MDM2 Promoter SNP344T.A (rs1196333) Status Does Not Affect Cancer Risk Stian Knappskog 1,2 *, Liv B. Gansmo 1,2 , Pa ˚l Romundstad 3 , Merete Bjørnslett 4,5 , Jone Trovik 6,7 , Jan Sommerfelt-Pettersen 8 , Erik Løkkevik 9 , for the Norwegian Breast Cancer Group trial NBCG VI " , Rob A. E. M. Tollenaar 10 , Caroline Seynaeve 11 , Peter Devilee 12,13 , Helga B. Salvesen 6,7 , Anne Dørum 5,14 , Kristian Hveem 3 , Lars Vatten 3 , Per E. Lønning 1,2 1 Section of Oncology, Institute of Medicine, University of Bergen, Bergen, Norway, 2 Department of Oncology, Haukeland University Hospital, Bergen, Norway, 3 Department of Public Health, Faculty of Medicine, Norwegian University of Science and Technology, Trondheim, Norway, 4 Department of Medical Genetics, Oslo University Hospital Radiumhospitalet, Oslo, Norway, 5 Faculty Division, The Norwegian Radium Hospital, University of Oslo, Oslo, Norway, 6 Department of Obstetrics and Gynecology, Haukeland University Hospital, Bergen, Norway, 7 Institute of Clinical Medicine, University of Bergen, Norway, 8 Director Naval Medicine, Royal Norwegian Navy, Bergen, Norway, 9 Division of Surgery and Cancer Medicine, Department of Oncology, Oslo University Hospital, Oslo, Norway, 10 Department of Surgery, Leiden University Medical Center, Leiden, The Netherlands, 11 Department of Medical Oncology, Family Cancer Clinic, Erasmus MC-Daniel den Hoed Cancer Center Rotterdam, Rotterdam, The Netherlands, 12 Department of Human Genetics, Leiden University Medical Center, Leiden, The Netherlands, 13 Department of Pathology, Leiden University Medical Center, Leiden, The Netherlands, 14 Department of Gynecological Oncology, Oslo University Hospital Radiumhospitalet, Oslo, Norway Abstract The MDM2 proto-oncogene plays a key role in central cellular processes like growth control and apoptosis, and the gene locus is frequently amplified in sarcomas. Two polymorphisms located in the MDM2 promoter P2 have been shown to affect cancer risk. One of these polymorphisms (SNP309T.G; rs2279744) facilitates Sp1 transcription factor binding to the promoter and is associated with increased cancer risk. In contrast, SNP285G.C (rs117039649), located 24 bp upstream of rs2279744, and in complete linkage disequilibrium with the SNP309G allele, reduces Sp1 recruitment and lowers cancer risk. Thus, fine tuning of MDM2 expression has proven to be of significant importance with respect to tumorigenesis. We assessed the potential functional effects of a third MDM2 promoter P2 polymorphism (SNP344T.A; rs1196333) located on the SNP309T allele. While in silico analyses indicated SNP344A to modulate TFAP2A, SPIB and AP1 transcription factor binding, we found no effect of SNP344 status on MDM2 expression levels. Assessing the frequency of SNP344A in healthy Caucasians (n = 2,954) and patients suffering from ovarian (n = 1,927), breast (n = 1,271), endometrial (n = 895) or prostatic cancer (n = 641), we detected no significant difference in the distribution of this polymorphism between any of these cancer forms and healthy controls (6.1% in healthy controls, and 4.9%, 5.0%, 5.4% and 7.2% in the cancer groups, respectively). In conclusion, our findings provide no evidence indicating that SNP344A may affect MDM2 transcription or cancer risk. Citation: Knappskog S, Gansmo LB, Romundstad P, Bjørnslett M, Trovik J, et al. (2012) MDM2 Promoter SNP344T.A (rs1196333) Status Does Not Affect Cancer Risk. PLoS ONE 7(4): e36263. doi:10.1371/journal.pone.0036263 Editor: Klaus Roemer, University of Saarland, Germany Received February 20, 2012; Accepted April 4, 2012; Published April 30, 2012 Copyright: ß 2012 Knappskog et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: The study was supported by grants from the Norwegian Cancer Society and the Norwegian Health Region West. The funders had no role in the study design, data collection and analysis, decision to publish or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: stian.knappskog@med.uib.no " The clinical investigators of the Norwegian Breast Cancer Group trial NBCG VI are listed in the Acknowledgments. Introduction The Mouse Double Minute 2 homolog (MDM2) is a key regulator of p53 as well as retinoblastoma protein function [1,2,3]. Thus, elevated MDM2 protein levels due to MDM2 gene amplification or other mechanisms have been regarded as an alternative to TP53 mutations diminishing p53 function in many human cancers [2,4,5,6,7]. In 2004 the group of A. Levine discovered a polymorphism SNP309T.G (rs2279744) in the MDM2 intronic P2 promoter [8]. SNP309G enhances MDM2 expression levels by increasing Sp1 transcription factor binding and was subsequently shown to be associated with increased risk and an early age at diagnosis of several malignancies [8,9,10]. While subsequent studies have confirmed an association between SNP309G and the risk of multiple cancer forms, the effect of this SNP seems to differ between ethnic groups: Thus, while most studies performed in Asian or Ashkenazi Jewish populations reports the SNP309G variant to enhance cancer risk many studies conducted in Caucasian populations have failed to reproduce a similar effect [11,12]. Recently, we reported a second polymorphism, SNP285G.C (rs117039649), located 24 base-pairs from SNP309 in the MDM2 P2 promoter. The SNP285C variant allele is observed among Caucasians only, in whom it forms a distinct SNP285C/309G haplotype accounting for about 12% of the SNP309G alleles [13]. SNP285C antagonizes the effect of SNP309G by reducing Sp1 transcription factor binding strength to the MDM2 promoter and is associated with a reduced risk for breast, ovarian and endometrial cancers [13,14]. Taken together, the data from the studies on SNP309 and SNP285 strongly indicate fine tuning of MDM2 P2 promoter PLoS ONE | www.plosone.org 1 April 2012 | Volume 7 | Issue 4 | e36263