Familial Cancer DOI 10.1007/s10689-014-9748-x
SHORT COMMUNICATION
Prevalence of the BLM nonsense mutation, p.Q548X, in ovarian cancer patients from Central and Eastern Europe Natalia Bogdanova • Alexandr V. Togo • Magdalena Ratajska • Wojtek Kluz´niak • Zalina Takhirova • Theresa Tarp • Darya Prokofyeva • Marina Bermisheva • Grigoriy A. Yanus • Tatiana V. Gorodnova • Anna P. Sokolenko • Alina Kuz´niacka • Amira Podolak • Maciej Stukan • Dominika Wokołorczyk • Jacek Gronwald • Danuta Vasilevska • Vilius Rudaitis • Ingo B. Runnebaum • Matthias Du¨rst • Tjoung-Won Park-Simon • Peter Hillemanns • Natalia Antonenkova • Elza Khusnutdinova Janusz Limon • Jan Lubinski • Cezary Cybulski • Evgeny Imyanitov • Thilo Do¨rk
•
Ó Springer Science+Business Media Dordrecht 2014
Abstract A nonsense mutation, p.Q548X, in the BLM gene has recently been associated with an increased risk for breast cancer. In the present work, we investigated the prevalence of this Slavic founder mutation in 2,561 ovarian cancer cases from Russia, Belarus, Poland, Lithuania or Germany and compared its frequency with 6,205 ethnically matched healthy female controls. The p.Q548X allele was present in nine ovarian cancer patients of Slavic ancestry (0.5 %; including one case with concurrent BRCA1 mutation). The mutation was not significantly more frequent in cases than in controls (Mantel–Haenszel OR 1.14, 95 % CI 0.49; 2.67). Ovarian tumours in p.Q548X carriers were mainly of the serous subtype, and there was little evidence for an early age at diagnosis or pronounced family history
of cancer. These findings indicate that the BLM p.Q548X mutation is not a strong risk factor for ovarian cancer.
N. Bogdanova Z. Takhirova T. Tarp D. Prokofyeva M. Bermisheva T.-W. Park-Simon P. Hillemanns T. Do¨rk (&) Department of Obstetrics and Gynaecology, Gynaecology Research Unit, Hannover Medical School, Hannover, Germany e-mail:
[email protected] M. Ratajska A. Kuz´niacka A. Podolak J. Limon Department of Biology and Genetics, Medical University of Gdansk, Gdan´sk, Poland
N. Bogdanova Clinics of Radiation Oncology, Hannover Medical School, Hannover, Germany N. Bogdanova N. Antonenkova N.N. Alexandrov Research Institute of Oncology and Medical Radiology, Minsk, Belarus A. V. Togo G. A. Yanus T. V. Gorodnova A. P. Sokolenko E. Imyanitov N.N. Petrov Institute of Oncology, St.-Petersburg, Russia A. V. Togo G. A. Yanus A. P. Sokolenko E. Imyanitov St.-Petersburg Pediatric Medical University, St.-Petersburg, Russia
Keywords Genetic susceptibility DNA double-strand break repair Chromosome breakage syndrome Ovarian carcinoma Bloom’s syndrome Founder effect
Introduction High-penetrance mutations in BRCA1 and BRCA2 genes are detected in about 5–25 % of all ovarian cancer patients [1–3]. Part of the remaining cases could be explained by mutations in other genes. The BLM gene encodes a helicase that associates with other repair proteins in a BRCA1-
W. Kluz´niak D. Wokołorczyk J. Gronwald J. Lubinski C. Cybulski International Hereditary Cancer Center, Department of Genetics and Pathology, Pomeranian Medical University, Szczecin, Poland Z. Takhirova D. Prokofyeva M. Bermisheva E. Khusnutdinova Institute of Biochemistry and Genetics, Ufa Scientific Center, Ufa, Russia D. Prokofyeva E. Khusnutdinova Department of Biology, Bashkir State University, Ufa, Russia M. Stukan Gdynia Oncology Centre, Department of Gynecologic Oncology, Gdynia, Poland
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N. Bogdanova et al. Table 1 Characteristics of seven ovarian cancer case–control series employed in this analysis Study name
Acronym
Country
Study type
Cases
Hannover-Jena Ovarian Cancer Study
HJO
Germany
Hospital based/ case–control
355
Hannover-Minsk Ovarian Cancer Study
HMO
Belarus
Hospital-based/ case–control
Hannover-Ufa Ovarian Cancer Study
HUO
Russia
Gdansk Ovarian Cancer Study
GDO
Gdansk-Vilnius Ovarian Cancer Study
Ctrls
Europeans (%)
Invasive cases (% serous)
Reference
531
99.5
350 (68)
13
201
1,205
99.0
201 (79)
3
Hospital-based/ case–control
253
1,069
100
252 (80)
11
Poland
Hospital based/ case–control
347
372
100
340 (76)
10
GVO
Lithuania
Hospital-based/ case–control
100
209
100
98 (80)
14
Szczecin Ovarian Cancer Study
SOC
Poland
Hospital-based/ case–control
578
1,726
100
578 (74)
12
St.-Petersburg Ovarian Cancer Study
SPO
Russia
Hospital based/ case–control
727
1,093
100
727 (89)
2
Study type, number of cases and controls, ethnicity and the proportion of serous histology is provided
associated genome surveillance complex [4, 5]. Its biallelic inactivation results in Bloom’s syndrome, a developmental disorder associated with genomic instability and high propensity towards cancer [6], while the presence of a germ-line mutation in the heterozygous state appears to confer an increased risk of breast carcinoma [7–9]. A nonsense mutation, p.Q548X, is recurrent in Slavic populations and has been associated with an about fivefold increased risk of breast cancer [7, 9]. It is presently unknown whether BLM mutations predispose to ovarian carcinoma. We were thus interested to determine the prevalence of the p.Q548X mutation in unselected ovarian cancer patients from Central and Eastern Europe.
