for other genetic variations, theremight be a different effect on lung cancer risk in other populations.Considering the high variability of genotype distribution and thedifferences in functional levels within and between populations,further studies are warranted to more precisely define sex- andpopulation-specific UGT2B17 gene regulation and function incancer.

Conflicts of interest statement

The authors have no conflicts of interest to disclose.

Acknowledgements

This study was financially supported by a grant of the InitiativeKrebsforschung of the Medical University of Vienna and the

Austrian Federal Ministry of Science and Research through theGen-AU-Child project (GZ 200.136/1 – VI/1/2005) to KV. MGwas awarded a TRHT fellowship of the European HematologyAssociation in 2010.

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