Patients and methods
Szczecin Ovarian Cancer Study and the Gdansk Ovarian Cancer Study from Poland, the Hannover-Minsk Ovarian Cancer Study from Belarus, the Gdansk-Vilnius Ovarian Cancer Study from Lithuania, and the Hannover-Jena Ovarian Cancer Study from Germany. The characteristics of these series are listed in Table 1, and some of them have been described in part previously [2, 3, 10–14]. Former studies included the analyses of BRCA1 and BRCA2 founder mutations as well as specific mutations in other breast cancer susceptibility genes such as CHEK2, ATM, NBN and PALB2. Identified carriers were not excluded in the present BLM mutational screening since evidence indicated that digenic heterozygosity may play a role in some breast cancer patients [9, 15]. All persons gave their informed consent prior to their inclusion in the study, and the study has been approved by the appropriate ethics committees at the participating institutions.
Patients We investigated patients from seven large case–control series, the St.-Petersburg Ovarian Cancer Study and the Hannover-Ufa Ovarian Cancer Study from Russia, the
D. Vasilevska V. Rudaitis Department of Gynecology, Hospital Santarisˇkiu˛ Clinics, Vilnius University, Vilnius, Lithuania I. B. Runnebaum M. Du¨rst Department of Gynecology, Jena University Hospital – Friedrich Schiller University Jena, Jena, Germany E. Imyanitov I.I. Mechnikov North-Western Medical University, St.-Petersburg, Russia
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Methods Genomic DNA was isolated from peripheral white blood cells by routine phenol–chloroform extraction. High resolution melting analysis of PCR amplicons from the BLM exon 7 that harbours the p.Q548X mutation, was performed as previously described [7, 9]. Cases and controls from Szczecin were genotyped using a real-time PCR-based TaqMan exonuclease assay as previously described [16]. Positive and negative controls were included in each assay, and mutation carriers detected in the screening were confirmed by direct sequencing using BigDye chemistry on a Genetic Analyser 3100 Avant (Applied Biosystems/Life Technologies).
Ovarian cancer patients from Central and Eastern Europe
Statistical analyses For a combined evaluation of the seven case–control association studies, stratified analyses were run and Mantel–Haenszel Odds Ratios and p values were calculated using EpiCalc 2000 (www.brixtonhealth.com/epicalc. html). A p value \0.05 was considered to be significant. Power calculations were performed using the PS Power and Sample Size Software (http://biostat.mc.vanderbilt.edu/ twiki/bin/view/Main/PowerSampleSize).
Results and discussion We combined seven case–control studies from Central and Eastern Europe to determine the frequency of the p.Q548X mutation in exon 7 of BLM among ovarian cancer patients and cancer-free female controls. The study comprised genomic DNA samples of ovarian cancer patients from Russia, Belarus, Poland, Lithuania and Germany (Table 1). The results of this mutational screening are summarised in Table 2 Frequency distribution of BLM*p.Q548X in ovarian cancer cases and controls from seven Central and Eastern European study centers
p values given for a two-sided analysis; bold face, combined Mantel–Haenszel odds ratio and p value a
This group included patients who are not native Russians (such as Bashkirs and Tatars) as well as patients with uncertain ancestry
Study center
Table 2. Heterozygosity for p.Q548X was confirmed in 4/980 patients from Russia, 1/201 patients from Belarus, 4/925 patients from Poland, but was not identified among 100 patients from Lithuania or 355 patients from Germany. Clinical features of the nine mutation carriers are summarised in Table 3. Most of the tumours were of the common serous subtype, and there was no particular tendency towards an early diagnosis or pronounced family history for breast or ovarian cancer, although one of the patients from St. Petersburg had been diagnosed with a breast carcinoma 8 years prior the diagnosis of ovarian cancer. The single BLM mutation carrier among the Byelorussian ovarian cancer cases had a first degree family history of ovarian cancer and also carried a BRCA1 mutation, c.4037delA (formerly known as 4153delA). This is a common BRCA1 mutation in ovarian cancer patients from Belarus [3], so the co-inheritance may be a chance finding. But the observation is in line with results from breast cancer studies that BLM truncations can coincide with BRCA1 mutations [7, 9, 15, 17]. The BLM mutation was further investigated for LOH in one serous ovarian
Prevalence of BLM*p.Q548X carriers
OR (95 % CI)
p
Cases
Controls
St. Petersburg
4/727 (0.6 %)
2/1,093 (0.2 %) [7]
3.02 (0.55; 16.52)
0.36
Minsk
1/201 (0.5 %)
2/1,205 (0.2 %) [9]
3.01 (0.27; 33.32)
0.91
Ufa, Russians
0/82
0/602 [9]
–
–
Ufa, non-Russiansa
0/171
0/467 [9]
–
–
Szczecin
1/578 (0.2 %)
10/1,726 (0.6 %)
0.30 (0.04; 2.33)
0.38
Gdansk
3/347 (0.9 %)
2/372 (0.5 %)
1.61 (0.27; 9.71)
0.94
Vilnius
0/100
1/209 (0.5 %)
0.00
0.71
Hannover/Jena
0/355
0/531
–
–
Slavic
9/1,935 (0.5 %)
16/4,998 (0.3 %)
1.14 (0.49; 2.67)
0.94
All combined
9/2,561 (0.4 %)
17/6,205 (0.3 %)
Table 3 Characteristics of BLM*p.Q548X carriers among ovarian cancer patients, listed by study center Patient ID
Ovarian cancer subtype
Age at diagnosis (years)
Tumor characteristics
Family history
St. Petersburg 1
Serous
32
G1, T3, M0, FIGO IIIC
None
St. Petersburg 2
Clear cell
40
T3c, M0, FIGO IIIC
None
St. Petersburg 3
Serous
74
G3, T2b, N1, M0, FIGO IIIC
None
St. Petersburg 4
Mucinous
55
G1, T3, M0, FIGO IIIC
Prior breast cancer in the patient
Belarus 1
Serous
55
n.d.
Ovarian cancer in patient’s mothera
Szczecin 1
Endometrioid
48
G3, T3a, FIGO IIIA
None
Gdansk 1
Serous
61
G3, FIGO IIIC
n.d.
Gdansk 2
Mucinous, borderline
73
n.d.
None
Gdansk 3
Serous
76
G2, FIGO IIIC, no LOH at BLM locus
None
n.d. not documented, LOH loss of heterozygosity a
Patient also carries BRCA1 mutation
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N. Bogdanova et al.
carcinoma from a Gdansk patient, and the analysis indicated that the tumour retained heterozygosity at this locus (Table 3). By comparison, the p.Q548X allele had been found in 2/1,205 healthy females from Belarus (0.2 %) [9], in 12/2,098 (0.6 %) of cancer-free women from Poland, in 1/209 (0.5 %) healthy females from Lithuania, and in 2/1,093 Russian female controls from St.-Petersburg (0.2 %) [7], while it was not detected in another 1,069 female controls, including 602 Russians, from Bashkortostan, nor was it observed in 531 healthy German females. Overall, the mutation frequencies in cases and controls were not significantly different, with a Mantel–Haenszel Odds Ratio 1.14 (95 % CI 0.49; 2.67, p = 0.94, analysis stratified by study and population according to Table 2). From these results and the lack of association, it is unlikely that BLM*p.Q548X confers a more than threefold increase in risk for ovarian cancer. With the present sample size, our study had about 86 % power to detect risks above 3, but just 42 % and 14 % power to detect 2- and 1.5-fold increases in risk, respectively. In summary, this collaborative case–control study did not reveal a significant role for p.Q548X as an ovarian cancer susceptibility allele, although minor risks could not be excluded and a modifying effect remains a possibility. The prevalence of this mutation in the general population appears to have its peak at about 1 in 200 in Poland, with a somewhat lower frequency of 1 in 500 in Belarus and Western Russia, and a very low occurrence in Germany or in the Volga-Ural region. Further studies will be required to refine the spectrum of malignancies associated with this and other mutations in the BLM gene. Acknowledgments We thank the patients for their participation and the many clinicians at hospitals in Belarus, Germany, Poland and Russia for their support of this work. NB was supported by an intramural Hannelore-Munke stipend at Hannover Medical School. The Hannover laboratory was furthermore supported by the Rudolf Bartling Foundation. The St.-Petersburg study and the Ufa study are supported by the Russian Foundation for Basic Research (Grants 12-04-00928, 12-04-00535, 12-04-97026, and 14-04-97088). The Gdansk study was supported by the Polish National Science Centre (Grant 2011/02/A/NZ2/00017). Conflict of interest
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